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S Sharma

Publications and source records attributed to S Sharma.

At least 379 records · Page 21Linked to original sources
PubMed2000

Purification of alkaline phosphatase from chicken intestine by three-phase partitioning and use of phenyl-Sepharose 6B in the batch mode.

Alkaline phosphatase from chicken intestine was purified from the crude preparation employing three-phase partitioning and by the use of phenyl Sepharose-6B in the batch mode. TPP uses a combination of ammonium sulphate and t-butanol to precipitate proteins from crude aqueous extracts. The precipitated protein forms interface between lower aqueous phase and upper organic solvent phase. The fold purification of the finally purified enzyme was 80 and the activity recovery was 61%. The sodium dodecyl sulphate-polyacrylamide gel electrophoresis analysis of enzyme showed considerable purification and its molecular weight was found to be around 67 kDa.

Alkaline Phosphatase↗
PubMed2000

Antibacterial activity of human neutrophil peptide-1 against Mycobacterium tuberculosis H37Rv: in vitro and ex vivo study.

The aim of the study was to investigate the activity of human neutrophil peptide (HNP)-1 to kill Mycobacterium tuberculosis H37Rv in vitro and ex vivo in the murine macrophage cell line J744A.1 on the basis of colony forming units. Macromolecular biosynthesis was studied by monitoring the incorporation of radioactive precursors into different macromolecules. The binding and localization studies were carried out with radioiodinated HNP-1 whereas the cytotoxicity of HNP-1 to macrophages was determined by trypan blue exclusion assay. A concentration dependent inhibition in the growth of M. tuberculosis H37Rv was observed in the presence of HNP-1. The minimum inhibitory concentration and median inhibitory concentration of HNP-1 were found to be 2.5 microg x mL(-1) and 0.8 microg x mL(-1). Treatment of both in vitro grown and phagocytosed mycobacterial cells with HNP-1 resulted in generalized inhibition in the macromolecular biosynthesis with maximum inhibition in deoxyribonucleic acid and lipid biosynthesis. HNP-1 exhibited equilibrium binding with respect to time and two-thirds of bound radioactivity was shown to be present inside the macrophages. Approximately 50% and 98% killing of intracellular mycobacteria was observed after 3 days of treatment with 5 microg x mL(-1) and 40 microg x mL(-1) of HNP-1, respectively. HNP-1 exhibited low cytotoxicity towards the macrophage cell line at the bactericidal concentration to mycobacteria. From the results of this study, it is concluded that human neutrophil peptide-1 possesses potent bactericidal activity against virulent mycobacteria in vitro as well as mycobacteria replicating within macrophages.

Animals↗
PubMed2000

Structure of isocitrate lyase, a persistence factor of Mycobacterium tuberculosis.

Isocitrate lyase (ICL) plays a pivotal role in the persistence of Mycobacterium tuberculosis in mice by sustaining intracellular infection in inflammatory macrophages. The enzyme allows net carbon gain by diverting acetyl-CoA from beta-oxidation of fatty acids into the glyoxylate shunt pathway. Given its potential as a drug target against persistent infections, we solved its structure without ligand and in complex with two inhibitors. Covalent modification of an active site residue, Cys 191, by the inhibitor 3-bromopyruvate traps the enzyme in a catalytic conformation with the active site completely inaccessible to solvent. The structure of a C191S mutant of the enzyme with the inhibitor 3-nitropropionate provides further insight into the reaction mechanism.

Amino Acid Substitution↗
PubMed2000

The habitual diet in rural and urban Cameroon.

OBJECTIVE: To evaluate the habitual diet of a rural and urban population in Cameroon, Central Africa. SETTING: An urban area-Cité Verte Housing District, Yaoundé (1058 subjects); and a rural area-three villages in Evodoula, Cameroon (746 subjects). SUBJECTS: Cameroonian men and women of African origin (1058 urban, and 746 rural), aged 24-74 y. METHODS: The habitual diet was estimated with an interviewer-administered food frequency questionnaire. MAIN OUTCOME MEASURES: Macro- and micronutrient intake. RESULTS: The intake of energy, fat and alcohol was higher in rural men and women than in urban subjects. In rural women, the intake of carbohydrates and protein was also higher. The intakes of fibre, iron, carotene, zinc, potassium, and of the vitamins C, D and E were all higher in rural men and women than in their urban counterparts. The intake of retinol was lower in rural subjects than in urban subjects. Eight of the 10 foods eaten in the highest amount and contributing most to energy intake differed between the rural and urban population. CONCLUSION: The habitual diet in rural Cameroon contains more fat and alcohol than the diet in urban Cameroon. The high physical activity in the rural area may explain the lower levels of the cardiovascular risk factors in this area compared to those of the urban dwellers. SPONSORSHIP: This work was supported by a grant from the European Union (contract no. TS3*CT92-0142) and by the Conseil Régional d'Ile de France and INSERM. European Journal of Clinical Nutrition (2000) 54, 150-154

Adult↗
PubMed2000

Chronic lymphocytic leukemia B cells are highly sensitive to infection by herpes simplex virus-1 via herpesvirus-entry-mediator A.

We found that chronic lymphocytic leukemic (CLL) B cells are highly sensitive to infection with vectors derived from replication-defective herpes simplex virus-1 (rdHSV-1). CLL B cells were found to express high levels of herpes virus entry mediator (Hve) A, but not HveC, the other known receptor for HSV-1. An HveA cDNA from CLL cells was found to encode Arg-->Lys and Val-->Iso substitutions at amino acids 17 and 241, respectively. Nevertheless, this cDNA encoded a functional receptor for HSV-1 when transfected into Chinese hamster ovarian (CHO) cells. Antibodies to HveA could block rdHSV-1 infection of CLL cells and HveA-transfected CHO cells with similar efficiencies in vitro. In contrast to B cells of normal donors, CLL B cells were resistant to the cytopathic effects of infection by rdHSV-1 and maintained high-level expression of the transgene for several days in vitro. We propose that this is due to the expression by CLL cells of the anti-apoptotic protein, bcl-2. Consistent with this, we found that transduction of HeLa cells with a retrovirus expression vector encoding bcl-2 rendered HeLa cells resistant to the cytopathic effects of rdHSV-1. HSV-1-derived vectors should be excellent vehicles for gene transfer into CLL B cells, allowing for its potential use in gene therapy for this disease. Gene Therapy (2000) 7, 1210-1216.

Animals↗
PubMed2000

Underreporting of energy intake in four populations of African origin.

OBJECTIVE: To investigate the frequency of dietary underreporting in four African populations in different geographic and cultural settings. SUBJECTS: Seven-hundred and forty three men and women from rural Cameroon, 1042 men and women from urban Cameroon, 857 men and women from Jamaica and 243 male and female African Caribbeans from the UK. Subjects who reported dieting or weight control were excluded. MEASUREMENTS: Habitual dietary intake was estimated with a quantitative food frequency questionnaire, developed specifically for each country. Underreporting was defined using three cut-off levels for energy intake/estimated basic metabolic rate (EI/BMRest), based on age, sex and weight, in each site. RESULTS: The EI/BMRest was highest in rural Cameroonian men at 3.07 (95% confidence interval: 2.97, 3.17) and women at 2.84 (2.74, 2.94), intermediate in urban Cameroon and Jamaica and lowest in the UK men and women at 1.44 (1.26, 1.62) and 1.41 (1.21, 1.61). This trend existed even after adjustment for age, BMI and education (P for trend<0.0001). The trend in the frequency of underreporting using the lowest cut-off level for EI/BMRest of 1.15 was 6% and 6% in rural Cameroon for women and men, respectively, 4% and 5% in urban Cameroon, 24% and 19% in Jamaica and 28% and 39% in the UK. With higher cut off levels this trend was similar. CONCLUSION: The results suggest that the frequency of dietary underreporting differs between societies and that Westernization may be one of the factors underlying this phenomenon.

Adult↗
PubMed2000

Local and systemic effects after adenoviral transfer of the murine granulocyte-macrophage colony-stimulating factor gene into mice.

Vectors encoding immunostimulatory genes are under investigation for their use as adjuvants for immunotherapy. Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a prominent candidate gene for this approach because this cytokine can prime immune responses to 'self' tumour or other weak antigens. Prior studies suggested that GM-CSF induces accumulation and differentiation of antigen-presenting cells, particularly dendritic cells that can initiate immunity. To evaluate this model in vivo, we performed i.m. and i.p. injections of an adenovirus vector encoding murine GM-CSF (Ad-mGM-CSF) and evaluated local and systemic effects. After intramuscular injection, local changes were characterized by the accumulation of myeloid cells, a subsequent infiltration of lymphocytes and then myonecrosis. Intraperitoneal injection also induced an accumulation of myeloid cells, an increase in CD3-positive T and a decrease in B220-positive B lymphocytes. Expression of the dendritic cell marker CD11c on 48 +/- 9% of the peritoneal cells (n = 6) along with high levels of surface MHC class II, a characteristic morphology, and endocytosis of FITC-dextran suggested in vivo differentiation of dendritic cells after i.p. injection of Ad-mGM-CSF. Systemic effects were observed after i.m. and i.p. injection of Ad-mGM-CSF. All mice developed hepatosplenomegaly resulting from extramedullary haematopoiesis. These changes were specific to GM-CSF as they were not seen in mice injected with an adenovirus vector without a transgene. Our observations indicate that adenoviral transfer of GM-CSF is a powerful tool for inducing local and systemic expansion of haematopoietic cells. The local expansion of myeloid cells displaying signs of dendritic cell differentiation, as characterized for the peritoneal cell compartment, can explain the potency of GM-CSF when used as an adjuvant in genetic immunotherapy.

Adenoviridae↗
PubMed2000

Effect of pre-emptive analgesia on self-reported and biological measures of pain after tonsillectomy.

This prospective, double-blind, randomized study assessed effect of pre-emptive peritonsillar block in 30 ASA-I children, aged 6-12 years, of both sexes, scheduled for tonsillectomy. Patients were divided into three groups: those in group I received a sham block, whereas peritonsillar blocks with bupivacaine 0.25% were given to the children before tonsillectomy (group II) or immediately after surgery had been completed (group III). Constant pain, pain on swallowing, blood glucose, serum epinephrine and norepinephrine concentrations were measured immediately after surgery and 4h after operation. Patients in group I experienced more pain (P< 0.05) than those in groups II and III, both in the immediate postoperative period and over the next 4 h. Patients in groups II and III experienced similar pain (P> 0.05). The pain experienced when water was swallowed was similar to that of the constant pain. The rise of serum norepinephrine concentration in group II was significantly less (P< 0.05) compared to groups I and III. We found both pre-emptive and postoperative block to be equally effective in treating postoperative pain, with pre-emptive block being more effective in preventing the rise in norepinephrine concentration.

Analgesics, Opioid↗
PubMed2000

Influence of neonatal rearing conditions on stress-induced adrenocorticotropin responses and norepinepherine release in the hypothalamic paraventricular nucleus.

Postnatal rearing conditions influence the development of hypothalamic-pituitary-adrenal (HPA) responses to stress in the rat. Thus, postnatal handling dampens HPA responsivity to stress, while prolonged periods of maternal separation have the opposite effect. HPA responses to stress are initiated by the release of corticotropin-releasing factor and/or arginine vasopressin from the neurones of the paraventricular nucleus of the hypothalamus (PVNh). A major source of input to the PVNh arises from brainstem noradrenergic neurones with signalling occurring via alpha1 adrenoreceptors. We examined the noradrenergic response to stress in the PVNh in adult animals exposed to daily periods of handling or maternal separation over the first 2 weeks of life using microdialysis in conscious animals. Maternal separation increased, while handling greatly decreased and norepinepherine responses to restraint stress in the PVNh as compared to non-handled controls; the same pattern was observed for plasma adrenocorticotropic hormone (ACTH) responses to stress. Rearing condition did not affect either alpha1 or alpha2 receptor levels in the PVNh. However, alpha2 receptor binding levels in the noradrenergic cell body regions of the locus coeruleus and the n. tractus solitarius were significantly increased in handled animals. These alpha2 receptors are principally located on noradrenergic neurones (i.e. autoreceptors) and inhibit noradrenaline release at terminal sites. The effects on alpha2 receptor levels could serve as a mechanism for the differences in stress-induced noradrenaline levels in the PVNh and in HPA activity among handled vs non-handled and maternal separation animals. Thus, early life events may serve to influence the differentiation of noradrenergic neurones and thus alter HPA responses stress in adulthood.

Adrenocorticotropic Hormone↗
PubMed2000

Characterization of a differential immunoscreen epitope of Plasmodium falciparum using combinatorial agents.

A differential serological screening of a lambdagt11 cDNA expression library has identified several clones, which react exclusively to sera samples from persons clinically immune to malaria but not to acute malaria patient sera. One such clone, IPf9, has a 315-bp cDNA insert, which was found to be conserved in different strains of the human and rodent malarial parasite Plasmodium falciparum and Plasmodium berghei, respectively. The induced expression product of IPf9 was used to generate polyclonal sera in rabbits. The IPf9 expression product was also screened with phage surface display combinatorial libraries to isolate reagents that specifically bound to the IPf9 product. The polyclonal antisera and the combinatorial reagents recognized a 50-kDa protein from P. falciparum, and a 53-kDa product from P. berghei. Immunofluorescence studies using asexual and sexual stages of P. falciparum showed the protein to be present within the parasite in each of the asexual and sexual stages. The combinatorial reagents showed a partial inhibition in the growth of P. falciparum in vitro. Mice infected with the P. berghei showed the presence of T-cells that exhibited lymphoproliferation when stimulated with the IPf9 protein. It is suggested that IPf9 protein is a conserved protein epitope, and may be relevant for a protective immune response to malaria.

Animals↗
PubMed2000

Enteroendocrine localization of GLP-2 receptor expression in humans and rodents.

BACKGROUND & AIMS: Glucagon-like peptide (GLP)-2, a product of the proglucagon gene, is expressed in enteroendocrine cells of the small and large intestine and is trophic to the gastrointestinal mucosa. GLP-2 also inhibits gastric acid secretion and emptying and up-regulates intestinal hexose transport. GLP-2 acts via binding to a single G protein-coupled GLP-2 receptor (GLP-2R), but the cellular targets for the diverse actions of GLP-2 remain unknown. METHODS: GLP-2R expression in rodent and human tissues was examined using a combination of Northern blotting, reverse-transcription polymerase chain reaction (RT-PCR), and immunocytochemistry. RESULTS: A single major GLP-2R messenger RNA transcript was detected by Northern blot analysis in rodent stomach, duodenum, jejunum, ileum, and colon, but not in rodent esophagus. GLP-2R expression was also detected by RT-PCR in RNA from the hypothalamus, brain stem, and lung. Immunocytochemical localization of human GLP-2R expression using specific antisera detected GLP-2R immunopositivity in subsets of endocrine cell populations in the epithelium of the stomach and both the small and large bowel. CONCLUSIONS: These findings suggest that enteroendocrine-derived GLP-2 acts directly on endocrine cells to induce one or more downstream mediators of GLP-2 action in the gastrointestinal tract.

Animals↗
PubMed2000

New gene and cell-based therapies for lung cancer.

Lung cancer is the leading cause of cancer-related mortality in men and women in the United States, in part, because of the poor treatment options available. New treatment strategies that specifically target discreet steps in the molecular and cellular pathogenesis of this disease are under development. This review highlights many of the basic defects that result in the cellular transformation and subsequent progression of lung cancer, and how the understanding of those fundamental defects lead to the formulation of rational gene-based or cell-based therapies.

Journal Article↗
PubMed2000

Shape-selective binding of geometrically-constrained bis-distamycins to a DNA duplex and a model Okazaki fragment of identical sequence.

The binding of ligands to nucleic acids is of great interest for the control of gene expression and other nucleic acid mediated processes. We have evaluated the binding of several geometrically-constrained bis-distamycins to a model Okazaki fragment [OKA], or a DNA duplex having identical base sequence [DD], using gel-shift assays, optical spectroscopy and differential scanning calorimetry. In the case of covalent attachment of two distamycins to a central benzene ring, a similar binding profile was observed for [DD] as was observed for [OKA] (para binds [K(app) > 10(6) M(-1)], meta binds only weakly). For a central pyridyl ring, however, clear distinction between the binding to [DD] and binding to [OKA] was observed. While none of the three meta isomers having a central pyridyl ring bound [OKA], two of them (MT-17 and MT-12) bound [DD] [K(app) > 10(6) M(-1)]. These results demonstrate subtle differences in lexitropsin shape and placement of electronegative atoms may result in selective binding to a nucleic acid duplex based both on base sequence and chemical composition. Selective binding to DNA duplexes may be useful for designing ligands that regulate transcription, but do not interfere in other nucleic acid mediated processes.

Base Sequence↗
PubMed2000

Failure of pentavalent antimony in visceral leishmaniasis in India: report from the center of the Indian epidemic.

In India, 320 patients with visceral leishmaniasis (209 in the state of Bihar and 11 in the neighboring state of Uttar Pradesh) received identical pentavalent antimony (Sb) treatment. Sb induced long-term cure in 35% (95% confidence interval [CI], 28%-42%) of those in Bihar versus 86% (95% CI, 79%-93%) of those in Uttar Pradesh. In Bihar, the center of the Indian epidemic, traditional Sb treatment should be abandoned.

Adolescent↗
PubMed2000

Comparative chemopreventive mechanisms of green tea, black tea and selected polyphenol extracts measured by in vitro bioassays.

Black tea extracts (hot aqueous, polyphenols and theaflavins) and green tea extracts (hot aqueous, polyphenols, epicatechin, epicatechin gallate, epigallocatechin and epigallocatechin gallate) were tested in nine standardized cell culture assays for comparative cancer chemopreventive properties. Most black and green tea extracts strongly inhibited neoplastic transformation in mouse mammary organ cultures, rat tracheal epithelial cells and human lung tumor epithelial cells. Nearly all tea fractions strongly inhibited benzo[a]pyrene adduct formation with human DNA. Induction of phase II enzymes, glutathione-S-transferase and quinone reductase, were enhanced by nearly all tea fractions, while glutathione was induced by only a few fractions. Ornithine decarboxylase activity was inhibited by nearly all the green tea fractions, but none of the black tea fractions. 12-O-tetradecanoylphorbol-13-acetate-induced free radicals were inhibited by most tea fractions. These results provide strong evidence of both anti-mutagenic, anti-proliferative and anti-neoplastic activities for both black and green tea extracts. Such anticancer mechanisms may well be responsible for the cancer preventive efficacies seen in both experimental and human studies.

Animals↗
PubMed2000

Reconstruction of a tibial defect with microvascular transfer of a previously fractured fibula.

A 43-year-old man sustained severe injuries to his lower limbs with extensive soft-tissue damage and bilateral tibial-fibular fractures. Acutely, the patient underwent external fixation and a free latissimus dorsi flap for soft-tissue coverage of the left leg. However, the tibia had a nonviable butterfly fragment that left a 7-cm defect after debridement. Subsequently, the contralateral fractured fibula was used as a bridging vascularized graft for this tibial defect. The transfer of a fibula containing the zone of injury from a previous high-energy fracture has not been reported. This case demonstrates the successful microvascular transfer of a previously fractured fibula for the repair of a contralateral tibial bony defect.

Adult↗