Q.1. -Twenty multiple choice questions.
Q.2. – Briefly describe the following-
a) Wobble hypothesis
b) Chemical carcinogenesis
c) Nucleotide excision repair
Q.3. – Write short notes on-
a) Protein sparing effect of carbohydrates
b) Bicarbonate buffer system of blood
c) 15 minutes PSP excretion test
d) Renal production and excretion of ammonia
Q.4. -a) Discuss in brief the causes, clinical manifestations and laboratory diagnosis of AIDS.
b) What is the cause of Orotic aciduria? Which pathway is most affected? Outline the steps of that pathway and explain the basis of treatment of this disorder.
Q.5.- Explain the principle, procedure and applications of –
a) DNA finger printing
c) Cloning of chimeric DNA
Q.6.- a) Discuss the significance of glucuronidation reactions of detoxification giving examples.
b) Describe the process of excessive uric acid production by abnormal activities of various enzymes. Explain the basis of using Allopurinol as a part of treatment of hyperuricemia.
c) Describe in brief the mechanism of chromosomal translocation in the causation of various cancers. Give suitable examples in support of your answer.
Multiple Choice questions
Q.1- Selenium poisoning can be cured by administration of:
a) Benzyl amines
b) p- Bromo Benzene
d) p- nitro benzaldehyde
Q.2- Atrial Natriuretic peptide (ANP) produces all except-
c) Decreased blood pressure
d) Increased GFR.
Q.3- All of the following are associated with metabolic acidosis except-
a) Lactic acidosis
c) Methanol poisoning
d) Prolonged diuretic therapy
Q.4- Feeding of raw egg white will result in the deficiency of –
Q.5- The ideal tumor marker for the diagnosis of pheochromocytoma is-
Q.6- If the p H of the blood is 7.4 then the ratio of [NaHCO3] to [H2CO3] will be-
d) none of the above.
Q.7- Prolonged administration of Acetazolamide (Diamox) as a diuretic in a patient of congestive heart failure will produce-
a) Acidic urine
b) Increase in NH4Cl excretion
c) Alkaline urine
d) none of the above.
Q.8- All are causes of water intoxication except-
a) Renal failure
b) Excessive administration of fluids
c) Deficiency of Aldosterone
d) Hypersecretion of Aldosterone
Q.9- The binding of the prokaryotic DNA dependent RNA polymerase to promoter site is inhibited by-
Q.10-The mutagenesity of a compound can be detected by-
b) Ame’s test
c) Western blotting
d) any of the above.
Q.11- A 20 year old man was diagnosed with abnormal form of β – globin (Hemoglobin Constant Spring) which is longer than the normal protein, which of the following point mutation is consistent with the abnormality?
d) GAC—> UAC
Q.12- Antipyrin breath test is undertaken to diagnose underlying-
d) Renal failure.
Q.13- Sudden increase in γ-GT in chronic Alcoholics indicates:
b) Intrahepatic cholestasis
c) Recent bout of drinking alcohol
d) All of the above.
Q.14- Urinary loss of Sodium occurs in-
a) Addison’s disease
b) Chronic renal failure
c) Diabetic keto acidosis
d) All of the above
Q.15- Carbonic anhydrase is absent in-
a) Parietal cells of stomach
d) Renal tubular epithelial cells.
State True or False
Q.16- High intestinal obstruction causes metabolic acidosis.
Q.17- If urine of specific gravity 1.020 or more contains 3.0gms NaCl/liter, salt depletion is present.
Q.18- Serum Alkaline phosphatase level is abnormally low in obstructive jaundice.
Q.19- BMR is increased in Acromegaly.
Q.20- Aspirin is mainly detoxified by oxidation.
Q.1- b, 2-b, 3-d, 4-a, 5-b, 6-c, 7-c, 8-c, 9-c, 10-b, 11-a,
12-a, 13-c, 14-d, 15-c, 16- true, 17-true, 18-false, 19-true,
Q2- Briefly describe
a) Wobble hypothesis- The mechanism by which the t RNAs can recognize more than one codon for a specific amino acid is described by the wobble hypothesis, in which the base at the 5’end of the anticodon (the first base of the anticodon) is not as spatially defined as the other two bases. Movement of that first base allows nontraditional base pairing with the 3’ base of the codon(the last base of the codon). This movement is called wobble and allows a single tRNA to recognize more than one codon. The result of the wobbling is that there need to be 61 codons coding for amino acids. (The examples of traditional and nontraditional base pairing are required to be given between the codon and the anticodon).
b) Chemical Carcinogenesis- A large number of chemicals have been incriminated as carcinogenic. Some of them are direct reacting and some are procarcinogens and are converted to ultimate carcinogens in the body. The examples of chemical carcinogens are as follows-
1) Polycyclic aromatic hydrocarbons-Benzpyrene and dimethyl benzanthracene.
2) Azo- dyes(Aromatic amines)- β nephthyl amines, N- methyl -4- amino azo benzene, 2-acetyl amino fluorine.
3) Nitrosamines and Amides-Dimethyl nitrosamines, Diethyl nitroso amines.
4) Naturally occurring compounds- Aflatoxin B1 produced by the fungus Aspergilus flavus.
5) Drugs- Alkylating and acylating agents e.g.- Cyclophosphamide, Busulfan.
diethyl stilbestrol, estrogen, Nitrogen Mustard, β- propiolactone.
6) Miscellaneous agents- Beryllium, Cadmium, Nickel, chromium, Arsenic, Asbestos, vinyl chloride, saccharine and cyclamates.
Mechanism of chemical carcinogenesis- the chemicals can be-
Direct reacting-which can interact directly with the target molecule.
Procarcinogens- are not chemically reactive. In the body after metabolism they are converted to ultimate carcinogens which are highly reactive. This process of conversion is called metabolic activation. The enzyme systems involved in metabolic activation are cytochrome P450 species present in the endoplasmic reticulum of cells.
DNA is the primary and the most important target of chemical carcinogens. Hence chemical carcinogens are mutagens. The damage to DNA occurs can be-
– binding covalently with DNA(also to RNA and proteins)
– Interaction with purine, pyrimidines and phosphodiester groups of DNA
– Most common site of attack is guanine .
C) Nucleotide Excision repair-
The exposure of a cell to ultraviolet light can result in the covalent joining of two adjacent pyrimidines(Usually Thymines) producing a dimer. These thymine dimers prevent the DNA polymerase to further replicate the DNA strand beyond the site of dimer formation. The dimers are excised in prokaryotes by Nucleotide excision repair. The similar mechanisms operate in human beings.
- Recognition of dimer by UV specific endonuclease- First a UV specific endonuclease recognizes the dimer and cleaves the damaged strand at the 5’side of the dimer.
- Excision of damaged DNA-Next an excision endonuclease recognizes the incision made by the endonuclease. The stretch of the DNA containing the dimer is removed and the gap is filled by using the sister strand as a template by DNA polymerase. The nick is sealed by DNA ligase.
- Xeroderma pigmentosum-The pyrimidine dimers can be formed in the skin cells of humans exposed to UV light. In the rare genetic disease Xeroderma pigmentosum, the cells can not repair the damaged DNA strand, resulting in extensive accumulation of the mutations and consequently skin cancers. The most common form of the disease is caused by the absence of the UV specific endonuclease.
Q.2- a) Protein sparing effect of carbohydrates-
Adequate amount of carbohydrates and fats in the diet may reduce the protein requirement. This may be due to metabolic products of carbohydrates such as oxaloacetate, pyruvate and alpha keto glutarate which provide the carbon skeleton for the formation of non essential amino acids through transamination. Carbohydrates reduce the requirement of the amino acids for gluconeogenesis. Both carbohydrates and fats are oxidized to produce energy and thus spare the proteins to be used for this purpose.
b)Bicarbonate buffer system of blood
This consists of a weak acid “Carbonic acid and its corresponding salt with a strong base (Sodium bicarbonate).
Normal ratio in blood – NaHCO3: H2CO3– 20:1
They are the chief buffers of blood and constitute the Alkali reserve of the body. Neutralization of the strong acids and non volatile acids entering the ECF is achieved by the bicarbonate buffers. Such acids, e.g.- HCl. H2SO4, Lactic acid, etc which are strong and volatile react with NaHCO3 component forming H2CO3. Thus a strong and non volatile acid is converted to a volatile and weak acid, which is eliminated through alveoli of lungs. Hence bicarbonate buffer system is linked directly with respiration.
Similarly when alkaline substance enters e.g. NaOH enters the ECF, it reacts with the acid component of the buffer system, and forming H2O.Thus the p H is maintained.
Advantages of bicarbonate system-
-It is present in very high concentration as compared to other buffers.
– Produces H2CO3 which is weak and volatile and CO2 ie exhaled out.
Thus it is a very good physiological buffer and acts as a front line defence.
Disadvantage of bicarbonate buffer
As a chemical buffer it’s a weak buffer as pKa is further away from the physiological pH
c) 15 minutes PSP excretion Test
This test is based on the fact that in the first 15 minutes 30 -50 % of the injected dye is excreted in urine which is an index of the adequate renal functions.
Test and interpretations-
When 1 ml of PSP(6mg) is injected IV, normal kidney will excrete 30-50% of the dye in first 15 minutes.
Excretion of less than 23% of the injected dye during this period regardless of the amount excreted in 2 hours indicates impaired renal functions.
It is also used to determine the function of each kidney separately. Here the appearance time and the rate of excretion of the dye are important. Increase in appearance time and the decrease in excretion rate indicate impaired renal functions.
c) Renal production and excretion of ammonia-
Ammonia mechanism operates in the distal tubular cells, for the elimination of H+ ions and the conservation of Na+.
Sources of Ammonia-
1) Ammonia is produced by the hydrolysis of Glutamine by the enzyme Glutaminase which is present in these cells.
2) In addition NH3 is also produced from other amino acids by the oxidative Deamination by L- amino acid oxidase.
3) It can also be formed from the glycine by the Glycine oxidase enzyme.
The NH3 thus formed forms Ammonium ion by combining with H+ ions and ammonium ion can exchange Na+ from NaCl.
The ammonia production is greatly increased in metabolic acidosis and is reduced in alkalosis. The activity of the Glutaminase enzyme is also enhanced in acidosis.. This mechanism is important for the conservation of the fixed bases.
Q4- a) AIDS is caused by a virus called Human Immuno deficiency virus. HIV belongs to the retrovirus group. They are RNA containing viruses that replicate with the help of the reverse transcriptase enzyme.75% of the total patients get this infection through sex. In about 15% of the patients the disease is transmitted through blood. Virus may be transmitted from mother to fetus through placenta. About 30% of the infants born of HIV positive mothers may get the infection.
Lymphadenopathy and fever may be seen at the end of the second stage. As the name indicates the basic defect lies in the immunodeficiency. This is mainly because the T helper cells are decreased in number. Since the immunity id deficient, non pathogenic micro organism enter the body and produce lesions in the skin, gastro intestinal tract, lungs, urinary tract and brain. Gastroenteritis and tuberculosis are the predominant pattern inIndia. In all the cases there is weight reduction and Kaposi’s sarcoma is also commonly associated with AIDS.
1) The antibodies in the blood are detected by ELISA test, but there is a chance of false positive test, so all the positive cases are confirmed by Western Blot analysis. In ELISA the antibody against only one antigen (gp120) is tested, so there is probability of false result.
2) In Western Blot analysis, antibodies against 6 different components of the virus are analyzed, so its confirmatory.
3) T helper count is lowered. The normal level is more than 400/cmm. In AIDS patients the level is always is below 300/cmm. As the disease progresses the, the T helper count is correspondingly is lowered.
4) In the last stages the p24 antigen starts rising.
5) By Reveres transcriptase polymerase chain reaction, or by real time PCR, the number of HIV particles in blood can be estimated, value of less than 5000 copies per ml of blood has good prognosis, while a count more than 100, 000 per ml means a very bad prognosis.
c) Orotic aciduria
This condition results from absence of either or both of the enzymes, OPRTase and OMP decarboxylase. It is an autosomal recessive disease, characterised by retarded growth and megaloblastic anemia. The rapidly growing cells are most affected, hence there is anemiaCrystalsare excreted in urine, which may cause urinary tract obstruction. Due to the lack of feed back inhibition orotic acid production is excessive. The condition can be treated by feeding cytidine or Uridine. The may be converted to UTP which can act as feed back inhibitor.
This defect is associated with the pathway of Denovo pyrimidine synthesis.
Steps are to be mentioned.
Q5- a) DNA finger printing
There are tandem repeats (VNTR) in chromosomes. The number of such repeats sites varies from person to person, but the number is unique for a given person. Therefore it serves as a molecular finger print. It is also known as a DNA profile. Probability of similarity between two persons is only 1 in 3×1010persons. DNA probes have been developed, so that at low stringency, they hybridize to a number of loci to produce individual specific finger prints. The technique is used identify the criminals and also used in disputes of parenthood. DNA can be isolated from the stains on clothing made by blood or semen stained even several years ago.
DNA finger printing is more applied in medico legal cases.
This Technique is widely used for detecting antigens, antibodies, estimating hormones, growth factors and tumor markers etc. This test is commonly employed to detect antigens or antibodies present in very small concentration in tissues or blood.
Antigen detection by ELISA Method
The specific antibody is fixed to the well of the micro titer plate. The patient’s serum is added to the well and incubated for 30 minutes at 37 degree centigrade. By this time if the serum contains antibodies it would have fixed to the well. Excess antigen and unwanted proteins are washed out and the antibody tagged with hose radish peroxidase is added. If the antigen is already fixed, the Ab-HRP conjugate will be fixed to the well. Then a color reagent containing hydrogen peroxide is added to the well. Color is developed. The development of the color indicates that the Ag is originally present in the patient’s serum. The intensity of the color formed is proportional to the amount og the antigen present in the given sample.
Based on the specific affinity the antibodies can also be detected by this method.
c) Cloning of chimeric DNA- cloning allows the production of a large number of identical molecules. The steps involved are-
1) Preparation of chimeric DNA molecules
2) Transfection of the vector in to the host
3) Selection of the colony carrying the desired genes
4) Amplification of the gene.
All steps are to be elaborated with diagrams. The applications of this technology in the field of medicine, agriculture and industrial fields are to be discussed in brief.
Q.6- a) Glucuronidation reactions of detoxifications
These reactions are for the detoxification of toxic substances. UDP Glucuronic acid is formed in the uronic pathway. Glucuronyl transferase is the enzyme responsible for the detoxification reactions. Various substances that are conjugated with Glucuronic acid are as follows-
1) Bilirubin to form bilirubin diglucuronide.
2) Aromatic acids e.g.- Benzoic acid
3) Phenols and other secondary and tertiary aliphatic alcohols
4) Morphine, menthol, acetanilide, sulfa, pyridine etc.
5) Hormones like thyroid hormones, derivatives of steroids etc
b) Hyperuricemia by abnormal activities of enzymes
The activities of PRPP synthetase, amido transferase, HGPRTase, Glucose-6 phosphatase and Glutathione reductase are to be mentioned.
The use of Allopurinol as a competitive inhibitor of Xanthine oxidase is to be mentioned.
d) Chromosomal Translocation- Most of the tumor cells show chromosomal abnormality, in which a piece of one chromosome is split off and joined to another chromosome. If the second chromosome also donates the piece to the first then is is called reciprocal translocation.
The examples of Burkitt’s lymphoma, chronic granulocytic leukemia are to be mentioned with diagrams.
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