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Biomedical subjects

B B Aggarwal

Publications and source records attributed to B B Aggarwal.

At least 145 records · Page 8Linked to original sources
PubMed1994

Inhibition by all-trans-retinoic acid of tumor necrosis factor and nitric oxide production by peritoneal macrophages.

Besides their growth-inhibiting and differentiation-inducing properties, retinoids have been shown to exert immunomodulatory and anti-inflammatory functions by mechanisms that are not well understood. Tumor necrosis factor-alpha (TNF), a cytokine produced by mononuclear phagocytes, has been shown to be an important mediator of endotoxin-induced septic shock, cachexia, bone resorption, and inflammation. Nitric oxide may also have a role in septic shock, hypotension, and vasodilatation. In this study, we examined the effects of retinoids on the production of TNF and nitric oxide by murine peritoneal macrophages. Of the various retinoids studied, all-trans-retinoic acid (RA) was most potent; it almost completely inhibited the production of TNF by macrophages activated with endotoxin and interferon-gamma. The inhibitory effect was dependent on the dose and duration of RA exposure to macrophages. RA also blocked phorbol ester-induced TNF production in a macrophage cell line (RAW 264.7). Besides TNF, the retinoid suppressed the production of nitric oxide from activated peritoneal macrophages. The importance of these results in relation to controlling various harmful effects of cytokines released by activated macrophages is discussed.

Animals↗
PubMed1994

Suramin inhibits tumor cell cytotoxicity mediated through natural killer cells, lymphokine-activated killer cells, monocytes, and tumor necrosis factor.

Suramin, a polysulfonated naphthylurea, is an antitrypanosomal and antifilarial drug. Because of its anti-reverse transcriptase activity and antiproliferative activity, suramin is also used for the treatment of acquired immunodeficiency syndrome and cancer. In spite of these uses, very little is known about its effects on the immune system. In this report, we investigated the effects of suramin on peripheral blood mononuclear cells. We found that natural killer (NK) cell-mediated cytotoxicity against human erythroblastoid cell line K562 was completely inhibited by suramin in a dose-dependent manner. It also completely blocked lymphokine-activated killer (LAK) cell-mediated cytotoxicity against the human B lymphoblastoid cell lines Raji and Daudi. The cytotoxicity against the human melanoma tumor cell line A-375 mediated by unstimulated and stimulated monocytes was also suppressed by suramin. Maximum inhibition of monocyte-mediated cytotoxicity was observed when suramin was present during both the activation and the effector phases of cytotoxicity. Besides its effects on cell-mediated cytotoxicity, suramin also inhibited the cytotoxic effects of tumor necrosis factor (TNF) against different tumor cell lines. Furthermore, we found that suramin interferes with the binding of TNF with its receptor. Thus our results indicate that suramin overall downregulates the immune system by inhibiting cell-mediated and TNF-mediated cytotoxicity against different tumor cells.

Animals↗
PubMed1994

Effect of tumor necrosis factors, interferons, interleukins, and growth factors on the activation of NF-kappa B: evidence for lack of correlation with cell proliferation.

The nuclear transcription factor NF-kappa B has been identified as a critical component in signal transduction pathways. We used an electrophoretic gel mobility shift assay to examine the activation of NF-kappa B in human U-937 cells treated with tumor necrosis factor (TNF), lymphotoxin (LT), interferons (IFN)-alpha, IFN-beta, and IFN-gamma, interleukins (IL)-1 beta, IL-4, and IL-6, leukemia inhibitory factor (LIF), basic fibroblast growth factor (FGF), granulocyte-macrophage colony-stimulating factor (GM-CSF), and transforming growth factor-beta (TGF-beta). Only TNF, LT, and IL-1 activated NF-kappa B. Since interferons have been shown to induce TNF receptors and potentiate TNF-mediated cellular responses, we also measured the effect of interferons on TNF-induced activation of NF-kappa B. Under our conditions, all three IFNs potentiated the cytotoxic effects of TNF but had no effect on the TNF-dependent NF-kappa B activation. These results suggest overall that the activation of NF-kappa B is not a generalized mediator of signal transduction of most cytokines and also that NF-kappa B activation is not sufficient for antiproliferative effects mediated through certain cytokines.

Base Sequence↗
PubMed1993

P80 form of the human tumor necrosis factor receptor is involved in DNA fragmentation.

Two different types of TNF receptors with molecular masses of 60 kDa (p60) and 80 kDa (p80) have been identified. TNF is known to cause DNA fragmentation in certain tumor cell lines but the role of p60 and p80 in this action is not understood. In the present study, we examined the role of these receptors in TNF-induced DNA fragmentation. Treatment of U-937 cells with phorbol ester caused downregulation of both types of TNF receptors and this was accompanied by disappearance of the TNF-induced DNA fragmentation. The removal of phorbol ester led to two time-dependent events: (1) the rapid regeneration of the p80 form but not the p60 form of the TNF receptor; and (2) the reappearance of TNF-induced DNA fragmentation. These results suggest that the p80 receptor could mediate the TNF-induced DNA fragmentation.

Cells, Cultured↗
PubMed1993

Okadaic acid induces down-modulation and shedding of tumor necrosis factor receptors. Comparison with another tumor promoter, phorbol ester.

Protein kinase C modulates the receptor for tumor necrosis factor (TNF) in wide variety of different cell types. However, there is no information about the role of phosphatases in the regulation of the TNF receptor. In this report, we investigated the effect of okadaic acid, an inhibitor of serine/threonine phosphatases, on TNF receptors in U-937 cells, a human histiocytic lymphoma cell line. In our study okadaic acid induced a dose- and time-dependent down-modulation of TNF receptors. On exposure of cells to 0.5 microM okadaic acid for 60 min at 37 degrees C, a complete down-regulation of the receptors was observed, but no modulation occurred at 4 degrees C. Scatchard analysis of the binding data on U-937 cells revealed that okadaic acid caused a decrease in the high affinity cell surface receptor number without a significant change in the affinity constant. The down-regulation of the TNF receptor by okadaic acid was not specific to U-937 cells, as it was also observed with several other cell types. Okadaic acid had no significant effect on the internalization of the receptor, but it did induce a shedding of the TNF receptor from its cell surface. On exposure of cells to okadaic acid, a dose- and time-dependent increase in the 40-kDa polypeptide was detected in the medium with anti-p80 antibodies by Western blot analysis. The secreted product was also found to bind TNF. The mechanism by which okadaic acid down-modulates the TNF receptor appears to be quite different from that of phorbol ester. First, okadaic acid synergistically potentiated the effect of phorbol ester. Second, the phorbol ester-mediated down-modulation could be blocked by H-7 and staurosporine, well known protein kinase C inhibitors, but these inhibitors had no effect on okadaic acid-mediated response. Third, phorbol ester-mediated down-modulation of the TNF receptor was reversible after removal of the agent, but the response mediated through okadaic acid was irreversible. Fourth, the TNF receptor was down-regulated by okadaic acid but not by phorbol ester in cells depleted of protein kinase C. Thus, overall, our results demonstrate that serine/threonine phosphatases can down-modulate and induce shedding of TNF receptors by a mechanism distinct from that of protein kinase C.

Down-Regulation↗
PubMed1993

Lymphotoxin is an autocrine growth factor for Epstein-Barr virus-infected B cell lines.

Because human lymphotoxin (LT) was originally isolated from a lymphoblastoid cell line, we investigated the role of this molecule in three newly established Epstein-Barr virus (EBV)-infected human B cell lines. These lines were derived from acute lymphoblastic leukemia (Z-6), myelodysplastic syndrome (Z-43), and acute myelogenous leukemia (Z-55) patients who had a prior EBV infection. Each lymphoblastoid cell line had a karyotype that was different from that of the original parent leukemic cells, and all expressed B cell, but not T cell or myeloid surface markers. In all three lines, rearranged immunoglobulin heavy chain joining region (JH) bands were found, and the presence of EBV DNA was confirmed by Southern blotting. Z-6, Z-43, and Z-55 cell lines constitutively produced 192, 48, and 78 U/ml LT, respectively, as assessed by a cytotoxicity assay and antibody neutralization. Levels of tumor necrosis factor (TNF) were undetectable. Scatchard analysis revealed that all the cell lines expressed high-affinity TNF/LT receptors with receptor densities of 4197, 1258, and 1209 sites/cell on Z-6, Z-43, and Z-55, respectively. Furthermore, labeled TNF binding could be reversed by both unlabeled TNF, as well as by LT. Studies with p60 and p80 receptor-specific antibodies revealed that the three lines expressed primarily the p80 form of the TNF receptor. When studied in a clonogenic assay, exogenous LT stimulated proliferation of all three cell lines in a dose-dependent fashion at concentrations ranging from 25 to 500 U/ml. Similar results were obtained with [3H]TdR incorporation. Monoclonal anti-LT neutralizing antibodies at concentrations of 25-500 U/ml inhibited cellular multiplication in a dose-dependent manner. It is interesting that in spite of a common receptor, TNF (1,000 U/ml) had no direct effect on Z-55 cell growth, whereas it partially reversed the stimulatory effect of exogenous LT. In addition, TNF inhibited Z-6 and Z-43 cell proliferation, and its suppressive effect was reversed by exogenous LT. Both p80 and p60 forms of soluble TNF receptors suppressed the lymphoblastoid cell line proliferation and their inhibitory effect was partially reversed by LT. Our data suggest that (a) LT is an autocrine growth factor for EBV-transformed lymphoblastoid B cell lines; and (b) anti-LT antibodies, soluble TNF/LT receptors, and TNF itself can suppress the growth of lymphoblastoid cells, probably by modulating or competing with LT.(ABSTRACT TRUNCATED AT 400 WORDS)

Adult↗
PubMed1993

Regulation of two forms of the TNF receptors by phorbol ester and dibutyryl cyclic adenosine 3',5'-monophosphate in human histiocytic lymphoma cell line U-937.

Recently the cDNA for two different forms of TNF receptor, with gene products of molecular masses of 60 and 80 kDa, have been cloned. In the present report, we investigated the effects of phorbol ester and dibutyryl cAMP on the regulation of the transcript for each type of TNF receptor in U-937 cells. Our results indicate that exposure of these cells to either phorbol ester or dibutyryl cAMP increases the steady state mRNA levels of the 80 kDa form. This effect is dose- and time-dependent. The induction of the p80 receptor transcript by PMA and dibutyryl cAMP was additive suggesting independent mechanisms of induction. Under identical conditions, both agents failed to induce the transcript for the p60 form of the TNF receptor. As demonstrated by actinomycin D pulse-chase experiment, the mRNA for the p80 receptor was found to be highly stable with an approximate half-life of 16 h. No significant change in the half-life was observed when cells were treated with phorbol ester. The mechanisms by which phorbol ester and dibutyryl cAMP induce the upregulation of p80 receptor mRNA appear to be different. Induction of receptor transcript by cycloheximide suggests the presence of a labile repressor protein. Interestingly, the effect of cycloheximide on the induction of the p80 mRNA was found to be additive with that of dibutyryl cAMP but not with phorbol ester. 1-(5-Isoquinolinylsufonyl)-2-methylpiperazine (H7) and N[2-(methylamino) ethyl]-5-isoquinolinesulfonamide (H-8), inhibitors of protein kinase C and protein kinase A, respectively, both inhibited the phorbol ester-mediated induction of the p80-transcript but not that mediated through dibutyryl cAMP. Since dibutyryl cAMP undergoes intracellular dissociation into cAMP and butyric acid, we found that exposure of cells to sodium butyrate alone could induce p80 mRNA in a dose-dependent manner, thus suggesting the role of histone hyperacetylation. Furthermore forskolin treatment, an intracellular inducer of cAMP, increased the receptor transcript level whereas isobutylmethylxanthine, an inhibitor of phosphodiesterase, had no effect. Interestingly, while the p80 form of the TNF receptor mRNA levels was elevated by both phorbol ester and dibutyryl cAMP, only dibutyryl cAMP increased the TNF binding; phorbol ester treatment decreased the binding activity. Thus, our results demonstrate that the genes for the two forms of TNF receptors are differentially regulated. Furthermore, the mechanism of regulation by PMA differs from that by dibutyryl cAMP.

1-Methyl-3-isobutylxanthine↗
PubMed1993

Induction of chromosomal rearrangement by tumor necrosis factors produced by primary B lymphoblastoid cells derived from cancer patients.

We have previously reported the isolation of the cytotoxic factors, tumor necrosis factor-alpha (TNF-alpha), from monocytes and lymphotoxin or TNF-beta, from an established human B lymphoblastoid cell line. In the current study, we examined the production of cytotoxic factors by primary lymphoid cells derived from patients with different malignancies and correlated it with the chromosomal abnormalities in producer cells. Lymphoid cell lines were established from 78 untreated patients with breast carcinoma, 2 with cervical carcinoma, 24 with colon carcinoma, 12 with prostate carcinoma, 3 with melanoma, and the remaining from asymptomatic family members. Lymphoid cells from all 119 patients and their asymptomatic family members constitutively produced as much as 240 units/ml of TNF-beta in vitro. In contrast, TNF-alpha was produced by lymphoid cells from only 6 patients; a related cytokine with similar biological properties to TNF-beta. Sixty-five of these 119 samples were also analyzed for chromosomal abnormalities by standard cytogenetic technique. The production of TNFs was accompanied with varying degrees of chromosomal abnormalities in the cells from all patients. These included both structural and numerical abnormalities. We also examined the direct effect of TNF-beta on the induction of chromosomal abnormalities in growing cultures of both T and B lymphocytes. Our preliminary results indicate that treatment of lymphocytes with this cytokine induced chromosomal rearrangements in a dose-dependent manner. Thus, we provide for the first time both indirect and direct evidence that a soluble mediator such as TNF-beta can induce chromosomal alterations commonly associated with different types of tumors.

B-Lymphocytes↗
PubMed1992

Alpha-lipoic acid is a potent inhibitor of NF-kappa B activation in human T cells.

Acquired immunodeficiency syndrome (AIDS) results from infection with a human immunodeficiency virus (HIV). The long terminal repeat (LTR) region of HIV proviral DNA contains binding sites for nuclear factor kappa B (NF-kappa B), and this transcriptional activator appears to regulate HIV activation. Recent findings suggest an involvement of reactive oxygen species (ROS) in signal transduction pathways leading to NF-kappa B activation. The present study was based on reports that antioxidants which eliminate ROS should block the activation of NF-kappa B and subsequently HIV transcription, and thus antioxidants can be used as therapeutic agents for AIDS. Incubation of Jurkat T cells (1 x 10(6) cells/ml) with a natural thiol antioxidant, alpha-lipoic acid, prior to the stimulation of cells was found to inhibit NF-kappa B activation induced by tumor necrosis factor-alpha (25 ng/ml) or by phorbol 12-myristate 13-acetate (50 ng/ml). The inhibitory action of alpha-lipoic acid was found to be very potent as only 4 mM was needed for a complete inhibition, whereas 20 mM was required for N-acetylcysteine. These results indicate that alpha-lipoic acid may be effective in AIDS therapeutics.

Acetylcysteine↗
PubMed1992

Interferon-gamma induces cell surface expression for both types of tumor necrosis factor receptors.

Interferons are known to potentiate various biological effects of tumor necrosis factor (TNF). Recently, two different types of TNF receptors with molecular masses of 60 kDa (p60) and 80 kDa (p80), primarily expressed by epithelial cells and myeloid cells, respectively, have been identified. In the present report, we examined the effect of interferon-gamma (IFN-gamma) on each type of TNF receptor. Our results indicate that IFN-gamma induces TNF receptors on both myeloid (e.g. HL-60) and epithelial cells (e.g. HeLa). Furthermore, by using antibodies specific to each type of receptor, we demonstrate that both TNF receptors are equally inducible by IFN-alpha, IFN-beta and IFN-gamma. Thus, the increase in TNF receptors by interferons may play a role in their synergistic cellular response.

Cell Line↗
PubMed1992

Modulation of two forms of tumor necrosis factor receptors and their cellular response by soluble receptors and their monoclonal antibodies.

Recently, two different receptors for human tumor necrosis factor (TNF) with molecular masses of 60 kDa (p60) and 80 kDa (p80) have been identified. In this report, we investigated the effect of the soluble forms of these receptors and monoclonal antibodies against them on ligand interaction, receptor down-regulation, and mediation of cellular response in U-937 cells. Our results indicate that p60 and p80 constitute 20-30 and 60-80% of the total TNF-binding sites on U-937 cells, respectively. However, by cross-linking, only the p80 form of the receptor could be detected. In contrast to unlabeled TNF, the anti-p60 and anti-p80 antibodies together only partially inhibited ligand binding, and this inhibition was not additive. Lack of additive inhibition of binding was found to be not due to stereo-chemical hindrance. TNF binding to cells can be completely displaced by soluble forms of either the p60 or p80 receptor. However, 100-fold more of the p80 than the p60 form of the soluble receptor is needed for equivalent displacement. Under optimum conditions, TNF and the anti-p80 and anti-p60 antibodies down-regulated 30, 80, and 20% of the TNF receptors, respectively. The anti-p60 and anti-p80 antibodies down-regulated not only their own receptors, but also reciprocal receptors, suggesting a cross-communication between the p60 and p80 forms of the TNF receptor. In spite of inhibiting as much as 80% of TNF binding, none of the receptor antibodies significantly inhibited the cytotoxic response to TNF in U-937 cells. Soluble forms of both receptors, however, completely abrogated the cellular response to TNF. Thus, overall, our results indicate that the antibodies against both receptors together inhibit the majority of the receptor-ligand interaction without any significant effect on the biological response to TNF.

Antibodies, Monoclonal↗
PubMed1992

Human immunodeficiency virus type 1 tat gene up-regulates interleukin 4 receptors on a human B-lymphoblastoid cell line.

The human immunodeficiency virus type I (HIV-1) regulatory gene, tat III, is a powerful trans-activator of gene expression from the viral long terminal repeat and is essential for HIV replication. In addition, tat III protein has been shown to be immunosuppressive as indicated by the inhibition of antigen mediated T-cell proliferation. To further test whether tat III might play a direct role in the immunosuppressive effects of HIV-1 in addition to its role in virus replication, we examined the regulation of interleukin 4 (IL-4) receptors on a human B-lymphoblastoid cell line (Raji) transfected with HIV-1 tat gene (Raji-tat III). We used radioligand receptor binding analysis for cell surface expression and Northern blot analysis for the expression of human IL-4 receptor gene in Raji-tat III cells. Control Raji cells expressed 1383 +/- 361 (SE; n = 3) IL-4 binding sites/cell with a dissociation constant (Kd) of 144 +/- 27 pM (n = 3). However, Raji-tat III cells expressed about three times higher IL-4 receptors (4000 +/- 633 IL-4 binding sites/cell; P less than 0.03 compared to Raji cells) with a similar Kd of 273 +/- 90 pM (n = 3; P greater than 0.05 compared to Raji cells). Whereas both Raji and Raji-tat III cells exhibited a single mRNA species (approximately 4 kilobases) of IL-4 receptors by Northern blot analysis, the mRNA level was about 3-fold higher in Raji-tat III cells compared to Raji cells. Cycloheximide inhibited the expression of IL-4 receptors by 50% in about 2 h in both cell types indicating both the half-life of IL-4 receptors and the requirement for protein synthesis for the tat III up-regulation of IL-4 receptors. Since IL-4 under certain circumstances has been shown to be immunosuppressant, our observation that the HIV-1 tat gene up-regulates IL-4 receptors suggests the possibility that the immunosuppressive effects of HIV-1 are mediated at least in part through IL-4 receptors.

Cell Line↗
PubMed1992

Phosphatase inhibitors modulate the growth-regulatory effects of human tumor necrosis factor on tumor and normal cells.

Tumor necrosis factor alpha (TNF-alpha) has been shown to inhibit the growth of tumor cells and stimulate the growth of certain normal cells in vitro. The mechanism by which TNF exerts its cell growth-regulatory effects is not known. In this report, we investigated the effects of phosphatase inhibitors on the cell growth-inhibitory effects of TNF on L-929, a highly sensitive murine connective tissue tumor cell line, and on the growth-stimulatory effects of TNF on normal human fibroblasts. The antiproliferative effects of TNF on L-929 cells were inhibited by orthovanadate, an inhibitor of phosphotyrosine phosphatases, in a dose-dependent manner. Okadaic acid, which is a specific inhibitor of phosphoserine- and phosphothreonine-specific phosphatases, also blocked the growth-inhibitory effects of TNF, suggesting that TNF may function through the activation of certain phosphatases. These inhibitors had no effect on TNF receptors. Addition of phosphatase inhibitor, even 12 h after the treatment of cells with TNF, was sufficient to block the antiproliferative effects of the cytokine, suggesting that the inhibitor is acting at a late event in the pathway of action of TNF. Cells were protected by orthovanadate from the cytotoxic effects of TNF even in the presence of actinomycin D or cycloheximide, thus indicating the lack of a requirement for de novo protein synthesis. Orthovanadate altered the cell morphology from flat spindle shapes to rounded ones. Besides anticellular effects, a phosphatase inhibitor also suppressed the proliferative effects of TNF on human fibroblasts. These results thus suggest that phosphatases may be needed for both proliferative and antiproliferative effects of this cytokine. This is the first report to suggest that phosphatases play a role in the growth-regulatory action of TNF.

Animals↗
PubMed1992

Inhibition of ligand binding and antiproliferative effects of tumor necrosis factor and lymphotoxin by soluble forms of recombinant P60 and P80 receptors.

Tumor necrosis factor (TNF) is one of the mediators of inflammatory responses. Recently, the cDNA for two distinct receptors of TNF with predicted molecular masses of 60 kDa and 80 kDa, respectively, were isolated. In this report, we compare the inhibitory effects of these two forms of recombinant soluble TNF receptors (extracellular domains) on the ligand binding and on the antiproliferative effects of TNF and lymphotoxin (LT) in a human histiocytic lymphoma cell line (U-937). Our results show that the soluble form of the p60 receptor is approximately 100-fold more potent than the soluble form of the p80 receptor in inhibiting both the antiproliferative effects of TNF as well as in blocking TNF binding to U-937 cells. In contrast, the antiproliferative effects of LT and its binding to cells is inhibited equally by both the p60 and p80 forms of the soluble receptor. Thus, overall our results indicate that the two soluble receptors differ in their ability to inhibit TNF and LT. The impotance of these soluble receptors in blocking the harmful effects of TNF and LT is discussed.

Cell Division↗
PubMed1992

Induction of interleukin-2 receptor by tumor necrosis factor alpha on cultured ovarian tumor-associated lymphocytes.

We have recently reported that autologous tumor-specific cytotoxic T lymphocyte (CTL) lines and clones can be developed from lymphocytes infiltrating ovarian malignant ascites (TAL). In this study, we investigated the biological effects of tumor necrosis factor alpha (TNF alpha) in the induction, expansion, long-term proliferation and lytic function of CD8+ TAL. TNF alpha up-regulated the IL-2 receptor (IL-2R) alpha chain (Tac antigen) on the surface of CD3+ CD8+ CD4- TAL, enhanced the proliferation of autologous tumor-specific CTL, and potentiated their lytic function in long-term cultures. Furthermore, in the induction and expansion phase of CD8+ TAL, the presence of TNF alpha was associated with a selective increase in CD8+ IL-2R+ (Tac+) cells, and subsequent decrease in CD4+ IL-2R+ (Tac+) cells. These results suggest that the observed facilitation of the outgrowth of CD8+ cells in TAL cultures may be due, at least in part, to the up-regulation of IL-2R, and indicate the usefulness of TNF alpha in the analysis of signalling in autologous tumor-reactive CTL.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1992

Microtiter plate radioreceptor assay for tumor necrosis factor and its receptors in large numbers of samples.

We developed a rapid radioreceptor assay for tumor necrosis factor (TNF) and its receptors by utilizing serum-coated 96-well plates. This assay has an advantage over the traditional tube assay in speed, number of simultaneous samples, and amount of material needed for the assay. As many as 200 samples were assayed in less than 3 h. The usefulness of this assay to determine TNF and its cell surface receptors and to screen for TNF inhibitors in large numbers of biological fluids from patients is demonstrated. This assay may also prove useful for determining the activity of competitive antagonists of TNF.

Cell Line↗