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Biomedical subjects

S K Goswami

Publications and source records attributed to S K Goswami.

At least 37 records · Page 2Linked to original sources
PubMed1993

MyoD transactivates angiotensinogen promoter in fibroblast C3H10T1/2 cells.

MyoD is one of the regulatory genes that causes activation of the expression of muscle specific genes in fibroblasts through interaction with a cis-acting DNA element CANNTG (E-box). We show here that forced expression of MyoD in mouse fibroblast C3H10T1/2 cells induces the expression of the liver specific rat angiotensinogen gene promoter. In the absence of MyoD, the angiotensinogen promoter is not expressed in C3H10T1/2 fibroblasts. Cotransfection of MyoD has no significant effect on the angiotensinogen promoter activity in the liver-derived HepG2 cells, suggesting that the regulatory programs of HepG2 and C3H10T1/2 cells are different, at least in terms of the requirement for MyoD-mediated transactivation of the target angiotensinogen promoter.

Angiotensinogen↗
PubMed1992

Isolation and characterization of a macrophage-derived high molecular weight protein involved in the regulation of mucus-like glycoconjugate secretion.

Pulmonary macrophages release a variety of mediators that are involved in inflammatory processes and probably are involved in respiratory mucus secretion. Conditioned media obtained from activated pulmonary macrophages were found to contain a protein that functioned as a secretagogue for mucus-like glycoconjugate (MLGC) in an in vitro bioassay. A human macrophage-derived hybridoma cell line, HB-63, exhibited the same properties and was very useful in obtaining large amounts of the protein for purification and characterization. With ultrafiltration membranes and gel electrophoresis, the protein isolated from the conditioned media of zymosan- or lipopolysaccharide-treated cells was found to have a molecular weight of approximately 68 kd. The purified protein obtained from hybridoma cells and from pulmonary macrophages exhibited strong biologic activity when it was used to stimulate MLGC secretion, both in human airway explants and in an in vitro human secretory epithelial cancer cell line. The proteins from both sources were found to have similar amino acid compositions. Preliminary results indicate the presence of the 68 kd protein in the bronchoalveolar lavage fluid of a patient with severe chronic bronchitis and mucus hypersecretion. The role of this novel protein in the lungs is, so far, speculative. The 68 kd protein may be a useful tool for studying the biosynthesis and regulation of MLGC secretion and hypersecretion.

Amino Acids↗
PubMed1992

fos-mediated repression of cardiac myosin light chain-2 gene transcription.

The transcription of the chicken cardiac myosin light chain-2 (MLC-2) promoter containing a 1.3 Kb 5'-flanking DNA segment is repressed upon co-transfection with an expression vector (pMMV) containing the proto-oncogene fos in embryonic chicken cardiac muscle cells in culture. Similar concentrations of co-transfectants containing other genes e.g. luciferase were ineffective. To identify the DNA element(s) in MLC-2 gene that responds to fos-mediated inhibition, 5'-sequential deletion mutants of MLC-2 promoter were tested in a transient transfection assay. A mutant, in which the 5' distal sequence was deleted upto -1200 bp upstream of the mRNA start site was sensitive to fos inhibition, but the mutant containing -1130 bp was not, suggesting that a fos responsive element (FRE) is located between -1130 to -1200 bp upstream of the transcription initiation site. The same FRE sequence was also responsive to fos-inhibition in chicken skeletal muscle cells as well. Since over-expression of fos is implicated in repression of myogenic process, the selective inhibition of MLC-2 promoter activity by fos and identification of FRE sequence potentially important in understanding the relationship between myogenesis and the oncoprotein-mediated signal pathway(s).

Animals↗
PubMed1991

Mucus secretagogue production by a human macrophage hybridoma.

A pulmonary macrophage-monocyte-derived mucus secretagogue (MMS) oligopeptide has been previously reported to induce mucus secretion in an in vitro model system with human airway explants and secretory epithelial cells. To understand the possible role of macrophages in the regulation of secretion of mucus, our laboratory has used a series of human macrophage hybridomas that were generated by fusing an hypoxanthine guanine phosphoribosyl transferase-deficient promonocytic line, U937, with macrophages obtaining by maturing monocytes in Teflon bags. The cell lines were proven to be true hybridomas by acquisition of donor class I antigens, additional chromosomes, as well as macrophage specific (maximum velocity) not present on the U937 parent line. One clone, clone 63, produced large amounts of an oligopeptide with an approximate molecular weight of 2000, which was identified from culture supernatants by ultrafiltration, chromatography, isoelectric focusing, and Western blot. Processed clone 63 supernatant had biologic activity causing increased secretion of radiolabeled glycoconjugate in both cultured airways and secretory epithelial cells. Immunoblot analysis with a polyclonal rabbit antisera generated against MMS was positive, and Western blot analysis produced a band at approximately 2000 daltons, consistent with the previously described MMS. MMS secretion could be stimulated by zymosan and lipopolysaccharide and inhibited by both cycloheximide and erythromycin. Dexamethasone had a different effect, appearing to stimulate MMS production intracellularly but inhibiting its release once it was synthesized. The availability of cloned hybridomas allows for study of the regulation of mucus secretagogue production as well as purification of molecular species and provides a valuable tool for the study of mucus secretion.

Cell Line↗
PubMed1991

Respiratory mucus hypersecretion (bronchorrhea): a case discussion--possible mechanisms(s) and treatment.

The mechanism(s) underlying mucus hypersecretion (bronchorrhea) and the treatment of this condition are poorly understood. We have previously demonstrated that erythromycin inhibited mucus secretion from human airways and from secretory epithelial cells in vitro. We encountered a patient with airway obstruction marked by severe bronchorrhea, who previously had responded only to inhaled bronchodilators and high-dose prednisone. Many attempts to wean him from prednisone had failed. During the course of his disease, he had developed an IgG antibody to vasoactive intestinal peptide, had increased amounts of mucus secreted by his respiratory epithelial cells, and demonstrated hyperreactive airways as measured by methacholine challenge provocation test. Erythromycin was added to his therapy. The effect of erythromycin treatment was quite dramatic and included clinical and laboratory improvement. After a short trial of erythromycin, the patient tolerated low, every-other-day doses of prednisone. there was a significant reduction in the volume of his bronchorrhea, a major decrease in the epithelial mucins in his total expectorated mucus, complete inhibition of his airway hyperresponsiveness to inhaled methacholine, and significant reduction in the level of IgG antibody to vasoactive intestinal peptide. This response was specific for erythromycin since other antibiotics did not have any clinical, biochemical, or physiologic effects. We conclude that erythromycin may play a role in the treatment of patients with bronchorrhea and may have a steroid-sparing effect. Additional studies with larger numbers of patients are indicated.

Aged↗
PubMed1991

Mechanism of selective translation of vaccinia virus mRNAs: differential role of poly(A) and initiation factors in the translation of viral and cellular mRNAs.

We have recently demonstrated that the poly(A) moieties of short RNAs obtained from both in vitro transcription and from vaccinia virus (VV)-infected cells exhibit dissimilar effects on the in vitro translation of cellular and VV mRNAs (R. Bablanian, G. Coppola, P. Masters, and A. K. Banerjee, Virology 148:375-380, 1986; M. J. Su and R. Bablanian, Virology 179:679-693, 1990). In the present study, we have investigated the roles of poly(A), m7GTP, and initiation factors in the mechanism of selective translation of VV mRNAs. The effects of unfractionated poly(A) [termed poly(A)un, with various chain lengths up to 3,000 nucleotides] and a 150- to 300-nucleotide fraction of synthetic poly(A) [termed poly(A)150-300] on the translation of HeLa cell mRNAs and early and late VV mRNAs were studied. Both the poly(A)un and the poly(A)150-300 completely inhibited the translation of HeLa cell mRNAs obtained from total cytoplasmic RNA in the nuclease-treated reticulocyte lysates. Viral mRNAs from total cytoplasmic RNA also were slightly inhibited (15 to 38%) by the poly(A)un, whereas the poly(A)150-300 had no significant effect on their translation. The translation of oligo(dT)-cellulose-selected HeLa mRNAs was as sensitive to inhibition by poly(A)150-300 as the mRNAs found in total cytoplasmic RNA. However, the translations of oligo(dT)-cellulose-selected viral mRNAs become more sensitive to the inhibitory effect of poly(A)150-300 than the translations of viral mRNAs found in the total cytoplasmic RNA. Both HeLa and VV mRNAs became more resistant to the poly(A)-mediated inhibition when these mRNAs were deadenylated, but the relative resistance to inhibition by poly(A)150-300 of deadenylated VV mRNAs was much greater than that of HeLa cell mRNAs. The translation of VV mRNAs was significantly less inhibited than the translation of HeLa mRNAs when the cap analog, m7GTP, was added to the cell-free system. The inhibition of HeLa cell mRNA translation by both poly(A)un and poly(A)150-300 was completely restored when poly(A)-binding protein (PAB) was added to the cell-free translational system. The addition of eukaryotic initiation factor 4A (eIF-4A) did not restore translation when poly(A)un was used to inhibit translation; however, inhibition by poly(A)150-300 was significantly reversed by this initiation factor. The reversal of poly (A)-mediated inhibition of HeLa cell mRNA translation was additive when PAB was used together with eIF-4A. Early VV mRNA translation was only slightly inhibited by poly(A)un (15%), and this inhibition was completely reversed by either PAB or eIF-4A.(ABSTRACT TRUNCATED AT 400 WORDS)

Carrier Proteins↗
PubMed1991

Tissue-specific transcription of the cardiac myosin light-chain 2 gene is regulated by an upstream repressor element.

Physiological expression of the cardiac muscle myosin light-chain 2 (MLC-2) gene in chickens is restricted to cardiac muscle tissue only, at least during the late embryonic to adult stages of development. The mechanism by which cardiac MLC-2 gene expression is repressed in differentiated noncardiac muscle tissues is unknown. Using sequential 5'-deletion mutants of the cardiac MLC-2 promoter introduced into primary skeletal muscle cells in culture, we have demonstrated that a 89-bp region, designated the cardiac-specific sequence (CSS), is essential for repression of cardiac MLC-2 expression in skeletal muscle. Removal of the CSS sequence alone allows transcription in skeletal muscle cells without affecting the transcriptional activity of the promoter in cardiac muscle cells. DNase I footprinting and gel shift assays indicate that protein binding to sequences in the CSS domain occurs readily in nuclear extracts obtained from skeletal muscle but not in extracts isolated under identical conditions from cardiac muscle. Thus, it appears that a negative regulatory mechanism accounts for the lack of expression of the cardiac MLC-2 gene in skeletal muscle and that the CSS element and its binding proteins are important functional components of the regulatory apparatus which ensures the developmental program for cardiac tissue-specific gene expression.

Animals↗
PubMed1990

Erythromycin inhibits respiratory glycoconjugate secretion from human airways in vitro.

Erythromycin and other antibiotics have been used empirically in the treatment of patients with chronic obstructive pulmonary disease (COPD). We studied whether this empirical role of antibiotics might not be related to a possible direct effect on respiratory glycoconjugate (RGC) secretion. The effect of erythromycin on RGC secretion and hypersecretion was studied in an in vitro preparation of human airways that were secreting [3H]glucosamine respiratory glycoconjugate (RGC), and on a human endometrial adenocarcinoma cell line secreting a glycoconjugate (tumor glycoconjugate = TGC) chemically similar to the RGC secreted by the airways. Erythromycin at 10(-5) M reduced RGC secretion by 35 +/- 4% (n = 9, p less than 0.001) in both human airways and the adenocarcinoma cells, and was increasingly active in the pharmacologic range of 10(-7) to 10(-4) M. The inhibitory effect of erythromycin was maximal within 16 h and was still evident 34 h after incubation. Erythromycin was noted to reduce both spontaneous (baseline) and stimulated RGC secretion (by histamine and methacholine) from airways in culture. The blocking effect appeared to be more selective for histamine than methacholine. These effects were not associated with any toxicity to the tissues and were not associated with the inhibition of protein synthesis. Dexamethasone also inhibited RGC release in both assay systems and exhibited dose-related effects in the physiologic ranges (10(-9) to 10(-5) M). When administered together, erythromycin and dexamethasone had an additive inhibitory effect on RGC secretion (68.0 +/- 3.0%, n = 7, p less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

Anti-Bacterial Agents↗
PubMed1989

Platelet-activating factor stimulates secretion of respiratory glycoconjugate from human airways in culture.

Platelet-activating factor (PAF) is a naturally occurring phospholipid that acts as a potent mediator of inflammation and bronchoconstriction. Since mucus secretion accompanies many pulmonary-allergic reactions, we examined the effect of PAF on respiratory glycoconjugate (RGC) release from human airways in vitro. PAF, in concentrations of 5 to 100 ng/ml, induced a specific, dose-dependent release of [3H]RGC from human airways in vitro (range of 15% to 120% increase above control, p less than 0.001; n = 8). Time-course studies revealed that RGC release reached its peak level by 60 minutes, and by the end of 4 hours, decreased almost to the baseline level, suggesting a stimulatory effect on secretion rather than synthesis. PAF analog, RO 19-3704, which is a PAF-receptor antagonist, inhibited RGC secretion mediated by PAF in a dose-dependent manner with an inhibitory concentration of 50% of 70 ng/ml. BW 755C and nordihydroguaiaretic acid, but not indomethacin, inhibited RGC release by PAF. LY 171883, a specific leukotriene D4-receptor antagonist, totally inhibited the release of RGC by PAF. Similar results were observed with FPL 55712. PAF-treated airways generated peptidoleukotrienes significantly above control airways in association with enhanced RGC secretion. This enhanced effect on RGC secretion was specifically and significantly blocked by LY 171883. Atropine (10(-5) mol/L) augmented the secretagogue activity of PAF, whereas dexamethasone (10(-5) mol/L) inhibited it. These data indicate that PAF is a significant RGC secretagogue that affects the RGC-secreting cells via the lipoxygenase pathway of arachidonic acid metabolism and the generation of leukotrienes in the airways, but not through the cholinergic receptors on the secretory cells.

Arachidonic Acids↗
PubMed1987

Selective inhibition of protein synthesis by synthetic and vaccinia virus-core synthesized poly(riboadenylic acids).

Studies were undertaken to compare the effect of poly(A)s from various sources on selective inhibition of protein synthesis in the reticulocyte lysate system programmed with viral and cellular mRNAs. RNA synthesized in vitro by vaccinia virus cores in the presence of only ATP inhibited overall HeLa cell polypeptide synthesis by over 80% with a minimal effect on translation of vaccinia virus mRNAs. Hybridization of the [alpha-32P]AMP-labeled RNA made in vitro by vaccinia virus cores in the presence of only ATP, showed no complementary to HindIII restriction fragments of vaccinia virus DNA indicating that the in vitro product was poly(A). Fractionation of synthetic and core-synthesized poly(A) into three size classes showed that the larger the size of poly(A), the greater its inhibitory activity of protein synthesis in the cell-free system. Inhibition of translation of mRNAs from vaccinia virus-infected HeLa cells was also observed in the presence of poly(A). However, virus-induced polypeptide synthesis was more resistant to the effect of poly(A) than were cellular polypeptides. Oligo(dT) when added to the reticulocyte lysate system was capable of reversing the inhibition of protein synthesis caused by both core-synthesized poly(A) and core-transcribed RNAs. These results indicate that poly(A) synthesized by the virion-associated enzyme has inhibitory properties similar to those of synthetic poly(A).

Cell-Free System↗
PubMed1985

Prostanoid synthesis in peripheral nerve.

The transformation of [1-14C]arachidonic acid into radiolabeled prostanoids was studied with homogenates and desheathed sciatic nerves of rats and frogs. All of the preparations studied were shown to synthesize prostaglandins; the specific prostanoids made were characterized by their migration on thin-layer chromatograms in three separate solvent systems. Both desheathed rat nerve and homogenates synthesize prostaglandin E2, prostaglandin F2 alpha, prostaglandin D2, 6-ketoprostaglandin F1 alpha and thromboxane B2. With preparations from frog nerve, prostaglandin E2 was the major prostanoid product formed. Several conditions were able to modulate the production of prostaglandin E2 with desheathed frog nerve. Electrical stimulation at high frequency (100 Hz) for 30 min increased the formation of labeled prostaglandin E2. Inclusion of glutathione also affected prostaglandin E2 formation. A lower concentration (0.1 mM) stimulated prostaglandin synthesis, while 1 mM glutathione was partially inhibitory. In both the rat and frog system, prostanoid synthesis was suppressed by indomethacin and aspirin.

Animals↗
PubMed1985

Inhibitory effects of tannic acid and benzophenone on soybean lipoxygenase and ram seminal vesicle cyclooxygenase.

Soybean lipoxygenase and cyclooxygenase from ram seminal vesicles were inhibited by tannic acid and the apparent ID50's were 7.5 x 10(-7) M and 6 x 10(-6) M, respectively. The inhibition of lipoxygenase by tannic acid was noncompetitive. Benzophenone inhibited cyclooxygenase, and the ID50 was 8 x 10(-7) M. The inhibition constant, Ki values of both tannic acid and benzophenone are presented.

Animals↗
PubMed1985

Effect of electrical stimulation on phosphoinositide metabolism in rat sciatic nerve in vivo.

The metabolism of phosphoinositides in rat sciatic nerves in vivo during electrical stimulation was studied. Nerves were prelabeled by injection of [2-3H]-myo-inositol alone for periods of 2 and 20 h or together with [32P]orthophosphate for 2 h and then electrically stimulated (100 Hz) for 5 or 20 min. Contralateral unstimulated nerve served as the control. When tritiated myo-inositol was used alone for prelabeling the nerves, approximately 6% and 14% of the label was incorporated into lipids after 2 h and 20 h, respectively. Both 5 and 20 min of electrical stimulation caused an insignificant change in the percentage of radioactivity recovered in lipids from the nerves prelabeled with either myo-inositol or with a mixture of myo-inositol and phosphate. The proportion of label associated with phosphoinositides of nerves prelabeled with myo-inositol for both 2 h and 20 h showed an increase in phosphatidyl-inositol-4-phosphate at the expense of phosphatidylinositol in stimulated nerves. Similar results were obtained with nerves prelabeled for 2 h with a mixture of [32P]orthophosphate and [2-3H]myo-inositol. No significant changes in the radioactivity associated with water-soluble inositol phosphates were found in stimulated versus control nerves.

Animals↗
PubMed1983

Dietary trilinoelaidate: effects on the in vitro biosynthesis of different prostaglandins in brain, lung and stomach fundus of rats.

The effects of dietary trilinoelaidate on the biosynthesis of different prostaglandins (PG's) from [1-14C]-arachidonic acid by brain, lung, and stomach fundus homogenates of rats were determined. Rats were fed diets containing hydrogenated tallow (essential fatty acid deficient), trilinoelaidin at 0 (control), 10 and 50 weight percent of the total dietary fat and sacrificed at the end of eleven weeks. Brain, lung and stomach fundus of rats receiving trilinoelaidin at a level of 50 weight percent of the total dietary fat and the essential fatty acids deficient diet showed increased activity for the in vitro conversion of exogenous arachidonic acid to PG's compared to controls.

Animals↗
PubMed1982

Determination of prostaglandins and thromboxane as their pentafluorobenzyl-trimethylsilyl derivatives by electron-capture gas chromatography.

The optimization of the parameters affecting the chromatographic properties and separation of prostaglandin pentafluorobenzyl derivatives by gas chromatography using electron-capture detection is described. The effects of composition and flow-rate of carrier gas, temperatures of detector and column, and nature of stationary phases on the detector response to different pentafluorobenzyl (both oxime and ester) trimethylsilyl ether derivatives of prostaglandins were systemically examined. The stability of some selected prostaglandin derivatives at -20 degrees C was also determined. After standardizing these parameters, prostaglandins and related compounds from biological samples, e.g. semen, rat aorta, dog serum and trout gill were successfully analyzed. Identification of prostaglandins was confirmed by gas chromatography-mass spectrometry.

Animals↗