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Biomedical subjects

M Mukherjee

Publications and source records attributed to M Mukherjee.

At least 55 records · Page 3Linked to original sources
PubMed1999

Enhancement of lipoprotein lipase activity by tissue factor pathway inhibitor.

The effect of a basic synthetic peptide, representing the C-terminal region of tissue factor pathway inhibitor (TFPI - Lys254 - Met276), as well as that of the whole protein, on the activity of lipoprotein lipase (LPL) is described. The activity of bovine LPL was measured by chromogenic assay using a water-soluble chromogenic substrate, p-nitrophenyl butyrate. Five and 10 microM concentrations of the peptide increased Vmax of bovine LPL by 48.9% and 85.6% respectively as compared with the buffer control without affecting Km. Poly l-lysine, though positively charged did not have any effect, suggesting the importance of the amino acid sequence of the test peptide. On the other hand, 0.25, 0.5 and 1.0 mM n-butyric acid - a product of LPL catalysis in the chromogenic assay, when added to the incubation mixture decreased Vmax non competitively by 22.8%, 40.4% and 63% respectively as compared with buffer control, confirming the known product inhibition of LPL. A 100-fold molar excess of n-butyric acid produced inhibition of the LPL reaction as compared with the synthetic peptide which produced potentiation, suggesting a 1:100 stoichiometric interaction of the peptide with n-butyric acid. At a fixed concentration of 0.25 mM substrate, 10 nM full length recombinant TFPI, containing the basic C-terminal domain, increased velocity of LPL reaction by 39.4% as compared with buffer control. The same concentration of two-domain recombinant TFPI (TFPI1-160) had no effect. It is possible that negatively charged n-butyric acid is sequestered by the positively charged peptide or the basic region of recombinant full length TFPI. Relieving of product inhibition could then be a possible mechanism of the observed potentiation of bovine LPL activity by the basic peptide or full length recombinant TFPI. The 39.4% increase in reaction velocity of LPL catalysis produced by 10 nM full length recombinant TFPI was comparable to 38.9% increase produced by 5 microM of the basic peptide under the same conditions. A further increase of 78.7% was brought about by 10 microM concentration of the same peptide. The reason for about 500-fold increase in the potency of the whole protein as compared with that of the peptide is not clear. It is possible that in its tertiary conformational state, the whole protein is able to sequester product and relieve product inhibition more effectively than the short linear peptide. Rabbit polyclonal antiserum against the basic peptide partially inhibited LPL activity of human post heparin plasma, measured by radioenzymatic assay using triolein substrate. Since post heparin plasma contains full length TFPI, binding of the added antibody to its basic C-terminus and hence the relative unavailability of latter for product sequestration (oleic acid in this case) could explain the observed inhibition of human LPL activity by antibody against the peptide. Thus by enhancing lipase activity, full length TFPI may facilitate hydrolysis of triglyceride and concomitantly lower factor VII coagulant activity as demonstrated earlier, particularly after heparin injection when both TFPI and LPL are released in circulation.

Amino Acid Sequence↗
PubMed1998

Development of in vitro screening system for assessment of antifilarial activity of compounds.

Evaluation of antifilarial activity of new potential agents in vivo is extremely time consuming and uneconomic. In the present study effort has been made to develop an in vitro screening method using Acanthocheilonema viteae, a subcutaneously dwelling rodent filariid with anaerobic metabolic characteristics like human filariids, W. Bancrofti/Brugia malayi as test parasite. Motility test and tetrazolium (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide, MTT) based colorimetric assay were used as parameters in in vitro assay. Results showed that 92.3% of compounds (in vivo active) could be picked up in the in vitro assay when both adults and microfilarae (mf) were used simultaneously. Mf and adult stages separately detected, respectively, 84.6 and 69.2% of in vivo active compounds. The adults and mf separately and both the life stages together exhibited, respectively, 80.0, 50.0 and 80.0% false positive results in the in vitro test with in vivo inactive compounds. It is felt that mf stage when used in in vitro test using motility and MTT assays as parameters would be useful in primary screening of new potential filaricides.

Animals↗
PubMed1998

Unexpected electrophoretic migration of RNA with different 3' termini causes a RNA sizing ambiguity that can be resolved using nuclease P1-generated sequencing ladders.

It has been widely believed that the electrophoretic migration difference of otherwise identical RNAs with a P versus OH terminus would be the same as occurs for DNA, a fairly reproducible approximately 1/2 nucleotide (nt) offset. RNA with a 5'-OH indeed migrates </=1 nt slower than if it had a 5'-P. Surprisingly, however, RNA with a 3'-OH terminus (generated by many cellular RNases of interest) migrates anywhere from approximately 1/4 to approximately 2 nts slower than the otherwise identical molecule with a 3'-P or 2', 3'-cyclic-P terminus (as present on standard RNase-generated sequencing ladders). This previously unrecognized variability in electrophoretic migration offset causes a 1-2 nt ambiguity in a commonly used method of RNA size determination. We also show two ways to overcome this problem and enable rigorous sizing of 3'-OH terminating RNAs. Most convenient is to use sequencing standards generated by nuclease P1, which is generally sequence-nonspecific but we show becomes G-specific or A-preferential under certain reaction conditions.

Animals↗
PubMed1998

Cellular immune response of Mastomys and gerbils in experimental filariasis.

OBJECTIVE: To determine mitogenic and antigen-specific cellular immune responses of two species of rodents, viz. Meriones unguiculatus and Mastomys coucha to assess the usefulness of the A. viteae/Mastomys model for cellular immune studies in experimental filariasis. METHODS: Lymphocyte blast transformation test (LTT) using spleen cells of normal and A. viteae infected animals. RESULTS: The proliferative response of gerbils was much higher than that of Mastomys to both ConA and filarial antigens. Cells of both species of rodents did not respond to microfilarial (mf) antigen, however, their mitogenic response differed during infection. Some degree of nonspecific suppression was observed in gerbils during prepatent and patent stages of infection, while Mastomys revealed highest proliferation during patent microfilaraemia. Mastomys cells did not respond to adult or mf antigen, while adult-specific proliferation was detected in the case of gerbils. CONCLUSION: The A. viteae/gerbil model shows more similarity to human filarial infection regarding cellular immune response. Markedly low responsiveness of a high percentage of Mastomys and wide variations in the cellular response to nonspecific mitogen limit the usefulness of Mastomys coucha in immunological studies, especially cellular immunity.

Animals↗
PubMed1998

Chronic fatigue syndrome: physical and cardiovascular deconditioning.

We investigated whether chronic fatigue syndrome (CFS) patients have physical and/or cardiovascular de-conditioning, in 273 CFS patients and 72 healthy controls. We used laboratory tests to assess haematological, biochemical, endocrinological and immunological systems. The cardiovascular system was assessed by echocardiography and carotid echography. Body composition was determined by dual energy X-ray absorptiometry (DEXA). CFS patients had smaller left ventricular end systolic (p < 0.001) and diastolic (p = 0.008) dimensions but thinner posterior walls (p = 0.02) than corresponding values in healthy controls. Left ventricular mass was also reduced in CFS patients (p = 0.006). Both maximum (p < 0.001) and minimum (p < 0.008) diameter of the carotid artery were smaller in CFS patients. The laboratory screening tests showed significant differences in serum albumin (p = 0.05), phosphate (p = 0.02), HDL-cholesterol (p = 0.03), HDL:total cholesterol ratio (p = 0.01), triglycerides (p = 0.02), neutrophils (p = 0.01) and thyroid-stimulating hormone (p = 0.04) between CFS patients and controls. Male CFS patients had an increased percentage of fat mass compared with healthy male subjects (p = 0.02). This large group of CFS patients had evidence of physical and cardiovascular de-conditioning, suggesting that in these patients a graded exercise programme could lead to physical reconditioning and could increase their ability to perform physical activities.

Adult↗
PubMed1998

Thrombogenicity of heparin and non-heparin bound arterial prostheses: an in vitro evaluation.

The effect on graft thrombogenicity of binding heparin to the luminal surface of prosthetic arterial grafts was investigated. Venous blood was obtained from healthy volunteers and exposed for 30 minutes to tubular segments of standard knitted dacron, polytetrafluoroethylene (PTFE) and a recently introduced heparin-bound knitted dacron graft. After this exposure the fibrinogen level of each sample was measured. The median (range) fibrinogen levels (expressed as a percentage of that in unexposed blood samples) were: standard dacron 3.5% (0-5.4%); PTFE 95.5% (0-121.1%); and heparin-bound dacron 79.8% (3.8-109.6%). Fibrinogen levels in the standard dacron group were significantly less than that of the PTFE and heparin-bound dacron groups (P < 0.05). No significant difference was found between the fibrinogen levels of the PTFE and heparin-bound dacron groups (P = 0.35). These findings suggest that heparin binding significantly reduces fibrinogen consumption and hence may reduce graft thrombogenicity.

Anticoagulants↗
PubMed1998

Central cooling effects in patients with hypercholesterolaemia.

1. A prospective study has been carried out, and 68 patients with hypercholesterolaemia have been investigated to study the effects of central cooling on serum lipid levels. 2. Central cooling was obtained by the exposure of the whole body to cold water. All patients were trained to gradually reduce the water temperature from 22 to 14 degrees C and to increase the time of exposure from 5 to 20 min over a period of 90 days. The 33 male and 35 female patients were aged between 40 and 60 years at entry with total cholesterol of 6.0 mmol/l or greater and low-density lipoprotein (LDL)-cholesterol of 4. 0 mmol/l or greater. Thyroid-stimulating hormone, free thyroxine (FT4), total T3, total cholesterol, LDL-cholesterol, high-density lipoprotein (HDL)-cholesterol, triacylglycerols and total fat mass (determined by dual-energy X-ray absorptiometry scan) were obtained at baseline and after 3 months treatment with hydrotherapy. 3. Central cooling obtained by hydrotherapy results in a median fall in tympanic temperature from 0.2 degrees C (P<0.001) to 0.8 degrees C (P<0.001). We have observed in these patients a significant reduction in total cholesterol (-0.2 mmol/l, P=0.006) and LDL-cholesterol (-0.2 mmol/l, P=0.004). Serum FT4 level was higher than baseline results in 30 of these hypercholesterolaemic patients (15.5 pmol/l to 17.3 pmol/l) and there was no significant change in serum thyroid-stimulating hormone and total T3. 4. In conclusion, in our patients with hypercholesterolaemia we have observed a significant reduction of total cholesterol and LDL-cholesterol after body temperature regulation.

Adult↗
PubMed1998

Association of overall adiposity rather than body mass index with lipids and procoagulant factors.

The association between obesity and risk of coronary artery disease is well established. The distribution of body fat was shown to be related to serum lipids and lipoproteins in a group of healthy men, but the association between body fat and haemostatic factors is less clear. The aim of the present study was to determine the association of overall adiposity (OVRAD, percent total fat mass contributing to body weight) and body mass index (BMI, weight/height2) with lipids and haemostatic factors in order to evaluate which of these was more associated with circulating procoagulant factors. The total fat mass was estimated by dual-energy X-ray absorptiometry (DEXA) and OVRAD computed for 28 male and 36 healthy female subjects, whose median age were 44.2 years and 48.4 years respectively. In addition, the BMI was computed for each of them from their weight and height measurements. Fasting samples were analysed for serum lipids (total, HDL- and LDL-cholesterol and triglyceride) and plasma fibrinogen, factor VII coagulant (FVII:C) activity, tissue plasminogen activator (t-PA) and plasminogen activator inhibitor-1 (PAI-1) activities. The men and women had similar median BMI (23.9 kg/m2 and 23.1 kg/m2 respectively), but the median fat mass of women (19.6 kg) was higher than that of men (16.9 kg). Age, BMI and OVRAD exhibited statistically significant correlations with lipids and haemostatic factors in both men and women. However, when BMI was adjusted for age and OVRAD, the statistically significant associations were no longer apparent in men or women. In contrast, OVRAD adjusted for age and BMI still exhibited statistically significant associations with FVII:C activity (R = 0.38, p = 0.05), triglyceride (R = 0.51, p = 0.008), LDL-cholesterol (R = 0.45, p = 0.02) and HDL/Total cholesterol ratio (R = -0.63, p <0.001). It is concluded that OVRAD, a fat mass-based index, rather than BMI, a weight-height based index, is better associated with circulating coronary risk factors.

Adult↗
PubMed1997

Importance of laccase in vegetative growth of pleurotus Florida.

Mycelial culture of Pleurotus florida produced highest extracellular laccase in optimum growth medium. At least two laccases (L(inf1) and L(inf2)) were shown to be present in the culture filtrate. Low-laccase-yielding mutants with impaired L(inf2) activity had poor mycelial growth and could not form fruit body, whereas the revertants from the same mutants were similar to the parent in mycelial growth and fruit body formation.

Journal Article↗
PubMed1997

The routine determination of the endogenous thrombin potential, first results in different forms of hyper- and hypocoagulability.

The area under the thrombin generation curve (the endogenous thrombin potential; ETP) has been proposed as a parameter for plasma-based hypercoagulability and to monitor anticoagulant treatment. We present an ETP assay for the routine laboratory using a centrifugal analyser. Throughput is 30 samples/h, within and between run imprecision is 4-5.6%. Suitable substrates were developed for the ranges of 10-500% and 2-100% of normal. Independent of tissue factor concentration (if > 4 pM), the normal value of the extrinsic ETP is 384.8 +/- 51.7 nM.min. The intrinsic ETP, triggered by ellagic acid, is 414 +/- 41 nM.min. The ETP is decreased to 15 and 35% of normal by oral anticoagulation (INR 2.5-4.0) and by heparin administration (APTT 1.5-2.5 x control). The ETP is increased in untreated subjects with congenital antithrombin deficiency and in women using oral contraceptives. In deep vein thrombosis (phlebographically confirmed), it is increased by 29.4% (extrinsic) and 53% (intrinsic). In (angiographically assessed) coronary artery disease the increase is by 10% and 17% respectively.

Adolescent↗
PubMed1997

Decrease in factor VII coagulant activity during percutaneous transluminal coronary angioplasty by heparin-mediated lipolytic action.

Levels of factor VII coagulant activity (FVII:C) and two-chain factor VIIa antigen (FVIIa:Ag) were measured in ten patients before and up to 6 h after receiving a bolus of heparin during percutaneous transluminal coronary angioplasty (PTCA). A significant and sustained post-heparin fall in the level of FVII:C was observed (approximately 30%) without any change in the level of FVIIa:Ag. The level of tissue factor antigen within the circulation remained unchanged. The observed decrease in FVII:C coincided with a significant decrease in triglyceride levels presumably due to lipoprotein and hepatic lipase released by the heparin. These findings appear to demonstrate a lipid (triglyceride) dependence of FVII:C. Thus, heparin may act indirectly as antithrombotic agent by limiting a lipid-dependent activation of the extrinsic pathway of coagulation.

Aged↗
PubMed1997

Optimization of test conditions for development of MTT as in vitro screen.

The quick and easy method of tetrazolium based colorimetric assay with MTT [3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] was used to test the viability of the adult parasites of a rodent filariid Acanthocheilonema viteae in vitro. The ideal conditions required for antifilarial screening were determined by correlating the MTT reduction ability of worms with their size and age in the vertebrate host, also the duration of incubation and temperature of the in vitro culture. It was observed that the worms collected from the host after 90 days of L3 (infective larvae) exposure were not suitable for in vitro screen as they could not reduce MTT to that extent as the worms of early infection. Healthy and full grown worms and also those incubated at 37 degrees C for 16 hr or more caused maximum MTT reduction. Thus, it is recommended to select healthy adult filariids of proper age and size (male > 3.5 cm; female > 7.0 cm). The incubation temperature of the in vitro culture system needs to be adjusted to 37 degrees C and parasites might be exposed to drugs upto 24 hr without much alteration in MTT reduction of untreated controls.

Animals↗
PubMed1997

Influence of haloperidol on testicular functions in rat.

The study involved exploration of the role of dopamine antagonist haloperidol on the testicular functions of rat. Chronic administration of haloperidol (0.2 mg/kg/day/sc for 21 days) caused significant increase in brain DA and serum prolactin. At testicular level the treatment revealed atrophic degeneration of seminiferous epithelium indicating suppression of hypophyseal gonadotrophins and proves importance of dopaminergic control over prolactin release for normal functions of male gonad.

Animals↗
PubMed1996

Interference of an activity assay of tissue-type plasminogen activator in human plasma by endogenous factors.

The estimation of the activity of circulating tissue-type plasminogen activator (t-PA) using the Coaset t-PA or Spectrolyse/fibrin t-PA reagent kits involves incubation of plasma with excess plasminogen in the presence of human fibrin fragments, and detection of the plasmin generated by an end-point amidolytic assay. We examined the interference of endogenous inhibitors such as plasminogen activator inhibitor-1 (PAI-1) and alpha 2-antiplasmin, and that of an endogenous activator namely single-chain urokinase-type plasminogen activator (scu-PA) on the Coaset t-PA assay. An extra acidification of the samples already collected into an acidified anticoagulant raised the mean Coaset t-PA activity of 15 samples from 1.39 +/- 0.25 (mean +/- SEM) to 1.71 +/- 0.27 (P < 0.001). In twelve of these samples, the PAI-1 and alpha 2-antiplasmin activities were determined. The increase in the t-PA activity by acidification was found to correlate inversely with PAI-1 (r = -0.66; n = 12; P = 0.019) but not with alpha 2-antiplasmin, demonstrating that the extra acidification step released t-PA from the t-PA-PAI-1 complex, but had no influence on the endogenous alpha 2-antiplasmin in the assay system. Incubation with monoclonal antibody against alpha 2-antiplasmin increased the Coaset t-PA activity in a concentration-dependent manner, linearly up to 2 micrograms/ml (final) of the antibody, irrespective of the extra acidification, the maximal rise being 41.5% and 19.7% in an in-house and commercial plasma pools respectively. In contrast to alpha 2-antiplasmin antibody, monoclonal antibody against scu-PA decreased the Coaset t-PA activity in the in-house and commercial plasma pools again in a concentration-related manner demonstrating that scu-PA in addition to t-PA was being measured by this method. Incubation with an irrelevant antibody (anti-DNA monoclonal antibody) did not affect the Coaset t-PA values. The mean Coaset t-PA activity of 40 subjects decreased from 1.76 +/- 0.10 (mean +/- SEM) to 0.73 +/- 0.07 when incubated with 8 micrograms/ml (final) monoclonal antibody against scu-PA (P < 0.001). The 't-PA' activity in the absence of scu-PA antibody correlated with the u-PA antigen (r = 0.53; n = 40; P < 0.001), and the activity in the presence of scu-PA antibody correlated with the t-PA activity measured by the bio-functional immunosorbent assay (r = 0.86; n = 40; P < 0.001). Hence, unless the Coaset t-PA activity is measured with appropriate amounts of antibodies to alpha 2-antiplasmin and scu-PA, it is best to qualify the activity measured by this method as being that of total plasminogen activators.

Antibodies, Monoclonal↗
PubMed1996

Identification of persons at high risk of coronary heart disease--a mathematical formula based on biochemical, anthropometric and clinical markers.

A reliable method for identification of the subset of population predisposed to coronary heart disease (CHD) would aid a targetted implementation of intervention strategies. To this end, a mathematical formula was developed based on stepwise linear discriminant analysis. Age, body mass index, the number of associated coronary risk factors and a large number of biochemical markers were analysed by computerised discriminant analysis on a test sample of 203 subjects. Unstandardised canonical discriminant coefficients of statistically significant independent variables were used to derive the total discriminant score or the 'risk score'. The 'low-risk' persons not in need of immediate preventive measures of CHD could be distinguished from the 'high-risk' individuals with an almost 90% correctness. As compared with the existing methods such as clinical evaluation and cardiac stress test, the risk scores derived by the new method, and based chiefly on blood markers besides clinical and anthropometric variables, appeared to correctly predict the future coronary episodes in members of the test sample selected at random. The risk scores were also tested on a new sample of 50 subjects; while low scores were not associated with CHD, high scores in some patients were associated with myocardial ischemia. It appears that the preventive measures of CHD may be directed at people who have no clinical manifestations of CHD, but whose risk scores are greater than 0.1. On the other hand, if the score is less than -1.0, immediate preventive measures may not be necessary. If the score is between -1.0 and 0.1 (borderline), no immediate action may be taken but the score may be determined after six months, and action taken accordingly.

Age Factors↗
PubMed1996

Association of antibodies to heat-shock protein-65 with percutaneous transluminal coronary angioplasty and subsequent restenosis.

Heat-shock protein (HSP)-65 of mycobacterial origin has been implicated in the mediation of atherosclerosis by immune mechanisms. Any role of HSP-65 in mediating restenosis is however not clear. We determined the anti-HSP-65 antibodies in 28 patients, 25 male and 3 female, aged 35 to 78 years, with coronary artery disease (CAD) and undergoing percutaneous transluminal coronary angioplasty (PTCA). Of the 28 patients, 12 suffered restenosis five to seven months later. The serum levels of antibody were measured at baseline, immediately after PTCA, before discharge from the hospital, and at 6 weeks, 3 and 6 months after the performance of PTCA. The antibody levels were expressed in OD U/log titre, as explained in the text, and termed estimated OD or EOD. The control group consisted of 29 healthy volunteers, 16 male and 13 female, aged 24 to 59 years. The mean EOD of the patients at baseline was higher than that of the controls (0.60 +/- 0.12, SD; 95% CI 0.56-0.64 compared with 0.53 +/- 0.13; 0.48-0.58; p < 0.01). There were no correlation between age and EOD of patients, controls or both taken together, ruling out the influence of age on the EOD changes in the given age range. The mean antibody levels of the patients with CAD were similar whether or not they suffered subsequent restenosis (0.61 +/- 0.15; 0.53-0.69 in the patency group, and 0.60 +/- 0.10; 0.54-.066 in the restenosis group). However, the patients who did not develop restenosis had a drop in their antibody levels immediately after PTCA (0.51 +/- 0.14; 0.48-0.55; 2-tailed p = 0.029), and at discharge from the hospital (0.52 +/- 0.15; 0.44-0.60; p = 0.036) as compared with the baseline. This decrease of antibodies was not observed in the restenosis group. Furthermore, the anti-HSP-65 antibody levels remained slightly low throughout the 6-month follow-up in the patients with patent coronaries as compared with patients who restenosed, but the decrease was statistically not significant at 5% level at any stage. Besides the anti-HSP-65 antibodies, the levels of anticardiolipin (ACL) antibody were also measured in all the patients. The levels of the ACL antibody were found to be within the normal range before and at all stages after PTCA, ruling out the PTCA-associated change in the anti-HSP-65 antibody as a non-specific occurrence. Thus, a drop in the level of antibody against HSP-65 after PTCA seemed to be associated with a favourable outcome, and may serve as a useful prognostic marker of coronary angioplasty.

Adult↗