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Biomedical subjects

D Ghosh

Publications and source records attributed to D Ghosh.

At least 37 records · Page 2Linked to original sources
PubMed2001

Effect of vaginally administered (Ala(8,13,18))-magainin II amide on the morphology of implantation stage endometrium in the rhesus monkey (Macaca mulatta).

Intravaginal administration of an anti-microbial agent, (Ala(8,13,18))-magainin II amide, during blastocyst implantation inhibits pregnancy establishment in a dose-related manner in the rhesus monkey (Macaca mulatta). In the present study, mated female rhesus monkeys were vaginally inserted with tampons containing vehicle (Group 1; n = 5) and test agent (magainin, 0.5 mg/animal; Group 2; n = 6) on cycle day 20. Endometrial tissue samples were collected on Cycle Day 24 from all monkeys and processed for morphometric and ultrastructural analysis. Concentrations of estradiol-17beta, progesterone, and chorionic gonadotrophin in peripheral circulation were determined, which revealed that two monkeys in Group 1 were pregnant while no animals were pregnant in Group 2. Endometrial morphology, however, revealed histologic evidence of pregnancy in three out of the six magainin-treated animals. It appears that intra-vaginal administration of magainin II amide had a marginal effect on the implantation stage endometrium and the initiation of the implantation process in the rhesus monkey.

Administration, Intravaginal↗
PubMed2001

Purification, characterization and crystallization of human placental estrone/dehydroepiandrosterone sulfatase, a membrane-bound enzyme of the endoplasmic reticulum.

Estrone (E1)/dehydroepiandrosterone (DHEA) sulfatase (ES/DHEAS) catalyzes the hydrolysis of E1 and DHEA-sulfates releasing unconjugated steroids. ES is a component of the three-enzyme system that has been implicated in intracrine biosynthesis of estradiol, hence, proliferation of hormone dependent breast tumors. ES is bound to the membrane of the endoplasmic reticulum, presumably through multiple transmembrane and other membrane anchoring segments. The highly hydrophobic nature of the enzyme has so far prevented its purification to homogeneity in quantities sufficient for crystallization. We report here the purification, biochemical characterization and crystallization of the full-length, active form of the enzyme from the membrane bound fraction of human placenta. Our results demonstrate that the key to successful purification and growth of diffraction quality crystals of this difficult membrane bound enzyme is the exploitation of optimal solubilization and detergent conditions to protect the structural and functional integrity of the molecule, thereby preventing nonspecific aggregation and other instabilities. This work paves the way for the first structural study of a membrane bound human sulfatase and subsequent rational design of inhibitors for use as anti-tumor agents.

Amino Acid Sequence↗
PubMed2001

Ictal deafness--a report of three cases.

Though negative phenomena like motor inhibition, blindness or aphasia are described as an ictal manifestation of seizures, ictal deafness has not been reported so far. We observed transient ictal deafness in three cases of partial seizures. One of them had seizure spread to the temporal lobe to produce a complex partial seizure. Two of them have CT-detected lesions in the left temporal/parietal area. The other one had left temporal focus on EEG with a normal imaging study. The mechanism of such ictal negative phenomena is unclear. An epileptic focus around the primary auditory cortex, dampening its receptive ability may manifest as cortical deafness.

Adolescent↗
PubMed2001

Immunohistochemical localization of insulin-like growth factors I and II at the primary implantation site in the Rhesus monkey.

There are various cellular mediators which can affect the process of blastocyst implantation by regulating the proliferation and differentiation of conceptus and maternal endometrial cells. Insulin-like growth factors I (IGF-I) and II (IGF-II) are potent mitogenic and differentiation-promoting growth factors. However, the role of IGF peptides at implantation in primate species is not well understood. The objective of the present study was to immunohistochemically localize IGF-I and IGF-II peptides in trophoblast cells and maternal endometrial cells during lacunar and villous stages of placentation in the Rhesus monkey. Female animals (n = 10) were laparotomized on estimated days 13-16 after fertilization to collect primary implantation sites which were subjected to immunohistochemical staining for IGF-I and IGF-II peptides. Cell-type specificity for IGF-I and IGF-II was evident with a very low level of IGF-I peptide immunolocalized in trophoblast cells lining lacunae, and primary and secondary villi, while moderate to high amounts of IGF-II peptide were detected in lamellar syncytiotrophoblast cells lining lacunae, early villi and cell columns, as well as in migrating trophoblast cells in the extravillous compartment and in endovascular trophoblast cells. The observed presence of IGF-II peptide in differentiated lamellar syncytiotrophoblast cells during the very early stages of implantation and placentation in the Rhesus monkey may be important in their transition to this differentiated cell population. Maternal endometrial cells showed similar distribution profiles for IGF-I and IGF-II. In conclusion, we report differential distribution of IGF-I and IGF-II peptides in trophoblast cell populations at the feto-maternal interface during lacunar and villous stages of gestation in the Rhesus monkey.

Animals↗
PubMed2001

Repression of Ets-2-induced transactivation of the tau interferon promoter by Oct-4.

Oct-4 is a POU family transcription factor associated with potentially totipotent cells. Genes expressed in the trophectoderm but not in embryos prior to blastocyst formation may be targets for silencing by Oct-4. Here, we have tested this hypothesis with the tau interferon genes (IFNT genes), which are expressed exclusively in the trophectoderm of bovine embryos. IFNT promoters contain an Ets-2 enhancer, located at -79 to -70, and are up-regulated about 20-fold by the overexpression of Ets-2 in human JAr choriocarcinoma cells, which are permissive for IFNT expression. This enhancement was reversed in a dose-dependent manner by coexpression of Oct-4 but not either Oct-1 or Oct-2. When cells were transfected with truncated bovine IFNT promoters designed to eliminate potential octamer sites sequentially, luciferase reporter expression from each construct was still silenced by Oct-4. Full repression required both the N-terminal and POU domains of Oct-4, but neither domain used alone was an effective silencer. Oct-4 and Ets-2 formed a complex in vitro in the absence of DNA through binding of the POU domain of Oct-4 to a site located between the "pointed" and DNA binding domains of Ets-2. The two transcription factors were also coimmunoprecipitated after being expressed together in JAr cells. Oct-4, therefore, silences IFNT promoters by quenching Ets-2 transactivation. The POU domain most probably binds to Ets-2 directly, while the N-terminal domain inhibits transcription. These findings provide further evidence that the developmental switch to the trophectoderm is accompanied by the loss of Oct-4 silencing of key genes.

Animals↗
PubMed2001

Modulation of macrophage mannose receptor affects the uptake of virulent and avirulent Leishmania donovani promastigotes.

The effect of oxidants and the anti-inflammatory steroid dexamethasone on the attachment and internalization of virulent and avirulent Leishmania donovani promastigotes by the macrophage mannosyl fucosyl receptor was examined. Oxidants and dexamethasone are known to down- and upregulate the expression of the mannose receptor. Macrophages, when treated with 500 microM H2O2 at 37 C for 30 min, stimulate about 45% inhibition in uptake of an avirulent strain (UR6), and 30 and 25% inhibition for virulent strains AG-83 and GE-I, respectively. Treatment of macrophages with dexamethasone for 20 hr resulted in a stimulation in uptake of the parasite. When UR6 was used, a 3-fold increase in uptake was observed compared with the controls. Parasite uptake was also inhibited by the H2O2-generating system, glucose/glucose oxidase; inhibition was blocked by catalase. Treatment of macrophages either with H2O2 or dexamethasone did not affect the binding of the advanced glycosylation end product-bovine serum albumin (AGE-BSA), the ligand for AGE receptor of macrophages. Similarly, indirect evidence also shows that both types 1 and 3 complement receptors (CR1, CR3) are not affected by these treatments, indicating that, besides the mannosyl fucosyl receptor, other receptors are minimally altered in the identified condition. These results suggest that the up- and downregulation of the mannose receptor of macrophages may play a role in affecting L. donovani infection.

Animals↗
PubMed2001

Isolation of measles virus from cerebrospinal fluid of children with acute encephalopathy without rash.

OBJECTIVE: To determine the viral agent involved in cases of acute encephalopathy in children during an outbreak in Northern India. DESIGN: Virological and serological studies using serum and cerebrospinal fluid specimens from patients. METHODS: Serum and CSF specimens were tested by IgM ELISA for IgM antibodies to variety of viruses like Japanese encephalitis, West Nile, Dengue and Measles. The specimens were inoculated into Vero cell monolayer for virus isolation. The viral strains isolated were identified by indirect immunofluorescence test and qualitative in-vitro neutralization test using polyclonal and monoclonal antibodies to measles. Identity of the isolates was reconfirmed using RT-PCR method. RESULTS: Of the 28 specimens tested, 17 had IgM antibodies to measles. Commercial IgM ELISA kits confirmed the serological findings. Vero cell cultures yielded 4 isolates from CSF and 2 from serum specimens of six different patients. Cytopathic effect was typical of measles. Indirect imunofluorescence using polyclonal and monoclonal antibodies to measles HA protein, confirmed the measles etiology. Neutralization tests reconfirmed the measles strain isolation. RT-PCR amplified product was confirmed as measles. CONCLUSION: The isolation of measles virus from CSF and serum of children with acute encephalopathy without rash proved the etiological role of measles virus in this outbreak.

Acute Disease↗
PubMed2001

Bilateral subdural effusion and subcutaneous swelling with normally functioning csf shunt.

We report a child with hydrocephalus due to tuberculous meningitis who developed a subcutaneous fluid collection around the ventriculoperitoneal shunt tube entry point, after one month of shunting. On investigation, he had decompressed ventricles with bilateral fronto parietal subdural hygroma. Bifrontal burr hole drainage helped resolution of both subdural effusion and subcutaneous scalp swelling. This complication is unique and its pathogenesis has been postulated.

Antitubercular Agents↗
PubMed2001

Morphological and functional characteristics of rabbit uterine epithelial cells grown on free floating collagen gel.

In the present study isolated uterine epithelial cells from normal rabbits were maintained in culture on free floating rat-tail collagen matrix, and the morphological characteristics of these cells were examined. Additionally, the pattern of protein synthesis and secretion by rabbit uterine epithelial cells grown on free floating collagen gels following estradiol and/or progesterone treatment in vitro was examined. Isolated epithelial cells cultured on collagen gels in complete medium containing serum attached to form monlayers, and eventually the gels became free floating and contracted giving rise to luminal arrangements. These cells were cytokeratin positive epithelial cells and were ultrastructurally polarized. These cells also exhibited differential upregulation and down regulation in the synthesis and secretion of proteins in response to estradiol, progesterone, and estradiol plus progesterone. Additionally, a permissive action between progesterone and estradiol in the synthesis of two species of secretory proteins was observed. It however remains to be examined whether different species of proteins produced in vitro in response to estradiol and progesterone bear any association with physiological states in reproductive cycle in this species.

Animals↗
PubMed2000

Multiple conformations of catalytic serine and histidine in acetylxylan esterase at 0.90 A.

Acetylxylan esterase (AXEII; 207 amino acids) from Penicillium purpurogenum has substrate specificities toward acetate esters of d-xylopyranose residues in xylan and belongs to a new class of alpha/beta hydrolases. The crystal structure of AXEII has been determined by single isomorphous replacement and anomalous scattering, and refined at 0.90- and 1.10-A resolutions with data collected at 85 K and 295 K, respectively. The tertiary structure consists of a doubly wound alpha/beta sandwich, having a central six-stranded parallel beta-sheet flanked by two parallel alpha-helices on each side. The catalytic residues Ser(90), His(187), and Asp(175) are located at the C-terminal end of the sheet, an exposed region of the molecule. The serine and histidine side chains in the 295 K structure show the frequently observed conformations in which Ser(90) is trans and the hydroxyl group is in the plane of the imidazole ring of His(187). However, the structure at 85 K displays an additional conformation in which Ser(90) side-chain hydroxyl is away from the plane of the imidazole ring of His(187). The His(187) side chain forms a hydrogen bond with a sulfate ion and adopts an altered conformation. The only other known hydrolase that has a similar tertiary structure is Fusarium solani cutinase. The exposed nature of the catalytic triad suggests that AXEII is a pure esterase, i.e. an alpha/beta hydrolase with specificity for nonlipidic polar substrates.

Acetylesterase↗
PubMed2000

Familial hypertrophic cardiomyopathy mutations in the regulatory light chains of myosin affect their structure, Ca2+ binding, and phosphorylation.

The effect of the familial hypertrophic cardiomyopathy mutations, A13T, F18L, E22K, R58Q, and P95A, found in the regulatory light chains of human cardiac myosin has been investigated. The results demonstrate that E22K and R58Q, located in the immediate extension of the helices flanking the regulatory light chain Ca(2+) binding site, had dramatically altered Ca(2+) binding properties. The K(Ca) value for E22K was decreased by approximately 17-fold compared with the wild-type light chain, and the R58Q mutant did not bind Ca(2+). Interestingly, Ca(2+) binding to the R58Q mutant was restored upon phosphorylation, whereas the E22K mutant could not be phosphorylated. In addition, the alpha-helical content of phosphorylated R58Q greatly increased with Ca(2+) binding. The A13T mutation, located near the phosphorylation site (Ser-15) of the human cardiac regulatory light chain, had 3-fold lower K(Ca) than wild-type light chain, whereas phosphorylation of this mutant increased the Ca(2+) affinity 6-fold. Whereas phosphorylation of wild-type light chain decreased its Ca(2+) affinity, the opposite was true for A13T. The alpha-helical content of the A13T mutant returned to the level of wild-type light chain upon phosphorylation. The phosphorylation and Ca(2+) binding properties of the regulatory light chain of human cardiac myosin are important for physiological function, and alteration any of these could contribute to the development of hypertrophic cardiomyopathy.

Amino Acid Sequence↗
PubMed2000

Object-oriented transcription factors database (ooTFD).

ooTFD (object-oriented Transcription Factors Database) is an object-oriented successor to TFD. This database is aimed at capturing information regarding the polypeptide interactions which comprise and define the properties of transcription factors. ooTFD contains information about transcription factor binding sites, as well as composite relationships within transcription factors, which frequently occur as multisubunit proteins that form a complex interface to cellular processes outside the transcription machinery through protein-protein interactions. In the past year, a few additions and changes were made to this database and associated tools, which are accessible through the IFTI-MIRAGE web site at http://www.ifti.org/

Databases, Factual↗
PubMed2000

Anti-nidatory effect of vaginally administered (Ala8,13, 18)-magainin II amide in the rhesus monkey.

The hypothesis that timed application of a potent anti-microbial, anti-tumor agent like magainin peptides can inhibit blastocyst implantation was examined in the present study using the rhesus monkey as the primate model. Incidence of pregnancy, vaginal bleeding patterns, serum levels of progesterone, estrogen and monkey chorionic gonadotropin were examined following vaginal administration of (Ala8,13,18)-magainin II amide, a synthetic analogue of magainin 2, via tampon during days 20 to 26 of mated cycles. Implantation occurred in two out of three animals following administration of 0.25 mg magainin, while administration of 0.5 mg (Ala8,13,18)-magainin II amide resulted in inhibition of implantation in all females with no change in lengths of treatment cycles, and subsequent cycles. It appears from the present study that, besides being a local microbicidal agent, intravaginal administration of (Ala(8,13,18))-magainin II amide is a potential anti-implantation strategy for intercepting pregnancy.

Administration, Intravaginal↗
PubMed2000

Anti-nidatory effect of vaginally administered fumagillin in the rhesus monkey.

In the present study, fumagillin, which is an antibiotic with potent angiostatic activity secreted from Aspergillus fumigatus was administered intravaginally during peri-implantation stage in the rhesus monkey and its effects on ovarian function, blastocyst implantation and pregnancy outcome in the rhesus monkey were investigated. Female monkeys (n = 18) showing normal menstrual cycles were vaginally inserted with tampons containing fumagillin (0 mg/animal in group 1; 1 mg/animal in group 2; 2 mg/animal in group 3; 4 mg/animal in group 4) on cycle day 20 of the mated treatment cycle, and these were removed on day 26 of the treatment cycle. Pregnancy was found to occur in animals treated with 1 mg and 2 mg fumagillin. However, animals treated with 4 mg fumagillin remained non-pregnant along with decreased (p <0.001) concentration of progesterone in circulation during the luteal period compared with that in normal, non-mated, ovulatory cycle.

Administration, Intravaginal↗
PubMed2000

Steroid dehydrogenase structures, mechanism of action, and disease.

Steroid dehydrogenase enzymes influence mammalian reproduction, hypertension, neoplasia, and digestion. The three-dimensional structures of steroid dehydrogenase enzymes reveal the position of the catalytic triad, a possible mechanism of keto-hydroxyl interconversion, a molecular mechanism of inhibition, and the basis for selectivity. Glycyrrhizic acid, the active ingredient in licorice, and its metabolite carbenoxolone are potent inhibitors of human 11 beta-hydroxysteroid dehydrogenase and bacterial 3 alpha, 20 beta-hydroxysteroid dehydrogenase (3 alpha, 20 beta-HSD). The three-dimensional structure of the 3 alpha, 20 beta-HSD carbenoxolone complex unequivocally verifies the postulated active site of the enzyme, shows that inhibition is a result of direct competition with the substrate for binding, and provides a plausible model for the mechanism of inhibition of 11 beta-hydroxysteroid dehydrogenase by carbenoxolone. The structure of the ternary complex of human 17 beta-hydroxysteroid dehydrogenase type 1 (17 beta-HSD) with the cofactor NADP+ and the antiestrogen equilin reveals the details of binding of an inhibitor in the active site of the enzyme and the possible roles of various amino acids in the catalytic cleft. The short-chain dehydrogenase reductase (SDR) family includes these steroid dehydrogenase enzymes and more than 60 other proteins from human, mammalian, insect, and bacterial sources. Most members of the family contain the tyrosine and lysine of the catalytic triad in a YxxxK sequence. X-ray crystal structures of 13 members of the family have been completed. When the alpha-carbon backbone of the cofactor binding domains of the structures are superimposed, the conserved residues are at the core of the structure and in the cofactor binding domain, but not in the substrate binding pocket.

17-Hydroxysteroid Dehydrogenases↗
PubMed2000

Methods for analysis of multiple events in the presence of death.

The use of multiple events data in the design and analysis of clinical trials is becoming quite common. Several methods for analyzing such data have been proposed in the literature. However, they do not account for deaths in the analysis. In this article, an overview of available methods for modeling recurrent events and death is given. These methods are illustrated using data from an HIV clinical trial.

Clinical Trials as Topic↗
PubMed2000

Preimplantation embryo morphology following early luteal phase anti-nidatory treatment with mifepristone (RU486) in the rhesus monkey.

The ultrastructural characteristics of peri-implantation stage embryos recovered on day 6 after ovulation from rhesus monkeys with or without mifepristone (RU486) treatment during the early luteal phase were examined in the present study. Monkeys were randomly allocated to two groups; group 1 animals were injected s.c. with 2 ml vehicle (1:4, benzyl benzoate: olive oil, v/v, n = 21) and group 2 animals received a single dose of mifepristone (2 mg/kg body weight, w/v, n = 30) in the same volume of vehicle on day 2 after ovulation in mated cycles. On day 6 after ovulation, female monkeys of both groups were laparotomized and their reproductive tracts were flushed to retrieve preimplantation stage embryos. Embryos that showed frank degeneration or desynchrony on gross microscopical examination were not included in the present study. Preimplantation embryo growth on day 6 after ovulation was significantly (P < 0.05) affected in the morula-blastocyst transition stage in mifepristone-treated monkeys compared with that in the control group of monkeys. Ultrastructurally, administration of mifepristone on day 2 after ovulation depressed preimplantation stage embryo development, characterized by loss of cell polarity, lack of mitochondrial maturity, and lack of differentiation in trophoblast cells. Furthermore, preimplantation embryos from mifepristone-treated animals displayed a higher occurrence of inter-blastomere space, intra-cytoplasmic vacuoles, myelinoid bodies, accumulation of lipid droplets, lysosomes, lipofuscins, autophagosomes and multivesicular bodies. Collectively, it appears that the developmental potential of preimplantation embryos was significantly compromised in mifepristone-treated cycles.

Animals↗