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Revista Romn de Medicin Dentar

Vol. XVI nr. 1/2013

Diagnosticul virusologic al VHC


n cavitatea bucal
prin metode minim invazive
(neinvazive)
The Oral Viral Diagnosis of HCV Infection
by Minimally Invasive (Non-Invasive)
Methods
Comnescu Cristian* Anton Gabriela**
CUPRINS

1.
2.
3.
4.

Introducere
Epidemilogia VHC
Istoria natural a virusului hepatitic C
Biologia moleculara a VHC
4.1. Proteinele structurale ale VHC
4.2. Proteinele non structurale
4.3. Genotipurile VHC
5. Ciclul viral al virusului hepatitic C
6. Micro-ARN ca markeri ai Infeciei cu VHC
7. Decelarea VHC n fluide ale cavitii bucale
8. Patogenia infeciei cu VHC
*As. Dr. UMF Carol Davila, Bucureti
**Cercet. 1 Dr. Instit.Virusologie tefan S. Nicolau

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TABLE OF CONTENTS

1. Introduction
2. HCV Epidemiology
3. Natural History of Hepatitis C Virus
4. Molecular Biology of HCV

4.1. HCV Structural Proteins

4.2. Nonstructural proteins

4.3. HCV Genotypes
5. HCV Lifecycle
6. MicroRNAs as HCV Infection Markers
7. HCV Detection in Mouth Fluids
8. HCV Infection Pathogenesis

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9. Noiuni de anatomia anului gingival


9.1. Parodoniul marginal structur
9.2. Componentele principale ale
parodoniului marginal
10. Gingia apect histologic
11. Aspectul clinic al gingiei sntoase
12. anul gingival
13. Lichidul anului gingival
14. Inflamaii gingivale
15. Material i metode
15.1. Acord informat i chestionar
15.2. Prelevare
16. Bibliografie
INTRODUCERE
Hepatita cu virus C se constituie ca o epidemie
globala, estimndu-se c peste 170 de milioane de
persoane prezint boala cronic cu acest virus.1 Aceast
infecie este decelat la circa de 5 ori mai multe persoane
dect cele infectate cu HIV. Datele epidemiologice
incomplete fac ca numai n Statele Unite numrul
persoanelor potenial infectate cu acest virus s fie de
dou ori mai mare dect cel estimat.2
n rile dezvoltate a fost instituit msura de testare a
sngelui, ceea ce a micorat considerabil riscul infectarii
post transfuzionale. Cazurile noi ns care apar se pot
explica prin faptul c infectarea predominant este
datorat consumului de droguri intravenoase i mai puin
altor ci de expunere.
Majoritatea cazurilor cu infecie VHC conduc la
cronicizare, acesta patologie devenind o indicaie major
a transplantului de ficat.
Chiar i dup doua decade de la descoperirea sa, VHC
(virusul hepatitic C) continu s reprezinte o problem
de sntate public, WHO estimnd c minim 3% din
populaia globului prezint infecie cronic cu acest
virus.3, 4
n Romnia, infecia cu VHC reprezint o problem
major, prevalena sa n populaia general nefiind bine

9. Gingival Sulcus - Anatomy Notions


9.1. Periodontium Structure

9.2. Main Components of the Periodontium

10. Gingiva - Histological Aspect


11. Clinical Aspect of healthy Gingiva
12. Gingival Sulcus
13. Gingival Crevicular Fluid
14. Gingival Inflammation
15. Material and Methods

15.1. Informed Consent and Questionnaire

15.2.Sampling

16. Bibliography
INTRODUCTION
Hepatitis C virus has become a global epidemic,
causing chronic infection in almost over 170 million
people worldwide. 1 This means about 5 times as
many people as are infected with HIV. Incomplete
epidemiologic data make that only in the United States
the number of people who might be infected with this
virus is twice higher that the estimated number.2
In the developed countries, blood screening was
introduced, which significantly decreased the risk of
post-transfusion infection. However, the new occurring
cases may be explained by that infection is mainly spread
by intravenous drug use and less frequently via other
ways of exposure.
In most cases, HCV infection leads to chronic
hepatitis, which is a major indication for liver transplant.
Even after two decades since its discovery, HCV
(hepatitis C virus) continues to be a public health
concern, and WHO estimates that a minimum of 3%
of the worlds population is chronically infected with
this virus.3, 4
In Romania, HCV infection is a major cause of
concern and its prevalence in the general population

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precizat. Un studiu realizat n 2001 n Transilvania


meniona o prevalen de 6.28%5 n timp ce un alt studiu
realizat ntre 2006-2008 arta c prevalen anti-VHC
n populaia din ara noastr este de 3,23%. Cele mai
mici rate au fost nregistrate n Transilvania i Banat
(2.63%) iar cele mai mari n Moldova (4,25%) i n
Tulcea ( 7,19%).6
Dei VHC este detectat i intit de mecanismele
imune ale gazdei, prin dezvoltarea unor strategii
virusul stabilete i menine infecii persistente. Cea
mai important strategie de eludare a sistemului imun
const n abilitatea virusului de a se adapta prin
modificarea secvenei nucleotidice a genomului viral,
fapt care permite supravieuirea virusului prin selectarea
variantelor rezistente la presiunea drogurilor antivirale.
Modificarea secvenei nucleotidice se realizeaz prin
mutaii sau quasi-specii.7 Infeciile pesistente cu VHC
conduc adesea la ciroz i carcinom hepatocelular. Nu
exist nc un vaccin profilactic, cauza constituind-o
n pricipal rata mare a mutaiilor ntlnite la genomul
acestui virus.
Studiile moleculare asupra VHC au inceput n 1989 i
au continuat cu clonarea genomului viral. Experimentele
de clonare i secveniere au permis extinderea studiilor n
domeniu, studii mpiedicate mult timp de imposibilitatea
propagrii virusului n culturi celulare.8 De altfel,
dezvoltarea recent a unor sisteme robuste de cultivare
a virusului deschide oportuniti n descifrarea
mecanismelor moleculare virale i identificarea unor
noi inte terapeutice.9
Virusul hepatitic C este ncadrat n familia
Flaviviridae i este transmis prin expunere parenterala.
VHC are genom ARN monocatenar (cu catena pozitiv).
Pe baza secvenei nucleotidice a ARN viral s-au descris
6 genotipuri distincte ale acestui virus, cu numeroase
subtipuri n fiecare genotip. Lista genotipurilor
identificate n cazuri cu hepatita C cronic include
genotipurile 1(1a, 1b); 2 (2a, 2b, 2c i 2d); 3 (3a, 3b,
3c, 3d, 3e i 3f); 4 (4a, 4b, 4c, 4d, 4e, 4f, 4g, 4h, 4i i
4j); 5a i 6a. Prevalena acestor genotipuri este corelat
cu anumite arii geografice.10 Genotipul 1 este cel mai

16

is largely unknown. A study performed in 2001 in


Transylvania evidenced a 6.28%5 prevalence rate while
another study conducted between 2006 and 2008 showed
a 3.23% prevalence rate of anti-HCV in our countrys
population. The lowest HCV prevalence (2.63%) was
found in Transylvania and Banat and the highest rates
in Moldavia (4.25%) and Tulcea (7.19%).6
Although HCV is detected and targeted by host
immune mechanisms, by evolving multiple strategies,
the virus and establishes and maintains a persistent
infection. The most important strategy to evade the
immune system is its ability to adapt by changing the
nucleotide sequence of the viral genome, which allows
the virus to survive by selecting for the variants resistant
to antiviral drugs. The change in the nucleotide sequence
takes place by mutations or quasispecies.7 Persistent
HCV infection often leads to cirrhosis and hepatocellular
carcinoma. At present, there is no preventive vaccine,
the cause being mainly the high rate of mutations seen
in the genome of this virus.
Molecular studies of HCV began in 1989 and
continued with the cloning of the viral genome. Cloning
and sequencing enables extended studies in the field,
which were hampered for many years by the inability to
propagate the virus in cell cultures.8 Besides, the recent
development of robust HCV cell culture systems opens
opportunities to decoding the viral molecular mechanism
and identifying new therapeutic targets.9
Hepatitis C virus is classified within Flaviviridae
and is transmitted throughparenteral exposures. HCV
contains (positive) single-strand RNA genome. Based
on the nucleotide sequence of viral RNA, six distinct
genotypes have been identified, with multiple subtypes in
each genotype class. The list of the genotypes identified
in cases with chronic hepatitis C includes genotypes
1(1a, 1b); 2 (2a, 2b, 2c and 2d); 3 (3a, 3b, 3c, 3d, 3e
and 3f); 4 (4a, 4b, 4c, 4d, 4e, 4f, 4g, 4h, 4i and 4j); 5a
and 6a. The prevalence of these genotypes is correlated
with certain geographical areas.10 Genotype 1 is the

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frecvent ntlnit n SUA dar el este i cel mai dificil


de tratat. Genotiparea viral are importana pentru
instituirea terapiei, stiut fiind ca indivizii cu genotipurile
2 i 3 rspund de circa 3 ori mai bine la terapia cu
interferon alfa comparativ cu cei infectai cu genotipul 1.
inta infeciei virale o reprezint hepatocitul (dar
virusul nu este nc recunoscut a avea efect citopatic).
Afectarea parenhimului hepatic este consecina
rspunsului imun al gazdei cu dezvoltarea ulterioar
a infiltratului inflamator. n general, un pacient este
infectat cu un singur genotip dar poate prezenta o
multitudine quasi-specii. Abilitatea acestor quasispecii de a suferi rapid mutaii cu repercusiuni asupra
imunitii la tratamentul administrat, explic dificultaile
de tratament ale hepatitei cronice cu virus C.
n prezent, datorit gradului mare de variabilitate
genomic determinat de mutaii, nu exist vaccin un anti
VHC i nicio terapie viral specific. Tratamentul uzual
este o terapie prelungit cu interferon alfa n asociere cu
ribavirin, a crui eficien depinde i de genotipul viral.
Mecanismele exacte de ptrundere a virusului n
celulele hepatice pe calea receptorilor, rmn nc
neelucidate, VHC avnd de asemenea capacitatea de
transmitere direct ntre celule.
VHC are un ciclu de via foarte complex, care
nefiind n totalitate elucidat face ca terapia infeciei cu
virusul hepatitei C s fie nc dificil de efectuat i noi
tratamente greu de gsit.
Genomul virusului hepatitei C (genom viral ARNss,
avnd poaritate pozitiv) servete n mod direct ca ARN
mesager pentru sinteza unui precursor poliproteic.
Acesta va fi clivat ulterior n proteine virale. Strategia
acesta aparine unui supergrup de virusuri ARNss care
include att enterovirusurile umane (virusul hepatitei
A), ct i o serie de patogeni cu importana n medicina
veterinar (pestivirusuri: virusul holerei la porci, virusul
diareei la bovine).37
Virusul hepatitei C nu este considerat citopatic n
ciuda faptului c infecteaz direct hepatocitele, deoarece
afectarea parenchimal este o consecin a rspunsului
imun al gazdei i a infiltratului inflamator.11

most common type of hepatitis C genotype in the United


States and the most difficult to treat. Viral genotyping
is important to determine the therapy, since individuals
with genotypes 2 and 3 are almost 3 times more likely
than individuals with genotype 1 to respond to therapy
with alpha interferon.
The virus targets hepatocytes (but the virus is not yet
known to have cytopathic effect). Hepatic parenchyma
damage is the hosts immune response with the
subsequent development of the inflammatory infiltrate.
Generally, patients are only infected with one genotype,
but they may have a multitude of quasi-species. These
quasi-species have the ability to mutate very quickly and
become immune to current treatments, which explains
why chronic hepatitis C is so difficult to treat.
At present, there is neither an anti-HCV vaccine
nor a specific antiviral therapy, due to the high level
of genomic variability caused by mutation. The usual
treatment is a long-acting interferon-alpha, given in
combination with ribavirin, the efficiency of which
depends on the viral genotype.
How exactly the virus enters liver cells through
receptors is still unknown, as HCV is also capable
ofdirect cell-to-cell transmission.
The nature of the HCV lifecycle is very complex,
which as it is not fully explained makes treatment
of Hepatitis C virus infection still difficult and new
treatments hard to find.
The hepatitis C virus genome (Positive-sense ssRNA)
serves as direct RNA messenger in the synthesis of
a polyproteic precursor. This will be subsequently
split into viral proteins. This strategy belongs to a
subclass of ssRNA viruses which includes both human
enteroviruses (hepatitis A virus) and a series of pathogens
of importance in veterinary medicine (pestiviruses: hog
cholera virus, bovine viral diarrhea).37
The hepatitis C virus directly infects the hepatocyte
yet it is considered to be noncytopathic, as the hepatic
parenchymal damage occurs as a consequence of the
hosts immune response and inflammatory infiltrate.11

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EPIDEMILOGIA VHC
Cele mai recente rapoarte WHO estimez prevalena
infeciei HCV la 3% din populaia globului, prin
complicaiile produse fiind cauza principal a
recomandrilor de transplant hepatic.12 CDC estimeaz
ca aproape 4 milioane de americani au fost expui
virusului hepatitic C, peste jumtate dintre aceti
prezentnd infecii cronice. Mai mult, se raporteaz
anual circa 26000 de noi cazuri iar 12000 de persoane
decedeaz ca urmare a cirozei sau carcinomului
hepatocelular. 13 Dei VHC este endemic la nivel
global, distribuia sa nregistreaz o mare variabilitate
geografic, rile africane i asiatice avnd prevalena cea
mai mare. Comparativ, n tarile vest i nord- europene, n
America de Nord i Australia se raporteaz o prevalen
sczut.14, 15 n afara diferenelor geografice exist i
diferene temporale n patternurile de infecie cu VHC.
Spre exemplu, dei n SUA, Australia, Spania prevalena
infeciei este similar, (1.0%-1.9%), n SUA prevalena
cea mai mare a persoanelor infectate se nregistreaz
ntre 30-49 ani (circa 2/3 dintre cei infectai) ceea ce
indic faptul c transmiterea a aprut n ultimii 20-40
de ani la tineri.16
Factorii de risc care au fost asociai infeciei cu VHC
sunt transfuziile cu snge netestat, utilizarea drogurilor
injectabile sau a unor proceduri asociate actului medical
(chirurgicale). n rile n curs de dezvoltare, n care
seroprevalena crescut este corelat cu vrsta, se
consider ca actul medical necorespunztor (transfuzii,
injecii) a fost factorul major de transmitere a VHC17
i de meninere a unor arii hiperendemice izolate.18
Reutilizarea seringilor de sticl n cursul campaniei de
tratare a schisostomiasei n Egipt pare a fi responsabil
de transmiterea iatrogenic pe scara larg a acestui
patogen19. Comportamentul sexual este uneori asociat
cu cresterea riscului de infecie dar fr existena unor
dovezi clare.20 Transmiterea sexual pare a fi mai puin
incriminat, dar nu se poate preciza cauza: fie ncrcarea
virala n fluidele i esutul genital este mic fie celulele
tractului genital nu constituie o int viral.21 Cu toate
acestea, exist date care incrimineaz practicile sexuale
ca mijloc de transmitere a infeciei, procentele varind
ntre 2 i 22%.22 Asociaia American pentru Studiul

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HCV EPIDEMILOGY
The most recent WHO reports estimate a 3%
prevalence rate of HCV infection in the worlds
population, the complications occurred being the leading
cause of recommendations of liver transplantation.12
CDC estimates that almost 4 million Americans have
been exposed to the Hepatitis C virus, and half of them
are chronically infected. Furthermore, almost 26000
new cases are reported each year and 12000 people
die from cirrhosis or cirrhosis and hepatocellular
carcinoma.13 Although HCV is endemic worldwide,
there is a large degree of geographic variability in its
distribution, the African and Asian countries having the
highest prevalence rates. Comparatively, the Western
and North-European countries, the North-American
countries and Australia report a low prevalence.14, 15
There are both geographic and temporal differencesin
the patterns of HCV infection. For example, although in
the United States, Australia, Spain similar prevalence of
HCV infection (1.0%-1.9%) are reported, in the Unites
States prevalence is highest among persons 30-49 years
old (almost two-thirds of those infected), which indicates
that transmission occurred during the last 20-40 years
among young adults.16
The risk factors related to HCV infection are
unscreened blood transfusions, injectable drug use or
medical (surgical) associated procedures. In developing
countries, where high seroprevalenceis age-dependant,
unsafe medical procedures (transfusions, injections)
were a major risk factor for HCV transmission17 and
persistence in isolated hyperendemic areas.18 In Egypt,
glass syringes reused in the schistosomiasis treatment
campaign seem to be responsible for the large-scale
iatrogenic transmission of this pathogen.19 Sexual
behavior is sometimes associated to the high risk of
infection but without having any clear evidence.20
Sexual transmission seems to be less incriminated,
but no cause can be identified: either the viral load in
fluids and genital tissue is low or the genital duct cells
are not targeted by the virus.21 However, there is data
incriminating sexual practices as means of transmission
of the infection, with rates varying between 2 and
22%.22 The American Association for the Study of Liver

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Afeciunilor Hepatice, AASLD, afirma c n cuplurile


monogame nu este necesar modificarea obiceiurilor
sexuale, n sensul protejrii, cu scopul prevenirii
transmiterii de la un partener la altul, deoarece riscul
de infecie ar fi mic.
Transmiterea materno-fetal este de asemenea
raportat dar cel mai des este asociat coinfeciei cu
HIV.23 Se pare c virusul exist n saliva persoanelor
infectate dar datele epidemiologice arat c, contactul cu
acest fluid nu constituie un mod eficient de transmitere.24
n schimb, transmiterea nosocomial prin instrumentar
medical (colonoscop, dializ, instrumentar chirurgical)
este bine documentat.25, 26 n unele ri, interveniile
stomatologice constituie un factor major de risc pentru
transmiterea VHC. Contaminarea echipamentului
stomatologic s-a dovedit a fi cauza transmiterii virusului
n 17,94% din cazuri ntr-un studiu realizat n Pakistan.
Acelai studiu raporteaz c prevalena anti-HCV i a
infeciei active ntr-un grup de subieci supui chirurgiei
stomatologice era de 14,28%, cauza constituind-o
instrumentarul sterilizat necorespunztor.27
n pofida mijloacelor de sterilizare, contaminarea
viral a instrumentarului stomatologic poate constitui
nc o surs de infectare. Un studiu efectuat n China
arat c dup sterilizarea cu glutaraldehid 2%, se reduce
masiv contaminarea n instrumentarul stomatologic dar
nu total.28
Un studiu efectuat pe 286 pacieni cu hepatit cronic
cu virus C spitalizai n Iai ntre anii 2004 i 2007 a
urmrit investigarea factorilor de risc poteniali pentru
infectarea cu virusul hepatitei C n regiunea de nordest a Romniei. Pacienii inclui n studiu au declarat
existena n antecedente a unor factori de risc poteniali n
35,66% din cazuri, cei mai frecveni fiind: antecedentele
transfuzionale (32%), interveniile stomatologice
(25,5%) i procedurile invazive cu scop diagnostic sau
terapeutic (20%). neparea accidental a fost la raportat
la 5,6% din cazuri. Pacienii peste 40 ani au prezentat
o rat mai mare de raportare a factorilor de risc fa de
cei tineri (p=0,038).29
Factori de risc pentru hepatita C: (aa cum sunt
prezentai de de Douglas L et al sunt:
infectarea mamei nainte de a nate
transfuzii nainte de 1992

20

Diseases, AASLD, states that monogamous couples


do not have to change their sexual habits, in terms
of protection, in order to prevent partner-to-partner
transmission, as the infection risk would be low.
Maternal-fetal transmission is also reported but
is most frequently related to HIV co-infection. 23
It seems that the virus is present in the saliva of
HCV-infected patients, however epidemiologic data
show transmission efficiency by contact with this
type is low. 24 In exchange, nosocomial transmission
by surgical instruments (colonoscope, dialysis,
surgical instruments) is well documented. 25, 26
Dental interventions are a major risk factor for HCV
transmission in several countries. Contaminated
dental equipment appears to be the factor for
HCV transmission in 17.94% of cases in a study
conducted in Pakistan. The same study reports a
14.28% prevalence of anti-HCV and active infection
in a dental surgery group of students as a result of
improperly sterilized instruments.
Regardless the means for sterilization, viral
contamination of dental instruments may still represent
a source for infection. A study conducted in China
shows that after sterilization with 2% glutaraldehyde,
instrument contamination has been significantly reduced,
however not completely.28
A study conducted on 286 patients with chronic
hepatitis C hospitalized in Iasi between 2004 and 2007
aimed to investigate the potential risk factors for HCV
infection in the north-eastern part of Romania. The
patients included in the study reported potential risk
factors in the past in 35.66% of cases, and the most
frequent were: transfusions (32%), dental procedures
(25.5%) and invasive diagnostic or treatment procedures
(20%). Accidental needlestick injury was reported in
5.6% of cases. Patients older than 40 years had a
significant higher rate of declared risk factors compared
with younger patients (p=0.038).29
The risk factors for hepatitis C: (as presented by
Douglas L et al are:
Birth to a mother infected with positive antibodies
for HCV
Blood transfusions before 1992
Hemodialysis

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hemodializa
tatuaje efectuate
utilizarea de droguri
multiplii parteneri sexuali
nepturi sau expuneri mucozale (ca factor de risc
pentru personalul medical)
transplant de organe efectuat inainte de 1992
sex cu persoane infectate cu VHC
tratament pentru hemofilie efectuat nainte de 1987
n unele cazuri ns, factorii de risc nu au putut fi
identificai.
Dup cum afirma Douglas L. Senecal, PA-C; Joseph
Morelli n articolul Hepatitis C virus infection: A current
review , OCTOBER 2007 20(10) www.jaapa.com
JAAPA, istoria natural a virusului ghideaz informarea
pacientului i decizia unei terapii. Se arat c procentul
pacienilor infectai cu VHC cu vindecare spontan este
cuprins ntre 10 i 20%, n timp ce cronicizarea este de
80 pn la 90%; infectarea cronica conduce la ciroz
(pe parcrsul unei perioade ce 20-30 de ani) n cazul a
20-30% dintre cazurile clinice.
Faptul c ncrcarea viral poate s varieze
considerabil pe parcursul diferitelor momente ale zilei,
precum i faptul c nu indic evoluia hepatitei, fac ca
acest marker viral s nu fie eficient n monitorizarea
infeciei.
Evoluia pacienilor cu ciroza datorat infeciei VHC
este sever, n sensul c riscul de decompensare este de
aproape 4% pe an; acesteia i se asociaz sngerari ale
varicelor, encefalopatie hepatic, disfuncii pulmonare
sau renale, icter, hidroperitoneu. Se estimeaz c,
din fericire, numai un procent de 25% din numrul
pacienilor cu ciroz hepatic se decompenseaz
ajungnd s necesite transplant.
O corelaie ntre viremie i inflamaia lobular a
fost realizat folosind testul bDNA pentru msurarea
cantitii de ARN VHC30.

History of tattoos
History of cocaine use
Multiple sexual partners
Needlestick or mucosal exposure (in a health care
worker)
Organ transplant before 1992
Sex with an HCV-infected patient
Treatment for hemophilia before 1987
however, in some cases, the risk factors remained
unidentified.
According to Douglas L. Senecal, PA-C; Joseph
Morelli in article Hepatitis C virus infection: A current
review, OCTOBER 2007 20(10) www.jaapa.com
JAAPA, the natural history of HCV infection guides
patient education and the decision of whether a patient
should undergo therapy. It shows that in patients infected
with HCV, 10% to 20% will have spontaneous clearance
and chronic infection will develop in the other 80% to
90%; the chronic infection will lead to cirrhosis(over
a period of 20 to 30 years), in 20% to 30% of patients.
The fact that it can significantly fluctuate throughout
the course of a day and that it is not an indication
of progression to hepatitis makes this viral marker
inefficient in infection monitoring.
The history of patients with HCV-induced cirrhosis
is alarming in that there is a 4% risk per year that
decompensation will occur; this is accompanied
by bleeding from varices, hepatic encephalopathy,
pulmonary or renal dysfunction, jaundice, ascites.
Fortunately, only 20% of all patients with cirrhosis will
ever decompensate and require transplantation.
A correlation between viremia and lobular
inflammation was made using the bDNA test to measure
the HCV RNA load. 30

ISTORIA NATURAL A VIRUSULUI HEPA


TITIC

NATURAL HISTORY OF HEPATITIS C VIRUS

Dintre pacienii infectai cu VHC, n 10-20% din


cazuri apare clearance-ul, iar n 80-90% dintre cazuri
apar infecii cronice. Circa 20-30% dintre infeciile
cronice progreseaz in 20-30 de ani spre ciroz.31

Among patients infected with HCV, 10% to 20% will


have clearance and 80% to 90% will develop chronic
infection. The chronic infection will progress to cirrhosis
over a period of 20 to 30 years, in approximately 20%
to 30% of patients. The progression of fibrosis is more

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Progresia fibrozei este mai rapid n cazul subiecilor


n vrsta la momentul infectrii, al celor cu steatoz
hepatic, alcoolicilor sau al celor coinfectai cu HIV.
Istoria natural a pacientilor cu ciroza indus de VHC a
artat c odat boala instalat, riscul de decompensare
i declanare a cancerului hepatic creste anual cu 3.9%.
BIOLOGIA MOLECULAR A VHC
VHC este un virus ARN nrudit cu virusul hepatitei
G umane, cu virusul febrei galbene i virusul Dengue.
inta natural a VHC o constitue hepatocitul i, posibil,
limfocitul B. Replicarea viral este rapid, estimndu-se
c mai mult de 10 trilioane de virioni se produc zilnic,
chiar i n infecia cronic. Replicarea se realizeaz
de ctre o ARN polimereza ARN dependent creia
i lipsete funcia de proofreading (citirea corecta a
matriei) fapt care determin apariia unor qvasispecii
diverse dar nrudite, la aceeai persoan infectat;
aceste qvasispecii reprezint o provocare major pentru
rspunsul imun al gazdei.32
Genomu l v i r a l ( A R N m o n o c a t e n a r + ) d e
aproximativ 9.6kb este localizat n miezul (30-35 nm)
anvelopat (Fig. 1). Genomul viral nu intr n nucleul
celulei gazda iar replicarea are loc n citoplasma
hepatocitelor.

rapid in patients who are older at the time of infection,


have hepatic steatosis, consume alcohol or infected with
HIV. The natural history of patients with HCV-induced
cirrhosis showed that once this condition diagnosed, the
risk is as high as 3.9% per year that decompensation and
liver cancer will occur.
MOLECULAR BIOLOGY OF HCV
HCV is a RNA virus related to the hepatitis G virus,
dengue virus and yellow fever virus. The natural targets
of HCV are hepatocytes and, possibly, B lymphocytes.
Viral replication is rapid, more than 10 trillion virion
particles being estimated to be produced each day, even
during the chronic phase of infection. Replication is
made by a RNA dependant RNA polymerase lacking
the proofreading ability, which leads to the occurrence
of different yet related quasispecies in the same infected
patient; these quasispecies are a major challenge for the
hosts immune response.32
The single-strand viral RNA (positive-sense), which
is approx. 9.6 kb, is encapsulated in a (30-35 nm) inner
core (Fig. 1). The viral genome does not enter the cell
nucleus and replication occurs in the cytoplasm of
hepatocytes.

FIG 1. Structura VHC: proteina core (miez) interacioneaz cu genomul ARN viral pentru a forma nucleocpsid. Doua glicoproteine asociate anvelopei (E1 i E2) sunt nvelite de o anvelop lipidic derivat de la celula gazd ( Sharma, 2010).
FIG 1. HCV structure: The core protein interacts with viral genomic RNA to form the nucleocapsid.
Two membrane-associated envelope glycoproteins (E1 and E2) are embedded in a lipid envelope
which is derived from the host (Sharma, 2010).

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Organizarea genomului HCV este prezentat n Fig.


2. Genomul RNA const dintr-o regiune 5 netranslatat
(UTR) care conine un situs pentru ribozomi (IRES),
o regiune genomic ce codific genele structurale i
nestructurale i regiunea 3 UTR. HCV este translatat
ca o poliprotein procesat proteolitic de proteazele
gazdei i de cele virale. Proteinele structurale (C- core;
E1- proteina 1 a anvelopei; E2- proteina anvelopei) sunt
localizate n captul amino-terminal n timp ce proteinele
nestructurale (NS2, NS3, NS4A, NS4B, NS5A i NS5B)
sunt localizate n extremitatea carboxi-terminal.
Proteina p7 (ion channel sau viroporin) este localizat la
jonciunea proteinelor structurale cu cele nestructurale.33

HCV genome organization is presented in Fig. 2.


The viral RNA consists of a 5 untranslated region
(UTR) containing the internal ribosome entry site
(IRES), followed by the genomic region for structural
and nonstructural genes and the 3 UTR. HCV is
translated as a polyprotein that is proteolytically
processed by host and viral proteases. The structural
proteins (C- core; E1- envelope protein 1; E2envelope protein 1) are located at the amino-terminal
end while the nonstructural proteins (NS2, NS3,
NS4A, NS4B, NS5A, and NS5B) are located at the
carboxyl-terminal end. The p7 protein (ion channel
or viroporin) is located at the junction of structural
and nonstructural proteins.33

FIG 2. Reprezentarea schematica a genomului VHC, a proteinelor structurale i nestructurale.


FIG 2. Schematic representation of HCV genome, structural and nonstructural proteins

Genomul viral are un singur cadru de citire


(ORF- open reading frame) flancat de segmentele
5 i 3 netranslatate (NTRs). Structurile ARN au un
rol important n replicarea viral iar regiunile nalt
conservate 5- i 3-NTRs conin elemente de control al
translaiei poliproteinelor virale i al replicrii. 5 UTR
(+) alctuit din circa 341 nucleotide conine IRES cu o
structur pliat sub forma a 4 motive stem-loop ( I, II,
III i IV). IRES este necesar pentru translaia ARN viral
realizat de ribozomii celulei gazd.34 Proteina core a
VHC interacioneaz cu 5-NTR (+), regiune care nu
conine semnalele de mpachetare a ARN. 3-UTR (+)

24

The viral genome contains a single ORF (open reading


frame) which is flanked by 5 and 3 non translated
segments (NTRs). RNA structures play a major role in
the multiplication of viruses and the 5- and 3-NTRs,
which are highly conserved, contain control elements,
for translation of the viral polyprotein and replication.
The 5 UTR (+) is cca. 341 nucleotides in length and
contains an IRES with a structure folded into 4 stemloop motifs (I, II, III and IV). The IRES is required for
the translation of viral RNA, which is carried out by
host cellribosome.34 HCV core protein interacts with
the 5-NTR (+), which does not contain RNA packaging

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lung de circa 200 nucleotide este implicat n replicarea


ARN. Aceast regiune conine 3 domenii distincte : (i) o
secven pol- (U/UC) de circa 80 nucleotide; (ii) o zon
variabil; (iii) o regiune invariabil de 98 nucleotide
cu 3 structuri stem-loops. Studii recente arat c aceste
domenii pliate din catenele negative sunt recunoscute de
polimeraza viral ca situsuri de iniiere pentru sinteza
catenei pozitive a genomului VHC.72
Interaciile ARN-ARN par a interveni n modularea
ciclului viral. n interiorul captului 3 al proteinei
nestructurale a fost descris un element cis-acting (CRE
sau SL9266 sau 5BSL3.2) a crui disrupere blocheaz
replicarea ARN.35
Moleculele necodificatoare microRNAs (miR)
sunt i ele implicate n controlul replicarii virale. n
mod particular, miR-122 este specific ficatului unde
se exprim la nivele ridicate. Recent, s-a raportat c
miR-122 se leag la 5-UTR al HCV ceea ce conduce la
reducerea translaiei virusului. Aceste studii au generat
interes, miR 122 putnd reprezenta o potenial int
terapeutic.36
Proteinele structurale VHC sunt hidrofobe ceea ce
faciliteaz legarea beta lipoproteinelor din plasma la
anvelopa virusului. Se formeaz astfel un fel de ecran,
care mascheaz accesul anticorpilor neutralizani,
contribuind astfel la persistena viral.
PROTEINELE STRUCTURALE ALE VHC

VHC codific o singur poliprotein (NH2-C-E1E2-p7-NS2-NS3-NS4A-NS4B-NS5A-NS5B-COOH)


de aproximativ 3010 aminoacizi. Proteinele structurale
(core, E1 i E2) ca i p7 proteine sunt eliberate din
poliproteina prin clivare de ctre peptidaza reticului
endoplasmatic al celulei gazd. Proteinele nestructurale
(NS2, NS3, NS4A, NS4B, NS5A i NS5B) sunt
clivate de proteazele virle NS2- 3 i NS3-4A. n urma
proteolizei celulare i virale sunt eliberate cel puin 10
proteine virale mature.
Proteina bazic core, multifuncional, formeaz
component structural a particulei virale i este
implicat n apariia steatozei hepatice i a cancerului

signals. The 3-UTR (+) is around 200 nucleotides and


is involved in RNA replication. Three different domains
can be recognized in this UTR: (i) a poly (U/UC) tract
with an average length of 80 nucleotides; (ii) a variable
region, and (iii) a virtually invariant 98 nucleotides
made up of 3 stem-loops. Recent studies have showed
that these folded domains from the negative strands are
recognized by the viral polymerase as the initiation site
for plus-strand synthesis of the HCV genome.72
RNA-RNA interactions appear to intervene in the
modulation of the virus lifecycle. Within the 3-end of
the non-structural protein, a cis-acting element (CRE or
SL9266 or 5BSL3.2) was discovered and its disruption
blocks RNA replication.35
Non-coding microRNA molecules (miR) are also
important in the control of viral replication. In particular,
miR-122 is specifically expressed and is found to be
abundant in the human liver. A recent discovery has
showed that miR-122 binds to the 5-UTR of HCV,
which reduces HCV translation. These studies have
generated a lot of interest in the miR-122 role as potential
therapeutic target.36
HCV structural proteins are hydrophobic in nature,
which facilitates binding of beta-lipoproteins from
plasma to the virus envelop. Thus, a king of screen is
formed, which hides the access of neutralizing antibodies
and contributes to viral persistence.
HCV STRUCTURAL PROTEINS

HCV encodes a single polyprotein (NH2-C-E1E2-p7-NS2-NS3-NS4A-NS4B-NS5A-NS5B-COOH)


which is approximately 3010 amino acids. The structural
proteins (core, E1 and E2) and the p7 protein are released
from the polyprotein after cleavage by host endoplasmic
reticulum signal peptidase. The non structural proteins
(NS2, NS3, NS4A, NS4B, NS5A, andNS5B) are cleaved
by viral proteases NS2- 3 and NS3-4A. After cell and
viral proteases, at least 10 mature viral proteins are
released.
The multifunctional core protein, which is highly
basic in nature, forms the structural component of the
virus particle and is implicated in the development of

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hepatic. Ea deriv n urma procesrii poliproteinei virale


de ctre proteazele celulare ale reticulului endoplasmatic
(RE) gazda.37 Cercetri recente au artat c aceast
protein se poate autoasambla, n membranele reticului
endoplasmatic, n particule VHC-like (HCVLPs).38 O
regiune a acestei proteine (cuprins ntre aminocizii
112 i 152) este esenial pentru asocierea cu RE i cu
membranele mitocondriale participnd astfel la reglarea
Ca2+ i a apoptozei. Mai mult chiar, prin interacia
cu proteinele chaperon ale mitocondriei (prohibitin),
proteina core crete stresul oxidativ. Una dintre funciile
proteinei core const n recrutarea proteinelor non
structurale la lipidele asociate membranelor, lipide
utilizate n replicarea viral.
Glicoproteinele de anvelopa E1 i E2 sunt
componente structurale care alctuiesc proieciile
exterioare ale particulei virale. Aceste proteine sunt
supuse unor modificri post-translaionale la captul
N-terminal n timpul translatrii n RE. E1 i E2
au rol important n intrarea virusului n celula prin
intermediul receptorilor, E2 avnd afinitate pentru
CD81, o tetraspanin exprimat n diferite tipuri de
celule, inclusiv hepatocitele i limfocitele B.39 CD81,
receptorul scavenger SR-BI, claudin-1 (CLDN) i
occludin (OCLN) sunt cei mai importani receptori
care mediaz intrarea virusului n celul; se pare ca
VHC utilizeaz i glicozaminoglicanii sau receptorii
de mic densitate de pe suprafaa hepatocitului c
factori de iniiere a atarii virusului la celula gazd.40
Lipoproteinele HDL joac i ele un rol activ n
intrarea HCV n celul. Un domeniu hipervariabil din
vecintatea regiunii amino-terminale a E2 (HVR-1)
este inta n neutralizarea anticorpilor.
PROTEINELE NON STRUCTURALE

n aceast clas intr proteinele NS2, NS3, NS4A,


NS4B, NS5A i NS5B, importante pentru replicarea
genomului viral. Proteina p7 poate forma canale ionice
necesare produciei de particule virale infecioase,
eveniment n care este implicat i colaborarea dintre
proteinele structurale i cele non structurale. p7
aparine familiei viroporinelor i poate oligomeriza

26

hepatocellular steatosis and liver cancer. It is generated


from a viral polyprotein which is processed by
cellular proteases in the host endoplasmic reticulum
(ER).37 Recent studies showed that this protein can
self-assemble in HCV-like particles (HCVLPs) in
ER-membranes..38 A region of this protein (between
amino acids 112 and 152) is essential for association
with the ER and also with the outer mitochondrial
membrane, and is thus involved with Ca2+ regulation
and apoptosis. Furthermore, this interaction with the
chaperon proteins of mitochondria (prohibitin), results
in an increased oxidative stress. One of the functions of
the core protein is recruitment of non structural proteins
to the lipid-associated membranes, lipids which are
used for viral replication.
Envelope glycoproteins E1 and E2 are structural
components constituting the outer projections of the
virion. These proteins undergo post-translational
modifications at the N-terminal end while being
translated in the ER. Both, E1 and E2 play a major
role in HCV entry in cells through receptors, E2 having
the binding sites for CD81, a tetraspanin expressed
on various cell types including hepatocytes and B
lymphocytes. 39 CD81, scavenger receptor SR-BI,
claudin-1 (CLDN) and occludin (OCLN) are the most
important receptors that mediate HCV entry in cells; it
appears that HCV also uses glycosaminoglycans or low
density receptors on hepatocytes as initial attachment
factors to the host cell.40 HDL lypoproteins also play
an active role in HCV entry in cells. A hypervariable
domain near the amino terminus of E2 (HVR-1) is the
target for neutralizing antibodies.
NON STRUCTURAL PROTEINS

This class covers proteins NS2, NS3, NS4A, NS4B,


NS5A and NS5B, which are important for replication
of the viral genome. The protein, p7, can form ion
channels, required for the production of infectious virus
particles, which involves an interconnection between
structural and the non structural proteins. The p7 protein

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dup incluziunea sa n membrana lipidic, dnd natere


canalelor ionice. Clivarea ntre p7 i NS2 conduce la
formarea unui precursor E2p7NS2 cu rol potenial n
reglarea ciclului celular.41
Proteinele p7 joac rol n producia de particule virale
dar unul neesenial.42
VHC NS2 este o protein transmembranar
hidrofob de 23Kda 43, o cistein proteaz necesar
pentru infectivitate.44 NS2 interacionez cu ea nsi
formnd homodimeri dar i cu alte proteine nestructurale
participnd la asamblarea particulelor virale.45 Dat fiind
rolul su n infectivitate, NS2 este o potenial int
terapeutic.
VHC NS3 este membru al superfamiliei de helicaze
DExH/D-box care funcioneaz ca o serin proteaz,
ARN helicaz i NTP-az. mpreun cu NS4A, NS3
este responsabil de clivarea la nivelul jonciunilor
NS3/4A, NS4A/4B, NS4B/5A i NS5A/5B dar este
implicat i n antagonizarea cilor de rspuns imun
inscut ale gazdei.45
VHC NS4A este un cofactor pentru serinproteaz NS3.
NS4A este localizat att pe reticulul endoplasmatic, ct
i pe mitocondrie, proteina fiind implicata nu numai n
replicarea virusului ct i n patogeneza viral, prin
afectarea funciilor celulare.45
HCV NS4B este o protein hidrofob de 27-KDa
care conine 4 domenii transmembranare. Proteina
conine reziduri de acid palmitoleic prezena acestora
avnd rol important n replicarea viral.46 Trebuie
menionat faptul c variaiile de secven n NS4B n
diferite izolate virale au un rol important n replicarea
VHC n cultur.
HCV NS5A prezint activitate de legare a ARN,
fiind prezent n forme fosforilate i hiperfosforilate.47
HCV NS5B este o ARN polimeraz ARN dependent.
Ea iniiaz sinteza catenei ARN complementare,
negativ, utiliznd catena pozitiv ca matri. Ulterior,
genereaz catene ARN pozitive din catena negativ.48

belongs to the viroporin family and can oligomerize


following its inclusion into a lipid membrane creating
ion channels. The cleavage between p7 and NS2 results
in the formation of an E2p7NS2 precursor, which may
have a role in cell lifecycle regulation.41
The p7 proteins play a role in the production of viral
particles, but which is not essential.42
HCV NS2 is a 23-kDa transmembrane hydrophobic
protein, a cysteine protease required for infectivity.44
NS2 interacts with itself forming homo-dimers but
also other non structural proteins, being involved in the
assembly of viral particles.45 Given its role in infectivity,
NS2 may be a therapeutic target.
HCV NS3 is a member of the superfamily 2 DExH/Dbox helicases, which functions as a serine protease, an
RNA helicase and a NTPase. Along with NS4A, NS3
is responsible for cleavage at the NS3/4A, NS4A/4B,
NS4B/5A and NS5A/5B junctions but it is also involved
in antagonizing host innate-immune pathways.45
HCV NS4A is a cofactor for NS3 serine protease.
NS4A is localized not only on the endoplasmic
reticulum, but also on mitochondria, the protein being
also involved in virus replication and viral pathogenesis
by affecting cellular functions.45
HCV NS4B is a 27-KDa hydrophobic protein which
contains 4 transmembrane domains. The protein contains
palmitoleic acid residues which appear to be essential
for viral replication.46 It should be noted that variation
in the NS4B sequence between different HCV isolates
play a major role in HCV replication in cell culture.
HCV NS5A has RNA-binding activity and is found
in phosphorylated and hyperphosphorylated forms.47
HCV NS5B is a RNA dependent RNA polymerase.
It initiates synthesis of complementary negativestrand RNA using the positive polarity as a template.
Subsequently, it generates positive-strand RNA from
the negative strand.48

GENOTIPURILE VHC

HCV GENOTYPES

Pe baza variaiei nucleotidice, VHC este clasificat n


6 genotipuri majore i n mai mult de 80 de subtipuri.

Based on nucleotide variation, HCV is classified in 6


major genotypes and over 80 subtypes. The hypervariable

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Cea mai mare variabilitate este concentrat n regiunea


hipervariabil a glicoproteinelor E1 i E2 iar cea mai
mic ntre regiunea netranslatat 5 (UTR), care conine
secvene specifice i structuri secundare eseniale
pentru replicare i translaie. Variabilitatea de secven
este determinat de absena activitii de proofreading
(citire corect) a ARN polimerazei ARN dependent.
Rata de ncorporare nlocuire a bazelor corecte este de
10-3 baze azotate/ genom/an. Toate genotipurile VHC
sunt hepatotropice i patogene dar exist diferene de
infectivitate i patogenicitate ntre genotipuri, cu efecte
diferite asupra progresiei spre ciroz i cancer hepatic.
Durata i eficacitatea tratamentului sunt dependente
de genotipul i ncrcarea viral. ntre genotipurile
virale exist variaie de secven de 30-50% iar ntre
subtipuri de 15-30%. Genotipurile 1a i 1b sunt comune
n Europa de vest, genotipul 3 este frecvent n India,
Nepal i Pakistan iar genotipul 4 este comun n Africa.
Genotipul 5 este raportat n Africa de sud iar genotipul
6 n Hong Kong i Asia de sud-est. Genotipul 3a este
cel mai frecvent ntlnit, fiind identificat la 50% dintre
pacienii din rile europene i din Asia (ex. India). n
Romnia, genotipul prevalent este 1b.
CICLUL VIRAL AL VIRUSULUI HEPATITIC C
Etapele ciclului viral sunt reprezentate schematic n
figura 3. Sunt prezentate topologia proteinelor structurale
i nonstructurale virale la nivelul membranei reticului
endoplasmatic (ER). Replicarea ARN VHC este ilustrat
alturi de translaia mediata de IRES, ca i procesarea
poliproteinelor virale.49

region of glycoproteins E1 and E2 is the most variable


part and the 5 untranslated region (UTR), which
contains specific sequences and secondary structures
essential for replication and translation, is the least
variable part. Variability in sequence is a consequence
of the lack of proofreading activity of RNA dependant
RNA polymerase. Correct bases were found to have
an incorporation-replacement rate of 10-3 nucleobases /
genome / year. All HCV genotypes are hepatotropicand
pathogenic, however different genotypes vary in their
infectivity and pathogenicity, with impact on progression
to cirrhosis and liver cancer. The duration and efficiency
of therapy depends on the genotype and viral load. A 30
to 50% variation in sequence was found between viral
genotypes and a 15 to 30% variation between subtypes.
Genotypes 1a and 1b are common in Western Europe,
genotype 3 in frequent in India, Nepal and Pakistan
and genotype 3 is common in Africa. Genotype 5 is
reported in South Africa and genotype 6 in Hong Kong
and South-East Asia. Genotype 3a is the most common
type in Eastern countries and Asia, being identified in
50% of the patients (e.g. India). In Romania, genotype
1b is the most common type.
HCV LIFECYCLE

HCV lifecycle steps are depicted schematically


in figure 3. The topology of HCV structural and
nonstructural proteins at the endoplasmic reticulum
(ER) membrane is shown. HCV RNA replication is
depicted along with the IRES-mediated translationand
viral polyprotein processing.49

MICRO ARN CA MARKERI AI INFECIEI


CU VHC

MicroRNAs AS HCV INFECTION MARKERS

Micro ARN sunt molecule mici necodificatoare


care controleaz translaia i transcripia a numeroase
gene. Numeroase date arat o dereglare a expresiei
acestor molecule n anumite afeciuni, ceea ce indic
potenialul lor terapeutic. Datorit stabilitii lor
precum i prezenei lor n aproape toate fluidele
organismului, miRNA pot deveni o nou clas de
biomarkeri non-invazivi.50

microRNAs are small non-coding RNA molecules,


which regulate translation and transcription of many
genes. Countless data shows a deregulation in the
expression of these molecules in certain diseases,
which is an indication of their therapeutic potential.
The stability of miRNAs and their presence in almost
all body fluids can be useful for developing a new class
of non-invasive biomarkers.50

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Fig. 3. Ciclul de via al VHC (Brass et al, 2006)


Fig. 3. HCV Lifecycle (Brass et al, 2006)

Studii recente au artat c expresia unor micromolecule


ARN (miRNA) precum miR-122, miR-34a, miR-16 i
miR-21 este n mod curent sczut n fibroza hepatic i
carcinomul hepatocelular. n plus, nivelele circulante ale
acestor poteniali markeri par a se corela cu severitatea
histologic a bolii hepatice la pacienii cu infecii
cronice cu VHC.51 miR-122 regleaz cile metabolice
n ficat, inclusiv biosinteza colesterolului52, precum i
replicarea i sinteza virionilor VHC.53 Descreterea
nivelului de expresie a miR122 s-a corelat cu prognoza
sever n tumorile hepatice54 dar, pe de alt parte,
o cretere a nivelului sau a fost raportat n liniile
celulare derivare din cancere hepatocelulare HCV
pozitive55. Spre deosebire de miR122, miR21 este
semnificativ crescut n cancerele hepatocelulare dar
i n cazurile de fibroz hepatic induse de VHC.56
Mai puin asociat cu VHC, miR34 este un mediator al
funciilor proteinei tumorsupresoare p53 i este crescut
n cancerul hepatic.57
Noi date despre prezena miRNa n vezicule eliberate
de celule precum i potentialul lor prognostic, ridic
posibilitatea utilizrii unor fluide precum saliva58, 59 i
urina ca materialele biologice de investigare.60

Recent studies showed that expression of some


RNA molecules (miRNAs), such as miR-122, miR-34a,
miR-16 and miR-21, is commonly deregulated in liver
fibrosis and hepatocellular carcinoma. In addition, the
circulating levels of these potential markers appear to
correlate with the hepatic histological disease severity
in patients with chronic hepatitis C infection.51 miR-122
regulates the metabolic pathways in the liver, including
cholesterol biosynthesis 52, as well as replication
and HCV virion synthesis.53 Reduced expression of
miR122 was correlated with hepatic tumors of poor
prognosis54 but, on the other hand, a high level was also
reported in HCV-derived HCC.55 Unlike miR122, the
level of miR21 is significantly high in hepatocellular
carcinoma but also in HCV-induced liver fibrosis.56
Less associated with HCV, miR34 is a mediator of the
tumor suppressorprotein p53 functions and is high in
liver cancer.57
New data about miRNa presence in cells independent
of vesicles and their potential for diagnosis raises the
possibility of using fluids, such as saliva58, 59 and urine
as biological investigation materials.60

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DECELAREA VHC N FLUIDE ALE CAVITII BUCALE


Identificarea virusului hepatitic C (VHC) n alte
fluide decat snge este important pentru confirmarea
unor posibile rute nonparenterale de transmitere
viral. Totui, rolul fluidelor orale n transmiterea
VHC rmne controversat. ntr-un articol publicat
n 2005, Ferreiro i colab 61 subliniau c studiile
moleculare care confirm prezenta ARN VHC n
saliv aduc argumente pentru transmiterea virusului
pe aceast cale, chiar dac potenialul lor infectant nu
poate fi cu precizie determinat. Un studiu realizat de
Montebugnoli i colab62 a demonstrat c testele EIA
deceleaz prezena anticorpilor anti-HCV n lichidul
crevicular n 80% din cazurile pozitive n plasm dar
nu deceleaz anticorpii specifici n saliv. Studiul
subliniaz c virusul i anticorpii pot ptrunde n
lichidul crevicular i de aici se poate rspndi prin
saliv. Un studiu comparativ al nivelelor anticorpilor
specifici virali n saliv i lichid crevicular la
pacieni hemodializai VHC pozitivi63 a artat ca
seropozitivitatea n cele dou fluide a fost semnificativ
corelat (kappa=0.462; p<0.001). Rezultatele susin
conceptul c lichidul crevicular poate fi o semnificativ
surs de virus n saliv. Suzuki et al64 au determinat
cantitativ ARN VHC n saliv i lichidul crevicular al
pacienilor anti-HCV pozitivi. Autorii consemneaz c
78% dintre pacienii a cror saliva nu prezenta ARN
viral prezentau ncrcare viral n lichidul crevicular.
n plus, 77% dintre subieci prezentau ncrcare virala
mai mare n lichidul crevicular comparativ cu saliva.
Pe de alt parte, datele obinute de Lins i colab65 nu au
identificat nicio corelaie ntre prezenta ARN viral n
saliva i ncrcarea viral n plasm. Decelarea ARN
VHC n saliv ar putea constitui un argument ca acest
fluid biologic constituie o surs de infecie. Decelarea
ARN viral n specimene de saliv de la 32/61 pacieni
a permis o corelaie statistic cu ncrcarea viral
plasmatic (P<0.001), acesta fiind singurul parametru
predictibil.66

30

HCV DETECTION IN MOUTH FLUIDS


The search for hepatitis C virus (HCV) in fluids
other than blood is important for assessing possible
nonparenteral routes of viral transmission. However,
the role of oral fluids in HCV transmission remains
controversial. In an article published in 2005, Ferreiro
et colab 61 pointed out that the molecular studies
which confirmed HCV RNA presence in saliva bring
arguments for virus transmission on this route, even
if their potential for infection may not be accurately
determined. A study carried out by de Montebugnoli et
collab62 demonstrated that EIA tests detect the presence
of anti-HCV antibodies in crevicular fluid in 80% of
HCV-positive subjects in plasma but they do not detect
specific antibodies in saliva. The study shows that
the virus and antibodies can enter the crevicular fluid
and then spread via the saliva. A comparative study
of the levels of specific viral antibodies in saliva and
crevicular fluid in HCV seropositive hemodialysis
patients63 showed that seropositivity in the two fluids
was significantly correlated (kappa=0.462; p<0.001).
The results support the concept that the crevicular
fluid may be a significant source of HCV in saliva.
Suzuki et al64 we quantitatively determined HCV RNA
in saliva and crevicular fluid of anti-HCV-positive
patients. The authors note that 78% of the patients
whose saliva specimens were negative had HCV
RNA in their crevicular fluid. In addition, 77% of the
subjects had higher viral load in the crevicular fluid
than in the saliva. On the other hand, the data obtained
by Lins et collab65 found no correlation between the
viral RNA presence in saliva and the viral load in
plasma. HCV RNA detection in the saliva might be
an argument for considering this biological fluid a
source of infection. HCV RNA detection in saliva
specimens from 32/61 patients allowed for a statistical
correlation with plasma viral load (P<0.001), which
was the only predictable factor.66

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PATOGENIA INFECTIEI CU VHC


n majoritatea cazurilor infecia cu VHC este
considerat ca fiind o infecie auto-limitant. 50%
dintre subiecii infectai evolueaz spre cronicizare.
Se presupune c persistena infeciei virale cu virus
C ar putea fi asociat cu un anume genotip viral sau
cu reinfecia cu mai multe genotipuri diferite VHC.
Mecanismele responsabile de modificrile de la nivel
hepatic (mai ales n infecia cronic) nu sunt cunoscute
n pofida rezultatelor marcante obinute n epidemiologia
VHC i descifrarea biologiei moleculare a virusului.
n general, modificrile hepatice in vivo sunt induse
de infecia viral prin dou mecanisme: mediate
imunologic i citopatogenitate direct (avnd drept int
fie antigenele virale, fie autoantigenele).
NOIUNI DE ANATOMIA ANULUI GINGIVAL
PARODONIUL MARGINAL
STRUCTUR

HCV INFECTION PATHOGENESIS


In most cases, HCV infection is considered to be selflimited. 50% of the infected subjects progress to chronic
infection. HCV infection persistence might be associated
with a specific viral genotype or reinfection with many
different HCV genotypes. The mechanisms responsible
for the changes in the liver (especially chronic infection)
remain yet to be discovered although outstanding results
have been obtained in the HCV epidemiology and HCV
molecular biology.
Generally, in vivo hepatic changes are induced by
HCV infection by two mechanisms: immunologicallymediated and direct cytopathogeneity (targeting either
viral antigens or self-antigens).
GINGIVAL SULCUS - ANATOMY NOTIONS
PERIODONTIUM
STRUCTURE

Totalitatea esuturilor care asigur susinerea


i meninerea dinilor pe arcade poart numele de
parodoniu marginal. Superior, parodoniul marginal
este limitat de marginea gingival, care in mod normal
este situat n vecintatea coletului anatomic al dintelui;
exist nsa situaii n care nu este respectat acest nivel,
cum ar fi hiperpalziile gingivale i retracile gingivale
din numeroase cauze (de cele mai multe ori fiind nsoite
i de retracii ale osului alveolar).
Inferior, apical, limita nu este clar, fiind considerat
c se afl n dreptul proieciei apexului pe mucoas.
Limita ntre parodoniul marginal i cel apical, n cazuri
patologice, se face prin afectarea diferit a acestora,
prin anamneza, prin calea de afectare (endodintic,
desmodontal), precum i prin semnele clinice decelabile
la examinare.

All the tissues ensuring the support of and maintaining


the teeth in position on the arches bear the name of
periodontium. On the upper side, the periodontium is
limited by the gingival margin, which is normally located
in the proximity of the tooth cervix; however, there are
cases when that level is no longer valid, for instance
gingival hyperplasia and gingival retractions for various
reasons (most of the times being accompanied also by
retractions of the alveolar bone).
On the lower side, apically, the limit is not clear; it
is considered to be located next to the projection of the
apex on the mucous membrane. The limit between the
periodontium and the apex, in pathological cases, is given
by their different affectation, by anamnesis, based on the
means of affectation (endodontic, desmodontal), as well
as on the clinical signs detectable upon examination.

COMPONENTELE PRINCIPALE ALE


PARODONIULUI MARGINAL

MAIN COMPONENTS OF THE


PERIODONTIUM

Parodoniul marginal este mprit funcional n 2


componente:

The periodontium is functionally divided in 2


components:

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1. Parodoniul superficial, de nveli. Acesta este


constituit de gingie, cu cele 3 componente:
epiteliul gingival
corionul gingival
ligamentele supraalveolare
2. Parodoniul profund, de susinere; mai poart
denumirea de parodoniu funcional i are n structura sa:
cementul radicular
desmodoniul
osul alveolar
Reprezentarea acestor elemente este schematizat n
figura de mai jos.

GINGIA APECT HISTOLOGIC


esutul din care este format gingia este conjunctiv
(alctuiete corionul), acoperit de epiteliul pavimentos
pluristratificat.
Epiteliul gingival este format din:
epiteliul extern, oral
epiteliul intern, al antului gingival
epiteliul de jonciune (termenul se refer la interfaa
dinte-epiteliu)
Inseria epitelial se refer la celulele din epiteliul
joncional, situate mai apical de fundul antului gingival,
celule care intr n contact direct cu suprafaa dintelui.67
Celulele epiteliale situate mai apical = inseria
epitelial primar
Celulele epiteliale situate mai coronar = inseria
epitelial secundar67

1. The superficial periodontium, covering the teeth. It


is made up of the gingiva (gum), with its 3 components:
the gingival epithelium
the gingival chorion
the supraalveolar ligaments
2. The profound periodontium, with a role of support;
also named functional periodontium; includes in its structure:
the radicular cementum
the desmodontium
the alveolar bone
All those elements are schematically pictured in the
figure below.

GINGIVA - HISTOLOGICAL ASPECT


The tissue forming the gingiva is conjunctive (makes
up the chorion), covered in the stratified squamous
epithelium.
The gingival epithelium is formed of:
the external, oral epithelium
the internal epithelium, of the gingival sulcus
the junctional epithelium (the term refers to the
tooth-epithelium interface)
The epithelial insertion refers to the cells in the
junctional epithelium, located more apically from the
bottom of the gingival sulcus, which cells have direct
contact with the tooth surface.67
The epithelial cells located more apically = primary
epithelial insertion
The epithelial cells located more coronary =
secondary epithelial insertion 67

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Epiteliul oral prezit n majoritate keratinocite, celule


formatoare de keratin. Diferenierea apare din stratul
bazal acolo unde apar anumite filamente n citoplasma
celulelor, precursori ai keratinei; se numesc citokeratine
sau tonofilamente.
Forma tonofilamentelor este variat n funcie de
zona unde se afl: aplatizate n stratul cornos, turtite n
stratul granulos, pligonale n stratul spinos, cuboidale
alungite n stratul bazal.67
Epiteliul oral mai conine i melanocite; aceste celule
au prelungiri dendritice care produc un pigment protector
mpotriva radiaiei actinice i celule Langerhans cu
funcie de aprare fa de agresiunea microbian.67
Alctuirea epiteliului oral:
stratul bazal, geminativ, situat cel mai profund
stratul spinos
stratul granulos
stratul cornos, keratinizat, situat superficial, care
este descuamativ
Epiteliul antului gingival, intern acoper peretele
gingival al antului crevicular, fiind slab keratinizat sau
nekeratinizat. Acest aspect este important n patologia
zonei, atat ca mecanisme de apariie i evluie, ct i ca
tratament specializat.
Epiteliul joncional este singura zon din organism
unde o mucoas moale vulnerabil mecanic se ataeaz
mecanic de un esut dur, ambele fiind situate ntr-un loc
expus unei caviti deschise la mediul nconjurtor.67
Epiteliul de jonciune formeaz un manon pe
ntreaga circumferin a dintelui.

34

The oral epithelium is mostly made of cytokeratins,


keratin-forming cells. The differentiation occurs in the
basal layer, where certain filaments, keratin precursors,
appear in the cell cytoplasm; they are called cytokeratins.
The shape of cytokeratins varies depending on the
area where they are located: flattened in the horny layer,
oblated in the granular layer, polygonal in the spiny layer,
oblong and cubic in the basal layer.67
The oral epithelium also contains melanocytes; those
cells have dendritic extensions producing a protective
pigment against actinic radiations and Langerhans cells
with a defense role against the microbial aggression.67
The structure of the oral epithelium:
the basal layer, germinative, located in the deepest
places
the spiny layer
the granular layer
the horny, keratinized layer, superficially located,
which is squamous
The epithelium of the gingival sulcus covers the
inside of the gingival wall of the crevicular sulcus, being
poorly keratinized or non-keratinized. That aspect is
important for the pathology of the area, both in terms
of occurrence and evolution mechanisms, as well as in
terms of specialized treatment.
The junctional epithelium is the only region of
the body where a soft mucous membrane, which is
mechanically vulnerable, is mechanically attached to
a hard tissue, both being located in a spot which is
exposed to a cavity opening towards the surrounding
environment.67
The junctional epithelium forms a sleeve around the
entire circumference of the tooth.

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Se poate situa n mod fiziologic (figura urmatoare):


pe smal integral
pe smal i cement
pe cement integral

Figura de mai sus reprezint i stadiile erupiei


continue a dinilor. Dintre aceste stadii, n funcie de
variai factori (genetici, de edentaie, traumatici, periaj
incorect i altele), gingia se poate stabiliza la un anumit
nivel sau la altul.
Cea mai sntaos inserie este pe smal, deoarece
acesta este mult mai rezistent decat cementul la atacul
carios, n aceste cazuri fiind mai puin probabile aceste
afectri locale.
Imediat dup nceputul erupiei gingia contiun s
se deplaseze apical pe dinte pn i stabilizeaz nivelul
definitiv, cum este schematizat n figura urmtoare:

Physiologically, it can be located as follows (see the


figure below):
integrally on the enamel
on the enamel and the cement
integrally on the cement

The figure above also features the stages of the


continuous teeth eruption. Among those stages,
depending on various factors (genetic, edentation,
traumatic, incorrect brushing etc.), the gingiva may
stabilize at a certain level or at another.
The healthiest insertion is on the enamel, because it
is much more resistant than the cement to the attack of
the cavities, such cases making those local affectations
less likely.
Immediately after the beginning of the eruption, the
gingiva continues to move apically on the tooth until it
permanently stabilizes its level, as shown schematically
in the figure below:

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ASPECTUL CLINIC AL GINGIEI SNTOASE

CLINICAL ASPECT OF HEALTHY GINGIVA

n mod fiziologic gingia poate avea o gam variat


de culori n funcie de gradul de secreie a melanocitelor
din epiteliul oral. Coloraia difer n funcie de ras, de
condiii climatice. Ea este direct dependent de:
numrul melanoblastilor, care vor forma melanocitele
grosimea stratului epitelial
gradul de keratinizare i vascularizare
Urmtoarele aspecte sunt considerate normale din
punct de vedere al coloraiei:

Physiologically, the gingiva can have a varied range


of colors, depending on the degree of secretion of
melanocytes in the oral epithelium. The coloration differs
depending on the race and on the climatic conditions. It
directly depends on:
the number of melanoblasts which will form the
melanocytes
the thickness of the epithelial layer
the degree of keratinization and vascularization
The following aspects are considered normal from
the point of view of coloration:

Aspectul clinic sntos este dat de coaja de portocal


sau gravura punctat, cu o consisten destul de ferm,
ns uor depresibil.

The clinically healthy aspect is that of orange peel


or of dotted engraving, with a pretty firm consistence,
but slightly depressible.

ANUL GINGIVAL
Spaiul situat ntre gingie i dinte, pn la epiteliul
joncional, cu o adncime normal ntre 1 i 3 mm, avnd
un perete intern (dentar), unul extern (gingival), care se
ntlnesc n zona denumit baza anului, poart numele
de ant gingival.
LICHIDUL ANULUI GINGIVAL
Provine din venulele corionului gingival situat mai
apical de epiteliul sulcular. Este secretat continuu, n
cantiti mici.
Prin studii fcute pe animale s-a evidentiat ca
fluoresceina injectat intravenos apare n lichidul
crevicular dupa numai 3 minute.67
Rolul vizibil al lichidului crevicular este acela de
a ndeprta mecanic resturile mici alimentare sau alte
particule strine ce ar putea ajunge n antul ginival,
contribuind la sntatea local, precum i de a ajuta

36

GINGIVAL SULCUS
The space located between the gingiva and the tooth,
up to the junctional epithelium, with a normal depth
from 1 to 3 mm, having an internal (dental) wall, and an
external (gingival) wall, which meet in the area called
the base of the sulcus, bears the name of gingival sulcus.
GINGIVAL CREVICULAR FLUID
It comes from the venules of the gingival chorion,
located more apically from the sulcular epithelium. It is
continuously secreted, in small quantities.
Studies conducted on animals have shown that
the fluorescein intravenously injected appears in the
crevicular fluid after only 3 minutes.67
The visible role of the crevicular fluid is of
mechanically removing the small food leftovers or other
alien particles which could reach the gingival sulcus,
contributing to the local health, as well as helping to

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la vindecarea eventualelor microleziuni care ar putea


aprea la acest nivel.68
ns pe lang aceasta are rol n activarea
adeziunii inseriei epiteliale, datorit coninutului
de proteine plasmatice, precum i rol n activitatea
antimicrobian prin anticorpi, leucocite i factori
antimicrobieni.67
n anumite condiii se pot observa creteri ale
volumului de lichid crevicular, asa cum sunt ele descrise
de prof. Horia Dumitriu n cartea Parodontologie:
dimineaa
n cursul masticaiei
prin masaj gingival
prin periaj
n sarcin
n urma folosirii contraceptivelor
n cursul inflamaiei gingivale 67, 69
n perioada de vindecare dup tratament chirurgical
n timpul tratamentelor ortodontice70
Aceast ultim afirmaie este susinut i
documentat amplu de ctre Jonas Capelli Jr.; Rivail
Fidel Junior; Carlos Marcelo Figueredo; Ricardo
Palmier Teles n studiul Change in the gingival
fluid volume during maxillary canine retraction 70,
argumentnd faptul c micrile ortodontice, cu liza
i apoziie osoas dirijat prin forele aplicate pe
bracketi, determin n faza iniial o inflamaie care
se exprim i prin modificri de volum ale lichidului
crevicular.
Studiul a fost efectuat pe canini maxilari, la pacieni
cu vrste de 18 ani (mici variaii de cateva luni),
la pacieni fr ali factori care ar fi putut influena
modificri ale lichidului crevicular.
Graficul de mai jos prezint rezultatele obinute,
respectiv variaia volumului lichidului crevicular
cu timpul n funcie de presiune (rou) i tensiune
(negru).
Metoda de prelevare de lichid crevicular este cu
hrtie absorbant, fiind luate probe ata mezial ct i
distal de la acelai dinte la fiecare recoltare, imaginea
urmtoare:

the healing of any micro-lesions that could occur at


that level.68
However, apart from that, it has a role in activating
the adhesion of the epithelial insertion, due to the
content of plasmatic proteins, as well as a role in the
antimicrobial activity through antibodies, leucocytes
and antimicrobial factors.67
In certain conditions, one can notice increases in
the volume of crevicular fluid, as they are described by
Professor Horia Dumitriu in his book, Periodontology:
in the morning
during mastication
through gingival massage
through brushing
during pregnancy
after the use of contraceptives
during gingival inflamation67, 69
during the healing period after a surgical treatment
during orthodontic treatments70
This last statement is widely supported and
documented by Jonas Capelli Jr.; Rivail Fidel
Junior; Carlos Marcelo Figueredo; Ricardo Palmier
Teles in their study Change in the gingival fluid
volume during maxillary canine retraction 70 ,
arguing that orthodontic movements, with the bony
lysis and apposition steered by the forces applied
on the brackets, determines in the initial stage an
inflammation also expressed by changes in the volume
of the crevicular fluid.
The study was done on the maxillary canines, in
patients aged 18 y.o. (small variations of a few months),
who did not show other factors which could have
influenced the changes in the crevicular fluid.
The graph below shows the obtained results,
namely the variation in the volume of crevicular fluid
in time, depending on the pressure (red) and tension
(black).
The crevicular fluid sampling method uses absorbent
paper, samples being taken both medially and distally,
from the same tooth for each sampling, see the following
picture:

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Lichidul crevicular nu este doar un simplu transsudat


(filtrat fiziologic), ci este un infiltrat inflamator. 69
Aceasta implic faptul c apare ca un produs al unor
mecanisme locale de aprare activ.
Compoziia lichidului gingival este urmtoarea:67
aminoacizi
elemente celulare: leucocite PMN, limfocite,
monocite
albumine

38

The crevicular fluid is not just a simple transudate


(physiologically filtered), but is an inflammatory
infiltration.69 That means that it appears as a product of
certain local mechanisms of active defense.
The composition of the gingival fluid is as follows:67
amino-acids
cellular elements: PMN leucocytes, lymphocytes,
monocytes
albumins

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a1, a1, b i g globuline cu funcie de anticorpi:


imunoglobuline de tip IgA, IgG, IgM
Fibrinogen
Fibrinolizin
Fraciuni proteice ale complementului C3, C4
Glucide, de 3-4 ori mai multe ca n serul sanguin,
fiind rezultatul florei microbiene locale i nu al unui
mecanism metabolic de esut
Sistemul lactoperoxidazei cu rol n corectarea
pH-ului
Neutrofile ntregi sau fragmentate dup supran
crcarea cu bacterii i degranulare
Enzime lizozomale
Alte ezime: fosfataza acid i alcalin, bglucuroni
daza, catepsina, proteaze, lacticdehidrogenaza,
precum i lizozim.
Lizozimul este o enzim glucozidazic, care ac
ioneaz prin scindarea legturilor dintre N-acetilglucozamin i acidul acetilmuramic din component
peretelui bacterian.
Electrolit: Na, K, P, Ca
Uneori, atunci cnd se administreaz tratamente
pentru infecti i afeciuni generale, se concentreaz
(cu nivel mai mare dect n serul sanguin) la acest
nivel substane ca: tetraciclina, hidantoina i altele.

a1, a1, b and g globulins with the role of antibodies:


immunoglobulins of the IgA, IgG, IgM type
Fibrinogen
Fibrinolysin
Protean fractions of the C3, C4 complement
Carbohydrates, 3 to 4 times more than in the blood
serum, being the result of the local microbial flora
and not of a tissue metabolic mechanism
The system of lactoperoxidase with a role in correcting the pH
Neutrophils, entire or fragmented after the overload
with bacteria and degranulation
Lysosomal enzymes
Other enzymes: acid and alkaline phosphatase,
b-glucuronidase, cathepsin, proteases, lactic dehydrogenase, as well as the lysozime.
The lysozime is a glucosidase enzyme acting by
splitting the bonds between N-acetyl-glucosamine
and the acetyl-muramic acid in the composition of
the bacterial wall.
Electrolytes: Na, K, P, Ca
Sometimes, when treatments are given for general
infections and affections, at that level, substances
such as: tetracycline, hydantoin and others concentrate (at a higher level than in the blood serum).

Structura spaiului periodontal dup H.P. Mueller, 2005.


The structure of the periodontal space, by H.P. Mueller, 2005.

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OBIECTIVE

OBJECTIVES

Crearea unei baze de material biologic (snge,


saliv lichid crevicular) de la pacieni anti-VHC
pozitivi i negativi
Studiu comparativ asupra performanelor testelor
rapide versus metodele clasice n decelarea infeciei
cu VHC n lichidul crevicular, saliva, snge
Decelarea ARN VHC i determinarea ncrcrii
virale n probele biologice recoltate de la subieci
anti-VHC pozitivi
Rolul unor miRNA asociai VHC n lichidul
crevicular i saliv

Creating a base of biological material (blood, saliva,


crevicular fluid) from patients anti-HVC positive
and negative
Comparative study on the performances of rapid
tests versus classical methods in detecting the HVC
infection in the crevicular fluid, saliva, blood
Detecting HVC RNA and determining the viral load
in the biological samples taken from anti-HVC
positive subjects
The role of certain HVC-associated miRNA in the
crevicular fluid and saliva
STUDY OUTLINE

PREZENTAREA STUDIULUI
DATE CARE SUSIN STUDIUL

1
1. Montebugnoli L, Dolci G, Anti-HCV antibodies
are detectable in the gingival crevicular fluid of HCV
positive subjects, Minerva Stomatol. 2000 Jan-Feb 62
Autorii arat c testele ELISA realizate pe probe
de snge au decelat anticorpii virali specifici n 100%
din cazuri n timp ce anticorpii n saliv nu s-au putut
decela iar n lichidul antului gingival au fost detectai
n aproximativ 80% din cazurile clinice cunoscute
ca infectate; acest fapt se constituie ntr-un indicator
important: VHC ptrunde n cavitatea bucal pe calea
lichidului crevicular i de aici se rspndete mai departe
n saliv i apoi n afara cavitii bucale.
Studiul arat c lichidul crevicular reprezint o
alternativ viabil pentru detectarea anticorpilor anti
VHC. Mai mult dac se folosesc teste rapide ar fi posibil
o testare pe scar larg n cadrul cabinetelor de medicin
dentar.

1
2. Suzuki T, Omata K, Satoh T, Miyasaka T, Arai
C, Maeda M, Matsuno T, Miyamura T Quantitative
detection of hepatitis C virus (HCV) RNA in saliva and

40

DATA SUPPORTING THE STUDY

1
1. Montebugnoli L, Dolci G, Anti-HCV antibodies
are detectable in the gingival crevicular fluid of HCV
positive subjects, Minerva Stomatol. 2000 Jan-Feb 62
The authors have shown that the ELISA tests on
blood samples have detected specific viral antibodies
in 100% of the cases, while antibodies could not be
detected in the saliva, and in the gingival crevicular
fluid antibodies were detected in approximately 80%
of the clinical cases known as infected; that is an
important indicator: HVC enters the mouth in the
crevicular fluid and from there spreads into the saliva
and then outside the mouth via the saliva.
The study has shown that the crevicular fluid is
a valid alternative for the detection of anti HVC
antibodies. Moreover, if rapid tests are used, this
may be a useful routine procedure for use in dental
practices.

1
2. Suzuki T, Omata K, Satoh T, Miyasaka T, Arai
C, Maeda M, Matsuno T, Miyamura T Quantitative
detection of hepatitis C virus (HCV) RNA in saliva and

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gingival crevicular fluid of HCV-infected patients, J Clin


Microbiol. 2005 64

gingival crevicular fluid of HCV-infected patients, J Clin


Microbiol. 2005 64

Studiul VHC n fluidele corporale altele dect snge


este important pentru a determina i alte posibile ci
de transmitere, non parenterale. nc nu s-a stabilit cu
exactitate rolul lichidelor bucale n rspndirea infeciei.
n studiul mai sus menionat s-a efectuat o determinare
cantitativ a ARN ului VHC salivar i crevicular la
pacieni cunoscui ca fiind infectai. Dintre 18 pacieni
testai un numar de 14 (78%) au prezentat ARN viral doar
n lichidul crevicular i nu n saliv. 20 din 26 pacieni
(77%) au prezentat concentraie mai mare de ARN VHC
n lichidul crevicular dect n saliv, iar pacienii cu
cantiti serice mai mici de ARN viral nu prezentau i
n saliv cantiti detectabile.

The search for hepatitis C virus (HCV) in body fluids


other than blood is important when assessing possible
nonparenteral routes of viral transmission. However,
the role of oral fluids in HCV transmission has not been
yet precisely established. The above-mentioned study
included a determination of HCV RNA in saliva and
gingival crevicular fluid of known infected patients. Of
18 tested patients, 14 (78%) had viral RNA only in the
crevicular fluid and not in the saliva. 20 of 26 patients
(77%) had higher HCV RNA levels in their crevicular
fluid than in their saliva, and patients with low serum
viral RNA were less likely to have detectable quantities
in their saliva.

3. Gkhan Akgz, Murat nan Cengiz, lker


Keskiner, ereften Akgz, Murat Can, and Aydan
Akgz, Correlation of Hepatitis C Antibody Levels
in Gingival Crevicular Fluid and Saliva of Hepatitis
C Seropositive Hemodialysis Patients, International
Journal of Dentistry, Volume 2009 (2009), Article ID
247121 63

3. Gkhan Akgz, Murat nan Cengiz, lker Keskiner, ereften Akgz, Murat Can, and Aydan Akgz,
Correlation of Hepatitis C Antibody Levels in Gingival
Crevicular Fluid and Saliva of Hepatitis C Seropositive
Hemodialysis Patients, International Journal of Dentistry, Volume 2009 (2009), Article ID 247121 63.

n acest studiu a fost comparat prevalena anticorpilor


anti VHC salivari s creviculari la pacienii cu VHC sub
dializ, la studiu participnd 39 de cazuri clinice. Pentru
analiza anticorpilor s-a folosit testul ELISA Ortho HCV
3.0 SAVe. Au fost comparate statistic nivelurile de
anticorpi anti VHC n cele 2 lichide bucale cu nivelurile
din ser, cu urmatoarele rezultate:
anticorpi n ser 100%
anticorpi n lichid crevicular i saliv 15,4%
Studiul concluzioneaz c lichidul crevicular
reprezint o surs de ptrundere VHC n cavitate bucal.

That study compared the prevalence of HCV antibody levels in saliva, and gingival crevicular fluid
of HCV seropositive hemodialysis patients, the study
including 39 clinical cases. The ELISA Ortho HCV 3.0
SAVe test was used for antibody analysis. A statistic
comparison was made between the anti HVC antibody
levels in the 2 oral fluids and the seric levels, with the
following results:
antibodies in serum 100%
antibodies in crevicular fluid and saliva 15,4%
The study concludes that the crevicular fluid is a
source of HVC entering the mouth.

4. Lins L, Almeida H, Vitvisk L, Carmo T, Paran R,


Reis MG. Detection of hepatitis C virus RNA in saliva
is not related to oral health status or viral load, J Med
Virol. 2005 65

4. Lins L, Almeida H, Vitvisk L, Carmo T, Paran R,


Reis MG. Detection of hepatitis C virus RNA in saliva
is not related to oral health status or viral load, J Med
Virol. 2005 65

41

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Hepatita C reprezint o problem de sntate


la nivel mondial; este considerat cu transmitere
parenteral, ns virusul a fost identificat i n alte fluide
ale organismului.
ARN viral a fost izolat i din saliv, ns fr a se
face o legtur clar ntre prezena acestuia i patologia
oral. Studiul a analizat 50 pacieni VHC pozitivi i 25
persoane sntoase. ARN VHC a putut fi detectat n
saliv n toate cele 50 de cazuri clinice tiute ca fiind
afectai de hepatita C. n nici unul dintre cazurile de
pacieni sntoi nu a fost izolat i saliv.
Anamneza complet a cavitii bucale, incluznd
status dentar, leziuni mucozale, nu a permis o corelaie
ntre ARN viral n saliv, gradul de sntate bucal i
ncrcarea viral.
n concluzia studiului se afirm c prezenta n saliv
a ARN VHC este independent de ncrcarea viral i de
patologia cavitii bucale n cazul pacienilor analizai.
5. Marticic M, Poljak M, Kramar B, Seme K,
Brinovec V, Meglic-Volkar J, Zakotnik B, Skaleric
U, Detection of hepatitis C virus RNA from gingival
crevicular fluid and its relation to virus presence in
saliva, Journal of Periodontology, 200171
Studiul a avut drept scop cautarea unei posibile surse
de VHC n saliva i lichid crevicular i corelarea cu
parametri clinici, biochimici, histologici, virusologici
i de sntate oral n care se ncadrau pacienii
analizai. Studiul a fost desfurat pe 50 de cazuri
clinice la pacieni care prezentau viremie pozitiv i a
urmrit detectarea ARN viral prin PCR. n paralel s-au
investigat i ceilali parametri clinici i s-au corelat cu
rezultatele obinute. Rezultatele au artat c s-a detectat
ARN viral n lichidul crevicular n 59% dintre cazuri
(29/49), respectiv n saliv n 35% cazuri (17/48). n
cazul salivei s-a putut decela ARN viral acolo unde a
fost detectat i n lichidul crevicular i s-a observat c
era corelat strict cu prezena anumitor urme de snge
n saliv. Acolo unde mostrele de lichid crevicular
nu erau contaminate cu snge s-a putut detecta ARN

42

Hepatitis C is a worldwide public health problem and


its transmission is clearly associated with the parenteral
route, however, the virus has also been isolated from
other body fluids.
Viral RNA has also been detected in saliva, yet the
relationship between HCV and oral pathology is not
clearly understood. The study analyzed 50 anti-HCV
positive patients and 25 healthy people. HCV-RNA was
detected in all of the saliva samples from the 50 HCV
positive clinical cases. None of the saliva or serum
samples from the non-HCV group were positive for
HCV-RNA.
The complete mouth anamnesis, including the
dental status and the mucous lesions, did not allow any
correlation between the viral RNA in saliva, the degree
of oral health and the viral load.
In conclusion, the study suggests that the HCV-RNA
presence in saliva is independent of the viral load and
the oral pathology of the analyzed patients.
5. Marticic M, Poljak M, Kramar B, Seme K,
Brinovec V, Meglic-Volkar J, Zakotnik B, Skaleric
U, Detection of hepatitis C virus RNA from gingival
crevicular fluid and its relation to virus presence in
saliva, Journal of Periodontology, 200171
The purpose of the study was to search for a
possible HCV source in saliva, and crevicular fluid
and correlation with clinical, biochemical, histological,
virological, and oral health parameters. The study was
conducted on 50 positive viremic clinical patients
and aimed at detecting HCV RNA by PCR. The other
clinical parameters were investigated in parallel and
correlated with the results obtained. The results showed
that HCV RNA was detected in 59% (29/49) of the
crevicular fluid samples and in 35% (17/48) of the
saliva samples. In saliva samples, HCV RNA could be
detected only in cases which also had detectable HCV
RNA in the crevicular fluid samples and was strictly
related to the presence of blood in saliva. In crevicular
fluid samples with no blood detected, HCV RNA was

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viral numai atunci cand ncrcarea viral plasmatic


era mare (p=0.05).
n conlcuzie, s-a stabilit c, pe lng snge ,i lichidul
anului gingival este o surs real de VHC.
Autorii afirm c anumite mecanisme de la nivelul
cavitii bucale, neelucidate nc, ar explica discrepana
ntre valorile salivare i cele creviculare de ARN
detectate. Acesta prezinta un punct de pornire pentru a
se studia implicarea VHC n boala parodontal.

more often present in cases with higher plasma viral


load (P=0.05).
The results suggest that besides blood, the gingival
crevicular fluid is also a real source of HCV.
The authors state that unknown mechanisms in the
oral cavity might explain the discrepancies in RNA
detected values between saliva and crevicular fluid. This
is starting point for investigating the role of HCV in the
periodontal disease.

6. Gingival crevicular fluid harbors hepatitis C virus


RNA, Hepatitis Weekly, 14 Noiembrie 200572

6. Gingival crevicular fluid harbors hepatitis C virus


RNA, Hepatitis Weekly, 14 November 200572

Autorii articoului afirm c, la pacienii infectai, n


lichidul crevicular exista VHC. De asemenea cercettorii
japonezi care au elaborat studiul susin faptul c
identificarea unor fluide infectate, altele dect sngele,
este foarte important n detectarea unor posibile ci
de transmitere neparenterale; totui ei concluzioneaz
c rolul lichidului crevicular rmne controversat. n
cadrul studiului s-a urmrit determinarea cantitativ a
ARN VHC n saliv i lichid crevicular, la pacient care
prezentau anticorpi anti VHC. Majoritatea cazurilor,
78%, adica14 din 18, au prezentat ARN viral n lichidul
crevicular, dei n saliv nu prezentau. De asemenea
77%, adic 20 din 26, aveau niveluri mai mari ale ARN
viral n lichidul crevicular comparativ cu saliva.

The authors of the article state that HCV is present


in the crevicular fluid of infected patients. The Japanese
researchers also developed a study maintaining
that detection of infected fluids other than blood is
important for detecting possible nonparenteral routes
of transmission; however, they conclude that the role of
the crevicular fluids remains controversial. The purpose
of study was to quantitatively determine HCV RNA in
saliva and crevicular fluid in patients with anti-HCV
antibodies. Most patients, 78%, meaning 14 out of 18,
had HCV RNA in their crevicular fluid, although their
saliva samples were negative. Also, 77%, meaning 20
out of 26, had higher HCV RNA levels in their crevicular
fluid than in their saliva.

7. Hepatitis C And Saliva Transmission, HC Vets.


com, http://rense.com/general63/dew.htm, 18 Februarie
200573

7. Hepatitis C and Saliva Transmission, HC Vets.


com, http://rense.com/general63/dew.htm, 18 February
200573

Studiul s-a realizat n condiii stricte de recoltare


i igien care s nu afecteze rezultatele. Datele
obinute sugereaz c saliva pacienilor cu VHC poate
reprezeta un mediu de infectare, mai ales n cazul
pacienilor cu ncrcare viral mare i ingiena oral
deficitar, rezultatele obinute fiind prezentate la 43rd
Interscience Conference on Antimicrobial Agents and
Chemotherapy in Chicago on September 14th 2005.
Studiul s-a realizat pe probe de saliv colectate de la

The study was conducted under strict sampling


and hygiene conditions so that the results would not
be affected. The data obtained suggests that the HCV
patients saliva can be an infectious environment,
particularly if patients have a high HCV viral load and
poor oral hygiene, the results obtained being presented
to the 43rd Interscience Conference on Antimicrobial
Agents and Chemotherapy in Chicago on September
14th 2005. The study was conducted on saliva samples

43

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12 pacieni cu VHC, timp de 21 zile. Dintre cele 248 de


probe recoltate 21%, adic 52, au fost pozitive pentru
ARN VHC. Pentru decelare s-a folosit PCR capabil
sa deceleze concentraii de 43 000 copii/ml. 5 pacienti
nu aveau VHC detectabil deloc pe percursul ntregului
studiu, n timp ce nici unul dintre cei 7 pacieni care
prezentau VHC n saliv nu a avut cantiti decelabile
n fiecare zi (o medie de 7,3 zile, variind de la 1 la 13
zile n funcie de pacient). Pentru concentraia seric
sub 1 000 000 copii/ml nu se putea detecta n saliv nici
o ncrcare viral.
O alt corelaie direct s-a stablit ntre prezena
afectrilor gingivale i detectarea VHC n saliv, precum
i ntre sngerrile gingivale i ARN VHC salivar. O
corelaie invers a fost observat ntre periajul dentar i
detectarea n saliv a ARN VHC, astfel nct n cazurile
a 2 periaje zilnice scade probabilitatea de detectare
comparativ cu un singur periaj dentar pe zi.
n concluzie se afirm c n saliva pacienilor infectai
se poate detecta VHC o cauz posibil constituind-o
contaminarea cu micro particule de snge.
Exist alte cteva studii centrate pe idei asemantoare,
cum ar fi:
Hepatitis C virus quantification in serum and
saliva of HCV-infected patients, Gisele Barreto Lopes
Menezes, Fernanda Albuquerque Pereira, Csar Augusto
Barros Duarte, Theomira Mauadie Azevedo Carmo,
Hermes Pedreira da Silva Filho, Maria Alice Zarife,
Marco Aurlio Krieger, Eliana Almeida Gomes Reis,
Mitermayer G Reis, Mem Inst Oswaldo Cruz, Rio de
Janeiro, Vol. 107(5): 680-683, August 201274
Can Saliva Transmit Hepatitis C?, Nicole
Cutler, L.Ac., http://www.hepatitis-central.com/mt/
archives/2007/03/can_saliva_tran.html, 8 Martie 2007 2
Detection of hepatitis C virus RNA in saliva
samples from patients with seric anti-HCV antibodies,
Patrcia L. Gonalves, Carla B. Cunha, Solange C. U.
BusekII, Guilherme C. Oliveira, Rodrigo RibeiroRodrigues, Fausto EL Pereira, Spania Brazilia,
Brazilian Journal of Infectious Diseases, Februarie
2005 75

44

collected from 12 patients with HCV, over 21 days.


Among the 248 samples collected, 21%, that is 52,
tested positive for HCV RNA. For detection PCR testing
equipment was used, which can detect concentrations of
43,000 copies/ml. Five patients did not have detectable
HCV in their saliva on any day during the study period,
and none of the 7 patients who shed HCV in their saliva
did so every day (mean 7.30 days, range 1 - 13 days).
Below 1 000 000 copies/mL, no HCV viral load could
be detected in the saliva.
Another direct correlation was established between
the presence of gum diseases and HCV detection in the
saliva, as well as between gum bleeding and salivary
HCV RNA. A reverse correlation was established between brushing and HCV RNA detection in the saliva,
so in case of twice-daily brushing the likelihood for
detection decreases in comparison with single-daily
brushing.
The study concludes that HCV can be detected in
the saliva of infected patients, one of the reasons being
the contamination with microscopic amounts of blood.
There are several other studies conducted on similar
ideas, such as:
Hepatitis C virus quantification in serum and
saliva of HCV-infected patients, Gisele Barreto Lopes
Menezes, Fernanda Albuquerque Pereira, Csar Augusto
Barros Duarte, Theomira Mauadie Azevedo Carmo,
Hermes Pedreira da Silva Filho, Maria Alice Zarife,
Marco Aurlio Krieger, Eliana Almeida Gomes Reis,
Mitermayer G Reis, Mem Inst Oswaldo Cruz, Rio de
Janeiro, Vol. 107(5): 680-683, August 201274
Can Saliva Transmit Hepatitis C?, Nicole
Cutler, L.Ac., http://www.hepatitis-central.com/mt/
archives/2007/03/can_saliva_tran.html, 8 Martie 2007 2
Detection of hepatitis C virus RNA in saliva samples from patients with seric anti-HCV antibodies,
Patrcia L. Gonalves, Carla B. Cunha, Solange C. U.
BusekII, Guilherme C. Oliveira, Rodrigo Ribeiro-Rodrigues, Fausto EL Pereira, Spania Brazlia, Brazilian
Journal of Infectious Diseases, Februarie 2005 75

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Vol. XVI nr. 1/2013

Hepatitis B And C In Dentistry, Krasteva A., Vl.


Panov, M. Garova, R. Velikova, A. Kisselova, Z. Krastev,
Sofia Bulgaria, Journal of IMAB - Annual Proceeding
(Scientific Papers) 76
MATERIAL I METODE

Hepatitis B And C In Dentistry, Krasteva A., Vl.


Panov, M. Garova, R. Velikova, A. Kisselova, Z. Krastev,
Sofia Bulgaria, Journal of IMAB - Annual Proceeding
(Scientific Papers) 76
MATERIAL AND METHODS

ACORD INFORMAT

Pentru includerea n lotul de studiat, pacienii vor fi


informai despre investigaiile ce urmeaz a fi realizate
din produsele biologice recoltate i vor semna un acord
informat. Pentru estimarea factorilor de risc, pacienii
inclui n studiu vor fi solicitai s completeze un
chestionar.

INFORMED CONSENT

For participation in the research study group, the


patients will be informed about the investigations to be
conducted on the biological samples that were taken and
they will sign an informed consent. For an estimation
of the risk factors, the patients included in the research
study will be asked to complete a questionnaire.

FIA DE CONSIMMNT INFORMAT AL PACIENTULUI


Stimate pacient
Solicit acordul pentru includerea dumneavoastra intr-un studiu asupra posibilitilor de identificare a
virusului hepatitic C (VHC) in ser, sliva i lichid crevicular (lichidul anului gingival).
Studiul urmrete s verifice:
eficiena diferitelor metode de diagnostic a virusului hepatitic C
posibilitatea utilizrii lichidului crevicular ca surs de identificare a infeciei VHC
Din punctul dumneavoastra de vedere, acordul includerii n acest studiu implic:
- acceptarea completrii fiei ataate cu date ct mai precise
- acceptarea investigrii unor markeri ai virusului hepatitic C prin recoltare de snge, saliv i lichid
crevicular.
- acceptarea utilizrii (sub protecia anonimatului) de catre investigatori a unor informaii din
chestionarul ataat i a unor date personale (sex, vrst, factori de risc).
V precizm c aceste investigaii nu includ terapii experimentale sau prin metode neutilizate n practica
medical actual.
Tehnicile i riscurile sunt cele pe care medicul dumneavoastra vi le va preciza, consimmntul
dumneavoastr separat fiind necesar.

Data, numele i semntura pacientului
46

Data, numele i semnatura investigatorului

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PATIENTS INFORMED CONSENT FORM


Dear Patient,
We ask for your agreement to include you in a research study on the possibilities of detection of
hepatitis C virus (HCV) in serum, saliva and crevicular fluid (gingival crevicular fluid).
The purpose of this study is to see:
The efficiency of different methods for diagnosis of hepatitis C virus
The possibility of using the crevicular fluid as detection source of HCV infection
As far as you are concerned, your agreement to participate in this study involves:

- Your acceptance to complete the attached questionnaire as accurately as possible

- Your acceptance related to the investigation of some markers of hepatitis C virus by blood,

saliva and crevicular fluid sampling.


- Your acceptance for the researcher to use (anonymously) information from the attached
questionnaire, and personal data (sex, age, risk factors).
We specify that these investigations do not include experimental treatments or methods that are
not used in the current medical practice.
The techniques and risks are those specified by your doctor, and your consent is separately required.

Date, name and signature of the patient

Date, name and signature of the researcher

47

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Chestionar

Prin ntrebrile care urmeaz se ncearc estimarea prevalenei factorilor de risc pentru infecia HCV,
la pacienii care solicit asistena medical la clinica noastr. Pentru factorii de risc declarai/depistai
n acest interviu, esenial este s se estimeze data primei expuneri. Pentru aceste obiective, dac este
necesar utilizai liber reversul fiei notnd numrul rndului.
Date personale ale pacientului
Nume familie: _________________ Prenume : __________________Vrsta____ Sex (M/F): __
Locul naterii: ___________________________ Etnie: Roman Alta (numii): _______________
Status marital: Cstorit Necstorit Vduv Divorat
Copil
Tel: ______________________Mediu (U/R):____
Ocupaie i loc de munc: ______________________________________________________
Educaie pacient: (coli terminate): Universitar Liceal Scoal gimnazial
Venit (# main, cas n familia pacientului):_______________ Pacientul consum alcool (Da/Nu): ____
Mrime familie (# persoane care servesc masa simultan cu pacientul): ____#Aduli ____ #Copii ___
Instituie
Evenimente n experiena pacientului:
1. Transfuzie de snge ? (Da/Nu):___ Dac Da data (luna/an) efectuarii:___/____ [Verso > 1]
2. Transplant organe/esuturi ? (Da/Nu):___ Dac Da data (luna/an) efectuare:___/___ [Verso > 1]
3. Primete factori de coagulare ? (Da/Nu):___ Dac Da de cnd (luna/an) face tratament: ___/____
4. Dializ renal ? (Da/Nu):___ Dac dializ unic cnd: ___/___ Dac cronic dializat, de cnd: ___/___
5. Intervenie chirurgical ? (Da/Nu): ___ Dac mai multe cte ? ___minore(1)/majore(2) ___Vrsta prima__
6. Tratamente stomatologice ?Nu__(<5): ___ (<10)____(>10)_____ (luna/an) ultimul: ___/___
7. Multiple ((5) tratam. injectabile Nu__(<5): ___ (<10)____(>10)_____ (luna/an) ultimul: ___/___
Comportamente i riscuri
8. Droguri pe cale IV ? (Da/Nu):___Dac Da cnd a nceput (luna/an)? ___/___ [Clarific risc transmit!]
9. Nr. parteneri sexuali n anul 2012 (adult): __ Boli sex transm antecedente____care?______________
10. Lucreaz cu bolnavi sau n laborator serologie (Da/Nu): ____ [Dac Da clarific natura muncii riscul!]
11. Are tatuaje ? (Da/Nu): ___ [Dac Da clarific cnd i cum au fost facute]
12. Are cercei sau guri pentru cercei? (Da/Nu)____ [Dac Da clarific cnd i cum au fost fcute]
13. Utilizeaz frecvent servicii comerciale pentru manichiur, pedichiur sau brbierit (Da/Nu): ___
[Clarificai]
14. n familie utilizeaz n comun peria de dini sau/i aparatul de ras, (Da/Nu): ___ [Clarificai]
15. Alte boli hepatice__________Contact familial/profesional cu hep.virale________________

48

Data:

Semntura:

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Vol. XV1 nr. 1/2013

Questionnaire

The following questions attempt to estimate the prevalence of risk factors for HCV infection in patients
asking for healthcare in our clinic. For the risk factors stated/detected in this interview, an estimation of
the first exposure date is essential. For these objectives, you are required to use freely the overleaf, and
note down the row number.
Patients Personal Data
Last name: __________________ First name: _____________________Age____ Sex (M/F): __
Date of birth: ______________ Ethnic group: Romanian Other (to be specified): ____________
Marital status: Married Unmarried Widow(er) Divorced
Children
Tel.: ______________________Environment (U/R):____
Profession and place of work: ___________________________________________________
Patients education: (graduated schools): University High School Secondary School
Income (# car, house in the patients family):_____________ The patient drinks alcohol daily (Yes/No): __
Family members (# members eating at the same time with the patient): ____#Adults ____
#Children___ Institution
Events in the patients history:
1. Blood transfusion? (Yes/No):___ If Yes date (month/year):___/____ [Verso > 1]
2. Organ/tissue transplant? (Yes/No):___ If Yes date (month/year):___/____ [Verso > 1]
3. Does the patient receive clotting factors? (Yes/No):___ If Yes since when (month/year): ___/____
4. Renal dialysis? (Yes/No):___ If single dialysis, when: ___/___ If chronically dialyzed, since when: ___/___
5. Surgery? (Yes/No): ___ If several, how many? ___minor(1)/major(2) ___Age on the first surgery__
6. Dental treatments? No__(<5): ___ (<10)____(>10)_____ (month/year) last treatment: ___/___
7. Multiple ((5) injectable treatments No__(<5): ___ (<10)____(>10)_____ (month/year) last treatment: ___/___
Behaviors and risks
8. Injectable drugs? (Yes/No):___If Yes since when (month/year)? ___/___ [Clarify the transmission risk!]
9. No. of sexual partners in 2012 (adult): __
Sexually transmitted disease history ____which one? ______________
10. Does patient works with patients or in the serology lab? (Yes/No): ____
[If Yes clarify the nature of the work risk!]
11. Does she/he have any tattoos? (Yes/No): ___ [If Yes clarify when and how they were made]
12. Does the patient have pierces or wear earrings? (Yes/No)____
[If Yes clarify when and how they were made]
13. Does the patient use frequently commercial services such as manicure, pedicure or shaving (Yes/
No): ___ [ Clarify]
14. Does the patient share his/her toothbrush and/or razor with the other family members?
(Yes/No): ___ [ Clarify]
15. Other hepatic diseases __________ Family/professional contact with viral hepatitis________________

Date:

Signature:

49

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Material biologic: lichid crevicular. Locul de


prelavare a lichidului crevicular este lichidul antului
gingival. Recoltarea se efectueaz n cabinetul de
medicin dentar, cu instrumentar steril, pe conuri
de hrtie sterile, fr a contamina probele cu saliv
sau snge; n acest scop se izoleaz locul cu rulouri
absorbante de vat i se aspir cavitatea bucal
continuu. Recoltarea se face prin imbibiie a rulourilor,
cu rabdare, i aplicare atent pentru a nu provoca
sngerare la nivelul gingiei, acesta fiind de multe ori
deosebit de sensibil, n special la pacienii cu inflamaii
gingivale.

Biological material: Crevicular fluid. Crevicular fluid


samples will be taken from the gingival crevicular fluid.
Sampling will be conducted in the dental department,
using sterile instruments, on sterile paper cones, without
contaminating the samples with saliva and blood; for this
purpose the site is isolated with absorbent cotton rolls
and the mouth is continuously aspirated. The samples
are taken by roll imbibitions, with patience, and careful
application not to cause bleeding of the gum, which is
frequently extremely sensitive, especially in patients
with gingival inflammation.

n paralel cu recoltarea de lichid crevicular se


recolteaz i snge intrevenos pe anticoagulant (EDTA)
dar i saliv

The crevicular fluid samples are taken in parallel


with the IV blood samples, on anticoagulant (EDTA),
and saliva samples.

Metode:

Methods:

teste rapide pentru decelarea anticorpilor anti-HCV


teste ELISA pentru decelarea anticorpilor anti-HCV
metode moleculare (PCR) pentru evidentierea
ARN HCV
metode de cuantificare a numrului de copii virale
(qPCR).

Rapid test for detecting anti-HCV antibodies


ELISA tests for detecting anti-HCV antibodies
molecular methods (PCR) for highlighting HCV
RNA
methods for quantifying the number of viral copies
(qPCR).

Bibliografie
1. Ploss A, Dubuisson J, New advances in the molecular

identification of new treatment targets, Gut 2012;61(Suppl

biology of hepatitis C virus infection: towards the

1):i25ei35. doi:10.1136/gutjnl-2012-302048, 2012

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Vol. XV1 nr. 1/2013

2. Edlin BR., Perspective: test and treat this silent

14. Desenclos JC.; Epidemiology of hepatitis C. Revue du

killer,Nature 2011;474:S18e19, 2011

Praticien, 50: 106670.; 2000

3. Global surveillance and control of hepatitis C. Report

15. Law MG, Dore GJ, Bath N, et al. 2001, Modelling

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hepatitis C virus incidence, prevalence, and long-term

viral hepatitis prevention board, J Viral hepat ; 6 : 35-47,

sequelae in Australia, Int J Epidemiol 32: 717724, 2003

Antwerp, Belgium, 1999

16. Armstrong GL, Wasley A, Simard EP, McQuillan GM,

4. Alter, MJ, Epidemiology of hepatitis C virus infection,

Kuhnert WL, Alter MJ., The prevalence of hepatitis C virus

World J Gastroenterol; 13 : 2436-41, 2007

infection in the United States, 1999 through 2002, Ann

5. Molnar GB, Cocean S, Jebeleanu L, Popa S, Feder A.,

Intern Med; 144: 705-714, 2006

Evaluarea seroepidemiologica a infectiei anamnestice cu

17. Wasley A, Alter M, Epidemiology of hepatitis C:

virusul hepatitei C la populatia din 11 judete ale Romaniei,

geographic differences and temporal trends, Semin Liver

Rom J Infect Dis: VIII: 3-7, 2005

Dis; 20: 116, 2000

6. Gheorghe L, Csiki IE, Iacob S, Gheorghe C, Smira G, Regep

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