Regional differences in constituents of gall stones.

The pathogenesis of pigment and mixed gall stone formation remains elusive. The elemental constituents of gall stones from southern states of Tamil Nadu, Kerala and Karnataka have been characterized. Our aim was to determine the elemental concentration of representative samples of pigment, mixed and cholesterol gall stones from Andhra Pradesh using proton-induced X-ray emission (PIXE) using a 3 MV horizontal pelletron accelerator. Pigment gall stones had significantly high concentrations of copper, iron and lead; chromium was absent. Except for iron all these elements were significantly low in cholesterol gall stones and intermediate levels were seen in mixed gall stones. Highest concentrations of chromium was seen in cholesterol and titanium in mixed gall stones respectively; latter similar to other southern states. Arsenic was distinctly absent in cholesterol and mixed gall stones. The study has identified differences in elemental components of the gall stones from Andhra Pradesh.

Apolipoprotein E polymorphism and coronary heart disease.

Apolipoprotein E is a constituent of various lipoproteins and plays an important role in the transport of cholesterol and other lipids among cells of various tissues. The gene is polymorphic with three alleles (epsilon2, epsilon3, and epsilon4) coding for isoforms E2, E3, and E4 and having different binding affinities for the apo E receptors. While the epsilon2 allele is associated with elevated triglyceride levels, epsilon4 allele is associated with increased cholesterol levels. Though several studies support the role of apo E polymorphism in CHD either directly or indirectly via its influence on lipid and lipoprotein levels, there are some studies, which show no association. With the increasing incidence of CHD among Indians, it becomes imperative to identify genetic markers that may predispose individuals to coronary events. It would be of importance to determine if apo E gene will become a usefuladjunct to assess cardiovascular risk profile when performing genetic studies in families.

Methylenetetrahydrofolate reductase gene mutation and hyperhomocysteinemia as a risk factor for coronary heart disease in the Indian population.

OBJECTIVE: The screening and therapeutic guidelines for the management of lipid abnormalities are reasonably well established. However, other risk factors like hyperhomocysteinemia (HCA) and single nucleotide polymorphisms involving the angiotensin converting enzyme (ACE) and angiotensinogen genes, various clotting factors etc., have yet to be established firmly as other causative factors of atherothrombotic disease. Our study was aimed at finding the relationship between HCA, folate, vitamins B12 levels, and mutations in the 5,10-methylenetetrahydrofolate reductase (MTHFR) and cystathionine beta-synthase (CBS) genes. METHODS: We studied 230 subjects, which included patients with angiographically documented coronary heart disease (CHD) (n=115) and controls (n=115) with no history of CHD. RESULTS: Elevated levels of plasma homocysteine, above 18 nmoles/ml, were detected in 19.13% and 18.26% of our patients and controls, respectively. Homocysteine was significantly correlated to Apo A1 (r=0.51, p < 0.05) and Apo B (r=0.49, p < 0.05). The heterozygous MTHFR mutation was found to be 54.5% (12/22) in our patients with HCA. Of these, 31.8% (7/22) were deficient for plasma folate. Heterozygosity for T833C mutation in the CBS gene was observed in 9.99% (2/22) of our patients with HCA. Both these patients were also deficient for plasma folate and vitamin B12. CONCLUSION: In our study, heterozygosity for the thermolabile MTHFR mutation was found to be associated with hyperhomocysteinemia (HCA). This genetic predisposition to HCA could be risk factor for CHD and can be correlated with vitamin supplementation. To the best of our knowledge this is the first report from India on plasma homocysteine levels and its genetic aspect in patients with CHD.

Changes in the composition of erythrocyte membrane during storage of blood in di-(2-ethyl hexyl) phthalate [DEHP] plasticized poly vinyl chloride (PVC) blood storage bags.

Very little information is available on the changes in the erythrocyte membrane composition during storage of blood at 4 degrees C, particularly with respect to the glycosaminoglycans and glycoproteins. In view of this, a detailed study was carried out on the changes in the membrane proteins, glycosaminoglycans (GAG), carbohydrate components of glycoproteins, cholesterol, phospholipids and vitamin E in blood stored in glass bottles and a di-(2-ethyl hexyl) phthalate (DEHP) plasticized PVC bag (Penpol blood bag). Blood was collected in CPDA solution in glass bottles and in Penpol blood bags and kept at 4 +/- 1 degrees C. Analysis was made immediately after blood collection and after 28 and 42 days. Significant increase in the total protein in the erythrocyte membrane was observed during storage of whole blood in glass bottles and Penpol blood bag at 4 degrees C. This increase was progressively more with increase in storage time. Significant changes were also observed in GAG, carbohydrate components of glycoproteins, cholesterol, phospholipids and vitamin E in the erythrocyte membrane under these conditions. The protein:GAG ratio, protein:carbohydrate ratio, cholesterol:phospholipid ratio as well as protein:lipid ratio showed significant increase in the membrane. The extent of these changes was lower in the Penpol bag, indicating the stabilizing effect of DEHP on the erythrocyte membrane.

Clinical trial of benidipine in mild to moderate hypertension.

Thirty four patients with mild to moderate hypertension, were put on benidipine 4 mg/day after two weeks of placebo therapy. Twenty five patients completed the trial successfully for 4 mg benidipine. The blood pressure of 20 patients was controlled with benidipine 4 mg/day (effective rate 80%). Five patients with unsatisfactory control on 4 mg/day benidipine were put on 8 mg/day. Four of them were controlled and one was considered as failure (effective rate 80%). Most of the patients tolerated the drug well. Three patients had mild side effects like headache and heaviness in the head. One of them also had puffiness of face and body (on benidipine 8 mg/day) and was withdrawn from the study. One patient had mild constipation. We conclude that benidipine is well tolerated in the dose of 4-8 mg/day and is an effective antihypertensive agent for treatment of patients with mild to moderate hypertension.

Synthesis of NAD+ in erythrocytes incubated with nicotinic acid and the effect of di-(2-ethyl hexyl) phthalate (DEHP).

Synthesis of NAD+ from nicotinic acid by erythrocytes incubated in SAGM phosphate solution and effect of di-[2-ethyl hexyl] phthalate, a plasticizer commonly used in PVC blood/component storage bags, on this synthesis was studied. Erythrocytes are able to synthesise NAD+ in SAGM (sodium chloride, adenine, glucose, mannitol) phosphate solution and this synthesis was more in the presence of added nicotinic acid (optimum concentration 1 mM). The level of NAD+ decreased when the incubation period was increased from 24 to 48 hr. Glutamine had a deleterious effect on this synthesis, possibly due to the decrease in pH. Di-[2-ethyl hexyl] phthalate had an inhibitory effect on NAD+ synthesis when the cells were incubated in SAGM phosphate solution, either alone or in the presence of added nicotinic acid. There was significant decrease in the release of potassium and haemoglobin from the cells in the presence of nicotinic acid, indicating increased red cell stability.

Effect of DEHP [di-(2-ethyl hexyl) phthalate] on lipid peroxidation in liver in rats and in primary cultures of rat hepatocytes.

The effect of DEHP [di-(2-ethly hexyl) phthalate] on lipid peroxidation in the liver in rats and in primary cultures of rat hepatocytes incubated with it was studied. The doses of DEHP used in this study corresponded to the low levels of this substance leaching into blood stored in DEHP plasticised PVC bags. Increased activity of superoxide dismutase (SOD) and catalase, increased concentration of malondialdehyde (MDA) and conjugated dienes and decrease in the concentration of glutathione and vitamin E have been observed in the liver of rats administered DEHP. Primary cultures of rat hepatocytes incubated with DEHP also showed increase in the activity of these enzymes, increase in the concentration of MDA and decrease in vitamin E. These results indicate that DEHP promotes lipid peroxidation. Incorporation of vitamin E along with DEHP into the culture medium containing hepatocytes counteracted these effects.

Toxic effect of systemic administration of low doses of the plasticizer di-(2-ethyl hexyl) phthalate [DEHP] in rats.

DEHP [di-(2 ethyl hexyl) phthalate], a widely used plasticizer in blood storage bags, leaches out in appreciable amounts into blood (about 10 mg/100 ml) resulting in exposure of recipients of blood transfusion to this compound. Various reports indicate the toxicity of DEHP, particularly in liver and reproductive organs but all these studies used large doses (up to 2 g or more/Kg body weight) and oral route of administration which are not relevant to the intravenous administration during blood transfusion or the low amounts present in blood. We have studied changes in the activity of some important enzymes-gamma-GT, ALT, CPK, LDH, alkaline phosphatase, acid phosphatase, beta-glucuronidase and few other parameters like vitamin E, glutathione, serum albumin etc in rats administered low doses of DEHP (corresponding to transfusion of 2, 4, 6 and 10 units of blood). Histopathology of the organs has also been carried out. The results obtained indicate no serious toxic effects for DEHP at the level present in blood stored in DEHP plasticized blood bags as evidenced by the lack of any significant alteration in most of the biochemical parameters studied. Even in those cases where there was alteration (for e.g., decrease in the level of vitamin E) 24 hr after administration of DEHP, it returned to near normal level with in 72 hr to 7 days. No histopathological changes were observed in any of the organs at these levels of DEHP. It is concluded that DEHP did not cause any serious toxic effect even at doses corresponding to transfusion of several units of blood in a recipient.

Evidence for free-radical mediated injury during coronary artery bypass surgery.

Prolonged global ischemia followed by reperfusion during coronary artery bypass graft (CABG) surgery leads to a burst of oxygen free-radical generation and subsequent myocardial impairment. Blood samples were collected at different time periods from the right atrium of patients (n = 55) who underwent CABG surgery. Free-radical activity was measured by assays for thiobarbituric acid reactive species (TBARS), mainly malondialdehyde (MDA), at zero minute (before aortic cross-clamp), one minute and 10 minutes after declamping i.e. reperfusion. There was a significant increase (p < 0.001) in the level of MDA at one minute of reperfusion which continued to be elevated up to 10 minutes. This increase was significantly correlated with aortic cross-clamp time (r = 0.525, p < 0.05). The activity of CPK and CPK-MB was raised by two fold (p < 0.001) after release of the clamp. Plasma Troponin-T level was estimated in 10 patients during CABG surgery and a significant increase (p < 0.01) was observed at one minute as well as 10 minutes of reperfusion which correlated well (r = 0.81, p < 0.01) with the severity of ischemia. The concomitant rise in TBARS (marker of free-radical activity), CPK, CPK-MB and Troponin-T (indicators of myocardial damage) on reperfusion implies that there is free-radical mediated damage to the cardiac membrane during CABG surgery.

Dyslipidemia in patients with chronic renal failure and in renal transplant patients.

Indian studies on lipid profile abnormalities in chronic renal failure (CRF) have varied from no abnormalities at all to significant abnormality (hypertriglyceridemia and reduced HDL) as described in the Western literature. Moreover, there is no Indian study on the effect of renal transplantation on the abnormal lipid profile of CRF. The aim of our study was to determine the lipid profile of CRF patients on conservative treatment, end stage renal disease (ESRD) patients on maintenance hemodialysis (HD) treatment and renal transplant patients. We also looked at the effect of fish oil rich in polyunsaturated fatty acids (Max-EPA) on hypertriglyceridemia of CRF. The study included 4 groups; Gp I: control subjects (n = 9, age = 30 +/- 5 yrs), Gp II: CRF patients on conservative treatment (n = 9, age = 49 +/- 17 yrs), Gp III: ESRD patients on HD for at least 3 months (n = 19, age = 53 +/- 9 yrs), Gp IV: 3 months post-renal transplant patients (n = 9, age 31 +/- 11 yrs). The lipids and lipoproteins analysed include total cholesterol, HDL, LDL, triglycerides, Apo A1 and Apo B. It was observed that in Gp II patients triglycerides were significantly elevated (p < .05) and Apo A1/Apo B significantly abnormally lower (p < .001) compared to Gp I. In Gp IV patients, there was no significant difference in lipid profile compared to Gp I. With the use of Max-EPA in 5 patients with hypertriglyceridemia, there was a significant improvement in hypertriglyceridemia (p < .05). Our study suggests: 1) significant hypertriglyceridemia does develop in a majority of CRF patients. The abnormality probably improves with dialysis treatment and renal transplantation. 2) A lower Apo A1/Apo B ratio in CRF patients may account for higher risk of atherosclerosis. 3) Fish oil rich in polyunsaturated fatty acids improves hypertriglyceridemia of CRF.

Isolation of Legionella pneumophila from patients of respiratory tract disease & environmental samples.

In this preliminary study 45 patients suffering from pneumonia were screened for L. pneumophila. Also 17 random environmental samples viz., water from cooling system used for air-conditioning units and samples from respirator-humidifier were screened. Four of 45 (9%) clinical specimens and 13 of 17 (76%) environmental samples showed presence of L. pneumophila. This is the first report from India of the isolation on L. pneumophila.