1.

DIFFERENCES OF NECROSIS AND APOPTOSIS

Necrosis Apoptosis

Ada blebbing,
Morphology feature Loss of membrane integrity
No loss of membrane integrity

Dimulai dgn swelling Sitoplasma menyusut,

cytoplasm & mitochondria Nucleus condensed

Ends with fragmentation,

Ends with lysis
Formation of smaller body

Complete lysis,
Terbentuk apoptotic bodies
no vesicle formation

Ada regulation,
Biochemical feature No regulation
Usage of enzymatic step

Post-lytic DNA fragmentation Pre-lytic DNA fragmentation

Ada release caspase cascade

Ada release various factor dari

mitokondria ke sitoplasma

Physiological feature Affect group of cells Affect individual cells

Ada inflammatory response No inflammatory response

Disebabkan karena non- Akibat physiological

physiological disturbance disturbance

1
 2. CELL CYCLE

The cell cycle shown on the diagram visualized how cells reproduce.
The cycle goes from G1 S G2 mitosis / meiosis

Stage G1: At this stage, the cell that is going to reproduce is just growing into a bigger cell.
Before moving on to the next stage, here lies a checkpoint in which to see that the grown cell is
normal, such as no cell inhibition, no DNA damage, enough space for replication, etc.

Stage S: At this stage, the cells DNAs inside the nucleus are duplicated and form into
chromosomes.

Stage G2: At this stage, the cells DNAs are shaped in chromosomes and getting ready for cell
division. In this stage, here lies another checkpoint. This checkpoint looks for any DNA damage
or any wrong base pairings before moving on to cell division.

Mitosis/Meiosis

Mitosis: This is where cell division happens for autosomal cells (diploid cells reproducing
diploid cells). It started from the release of chromosomes from the nucleus (interphase) and the
formation of spindle fiber inside the cell (prophase). As the spindle is forming, the chromosomes
must line up at the center of the cell and attach to the spindle at their centromeres (metaphase).
After metaphase happened, here lies another checkpoint in order to see whether all spindle fiber
is attached to the centromeres. When all centromeres are attached, the spindle fiber will start to
pull the chromosomes to different polar, making the chromosomes separated into chromatids

2
(anaphase). After the separation of the sister chromatids, the nucleus started to form again while
the cells are now separating as well (telophase). The cell should separate perfectly in order for
reach success in cell cycle (cytokinesis).

Meiosis: This is where cell division happens for gametes (diploid cells reproducing haploid
cells). The basic of meiosis is similar with mitosis. The differences in meiosis is that it has 2
cycle of mitotic stage and involves exchanging genetic materials.

The basic of cell cycle was discussed above. By understanding cell cycle, therefore it is easier to
understand how cancer cells develop. There are 3 critical genes in controlling cancer cells, DNA
repair genes, proto-oncogenes/oncogenes, tumor suppressor genes.

DNA repair genes: These genes repair when DNA is broken.

Single strand damage:

o Mismatch repair for repairing mismatched base pairing
o Base excision repair for repairing damage due to point base mutation
o Nucleotide excision repair for repairing base that connects to the consecutive
base on the same strand
Double strand damage:
o Homologous recombinant end joining for repairing broken part of DNA
o Non-homologous recombinant end joining for repairing the loss base pair in
DNA sequence

Proto-oncogenes: This gene does not generate cancer cells, but when its mutated into oncogenes,
it generates cancer cells.

Tumor suppressor genes: This gene, particularly p53, suppressed the development of cancer.
When it is mutated, then there is nothing to suppress any cancer development. Therefore, cancer
cells might develop without inhibition.

3
 3. GENE EXPRESSION

1. Transcription
a. The synthesis of mRNA from a DNA template from 5 to 3 direction in
the nucleus
b. Factors of transcription:
i. DNA template
ii. Transcription factors
iii. RNA polymerase
iv. ATP
v. Nucleotides (NTPs)
c. Steps of transcription:
i. Initiation
1. Transcription factors (TFs) bind to promoter region(TATA
box)
2. RNA polymerase binds to the promoter
3. TFs hydrolyze ATP to obtain energy for DNA unwinding
4. RNA polymerase unwind the DNA template
ii. Elongation
1. Unwinding of dsDNA in the front
2. Synthesis of the RNA
3. RNA proofreading
4. Dissociation of RNA
5. Re-annealing of DNA behind the enzyme
iii. Termination
1. Dislocation of RNA strand from DNA template
2. Dislocation of RNA polymerase from DNA template

2. Processing / Splicing
a. RNA modifications which removes introns and joined exons
b. Conversion from primary transcript to mature RNA
c. Intron : DNA region within a gene that is not translated to a protein
Exons : DNA region within a gene that is translated to a protein
d. Steps of splicing:
i. Assembly of helper protein (ex. U1, U2AF, and BBP) at the
intron-exon borders defining the section to be cut
ii. Splicing factors act as a beacon to guide smaller nuclear ribo
proteins (snRNPs) to form a splicing machine called spliceosome.
iii. Spliceosome bring exons on the either side of intron together,
ready to be cut

4
 iv. Spliceosome cuts the 5 end of intron and loops it back on itself
to joint at a sequence of mRNA called branch site. This form
the lariat loop.
v. Spliceosome cuts the 3 end of intron releasing the lariat loop
and joining two exon ends together
vi. The edited mRNA and introns are released and the spliceosome
disassemble; the lariat loop also broken down

3. Translation
a. Synthesis of protein from mRNA template occurring in the ribosome of
cytoplasm
b. The initiator tRNA is formylated methionine (f-Met-tRNAiMet)
c. N-formyl methionine is the first amino acid of all proteins
d. Factors of translation:
i. Mature mRNA
ii. Ribosome (large and small subunit)
iii. tRNA
iv. Release factor
e. Translation steps:
i. Initiation
1. Messenger RNA binds to the ribosome
2. The initial tRNA binds the P site on the ribosome
ii. Elongation
1. Subsequent tRNAs with bound amino acids first enter at
the A site
2. mRNA pass along the ribosome in short spurts of 3
nucleotides at a time
3. The initial tRNA is moved to the E site and its amino acid
is transferred to the next amino acid on the P site.
Simultaneously, a new codon is presented on the A site.
4. The previous tRNA , which now no longer carries an amino
acid, leaves the E site and the next tRNA, with a
complementary anticodon, enters the A site.
5. The old tRNA paired with the previous codon is passed to
the P site then to E site as the amino acid it carried is
transferred to the growing amino acid chain
iii. Termination
1. Binding of release factor to stop codon
2. Releasing of tRNAs from the mRNA
3. Releasing of ribosome from the mRNA

5
 4. Post-Translational Modification (>200 modification available)
5. Protein Transport / Secretion

6
 4. DNA DAMAGE

Single strand DNA damage

Damage terjadi hanya pada salah satu dari 2 strand DNA. Dapat direpair

dengan cara :
o MMR (mismatch repair)

Dilakukan pada damage dimana terjadi kesalahan pasang (mismatch)

dari sequence nukleotida yg ada. Repair dilakukan agar tidak terjadi

mutasi permanen.
o BER (base excision repair)

Dilakukan untuk damage akibat alkilasi, deaminasi, atau oxidasi,

dimana terjadi perubahan sequence nukleotida skala kecil yg tidak

menyebabkan distorsi (non-helix distortion).

o NER (nucleotide excision repair)

Dilakukan pada damage akibat radiasi sinar UV, carcinogen, dimana

terjadi perubahan sequence nukleotida skala besar yg telah

menyebabkan distorsi (helix distortion). Repair dilakukan dgn

mengganti suatu region.

Double strand DNA damage

Damage terjadi pada kedua strand DNA yang ada. Dapat direpair dgn cara :
o Homologous end joining

Breakage akan direjoined menggunakan DNA ligation, merupakan

pathway cepat
o Non-homologous end joining

Menggunakan DNA pairing sbg template untuk hasil lebih akurat

namun lebih complex dan lama

7
 5. GENE HEREDITY PEDIGREE

1. Autosomal
a. Dominant
i. The allele is passed down generation to generation
ii. Both males and females may be affected
iii. Every affected person has an affected parent or even both
iv. Males can transmit this condition to both genders (and so can
woman)
v. Disease example:
1. Achondroplasia
2. Familial
hypercholesterolemia
3. Camptodactyly
4. Huntington disease
5. Etc.

b. Recessive
i. Affected individuals have both heterozygous parents or affected
ii. Presence of carrier
iii. Characteristic shown on homozygous recessive individual
iv. Males can transmit this condition to both genders (and so can
woman)
v. Both males and female can be affected
vi. Disease example:
1. Cystic Fibrosis
2. Albinism
3. Galactosemia
4. Sickle-cell anemia

2. Sex linked
a. X-linked dominant
i. Males and female may be affected
ii. Both son and daughter of affected woman has 50% risk of being
affected
iii. All daughter of affected male will surely affected
iv. No male to male transmission
v. Affected woman may have milder phenotype than affected man
vi. Disease example:
1. Ornithin Transcarbamylase Deficiency (OTCD)
2. Hyphysphothamic Rickets
3. Microphtalmia w/ linear skin defects
4. Oral facial digital type I syndrome

8
 b. X-linked recessive
i. Trait more frequent in males than females
ii. No male to male transmission
iii. All daughters of affected male are carriers
iv. Disease examples:
1. Hemophilia
2. X-linked severe combined immunodeficiency (SCID)
3. Androgen Insensitivity Syndrome
4. Color Blindness
5. Fragile X Syndrome
6. Muscular Dystrophy

c. Y-linked
i. Man to man transmission
ii. Only man is affected
iii. Gender determination
1. SRY present testes form
2. SRY absent ovaries form
iv. Fewer than two dozen genes identified

9
 6. 5 VARIANTS OF MEDELLIAN INHERITANCE

1. Incomplete Dominance
F1 hybrid have the appearance of between of the

parents phenotype

2. Codominance
Two or multiple allele are expressed in

heterozygous individuals

3. Epistasis
The expression of one gene affects the

phenotypic expression of

another gene

4. Pleiotropy
A single gene is responsible for

a variety of traits

5. Polygenic Inheritance
Two or more genes are responsible for a

single trait

7. ENZYME AS BIOCATALYST

Definition and Working Method

Enzymes are protein that speed up chemical reactions in a cell without
getting consumed. A special region on the enzyme, called the active site, has
a shape that fits with specific substrate molecules. An enzyme works by
binding to one or more specific molecules called reactants or substrates
which occur on the active site. The enzyme substrate complex can occur in

10
 lock & key model as well as in induced fit model. The interactions between
the substrate and the enzyme stresses or weakens some of the chemical
bonds in the substrates; these encourage the formation of the products. The
new product(s) is formed and released from the active site, the enzyme
assumes its original shape and free to work again.

Activation Energy (Ea)

Activation energy is the energy required for a chemical reaction to took place.
That means the lower the Ea, the faster the reaction will go and vice versa.
Enzyme works by lowering those Ea thus speeding up the reaction a lot more.
Analogically, it can be said that you want to go to point a to b while there is a
tall mountain in between. The journey must have been tough, but if you are
given a shovel and can dig through the mountain straight to point b then the
journey must be easier. In this case, the mountain represents the activation
energy and the shovel represent the enzymes.

Types

8. STEM CELL BASED ON DIFFERENTIATING ABILITY

Totipotent

11
 Each cell can develop into new individual
(ex. Cell from early embryo [1-3 days])

Pluripotent

Cells can form any (over 200 cell types)

Except placenta and other fetal supporting
cells

Multipotent

Cells have differentiated but can form a

number of other tissues; these cells are
committed to give rise of cells that have
specific functions (ex. Hematopoietic cell)

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 9. INDUCED PLURIPOTENT STEM CELL

Pluripotent stem cells are that generatedfrom differentiated (non-pluripotent) cells;

orapluripotent cells derived from dedifferentiatingand reprogramming cellswhose developmental
fate have beendetermined.

Challenge

1. Finding approaches for delivering reprogramming factors into nucleus

2. Difficult to yield high number iPSC adifferent cell different efficiency.
3. Minimizing DNA alteration on reprogramming.
4. Developing more safely programming for clinical application

Treatment

13
 1. Amyotropic Lateral Sclerosis (ALS)/Lou Gehrigs disease
2. HIV AIDS patients
Because HIV/AIDS can mutate and become immune totreatments at an accelerated pace,
most treatmentsinvolve a chemical cocktail
47 (give or take) different medications
Not a simple method
This treatment carries with it many complications
Patients play the side effect game
The chemical compounds are a burden on the body
Extremely large structures
Designed now so the body to not be able to break itdown
Intended to extend the length of treatment
3. Mesenchymal stem cells for Autism and Autism Spectrum Disorders (ADS)
4. Treatment of chronic spinal cord injured patients with autologous bone marrow-derived
hematopoietic stem cell transplantation
5. Cairo University conducted human trial of hematopoietic progenitor stem cells in SCI
o Results: Improved functions
o Not FDA approved
o MRI showed no tumors
o Awaiting 1 year follow up

14
 10. RESPIRATION (AEROBIC / ANAEROBIC)

o Glycolysis

Result: 2 ATP, 2 NADH, and 2 Pyruvate

o Krebs Cycle / Citric Acid Cycle (Aerobic)

Results: 2 ATP, 6 NADH, 2 FADH2, 4 CO2

o Electron Transport Chain

15
 In complex I, NADH is oxidized to NAD+, by reducing Flavin mononucleotide to FMNH2 in one
two-electron step. FMNH2 is then oxidized in two one-electron steps, through a semiquinone
intermediate. Each electron thus transfers from the FMNH2 to an Fe-S cluster, from the Fe-S cluster
to ubiquinone (Q). Transfer of the first electron results in the free-radical (semiquinone) form of Q,
and transfer of the second electron reduces the semiquinone form to the ubiquinol form, QH2.
During this process, four protons are translocated from the mitochondrial matrix to the
intermembrane space.
In complex II, additional electrons are delivered into the Quinone pool (Q) originating from
succinate and transferred (via FAD) to Q. Complex 2 is a parallel electron transport pathway to
complex 1, but unlike complex 1, no protons are transported to the intermembrane space in this
pathway. Therefore, the pathway through complex 2 contributes less energy to the overall electron
transport chain process.
In Complex III, two electrons are removed from QH2 site and sequentially transferred to two
molecules of cytochrome c, a water-soluble electron carrier located within the intermembrane
space. The two other electrons sequentially pass across the protein to the Qi site where the
quinone part of ubiquinone is reduced to quinol. In total four protons are translocated: two protons
reduce quinone to quinol and two protons are released from two ubiquinol molecules. Simply:

QH2 + 2 cytochrome c (FeIII) + 2 H+in Q + 2 cytochrome c (FeII) + 4 H+out

In complex IV, four electrons are removed from four molecules of cytochrome c and transferred
to molecular oxygen (O2), producing two molecules of water. At the same time, eight protons are
removed from the mitochondrial matrix (although only four are translocated across the
membrane), contributing to the proton gradient. The activity of cytochrome c oxidase is inhibited
by cyanide.

16
o Net Energy Production

Process ATP Comment

Glycolysis 2 ATP 2 ATP loss
Krebs Cycle 2 ATP -
Electron Transport Chain
Each glycolysis NADH use 1 ATP for mobility to
2 NADH 6 ATP
mitochondria
NADH in conversion of pyruvate to acetyl COA &
6 NADH 18 ATP
Krebs Cycle
2 FADH2 4 ATP -

1 NADH = 3 ATP | 1 FADH2 = 2 ATP

o Fermentation (Anaerobic)
Alcohol (Yeast & Bacteria)

Lactate (Humans and other mammals)

o Aerobic Vs. Anaerobic Result

Process Net ATP

Aerobic 36 ATP

Anaerobic 2 ATP

17
 11. RELATIONSHIP GLYCOLISIS, PPP, NUCLEOTIDE METABOLISM

18
 12. 4 MODELS OF CELL SIGNALING

1. Contact Dependent
Cells may communicate directly with their immediate neighbor through gap
junctions or cells can interact in a classic signaling manner, through cell
surface molecules. The signaling molecule is not secreted, but is bound to the
plasma membrane of the signaling celland interacts directly with the receptor
exposed on the surface of the target cell.

2. Paracrine
Signal diffuse through the extracellular fluid and act locally on nearby cells.
This will usually result in a signal concentration gradient, with the cells in the
local area responding differentially to the extracellular signaling molecule
according to the concentration they are exposed to (this is an important
strategy in development). In order to keep the effect contained, signaling
molecules involved in paracrine signaling are usually rapidly taken up by cells
or degraded by extracellular enzymes.

3. Neuronal / Synaptic
A faster and more specific form of cellcell signaling. Nerve cells, or neurons,
can convey signals across considerable distances to the next neuron in the
neuronal network within milliseconds. By contrast, blood-borne messages can
only operate as fast as blood circulates, but reach many more cellular targets
in different tissues. The transfer of information from one neuron to the next is
mediated by complex structures called synapses, which are essentially
formed by a presynaptic terminal (neuron 1), a synaptic cleft (the tiny gap
between the two neurons) and a postsynaptic membrane (neuron 2). When
electrical signals reach the end of a neuronal axon (the thin tube-like part of
neurons), molecules released from the axon can cross the physical gap
between cells and bind to receptors in the target cell.

4. Endocrine
Signals are transmitted over larger distances, for example from one organ,
such as the brain, to another, such as the adrenal gland. For long-distance
signaling, diffusion through the extracellular fluid is obviously inadequate. In
such cases, signaling molecules may be transported in the blood.

19
20
 13. INSULIN EFFECT

Insulin reacts depending on the concentration of blood glucose at one time. If the
concentration of blood glucose is above normal, the insulin signaling cascade will
happen.

1. Insulin receptor is triggered

2. The binding activates many protein cascade ultimately creating Shc and IRS
proteins
3. The IRS proteins then work alongside p85 and p110 to create PI3-kinase
4. The PI3-kinase is responsible for the translocation of GLUT4 from vesicle to
membrane
5. Glucose then can pass through the increasing GLUT4
6. Glucose converted to glucose-6-phosphate by hexokinase II which then used
for metabolism/storage
7. Simultaneously, Shc and IRS proteins work side by side for mitogenesis,
protein synthesis, glucose transport, and glycogen synthesis
8. As level of blood glucose in blood reach normality, the GLUT4 are
endocytosed back to vesicle

21
 14. CRITICAL CANCER GENE

DNA repair genes

o None, poor, or reduced DNA repair mechanism can result in
pathological effects of such as cancer or senescence. One studies show
that reduction in the ability of NHEJ increase the occurrence of
lymphoma (a type of cancer).
o If the amount of error exceeds the capacity of the cell to repair it, the
accumulation of errors can overwhelm the cell and result in early
senescence or cancer.

Proto-oncogenes
o Proto oncogenes are expressed during regulated growth as it
encodes:

Stimulation of cell Nonreceptor Tyrosine

mitosis Kinase
Growth factors Cell apoptosis

22
 o When mutated, the cell cannot control the cell cycle correctly and
stimulate more and more cell mitosis even when it is not needed which
in turn resulting in cancer
o If the cell cycle is a car, then oncogene is the gas pedal and when it is
mutated, it is similar to a gas pedal being pressed down all the time
resulting nonstop movement of the car
o
Tumor suppressor genes
o TSG, particularly p53, suppressed the development of cancer by
regulating cell cycle; TSG acts as a break whenever mistake occur in
the cell cycle thus preventing the spread of mutation.
o There are two copies of TSG and if one is mutated, usually normal
people can still prevent most cancer from forming by the remaining
normal TSG

o
15. TUMOR ANGIOGENESIS STEPS
o

o Angiogenesis is a necessary part of the process in the progression of

cancer from small, localized neoplasms to larger, growing, and
potentially metastatic tumors. To grow beyond 1 to 2 mm in diameter,
a tumor needs an independent blood supply, which is acquired by the
expression of growth factors that recruit new vasculature from existing
blood vessels (Figure 1). This process continues even as the tumor
matures. Thus, upregulation of angiogenesis is a key step in sustained
tumor growth and may also be critical for tumor.

1. Endothelial cell activation by growth factors (ex. VEGF and bFGF)

2. Degradation of the endothelial wall by certain proteinase (ex. MMPs)
3. Formation of branch point in vessel wall
4. Migration of the endothelial cells to the extracellular matrix towards the
angiogenic stimulus
5. Re-organization of endothelial cells to form tubules with a central lumen
6. Interconnection of the new tubules to form a branched network (anastomosis)
o