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Biomedical subjects

S Bagchi

Publications and source records attributed to S Bagchi.

48 records · Page 3Linked to original sources

Phosphorylation-dependent activation of the adenovirus-inducible E2F transcription factor in a cell-free system.

Adenovirus infection induces a large increase in the DNA binding activity of a cellular transcription factor that is utilized by the viral E2 promoter and termed E2F. Using cell-free extracts, we have developed an assay for the in vitro activation of DNA binding activity of E2F. E2F activity is undetectable in HeLa extracts but upon incubation with a fraction from adenovirus-infected cells, there is an ATP-dependent increase in E2F DNA binding activity. This increase does not occur using an equivalent fraction from dl312 (E1A-)-infected cells. Incubation of E2F with phosphatase inactivates E2F binding activity. Incubation of the phosphatase-inactivated E2F with an infected cell fraction restores E2F activity as does incubation with a known protein kinase. In contrast, incubation with an extract from mock-infected cells does not restore activity. We conclude that the DNA binding activity of E2F is regulated by phosphorylation in an E1A-dependent manner.

Adenovirus Early Proteins↗

DNA-binding activity of the adenovirus-induced E4F transcription factor is regulated by phosphorylation.

Previous experiments have identified E4F, an inducible cellular factor that binds to sequences in the adenovirus E4 promoter that are critical for E1A-dependent transcriptional activation. The E4F factor has been purified and shown to stimulate transcription in vitro from the E4 promoter. Analysis of the affinity-purified factor identifies a single polypeptide of 50 kD that has E4F-specific binding activity. E4F binding activity is also regulated during F9 cell differentiation and can be activated in differentiated F9 cells by viral infection. Furthermore, the activation process appears to involve a phosphorylation event, because treatment of E4F with alkaline phosphatase abolishes activity and incubation of the phosphatase-inactivated factor with an extract from virus infected cells restores activity.

Adenoviridae↗

Structure and expression of murine malic enzyme mRNA. Differentiation-dependent accumulation of two forms of malic enzyme mRNA in 3T3-L1 cells.

Many murine cells express two mRNAs with markedly different sizes (2.0 and 3.1 kilobases (kb)) that hybridize with cDNA probes for cytosolic malic enzyme ((S)-malate NADP+ oxidoreductase (oxaloacetate-decarboxylating, EC 1.1.1.40). A series of overlapping cDNA clones corresponding to 3129 nucleotides of malic enzyme mRNA was isolated and sequenced to determine the relationship between the two mRNAs and establish the primary structure of mouse malic enzyme. The larger mRNA has an open reading frame of 1716 nucleotides followed by a 3' untranslated region of 1348 nucleotides. The sequence of an exceptionally G/C-rich (88%) portion (65 nucleotides) of the 5' noncoding region was also established. An uncommon poly A addition signal (AUUAAA) is used during the processing of the 3.1-kb mRNA. The 2.0-kb mRNA results from the utilization of another poly A addition signal that truncates the 3' noncoding sequence by approximately 1 kb. The mRNA coding sequence indicates that the malic enzyme subunit contains 572 amino acid residues and has a Mr of 64,000. Two putative components of an NADP-binding domain are located between residues 100 and 165. During the differentiation of 3T3-L1 preadipocytes into adipocytes both the rate of synthesis and relative mRNA concentration for malic enzyme and another lipogenic enzyme, ATP-citrate lyase, are coordinately increased 5-7-fold. However, as preadipocytes approach confluence, the mRNA levels for both lipogenic enzymes transiently increase 3-4-fold, whereas the rates of synthesis of the two proteins are only slightly elevated. Thus, lipogenic enzyme expression is controlled at a pretranslational level during adipogenesis, but the accumulation of the same enzymes may also be subject to translational control in the fibroblast-like preadipocytes. In contrast, mRNA coding for a third enzyme required for lipogenesis, glycerol-3-phosphate dehydrogenase, is not detected in 3T3-L1 preadipocytes, but rapidly accumulates during adipocyte development.

Adipose Tissue↗

Demonstration of Crohn's disease tissue-specific proteins by enzyme-linked immunosorbent assay (ELISA).

Theories on the etiology of Crohn's disease have included extrinsic agents and intrinsic bowel wall defects. We sought to determine the presence of immunoreactive antigens specific to Crohn's disease tissue by modifying the enzyme-linked immunosorbent assay. Tissue proteins were extracted from four patients with Crohn's disease and from four normal segments of colon from patients with colonic cancer. These tissue extracts were further purified on Con A Sepharose 4B affinity column. The glycoproteins eluted from this column were adsorbed by polystyrene plates as antigen and tested against 85 sera from patients with Crohn's disease, ulcerative colitis, other diarrheal diseases, and normal subjects. Sera from 48 patients with Crohn's disease showed significantly greater recognition of Crohn's disease tissue glycoproteins than sera from 27 disease controls (P less than 0.0125) and 10 normal subjects. These Crohn's disease sera also showed preferential recognition of glycoproteins extracted from Crohn's disease tissue compared to glycoproteins from normal colonic tissue (P less than 0.0005). The nature of these immunoreactive proteins, whether extrinsic or intrinsic, is not yet known. The ELISA may help in further characterization of Crohn's disease tissue-specific glycoprotein(s) and to develop a clinically useful serological test.

Antigens↗

Isolation and characterization of Crohn's disease tissue-specific glycoproteins.

Two Crohn's disease tissue-specific proteins are identified and purified several thousand-fold from crude tissue extracts by different chromatographic procedures. The two proteins migrate in sodium dodecyl sulfate-polyacrylamide gel electrophoresis as 200-210-kilodalton and 150-160-kilodalton species. They are glycoproteins, as evidenced by their binding to concanavalin A-Sepharose 4B and positive staining with Schiff's reagent. Specific immunoreactivity of the two glycoproteins against Crohn's disease sera was demonstrated by immunotransblot analysis. All but one of the operative specimens of colon or small intestine (or both) from 13 patients with Crohn's disease contained either or both of the proteins; they were not detected in specimens of colon from 5 patients with ulcerative colitis, 1 patient with diverticulitis, 1 patient with ischemic colitis, from the normal bowel segments resected from 3 patients with colon cancer, and from two normal ileal tissue specimens. The two glycoproteins did not react with antihuman IgG, IgM, and IgA, suggesting that they are not immunoglobulins. The purified glycoproteins may provide important leads toward the understanding of the pathogenesis of Crohn's disease.

Adult↗

Detection and partial characterization of Crohn's disease tissue specific proteins recognized by Crohn's disease sera.

To identify immunoreactive proteins in Crohn's disease (CD) tissue, we examined intestinal tissues from eight patients with CD, seven patients with ulcerative colitis (UC) and four normal colon specimens from patients with colon carcinoma. Tissues were homogenized in 0.05 M Tris-buffered saline containing 2 mM phenyl-methyl-sulphonyl fluoride, and the cell free supernatants were incubated with sera from eight patients with CD, five with UC and five normal volunteers. Immune complexes formed in vitro were precipitated with pansorbin, washed with several detergents and analysed by SDS-polyacrylamide gel electrophoresis. When CD tissue was immunoprecipitated with CD sera, three major proteins of 160,000, 120,000 and 110,000 daltons were detected. Partial enrichment of these proteins were achieved when immune complexes formed in vitro were precipitated with polyethylene glycol followed by protein A-Sepharose 4B chromatography. Affinity gel chromatography and autoradiographic studies with purified serum IgG from CD patients further confirmed the presence of these immunoreactive proteins in CD tissue extracts. When similarly examined, these proteins were absent from UC and control tissue extracts after incubation with CD, UC or control sera. These studies suggest that CD tissue contains several proteins which are specifically recognized by CD patients' sera. Characterization of these immunoreactive proteins may provide an important lead in understanding the pathogenesis of CD.

Antibody Specificity↗

A study of intersectoral co-ordination in disaster management in flood prone districts of West Bengal.

Disaster management is essentially a multi-sectoral and multidisciplinary endeavor. The status of disaster preparedness, coordination among different sectors and its impact on disaster management performance were studied and compared in two flood prone comparable districts viz. Midnapore and Murshidabad of West Bengal. The perception of state level officers about important job responsibilities of other departments in relation to disaster management and its intersectoral co-ordination was found mostly satisfactory; but this desired status did not prevail at district level. Lack of co-ordination among some sectors at district levels persisted in Murshidabad district, indicating comparatively better intersectoral co-ordination in Midnapore district. Thus, 'early response following flood' the most important indicator of adequacy of pre-disaster preparedness was found significantly better in Midnapore district. On the contrary, due to liberal use of Radio transmission set for flood warning, a significantly higher proportion of affected families in Murshidabad district received early flood warning compared to Midnapore. Organisational aspects of disaster management need to be improvised with role clarity of different departments in relation to other sectors involved in this endeavor.

Disaster Planning↗

Disaster to development.

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Community Health Services↗

Microencapsulation of beta-galactosidase with Eudragit L-100.

Microcapsules containing beta-galactosidase (lactase) were prepared by solvent evaporation using the pH sensitive polymer, Eudragit L-100. Formulations were prepared using various polymer-enzyme ratios with total solids content of the internal phase using sucrose stearate as a droplet stabilizer. Particle size distributions were invariant to relative proportion of ingredients but were dependent on stirring conditions. Although sucrose stearate had no effect on particle size distribution, release rate or encapsulation efficiency, its presence at a minimum 2% level was necessary to ensure intact microcapsules. Encapsulation efficiencies were higher for formulations prepared with 15% compared to 10% total solid content. DSC results revealed an interaction between encapsulated Eudragit L-100-enzyme-sucrose stearate vs their physical mixtures. The enzyme activities of the freshly prepared product vs those stored under stressed condition (40 degrees C and 75% RH) were 68 and 40% of their pre-processing activity, respectively. In vitro dissolution showed no enzyme release at 1 h in acidic media but 80% of the lactase was released from the microcapsules over 2.5 h in pH 6.8 media, thus establishing the feasibility of lactase microencapsulation to retard enzyme release in an acidic environment and ensuring release at intestinal pH.

Calorimetry↗

Study of KAP of the private medical practitioners about national disease control programmes.

This study was carried out among the Private Allopathic Medical Practitioners (PMPs) at Khardah Municipal area (West Bengal) to find out their perception & practice about management protocol of diseases like acute respiratory infections and diarrhoel diseases in children, malaria and tuberculosis. Data was collected by interviewing the PMPs with pre-designed open-ended questionnaire. It was supplemented by analysing sample prescriptions of the same diseases. The study revealed that PMPs knowledge and practice were not at par with national guidelines. The need for periodic sensitization of PMPs regarding national disease control programme was emphasized.

Child↗