Junctures to the therapeutic goal of diabetes mellitus: Experience in a tertiary care hospital of Kolkata.

INTRODUCTION: The World Health Organization has declared India as the "diabetic capital" of the world. In controlling of such chronic, mostly asymptomatic disease, patients' role can't be overemphasized. AIMS: To assess the level of compliance to anti-diabetic therapies and to ascertain the determinants of non-compliance, if any. MATERIALS AND METHODS: A cross-sectional observational study was conducted for 3 months in a diabetic clinic of R G Kar Medical College and Hospital, Kolkata. Data were collected by interviewing the patients, examining their prescriptions and laboratory reports and anthropometry after obtaining informed consent. RESULTS: Blood report at the point of data collection revealed controlled glucose homeostasis in 38.93% patients but evaluation of past 3 months report showed only 24.3% had control over hyperglycemia. Glycemic control was seen to be positively related to short duration of disease, compliance to therapies, and high knowledge about diabetes. Compliance to therapies found in 32.22% of study subjects was in turn associated with short duration of disease. House-wives showed poor compliance; insulin treatment with or without oral-anti-diabetic agent showed better compliance. Knowledge of diabetes was significantly high among higher educated; poor among women, house-wives, and rural people. CONCLUSION: Patient-providers collaboration is to be developed through a patient-centered care model based on the mutual responsibility of both so that each patient is considered in the mesh of his/her other goals of life and helped to promote empowerment to take informed decision for behavioral change conducive to control the disease.

A study on habits of tobacco use among medical and non-medical students of Kolkata.

BACKGROUND: Age-old practice "using tobacco" is a well known major global concern as it victimizes all its lovers by a host of chronic noncommunicable diseases including cancer; all develop very slowly and silently, and can cause premature death. OBJECTIVES: To assess the pattern of tobacco use among the medical and nonmedical college students. MATERIALS AND METHODS: A cross-sectional descriptive study was carried out in Kolkata collecting anonymous data from 515 medical and 349 nonmedical college students of two medical and two general colleges, selected randomly. RESULT: Overall prevalence of tobacco use (18.3% vs 43.6%) and smoking (14.9% vs 40.7%) were significantly less in medical subjects, both across the sex and years of study. Lower rate of tobacco adoption at college level, higher quitting rate, correct knowledge regarding uselessness of filter attached with cigarette, and ill-effects of tobacco consumption were observed among medical participants. More nonmedical subjects were increasingly smoking compared to medical students. Filter-tipped cigarette was the top choice, and smoking was more prevalent mode of use among the nonmedical participants, most (62.3%) of whom were mild users. Curiosity was the top influencing factor for the initiation of tobacco use and two-third users wanted to quit. CONCLUSION: Although the mortal habits was comparatively less among medical students, the medical environment seemed to fail to curb the dreadful practice totally. Thereby it can be recommended that active behavior-changing communication is required for all sections of the society to tear out the social root of the problem instead of unimpressive vague health warnings in vogue.

Socioeconomic consequences of HIV/AIDS in the family system.

INTRODUCTION: HIV/AIDS can lead to poverty affecting particularly women and young people and can halt or reverse socioeconomic development of a country. OBJECTIVE: The objective of this study was to assess the socioeconomic consequences of HIV/AIDS within the family. MATERIALS AND METHODS: A cross-sectional descriptive study was carried out among patients admitted in in-patient department and those attending integrated counseling and testing centre (ICTC) of School of Tropical Medicine, Kolkata. Data were gathered by interviewing the patients by using a predesigned questionnaire. RESULTS: For prolonged duration and severity of disease, higher proportion of indoor patients reported loss of job, decreased family income, increased expenditure for care seeking, and faced greater economic consequences, reflected by selling assets. Loss of job was mainly due to illness (86.8%), disclosure of sero-status (13.2%), and predominantly among skilled workers. Assets were sold mainly to meet the cost of own illness for indoor patients, but more to meet the expenditure for husband's illness, in the case of ICTC patients. High school dropout seen in both groups was mainly due to economic reasons. HIV/AIDS status was known to other members of family for 84.8% of indoor patients out of which 15.4% experienced rejection by family members. Out of 72 ever married women indoor patients whose in-laws were aware of their HIV/AIDS status, 41.7%, 40.9%, and 33.33% reportedly were blamed for spouse's illness, and had strained relation with in-laws and spouse, respectively. CONCLUSION: Intensive behavior change communication and provision of care and support are required to curb AIDS-related stigma, discrimination, and to maintain physical, mental, and social wellbeing of people living with HIV/AIDS.

Construction and characterization of a series of vectors for Schizosaccharomyces pombe.

A set of vectors was created to allow cloning and expression studies in Schizosaccharomyces pombe. These vectors had a uniform backbone with an efficient Sz. pombe ARS, ARS3002, but different selectable markers--his3+, leu1+, ade6+ and ura4+. The vectors functioned efficiently as autonomously replicating plasmids that could also be converted into integrating vectors. The ura4+-containing vector was used to construct a Sz. pombe genomic library.

Evaluation of Ophthacare eye drops--a herbal formulation in the management of various ophthalmic disorders.

An open prospective multicentre clinical trial was conducted in patients suffering from various ophthalmic disorders namely, conjunctivitis, conjunctival xerosis (dry eye), acute dacryocystitis, degenerative conditions (pterygium or pinguecula) and postoperative cataract patients with a herbal eye drop preparation (Ophthacare) containing basic principles of different herbs which have been conventionally used in the Ayurvedic system of medicine since time immemorial. These include Carum copticum, Terminalia belirica, Emblica officinalis, Curcuma longa, Ocimum sanctum, Cinnamomum camphora, Rosa damascena and meldespumapum. These herbs reportedly possess antiinfective and antiinflammatory properties. The present study was undertaken to elucidate the role of this herbal product in a variety of eye ailments. Side effects, if any, were noted during the study. An improvement was observed with the treatment of the herbal eye drop treatment in most cases. There were no side effects observed during the course of the study and the eye drop was well tolerated by the patients. The herbal eye drop Ophthacare has a useful role in a variety of infective, inflammatory and degenerative ophthalmic disorders.

Identification of two transmembrane regions and a cytosolic domain of rat mitochondrial glycerophosphate acyltransferase.

The topography of rat glycerophosphate acyltransferase (GAT) in the transverse plane of the mitochondrial outer membrane (MOM) was investigated. Computer analysis of the amino acid (aa) sequence derived from rat mitochondrial GAT cDNA (GenBanktrade mark accession nos. and ) predicts the presence of two possible transmembrane domains (aa 473-493 and 574-594) separated by an 80-aa stretch (aa 494-573). To determine the actual orientation of the native protein, we prepared anti-peptide antibodies to three regions: one in between (aa 543-559) and the other two (aa 420-435 and 726-740) flanking the two putative transmembrane regions. Both immunoreaction and immunoprecipitation experiments employing intact and solubilized mitochondria indicate that regions on the N- and C-terminal sides of the transmembrane regions are sequestered on the inner surface of the MOM, while the region between the transmembrane domains is present on the cytosolic face of the MOM. Additionally, two green fluorescent protein (GFP) fusion proteins consisting of full-length GAT fused to GFP at either the C terminus or inserted 115 amino acids from the N terminus were also constructed to determine the orientation of the N and C termini. COS-1 cells expressing these fusion proteins were fractionated to obtain mitochondria. Protease digestion of intact and solubilized COS-1 cell mitochondria revealed that the GFP domains of these fusion proteins are sequestered on the inner side of the MOM. The present findings indicate that GAT is a dual-spanning, transmembrane protein adopting an inverted "U" conformation in the transverse plane of the MOM, where the N and C termini are sequestered on the inner surface of the MOM, while aa 494-573 are exposed on the cytosolic surface of the MOM.

Phosphatidic acid synthesis in mitochondria. Topography of formation and transmembrane migration.

The topography of formation and migration of phosphatidic acid (PA) in the transverse plane of rat liver mitochondrial outer membrane (MOM) were investigated. Isolated mitochondria and microsomes, incubated with sn-glycerol 3-phosphate and an immobilized substrate palmitoyl-CoA-agarose, synthesized both lyso-PA and PA. The mitochondrial and microsomal acylation of glycerophosphate with palmitoyl-CoA-agarose was 80-100% of the values obtained in the presence of free palmitoyl-CoA. In another series of experiments, both free polymyxin B and polymyxin B-agarose stimulated mitochondrial glycerophosphate acyltransferase activity approximately 2-fold. When PA loaded mitochondria were treated with liver fatty acid binding protein, a fifth of the phospholipid left the mitochondria. The amount of exportable PA reduced with the increase in the time of incubation. In another approach, PA-loaded mitochondria were treated with phospholipase A(2). The amount of phospholipase A(2)-sensitive PA reduced when the incubation time was increased. Taken together, the results suggest that lysophosphatidic acid (LPA) and PA are synthesized on the outer surface of the MOM and that PA moves to the inner membrane presumably for cardiolipin formation.

Kinetic analysis of cardiolipin synthase: a membrane enzyme with two glycerophospholipid substrates.

Mitochondrial cardiolipin synthase catalyzes the transfer of a phosphatidyl moiety from phosphatidyl-CMP (PtdCMP) to phosphatidylglycerol (PtdGro) in the presence of specific divalent cations. The synthase was solubilized from Saccharomyces cerevisiae mitochondria and purified about 300-fold. The partially enzyme was part of a medium-size, mixed micelle which had to bind to a foreign substrate/detergent micelle before catalysis could occur. The kinetics of cardiolipin synthase were studied by changing the molar fraction of substrate in the micelles. The enzyme obeyed Michaelis-Menten kinetics in relation to PtdCMP with a Km of 0.03 mol%. PtdGro caused sigmoidal kinetics with a low apparent affinity. It is speculated that it was involved in docking the enzyme to the substrate/detergent micelle. Cardiolipin synthase did not catalyze isotope exchange between [14C]CMP and PtdCMP, virtually excluding a ping-pong catalytic mechanism. Mg2+ stimulated the activity by increasing the turnover number rather than the substrate affinity, a mechanism which was also found for the Co(2+)-activation of rat liver cardiolipin synthase. It is concluded that a direct association of the metal ion and the enzyme forms the active cardiolipin synthase which has a very high affinity for PtdCMP and a lower affinity for PtdGro.

Purification and characterization of glycerophosphate acyltransferase from rat liver mitochondria.

Glycerophosphate acyltransferase (GAT) catalyzes the conversion of sn-glycerol 3-phosphate to lysophosphatidic acid (LPA), the first and committed step of triacylglycerol and phospholipid synthesis. In spite of the important regulatory roles GAT may play in this biosynthetic pathway, little information is available on the structure, biochemical properties, and regulation of GAT from eukaryotic cells. We solubilized GAT from rat liver mitochondrial membranes and purified it to an apparent homogeneity by hydroxylapatite chromatography, preparative isoelectric focusing, and gel filtration. The enzyme is composed of a single polypeptide of 85 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration chromatography of the native protein. The GAT activity was completely lost during the purification procedure and required addition of exogenous phospholipids for its reconstitution. Since a high phospholipid to detergent ratio was needed for full reactivation, it is concluded that GAT requires "lipid boundary" for reconstitution. The ability of different phospholipids to reactivate GAT decreased in the following order: phosphatidylglycerol (PG), phosphatidylethanolamine (PE), phosphatidylcholine (PC), asolectin, phosphatidylinositol (PI), phosphatidylserine (PS), and cardiolipin. 1,2-Dioleoyl derivatives of PG and PE were more effective in reconstituting the GAT activity than corresponding dipalmitoyl derivatives. The GAT activation was further increased by using a combination of PG and PE or PG and PC. Regardless of the phospholipid used for reconstitution, palmitoyl-CoA was the best acyl donor and LPA was the only reaction product.

Cardiolipin is synthesized on the matrix side of the inner membrane in rat liver mitochondria.

In the mitochondrial inner membrane, cardiolipin is a specific lipid component associated with various protein complexes. The assembly of such complexes has been studied, and it seems that most protein subunits enter the inner membrane from the matrix side, but nothing is known about the path of cardiolipin. In this paper, the topography of cardiolipin biosynthesis is investigated. Cardiolipin synthase, a membrane-bound protein, could not be released by sonication or 1 M KCl. In sucrose density gradient subfractionation, cardiolipin synthase co-migrated with the inner membrane marker cytochrome oxidase. no indication was obtained for a preferential localization of this enzyme at contact sites between the outer and inner membranes. Protease digestion experiments showed that cardiolipin synthase exposed protease-susceptible domains mainly to the matrix side of the inner membrane. In intact mitochondria, the Mn(2+)-dependent stimulation of cardiolipin synthesis was abolished when the Mn2+ influx into the matrix was blocked by ruthenium red. 1-Decanoyl-sn-glycero-3-phosphorylcholine, a water-soluble inhibitor of cardiolipin synthase, was only effective after disintegration of mitochondria. The metabolic precursor of cardiolipin, CDP-diacylglycerol, was synthesized by an inner membrane enzyme whose protease-susceptible domains were mainly exposed to the matrix side. It is concluded that cardiolipin is synthesized in the inner leaflet of the mitochondrial inner membrane.

Effect of disulfiram (DS) on mitochondria from rat hippocampus: metabolic compartmentation of DS neurotoxicity.

This experiment was designed to study the acute effects of disulfiram on mitochondrial enzymes in nonsynaptic and synaptic mitochondria from rat hippocampus. Cytochrome c oxidase, monoamine oxidase-B, glycerolphosphate acyltransferase and beta-hydroxybutyrate dehydrogenase were studied. Differences in enzyme activity were seen in controls. Cytochrome c oxidase activity was higher in synaptic mitochondria whereas glycerolphosphate acyltransferase activity was higher in nonsynaptic mitochondria. Mitochondria from disulfiram treated rats, particularly synaptic mitochondria, exhibited lower specific activities of cytochrome c oxidase and monoamine oxidase-B. These alterations were not limited to either the inner or outer mitochondrial membrane. Transmission electron microscopy revealed that mitochondria from disulfiram treated rats were severely altered in isolated preparations as well as in those from whole tissue. This study shows that disulfiram exerts a differential effect on mitochondrial subpopulations.

Regulation of mitochondrial and microsomal phospholipid synthesis by liver fatty acid-binding protein.

Recently we have detected and partially purified a 15-kDa cytosolic L-alpha-lysophosphatidic acid (LPA)-binding protein (LPABP), which stimulates export of LPA from mitochondria (Vancura, A., Carroll, M. A., and Haldar, D. (1991) Biochem. Biophys. Res. Commun. 175, 339-343). Now we have purified this protein to homogeneity. By Western immunoblot analysis, amino acid sequence analysis, and binding characteristics we have shown that LPABP is identical with liver fatty acid-binding protein (L-FABP). This protein binds LPA, and stimulates mitochondrial and microsomal glycerophosphate acyltransferase (GAT) and the export of LPA from both the organelles. The mitochondrially synthesized LPA exported by L-FABP can be converted to phosphatidic acid by microsomes. L-FABP also stimulates microsomal conversion of LPA to phosphatidic acid but strongly inhibits this reaction in mitochondria. However, in the absence of L-FABP mitochondria predominantly synthesize PA. Taken together, these findings are suggestive that L-FABP plays a major role in mitochondrial and microsomal phospholipid metabolism by regulating both the synthesis and utilization of LPA.

A lysophosphatidic acid-binding cytosolic protein stimulates mitochondrial glycerophosphate acyltransferase.

Rat liver cytosolic fraction caused up to five fold stimulation of mitochondrial glycerophosphate acyltransferase apparently by removing the lysophosphatidic acid formed by the acyltransferase. When mitochondria were incubated with palmityl-CoA, [2-3H]-sn-glycerol 3-phosphate and the cytosolic fraction and the supernatant fluid of the incubated mixture was passed through a Sephadex G-100 column, labeled lysophosphatidic acid eluted in three peaks with Mrs (i) 60-70 kDa, (ii) 10-20 kDa, and (iii) less than 5 kDa. Proteins, responsible for binding of lysophosphatidic acid in peaks (i) and (ii), were purified to near homogeneity as judged by electrophoretic analysis. The lysophosphatidic acid binding protein in peak (i) appears to be serum albumin and peak (iii) represents largely unbound lysophosphatidic acid. The 15 kDa protein, purified from peak (ii), bound lysophosphatidic acid, stimulated the acyltransferase and export of lysophosphatidic acid from mitochondria.

Experimental hepatic amoebiasis in inbred mice.

A model of hepatic amoebiasis in two inbred strains of mouse (Swiss and BALB/c, has been developed by introducing Entamoeba histolytica-infected hamster liver tissue in between the adjacent liver lobes of mouse. It is expected that this model might be useful in studying various parameters of host-parasitic interactions and experimental chemotherapy of amoebiasis with relative ease.

Export of mitochondrially synthesized lysophosphatidic acid.

We have previously demonstrated that the properties of mitochondrial glycerophosphate acyltransferase are in keeping with the asymmetric distribution of fatty acids found in naturally occurring cell glycerophospholipids. We are now examining if mitochondria can export lysophosphatidic acid and if it is converted to other phospholipids by the microsomes. Rat liver mitochondria were incubated for 3 min with [2-3H]-sn-glycerol 3-phosphate, palmityl-CoA, and N-ethylmaleimide in the acyltransferase assay medium. In the absence of bovine serum albumin in the medium, greater than 80% of the phospholipids sedimented with the mitochondria. In the presence of the albumin, the lysophosphatidic acid was present entirely in the supernatant fluid. The very little phosphatidic acid that was formed sedimented with the mitochondria. Addition of microsomes to the supernatant fluid followed by a further incubation of 5 min converted 61% of the lysophosphatidic acid to phosphatidic acid which sedimented with the microsomes. When mitochondria and microsomes were incubated together in the assay medium containing albumin and N-ethylmaleimide, the product contained more phosphatidic and less lysophosphatidic acid. When the subcellular components were reisolated by differential centrifugation, 70% of the phosphatidic acid sedimented with the microsomes and the lysophosphatidic acid stayed in the postmicrosomal supernatant. Thus, under appropriate conditions mitochondrially produced lysophosphatidic acid can leave the organelles and this phospholipid can be converted to phosphatidic acid by the microsomes.

Delayed cesarean section: neonatal outcome.

One hundred and twenty one consecutive cesarean sections producing single term, appropriate for gestational age neonates, out of which 85% were emergency cesareans, were included in this study. Fetal distress, nonprogress of labour, and cephalopelvic disproportion were major indications for surgery. The waiting period varied from less than 30 minutes to greater than 4 hours. More than 50% of neonates studied suffered from some problem. The morbidity increased significantly if cesarean section was delayed for more than two hours.