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Biomedical subjects

S Ganguly

Publications and source records attributed to S Ganguly.

At least 55 records · Page 3Linked to original sources
PubMed1996

Transcriptional regulation of the CYP2B1 and CYP2B2 genes by C/EBP-related proteins.

Cytochrome P450 (CYP) 2B1 and 2B2 are encoded by two closely related genes, CYP2B1 and CYP2B2, that are expressed at low levels in adult rat liver but are induced markedly by the administration of the drug phenobarbital (PB) or other structurally unrelated hydrophobic compounds to animals. Very little is understood about the molecular mechanisms that control both basal and induced transcription of these genes. We have identified two liver specific DNase I hypersensitive sites associated with the CYP2B1 and CYP2B2 (CYP2B) genes. One site, which maps to a region in the 5'-flanking region between -2.2 and -2.3 kb, became more resistant to DNase I cleavage in nuclei from PB-treated rats; the converse was true of the other hypersensitive site, which maps to the proximal promoter region between -0.05 and -0.15 kb. DNase I footprint analysis revealed three prominent and one weak footprinted regions in the promoter region in the vicinity of the proximal hypersensitive site. Using competitor oligonucleotides, we determined that one footprinted region (FT2), between -42 and -66 bp, is likely to represent a binding site for CCAATT enhancer binding protein (C/EBP) family members. Indeed, bacterial expressed recombinant C/EBP alpha bound at this site and formed a footprint pattern identical to the pattern observed with liver nuclear extract. In vitro transcription assays demonstrated that the FT2 site contributed strongly to promoter activity, since its mutation reduced transcription by 80%. Two other sites identified by footprint analysis (FT1 and FT3) are also required to maintain high basal transcription of CYP2B2 promoter constructs in an in vitro transcription assay. Transient transfection experiments confirmed the expectation that C/EBP alpha could activate the 1.4 kb CYP2B promoter constructs, with mutation of the FT2 site impairing both basal transcription and transactivation by exogenous C/EBP alpha.

Animals↗
PubMed1996

PROXIMAL: a database system for the efficient retrieval of genetic information.

We report on a query language, called PROXIMAL, for the efficient specification of complex queries over biological sequences. We present a data model for asking complex and structured questions of biological sequences, and show how the query language can be used. In addition, we describe a visual query metaphor intended for a biologist-friendly front end to the PROXIMAL query language.

Base Sequence↗
PubMed1996

Technetium-99m radiolabeled ouabagenin-cysteine conjugate: biological evaluation in animal models.

Two ouabagenin-cysteine conjugates were synthesized by condensing 3-beta monochloroacetyl and 3-beta, 11-alpha dichloroacetyl ouabagenin with cysteine. The resulting ligands were radiolabeled with technetium-99m (99mTc) to furnish a single homogenous 99mTc chelate in each case with good stability. The animal experiments with these 99mTc-labeled conjugates established the superiority of guinea pig over rat and rabbit as an animal model, as previously observed for other tritiated or radioidinated cardiac glycosides or aglycones. In biodistribution experiments in guinea pig, these 99mTc chelates showed a favorable heart to liver (and other nontarget organ) uptake ratio, comparable to that of recently reported 125I-digoxigenin iodohistamine-3-oxime. The low heart to blood ratio in animal experiments with ouabagenin derivatives could be attributed to the absence of 3-beta sugar residues in these molecules, which is in agreement with the previous observation reported in connection with radioiodinated digoxin and digoxigenin derivatives.

Animals↗
PubMed1996

Human metallothionein gene expression determined by quantitative reverse transcription-polymerase chain reaction as a biomarker of cadmium exposure.

Expression of the metallothionein (MT) gene in frozen human lymphocytes has been developed as a new molecular biomarker of heavy metal exposure. Workers at a Polish battery factory with high exposure to cadmium were monitored for airborne exposure and blood cadmium levels. A novel quantitative reverse transcription-PCR (RT-PCR) technique, making use of a homologous internal standard, was used to assess the level of MT-specific mRNA in frozen stored aliquots of blood samples taken from exposed and control workers. Results from this assay showed a statistically significant 2.5-fold increase in MT mRNA in exposed compared to control workers. The RT-PCR results also showed significant correlation with airborne cadmium, as registered on personal monitors and with blood cadmium levels. The results suggest that gene induction measured by quantitative RT-PCR is a promising approach for application as a biomarker of biologically effective dose in small samples of frozen tissues or cells.

Base Sequence↗
PubMed1995

Heavy chain dimers as well as complete antibodies are efficiently formed and secreted from Drosophila via a BiP-mediated pathway.

We have constructed a stable Drosophila cell line co-expressing heavy chain (HC) and light chain (LC) immunoglobulins of a humanized monoclonal antibody (mAb) that recognizes the F antigen of respiratory syncytial virus (Tempest, P. R., Bremmer, P., Lambert, M., Taylor, G., Furze, J. M., Carr, F. J., and Harris, W. J. (1991) Bio/Technology 9, 266-271. These cells efficiently secrete antibody with substrate binding activity indistinguishable from that produced from vertebrate cell lines. Significantly, the Drosophila homologue of the immunoglobulin binding chaperone protein (BiP), hsc72, was found to interact specifically with the immunoglobulin HC in an ATP-dependent fashion, similar to the BiP-HC interaction known to occur in vertebrate cells. This is, in fact, the first substrate ever shown to interact specifically with Drosophila hsc72. Most surprisingly, expression of heavy chains in the absence of LC led to the efficient secretion of heavy chain dimers. Moreover, this secretion occurred in association with hsc72. This dramatically contrasts with what is seen in vertebrate cells where in the absence of LC, HC remains sequestered inside the cell in stable association with BiP. Our results clearly suggest that Drosophila BiP can substitute for its mammalian counterpart and chaperone the secretion of active IgG. However, the finding that Drosophila BiP can also uniquely chaperone heavy chain dimers indicates mechanistic differences that may relate to the evolved need for retaining immature IgGs in vertebrates.

Animals↗
PubMed1995

Conversion of murine Fabs isolated from a combinatorial phage display library to full length immunoglobulins.

The use of combinatorial Ig libraries displayed on the surface of bacteriophage has advantages over traditional hybridoma techniques for the generation of mAbs but in many instances full length Igs may be more desirable than Fab fragments. Two murine Fabs reactive with the human complement component C5a, recovered from a combinatorial library, were converted to full length IgG2a mAbs. The VH and VL domains of these antibodies were removed from the bacterial expression vector used for the combinatorial library construction, and subcloned into individual mammalian expression vectors containing the corresponding Ig heavy and light chain constant regions. The subcloning relied on 5' restriction endonuclease sites encoded by the oligonucleotide primers originally used to amplify the Ig cDNAs and 3' sites conserved in CH1 and C kappa. These vectors were co-transfected into COS cells yielding full length IgG2a versions of the anti-C5a antibodies. The mAbs, purified from the culture supernatant, retained the full activity of the Fabs, binding specifically to and neutralizing human recombinant C5a. Refined versions of the mammalian expression vectors have been constructed for single step conversion of murine recombinant Fabs, recovered from combinatorial libraries, to IgG2a mAbs.

Animals↗
PubMed1995

Iodoacetylated ouabagenins: their syntheses, spectroscopic characterizations, and stability studies.

Ouabain shows high binding ability to myocardial Na+,K(+)-ATPase and, therefore, a suitably radiolabeled derivative of this compound may find use in myocardial imaging. In this pilot experiment we report the preparation of several chloroacetylated and iodoacetylated ouabagenins. These intermediates may possess the potential for conversion to 99mTc-labeled tracer by a reported procedure with which the imaging of myocardial Na+,K(+)-ATPase may be possible. Appropriate analytical and spectroscopic data for these intermediates are reported for the first time. To determine the stability of the iodoacetylated ouabagenins, corresponding 131I derivatives were synthesized which showed sufficient stability for incorporating a suitable radiometal-binding chelating moiety to these steroid molecules.

Acetylation↗
PubMed1995

Production of monoclonal antibodies in COS and CHO cells.

Recent advances in the generation of genetically engineered monoclonal antibodies have enhanced the importance of COS cells as expression systems for rapidly producing sufficient quantities of these proteins for preliminary biochemical and biophysical analysis. In order to meet the demand for clinical supplies, a gradual increase has occurred in the usage of dihydrofolate reductase negative (DHFR-) Chinese hamster ovary (CHO) cells for large-scale antibody production. Using a variety of mammalian expression vectors and selection/amplification protocols, CHO cell lines capable of producing monoclonal antibodies at levels exceeding 1 gl-1 can now be obtained in an almost routine fashion. For the applications of monoclonal antibodies to expand into additional therapeutic areas, however, a 5-10-fold increase over current highest expression levels may still need to be achieved.

Animals↗
PubMed1995

Effects of chemically-induced stress on blood viscosity of rabbits.

Disease-induced stress in a living system needs its characterization before quantification. We address this problem by studying the flow behaviour of blood of rabbits. The flow behaviour of rabbit blood, both normal and streptozotocin-induced diabetic was observed by plotting viscosity levels against known shear stress. The curves obtained from normal animals showed a curvilinear relationship between eta and tau, whereas the curves obtained from the blood of the induced diabetic animals showed a 'thixotropic' nature i.e. viscosity varies inversely with shear stress. The results obtained were (a) increased viscosity associated with increasing doses, (b) the lower the shear stress, higher was the viscosity and (c) significant variation of viscosity levels between normal and induced-diabetic rabbits observed by increased stress. The possible interpretation of the results in the light of rheological properties of blood is discussed.

Animals↗
PubMed1995

The prevalence and spectrum of colonic lesions in patients with cirrhotic and noncirrhotic portal hypertension.

Portal hypertension diffusely affects the gastrointestinal tract. The frequency and profile of distinct colonic mucosal lesions (portal colopathy) and rectal varices (RV; veins > 4 cm above the anal verge) is not well studied. Fifty consecutive patients with portal hypertension (25 with cirrhosis, 10 with noncirrhotic portal fibrosis [NCPF], and 15 with extrahepatic portal vein obstruction [EHPVO]) were assessed clinically and by upper and lower gastrointestinal (GI) endoscopy. Colorectal lesions were seen in 35 (70%) patients, significantly more often in bleeders than in nonbleeders. Rectal varices were detected in 22 (44%) patients; larger and more often seen in EHPVO (80%) than in cirrhosis (28%) and NCPF (30%) (P < .01) patients. Portal colopathy was seen in 26 (52%) patients, with nearly similar frequency in cirrhotics, NCPF, and EHPVO patients. Previous sclerotherapy or presence of gastric varices had little influence on the development of these lesions. An association (P < .01) was, however, seen between the presence of colopathy and portal gastropathy. Overt bleeding was seen in 8% and 4% of patients with RV and colopathy, respectively. In conclusion, our results demonstrate that colorectal lesions are present in about two thirds of patients with portal hypertension. Patients with portal hypertension and lower GI bleeding should be colonoscoped. Patients with extrahepatic portal vein obstruction may in turn benefit from baseline sigmoidoscopic examination to define the presence and size of rectal varices.

Adolescent↗