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Biomedical subjects

D Banerjee

Publications and source records attributed to D Banerjee.

At least 199 records · Page 11Linked to original sources
PubMed1990

A flow cytometric assay for target binding by NKH1A+ cells using a single laser system.

A modified method of analysis by flow cytometry has been adapted to the NK/target cell system to monitor conjugate formation within a mixed population. The two-parameter assay is based on the use of two colour fluorescent markers in which an indirect FITC-conjugated NKH1A antibody (green fluorescence) is used to detect NK-specific effector cells and an intracellular strain, hydroethidine (red fluorescence) is used to detect target cells. The two cell populations and their conjugates are discernible by the presence of both green and red fluorescence, using a single laser beam tuned to 488 nm. In comparison, a manual count conjugate assay was applied using a hemacytometer and fluorescence microscope. Of the two methods, the automated technique using flow cytometry provides a lower but more precise representation of conjugate formation, due to the avoidance of technical/observer error that is common to manual count assays. The cytometric method has proven to be reproducible and superior in consistency to the manual count assay and can be adapted to almost any effector/target system under investigation. In comparison with previous methods using dual laser, the single laser system provides advantages of cost and time efficiency due to setup simplicity, as well as improved availability.

Antibodies, Monoclonal↗
PubMed1990

Differential effect of retinoic acid on ADP and collagen induced platelet aggregation.

Retinoic acid (RA) was found to inhibit ADP induced but not collagen induced aggregation of human platelets and the differential action is related to intraplatelet Ca2+ reflux. RA was active at concentrations as low as 10(-7) M and required 20 min prior incubation with platelet suspension in order to inhibit aggregation by ADP. All the steps in ADP induced but not collagen induced platelet activation, viz. hydrolysis of phosphatidyl inositol, phosphorylation of 20, 47 and 250 kDa proteins as well as increased association of actin with Triton X-100 insoluble cytoskeletal matrix were inhibited by RA. RA when used as an agent for differentiation induction of cell progenitor is likely to affect the platelet aggregation and thereby the haemostatic process.

Adenosine Diphosphate↗
PubMed1990

Proteinaceous lymphadenopathy due to monoclonal nonamyloid immunoglobulin deposit disease.

We report two cases of lymph node enlargement due to massive extracellular nonamyloid immunoglobulin deposits that obscured the underlying cellular pathologic condition. In both cases, the deposits were demonstrated to be restricted to a single heavy and light chain, consistent with a monoclonal paraprotein, and cytoplasmic staining in the lymphocytes or plasma cells was identical to that of the paraprotein. The use of the protein A-gold technique was instrumental in revealing a monoclonal pattern in one case in which light microscopic immunohistochemistry did not reveal a clear-cut monoclonal pattern in the extracellular deposits. This case was subsequently shown to have multiple myeloma, while the second case has had an unusual history of hypocomplementemic vasculitis and normal bone marrow. Neither case had evidence of significant renal disease.

Adult↗
PubMed1990

HBV & HIV seropositivity in multi-transfused haemophilics & thalassaemics in eastern India.

Seropositivity to HBV (HBsAg) in multi-transfused patients of haemophilia A, haemophilia B, B thalassaemia and EB thalassaemia from Eastern India, was found to be 9, 0, 22.1 and 13 per cent respectively. HIV seropositivity was detected in patients of haemophilia A (4.4%) and B thalassaemia (0.8%) who received plasma components and packed cells periodically. Seropositivity to both HBsAg and HIV was found in one patient of haemophilia A. Serum alanine amino transferase (ALT), raised in multi-transfused thalassaemics suggests concurrent hepatitis which might have enhanced the transmission of viruses due to disturbed immune status. The universal voluntary blood donation programme, screening of blood for HBV and HIV by sensitive tests, early immunisation and periodic monitoring of HBV and HIV status are prerequisites for the management of transfusion dependent thalassaemia and haemophilia.

Blood Transfusion↗
PubMed1990

Effect of retinoic acid on adenosine diphosphate and collagen-induced alterations in enzymes of GSH-linked antioxidant defence system of human blood platelets in vitro.

Collagen stimulation of blood platelets resulted in significant increases in malondialdehyde (MDA) formation and activity of glucose-6-phosphate dehydrogenase (G6PDH) and a decrease in catalase and glutathione peroxidase (GPx). Retinoic acid (RA) pretreatment did not show any appreciable changes except for a decrease in G6PDH activity as compared with collagen alone. RA pretreatment of human blood platelets resulted in an increase in the activities of catalase and GPx, two important radical scavenging enzymes, with significant decrease in MDA formation when compared with ADP alone. It is suggested that RA has a significant effect on the antioxidant defence system in ADP stimulated platelets but not in the collagen stimulated platelets.

Adenosine Diphosphate↗
PubMed1989

Expression and secretion of chicken apolipoprotein AI in transfected COS cells.

A full-length chicken apolipoprotein A-I (apoAI) cDNA has been cloned into an expression vector, pRSVapoAI. This plasmid was transfected into a monkey kidney (COS-1) cell line in order to study apolipoprotein-lipid assembly. Chicken apoAI is the major apolipoprotein of chicken high-density lipoprotein (HDL), which is less complex in apolipoprotein content than the HDL of human plasma. The transient transfected COS-1 cells synthesized and secreted authentic plasma apoAI. Under serum-free medium conditions, COS cells secreted only proapoAI. A small portion (15%) of the secreted apoAI floated at a density 1.07-1.20 g/ml. Upon incubation with fetal bovine serum at 10 degrees C, a majority of the apoAI was recovered in the HDL density (1.06-1.20 g/ml) region. Secreted apoAI was labeled when transfected COS cells were incubated with [U-14C]palmitate, but the incorporation of radioactivity was not the result of fatty acid acylation through ester bond formation. These results indicate that heterologous COS-1 cells are capable of synthesizing and secreting apoAI, and that intracellular association of apoAI with lipids is not necessary for secretion.

Acylation↗
PubMed1989

Immunohistochemical localization of metallothionein in human testicular embryonal carcinoma cells.

The presence of high levels of metallothionein (MT) in developing mammalian cells is well documented. It has been suggested that the developmental profile and gene expression of MT is similar to that of the so-called oncodevelopmental gene products such as a-fetoprotein. In this study tissue sections of nine human embryonal carcinomas of the testis were tested by means of the avidin-biotin peroxidase complex for the presence of MT. The antigen was localized in variable amounts in the cytoplasm and nucleus in tumour cells in all cases. There was evidence that immunoreactivity was related to the histological growth pattern of tumour cells. These findings suggest that MT may be considered an oncodevelopmental product which could be useful as a tumour marker. In addition, the histology of these tumours might predict MT expression; this may prove of value in testing the hypothesis of MT-related emergence of drug-resistant cell lines in the course of treatment of tumours with metal-containing chemotherapeutic agents.

Adolescent↗
PubMed1989

T-cell-mediated protection of mice against virulent Mycobacterium tuberculosis.

We sought to protect CBA mice against tuberculosis using in vivo transfer of a T-cell line previously shown to be capable of I-A-restricted recognition of peritoneal macrophages infected in vitro with Mycobacterium tuberculosis. This line induces total bacteriostasis in vitro. In mice that received 500 rads of irradiation 48 h before infection, the T-cell line caused significant prolongation of life when given intravenously with a challenge dose of 5 x 10(6) organisms. Similar experiments with two other T-cell lines showed that these lines offered no protection. Bacterial load at the time of death was inversely related to the time of survival. Thus, death occurred at a lower bacterial load in adoptively protected mice, implying the contribution of an immunopathological component in these animals. The protective T-cell line, which was CD4+ CD8-, had no effect on the rate of growth of strain BCG in CBA nu/nu mice or M. tuberculosis in fully T-cell-deprived mice. This could indicate that CD8+ cells play a role in this system or that there is a need for the recruitment of interleukin 2-producing cells in the recipient. Experiments with monoclonal antibodies to selectively deplete T-cell subsets in normal CBA mice showed that depletion of CD4+ cells strikingly shortened survival, whereas depletion of CD8+ cells did not. However, CD8-depleted mice died with a lower bacterial load than those found in nondepleted controls, and the lesions in CD8-depleted mice were histopathologically distinct. These results suggest that the CD8+ cells either down-regulate bacteriostasis or cause immunopathology in this model and that it is the CD4+ cells that are the major protective subset in long-term protection experiments.

Animals↗
PubMed1989

A simple method for preparation of good quality cryoprecipitate.

Production of good quality cryoprecipitate from plasma was achieved, as assessed by quantitating Factor VIII coagulant activity (F VIII C) and Factor related antigen (F VIII R: Ag). The method used resulted in good recovery of FVIII in the cryoprecipitate containing F VIII C: Fibrinogen 0.82 +/- 0.015 IU/mg. The method therefore appears suitable for indigenous preparation of F VIII in standard blood banks as a replacement therapy, which is not expensive.

Centrifugation↗
PubMed1989

Detection of haemophilia A traits in carriers.

Obligate carriers of haemophilia A and normal women matched for age were studied in order to find a suitable parameter for detection of carriers of haemophilia A. The ratio of factor VIII coagulant activities (F VIIIC) to factor VIII related antigen (F VIII R : Ag) was 0.55 in obligate carriers and 1.0 in non-carrier females.

Adult↗
PubMed1989

Ferritin release by mononuclear cells in hereditary hemochromatosis.

An anomaly of the iron-loading disorder hereditary hemochromatosis is that bone marrow iron stores remain low until later stages of the disease. The possibility that this may be related to a disorder of reticuloendothelial ferritin metabolism was examined by studying ferritin release from mononuclear cells. Ferritin release was measured in peripheral blood mononuclear cells from four patients with hemochromatosis who had not received treatment, from six patients with hemochromatosis who had received treatment, and from 10 age- and gender-matched controls by using a modified hemolytic plaque assay. Ferritin release from the hemochromatotic cells was enhanced when compared with that of controls, and added iron stimulated ferritin release to a comparable degree in both groups. Enhanced ferritin release above matched control values was found both in cells from patients with hemochromatosis with partial phlebotomy who had high serum ferritin values and in cells from patients with hemochromatosis with full phlebotomy who had normal serum ferritin values. The increased ferritin release observed in these studies may signify abnormal reticuloendothelial iron metabolism in hemochromatosis.

Aged↗
PubMed1988

Retinoic acid inhibits phospholipid turnover and protein kinase C activity in RA-sensitive but not in RA-resistant cells.

Treatment with 10(-5) M retinoic acid causes loss of anchorage-independent growth in src-transformed RR1022 cells but not in ras-transformed KNRK cells. In an effort to elucidate the mechanisms underlying this difference, we investigated the effect of RA on phospholipid turnover and PKC activity in these two cell lines. 10(-5) M RA treatment caused a drastic inhibition of 32P incorporation into PI and PA and a large increase in 32P incorporation into PC in RR1022 cells. Similar treatment of KNRK cells yielded no change in PC or PA labelling and a much smaller decrease in PI labelling. Furthermore, 10(-5) M RA treatment causes a large decrease in PKC activity in RR1022 cells (35% of control) but only a small decrease in KNRK cells (78% of control). We suggest that these effects are part of an altered signal transduction pathway which mediates the differential effects of RA on anchorage-independent growth in these two cell lines.

Animals↗
PubMed1988

Flow cytometric analysis of the phenotypic changes in tumour cell lines following TPA induction.

Single-cell analysis by flow cytometry has enabled us to analyze the effects of a phorbol ester and known tumour promoter, TPA, on the phenotypes of four tumour lines. TPA is capable of triggering a variety of cellular alterations that can affect gene expression and the biochemical balance of intracellular events. We have investigated the effect of TPA on such properties as rate of proliferation, differentiation, expression of cell surface molecules, and susceptibility to natural killer (NK) cell-mediated cytolysis. Four human leukemia and lymphoma cell lines; K562, MOLT 4, Raji, and HL60, were studied in their response to TPA treatment. Based on measurements of the defined cellular properties, we have characterized the pleiotropic responses of each tumour cell line to the phorbol ester in relation to intensity and time of onset of each response. The effects of TPA are highly varied, ranging in time of onset from minutes to days, and in intensity from strong to weak within the four cell lines studied. However, within all the processes that are affected, the activation of protein kinase C appears to be a common initiating event of phorbol ester induction.

Antigens, Neoplasm↗
PubMed1988

Suppression of immunoglobulin production by germinal centre HNK-1+ CD3+ cells.

Germinal centre T cells co-expressing HNK-1 antigen have little lytic activity against NK targets (K562 cells). In order to determine whether these cells regulate B cell function, they were purified from human tonsils by panning on anti-HNK-1 antibody coated Petri dishes and co-cultured with autologous and allogeneic tonsillar T and B cells in the presence of pokeweed mitogen. At the end of 7 days of culture, supernatants were assayed for immunoglobulin concentration by ELISA. A dose-dependent suppression of both IgG and IgM production was demonstrated at ratios from 1:125 to 1:16 of HNK-1+ cells to B cells, but enhancement was observed at very low ratios (less than 1:500) or ratios exceeding 1:16 in some tonsil preparations. Similar results were obtained with peripheral blood HNK-1+ cells but without enhancement in some cases at the extremes of HNK-1+ cells to B cell ratios. The suppression was not MHC-restricted. These preliminary experiments indicate that germinal centre HNK-1+ cells may be intrafollicular suppressor cells.

Antibody Formation↗
PubMed1988

Transferrin receptor and 4F2 expression by NK-sensitive and NK-resistant tumour cell lines.

Upon stimulation with a phorbol ester and known tumour promoter, 12-O-tetradecanoylphorbol-13-acetate (TPA), the human erythroleukemia K562 cell line, a standard target for human natural killer (NK) cells, shows a significant reduction in expression of transferrin receptors (TfR) and becomes resistant to NK-mediated cytolysis. Prompted by the initial finding that expression of TfRs by K562 cells correlates with target sensitivity to NK cytotoxicity, we have investigated the role of two proposed NK-target structures, the TfR and 4F2 molecule in three other tumour cell lines which vary in their susceptibility to NK activity. We report that similar to K562 cells, MOLT 4, Raji and HL60 cells demonstrate phorbol-ester-induced down-regulation of TfR expression. The expression of TfRs alone does not determine target sensitivity. However, TfRs are probably located close to the NK-target structure in NK-sensitive cell lines. Based on the properties of phorbol ester induction, it is possible that the NK-target molecule is down-regulated in response to phorbol ester induction in a similar, if not identical manner to that of the TfR; thus, rendering NK-sensitive cells resistant to NK killing, after TPA exposure. Conversely, the target molecule is probably upregulated in MOLT 4 cells after TPA treatment as indicated by the acquired sensitivity of these cells to NK-mediated cytolysis. The expression of 4F2 molecules was not influenced by TPA treatment and does not correlate with NK sensitivity.

Antigens, Surface↗
PubMed1988

Localization of the McLeod locus (XK) within Xp21 by deletion analysis.

The McLeod phenotype is an X-linked, recessive disorder in which the red blood cells demonstrate acanthocytic morphology and weakened antigenicity in the Kell blood group system. The phenotype is associated with a reduction of in vivo red cell survival, but the permanent hemolytic state is usually compensated by erythropoietic hyperplasia. The McLeod phenotype is accompanied by either a subclinical myopathy and elevated creatine kinase (CK) or X-linked chronic granulomatous disease (CGD). Seven males with the McLeod red-blood-cell phenotype and associated myopathy but not CGD, one male with the McLeod phenotype associated with CGD, and two males known to possess large deletions of the Duchenne muscular dystrophy (DMD) locus were studied. DNA isolated from each patient was screened for the presence or absence of various cloned sequences located in the Xp21 region of the human X chromosome. Two of the seven males who have only the McLeod phenotype and are cousins exhibit deletions for four Xp21 cloned fragments but are not deleted for any portion of either the CGD or the DMD loci. Comparison of the cloned segments absent from these two McLeod cousins with those absent from the two DMD boys and the CGD/McLeod patient leads to the submapping of various cloned DNA segments within the Xp21 region. The results place the locus for the McLeod phenotype within a 500-kb interval distal from the CGD locus toward the DMD locus.

Adult↗
PubMed1987

Epidermal and transforming growth factors modulate secretion of a 69 kDa phosphoprotein in normal rat kidney fibroblasts.

Our study shows that the secretion of a major non-glycosylated, phosphoprotein of 69 kDa (pp69) is a specific marker for non-transformed NRK-49F cells. Treatment of NRK-49F cells with EGF alone or with different combinations of EGF plus TGF-beta modulates the secretion of pp69, suggesting its relationship with cellular proliferation. Antibody raised against pp69 recognizes, in addition to pp69, another major phospho-protein of 62 kDa (pp62) secreted by RR1022 and spontaneously transformed NRK-49F cells. Immunoprecipitation of total cell lysates from both NRK-49F and RR1022 cells with anti-pp69 antibody detected only pp69. These observations suggest a precursor-product relationship between pp69 secreted by non-transformed NRK-49F cells and pp62 secreted by transformed cells.

Animals↗