Biomedical subjects

M Nagarkatti

Publications and source records attributed to M Nagarkatti.

At least 19 recordsLinked to original sources

PubMed2001

Enhanced activation-induced cell death as a mechanism of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced immunotoxicity in peripheral T cells.

T cells upon activation undergo apoptosis, a process termed activation-induced cell death (AICD). In the current study, we investigated whether 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) increases AICD and whether this constitutes one of the mechanisms by which TCDD induces immunotoxicity. To this end, C57BL/6+/+, C57BL/6 gld/gld (Fas ligand-defective) and C57BL/6 lpr/lpr (Fas-deficient) mice were injected with TCDD (50 microg/kg body weight, ip) or the vehicle (corn oil) and with anti-CD3 mAbs into the footpads. 3 days later, inguinal and popliteal lymph node cells were harvested, pooled and enumerated. Cells were cultured in vitro with anti-CD3 mAbs and cell proliferation was measured. Also, such cells were studied for their ability to undergo apoptosis upon in vitro culture with either tissue culture medium alone or with anti-CD3 mAbs. The data demonstrated that lymph nodes from TCDD-treated wild-type (+/+) mice showed a decrease in cellularity and the T cells exhibited decreased responsiveness to anti-CD3 mAbs when compared to the vehicle-treated control group. Furthermore, such cells from TCDD-treated mice exhibited increased levels of apoptosis upon in vitro culture when compared to the cells from vehicle-treated mice. In contrast, activated lymph nodes from TCDD-treated C57BL/6 gld/gld and C57BL/6 lpr/lpr mice showed normal cellularity and T cell responsiveness to anti-CD3 stimulation when compared to the vehicle controls. In addition, the activated lymph node T cells from the TCDD-treated C57BL/6 gld/gld and C57BL/6 lpr/lpr mice failed to exhibit increased apoptosis when compared to the controls. The current study demonstrates that the immunotoxic effects of TCDD in activated peripheral T cells may result from increased AICD mediated through Fas-Fas ligand interactions.

Animals↗

PubMed2001

Evidence for estradiol-induced apoptosis and dysregulated T cell maturation in the thymus.

In an attempt to delineate the immunological alterations that may occur following treatment with estrogen, groups of C57BL/6 mice were treated with 75mg/kg body weight of beta-estradiol-17-valerate (E2) or the vehicle. The thymus from these mice were harvested on days 1, 4 and 7 following treatment. The thymocytes from E2-treated mice when cultured in vitro for 24h, showed increased levels of apoptosis when compared to controls. The apoptosis was demonstrable by both TUNEL assay and AnnexinV/propidium iodide (PI) staining. Also, thymic atrophy and increased apoptosis of thymocytes when cultured in vitro were seen when lower doses of E2 (5mg/kg) were administered. The thymus from E2-treated mice on days 4 and 7 also showed a decrease in the percentage of CD4(+)CD8(+) (DP) T cells and an increase in the percentage of CD4(-)CD8(-) (DN), CD4(+) and CD8(+) T cells. However, the total cellularity of all T cell subsets in the thymus was decreased following E2 treatment. Earlier studies from our laboratory and elsewhere have demonstrated that in thymocytes undergoing apoptosis, there is increased expression of surface markers including CD3, alphabetaTCR and CD44 with a simultaneous decrease in the expression of J11d. Similar changes were observed in thymocytes from mice on days 4 and 7 following E2 treatment. These data therefore confirmed that the thymocytes were indeed undergoing apoptosis following E2 treatment. Together, our studies suggest for the first time that estrogen may induce thymic atrophy by triggering apoptosis.

Animals↗

PubMed2001

CD44-deficient mice exhibit enhanced hepatitis after concanavalin A injection: evidence for involvement of CD44 in activation-induced cell death.

Administration of Con A induces severe injury to hepatocytes in mice and is considered to be a model for human hepatitis. In the current study, we investigated the role of CD44 in Con A-induced hepatitis. Intravenous administration of Con A (20 mg/kg) caused 100% mortality in C57BL/6 CD44-knockout (KO) mice, although it was not lethal in C57BL/6 CD44 wild-type (WT) mice. Administration of lower doses of Con A (12 mg/kg body weight) into CD44 WT mice induced hepatitis as evident from increased plasma aspartate aminotransferase levels accompanied by active infiltration of mononuclear cells and neutrophils, and significant induction of apoptosis in the liver. Interestingly, CD44 KO mice injected with similar doses of Con A exhibited more severe acute suppurative hepatitis. Transfer of spleen cells from Con A-injected CD44 KO mice into CD44 WT mice induced higher levels of hepatitis when compared with transfer of similar cells from CD44 WT mice into CD44 WT mice. The increased hepatitis seen in CD44 KO mice was accompanied by increased production of cytokines such as TNF-alpha, IL-2 and IFN-gamma, but not Fas or Fas ligand. The increased susceptibility of CD44 KO mice to hepatitis correlated with the observation that T cells from CD44 KO mice were more resistant to activation-induced cell death when compared with the CD44 WT mice. Together, these data demonstrate that activated T cells use CD44 to undergo apoptosis, and dysregulation in this pathway could lead to increased pathogenesis in a number of diseases, including hepatitis.

Adjuvants, Immunologic↗

PubMed2000

Growth of FasL-bearing tumor cells in syngeneic murine host induces apoptosis and toxicity in Fas(+) organs.

In the current study, we investigated whether the growth of FasL-bearing tumor cells would induce apoptosis and toxicity in organs that express high level of Fas. Sera from C57BL/6 +/+ (wild-type) mice injected with syngeneic FasL(+) tumors, LSA, or EL-4, showed significantly higher levels of soluble FasL than that from the nontumor-bearing mice. Furthermore, the soluble FasL was functional inasmuch as the sera from tumor-bearing mice were able to induce apoptosis in Fas(+) but not Fas(-) targets. Histopathologic studies and in situ TUNEL assay to detect apoptosis were carried out in C57BL/6 +/+ (Fas(+)) or C57BL/6 lpr/lpr (Fas(-)) mice injected with syngeneic LSA and EL-4 tumor cells. The morphology of the liver and thymus from tumor bearing C57BL/6 +/+ mice showed marked damage and tissue destruction. In contrast, the liver and thymus from tumor-bearing C57BL/6 lpr/lpr mice showed minimal damage. Furthermore, the tumor-bearing C57BL/6 +/+, but not the C57BL/6 lpr/lpr, mice exhibited significant apoptosis in the liver and thymus. The FasL responsible for toxicity was tumor derived rather than host derived; tumor-bearing C57BL/6 gld/gld (FasL-defective) mice also exhibited significant apoptosis in the liver and thymus. Together, these data suggested that the in vivo growth of FasL-bearing tumor cells can induce significant apoptosis and toxicity in Fas(+) tissues of the host. Such toxicity may be mediated by the soluble FasL produced by tumor cells. (Blood. 2000;95:2111-2117)

Animals↗

PubMed1999

Role of Fas-Fas ligand interactions in 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD)-induced immunotoxicity: increased resistance of thymocytes from Fas-deficient (lpr) and Fas ligand-defective (gld) mice to TCDD-induced toxicity.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a highly toxic environmental pollutant well known for its toxicity to the thymus. Recent studies from our laboratory demonstrated that TCDD induces apoptosis in thymocytes. In the current study, we investigated the mechanism of TCDD-induced apoptosis. Administration of a single dose of TCDD at 0.1, 1, 5, and 50 microg/kg body wt intraperitoneally, into C57BL/6 +/+ (wild-type) mice caused a dose-dependent decrease in thymic cellularity. In contrast, a similar treatment with TCDD, in Fas-deficient C57BL/6 lpr/lpr (lpr) or Fas-ligand defective C57BL/6 gld/gld (gld), mice failed to induce thymic atrophy at 0.1-5 microg/kg body wt of TCDD. In lpr and gld mice, significant thymic atrophy was seen only at 50 microg/kg body wt of TCDD. Injection of TCDD caused apoptosis only in wild-type but not in lpr or gld mice. The sera from TCDD-treated wild-type mice exhibited increased levels of soluble Fas ligand, inasmuch as incubation of Fas(+), but not Fas(-) cells with the sera, triggered apoptosis. Also, TCDD-induced apoptosis in thymocytes was inhibited both in vitro and in vivo by caspase inhibitors. TCDD treatment caused significant up-regulation in the expression of FasL but not Fas mRNA in the thymocytes of wild-type mice. Also, such thymocytes exhibited marked alterations in the surface markers, characteristic of cells undergoing apoptosis. In contrast, TCDD treatment caused minimal phenotypic changes in thymocytes from lpr and gld mice. Together, the current study demonstrates that Fas-Fas ligand interactions play an important role in TCDD-mediated induction of apoptosis and immunotoxicity.

Animals↗

PubMed1999

Evidence for the involvement of CD44 in endothelial cell injury and induction of vascular leak syndrome by IL-2.

At sites of chronic inflammation seen during infections, autoimmunity, graft-vs-host response, and cytokine therapy, endothelial cell injury is known to occur, the exact mechanism of which is unknown. In the current study we used IL-2-induced vascular leak syndrome (VLS) as a model to investigate whether cytotoxic lymphocytes use CD44 in mediating endothelial cell injury. Administration of IL-2 to wild-type mice triggered significant VLS in the lungs and liver. In contrast, in CD44 knockout (KO) mice, IL-2-induced VLS was markedly reduced in the lungs and liver. IL-2-treated wild-type and CD44 KO mice had similar levels of perivascular infiltration with lymphocytes in the lungs and liver. This suggested that the decrease in VLS seen in CD44 KO mice was not due to the inability of lymphocytes to migrate to these organs. Ultrastructural studies demonstrated extensive endothelial cell damage in the lungs and liver of IL-2-treated wild-type, but not CD44 KO, mice. Moreover, CD44-KO mice exhibited a marked decrease in IL-2-induced lymphokine-activated killer cell activity. The induction of VLS was dependent on the expression of CD44 on immune cells rather than endothelial cells because adoptive transfer of CD44+, but not CD44- spleen cells along with IL-2 into CD44 KO mice triggered VLS. The IL-2-induced VLS was blocked by administration of F(ab')2 of Abs against CD44. The current study demonstrates that CD44 plays a key role in endothelial cell injury. Blocking CD44 in vivo may offer a novel therapeutic approach to prevent endothelial cell injury by cytotoxic lymphocytes in a variety of clinical disease models.

Adoptive Transfer↗

PubMed1998

Role of spontaneous and interleukin-2-induced natural killer cell activity in the cytotoxicity and rejection of Fas+ and Fas- tumor cells.

In the current study, we investigated whether the naive, poly I:C or interleukin-2 (IL-2)-induced natural killer (NK)/lymphokine-activated killer (LAK) cells use perforin and/or Fas ligand (FasL) to mediated cytotoxicity. We correlated these findings with the ability of mice to reject syngeneic Fas+ and Fas- tumor cells either spontaneously or after IL-2 treatment. The spontaneous NK-cell-mediated cytotoxicity was primarily perforin based, whereas the poly I:C and IL-2-induced NK/LAK activity was both FasL and perforin dependent. L1210 Fas+ tumor targets were more sensitive than L1210 Fas- targets to poly I:C and IL-2-induced cytotoxicity in wild-type, gld/gld, and perforin knockout mice. When L1210 Fas+ and Fas- tumor cells were injected subcutaneously (sc) or intraperitoneally into syngeneic mice, Fas- tumor cells caused mortality earlier than Fas+ tumor cells. Also, approximately 20% of the mice injected sc with L1210 Fas+ tumor cells survived the challenge(>60 days), whereas all mice injected similarly with L1210 Fas- tumor cells died. When immunotherapy using IL-2 (10,000 U, three times/d for a week, followed by once/d for an additional week) was attempted in mice injected sc with tumor cells, IL-2 treatment was very effective against mice bearing L1210 Fas+ (40% survival) but not L1210 Fas- (0% survival) tumors. These data correlated with the finding that the LAK cells from IL-2-injected mice caused increased cytotoxicity against L1210 Fas+ when compared with L1210 Fas- targets. Also, L1210 Fas+ tumor-bearing mice showed increased tumor-specific cytotoxic T lymphocyte (CTL) activity when compared with those bearing L1210 Fas- tumor cells. Together our studies show for the first time that expression of Fas on tumor targets makes them more immunogenic as well as susceptible to CTL- and IL-2-induced LAK activity. The Fas+ tumor cells are also more responsive to immunotherapy with IL-2.

Animals↗

PubMed1998

Evidence for the involvement of Fas ligand and perforin in the induction of vascular leak syndrome.

Endothelial cell injury resulting in vascular leak syndrome (VLS) is one of the most widely noted phenomenons in a variety of clinical diseases. In the current study we used IL-2-induced VLS as a model to investigate the role of cytolytic lymphocytes in the cytotoxicity of endothelial cells. Administration of IL-2 (75,000 U/mouse, three times a day for 3 days) into BL/6 wild-type mice triggered significant VLS in the lungs, liver, and spleen. Interestingly, perforin-knockout (KO) mice exhibited a marked decrease in IL-2-induced VLS in all three organs tested. Also, Fas ligand-defective (gld) mice and Fas-deficient (lpr) mice exhibited decreased VLS in the liver and spleen, but not in the lungs. The decreased VLS seen in perforin-KO, gld, and lpr mice was not due to any defect in lymphocyte migration or homing to various organs because histopathologic studies in these mice demonstrated significant and often greater perivascular infiltration of lymphocytes compared with the IL-2-treated wild-type mice. Ultrastructural studies of the lungs demonstrated significant damage to the endothelial cells in IL-2-treated wild-type mice and decreased damage in perforin-KO mice. IL-2 administration caused up-regulation of CD44 in all strains of mice tested and triggered increased LAK activity against an endothelial cell line in wild-type and gld mice, but not in perforin-KO mice. The current study demonstrates for the first time that perforin and Fas ligand may actively participate in endothelial cell injury and induction of VLS in a variety of organs.

Animals↗

PubMed1998

Differential induction of apoptosis in activated and resting T cells by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and its repercussion on T cell responsiveness.

TCDD is well known for its immunotoxic effects on T cells, although the exact mechanism of toxicity remains unknown. In the current study, we investigated the effect of TCDD administration on resting and activated T cells within the same animal. To this end, C57BL/6 mice were injected intraperitoneally with either TCDD (50 microg/kg body weight) or the vehicle and were injected with anti-CD3 mAbs into the rear footpads to polyclonally activate T cells in the popliteal lymph nodes (LN). Axillary LN cells harvested from the same groups of mice served as a source of resting T cells. When the LN cells were tested for their proliferative responsiveness to stimulation with anti-CD3 mAbs in vitro, the activated popliteal LN, but not the resting axillary LN cells from TCDD-treated mice exhibited a significant decrease in responsiveness when compared to the vehicle controls. Inasmuch as TCDD has been shown to induce apoptosis in thymocytes, we addressed whether TCDD triggered apoptosis in LN cells, using the terminal deoxynucleotidyl transferase (TdT)-mediated FITC-dUTP nick end labeling (TUNEL) method. The axillary and popliteal LN cells from TCDD-treated mice failed to exhibit significant levels of apoptosis when freshly harvested. However, upon in vitro culture for 24 h with either tissue culture medium alone or with anti-CD3 mAbs, activated popliteal LN cells from TCDD-treated mice showed a significant increase in apoptosis when compared to similar cells from vehicle-treated mice. In contrast, resting axillary LN cells from TCDD-treated mice, similarly cultured in vitro, exhibited decreased levels of apoptosis when compared to the controls. Using a double-staining technique, the activated popliteal LN cells undergoing increased apoptosis in TCDD-treated animals were confirmed to be CD3+ T cells. Together, these data demonstrate that TCDD exerts differential effects on activated and resting T cells, even within the same animal, by inhibiting the proliferative responsiveness of activated, but not resting, T cells. Furthermore, this effect may be mediated by the ability of TCDD to induce increased apoptosis in activated, but not resting, T cells.

Animals↗

PubMed1998

Role of CD44 in CTL and NK cell activity.

CD44 is a widely distributed cell surface glycoprotein whose principal ligand has been identified as hyaluronic acid (HA), a major component of the extracellular matrix (ECM). Recent studies have demonstrated that activation through CD44 leads to induction of effector function in T cells and macrophages. At sites of chronic inflammation as seen in certain infections, autoimmune diseases, allograft rejection, graft-versus-host (GVH) disease and treatment of cancer patients with high doses of interleukin-2, significant damage to the endothelial cells has been known to occur, which leads to the toxicity or pathogenesis associated with the disease. The exact mechanism of endothelial cell damage is not clear, although, it has been widely speculated that immune cells may play a critical role. Studies from our laboratory have used interleukin-2 (IL-2) induced vascular leak syndrome (VLS) as a model to investigate the role of cytolytic lymphocytes in the direct cytotoxicity of endothelial cells. Cytotoxic T lymphocytes (CTL), double-negative (DN) T cells and natural killer (NK) cells upon activation express high levels of CD44 and mediate efficient MHC-unrestricted TCR-independent lysis following ligation of CD44. Such CD44-mediated cytotoxicity may play an important role in protection against viral infections and cancer. However, it could also cause non-specific tissue injury. For example, dysregulation in the interaction between activated cytotoxic lymphocytes expressing CD44 and endothelial cells bearing the appropriated ligand such as the hyaluronate (HA), could lead to endothelial cell lysis. Furthermore, such endothelial cell injury could lead to the pathogenesis associated with a variety of clinical diseases.

Animals↗

PubMed1998

Characterization of phenotypic alterations induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin on thymocytes in vivo and its effect on apoptosis.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a highly toxic environmental pollutant and is well known for inducing thymic atrophy in mice, although the exact mechanism of its action remains unclear. Recent studies from our laboratory demonstrated that TCDD induces apoptosis in thymocytes and that Fas- mice (lpr/lpr) were more resistant to TCDD-induced immunotoxicity when compared to the Fas+ wild-type mice. Inasmuch as induction of apoptosis is associated with alterations in adhesion molecule expression, in the current study we analyzed the expression of a variety of surface molecules on thymocytes treated with TCDD in vivo. Interestingly, in thymocytes from mice treated with a single dose of 50 micrograms/kg body wt of TCDD, there was a significant increase in the density of expression of CD3, alpha beta TCR, CD44, and IL-2R, and a decrease in the expression of J11d, CD4, and CD8 molecules when compared to the control thymocytes. These alterations were first visible 3 days after TCDD treatment and increased on Days 5 and 10 posttreatment. Furthermore, most of the alterations in the density of expression of various markers were dose dependent with minimal but significant changes at 0.1 microgram and maximum alterations at 50 micrograms/kg body wt of TCDD. At most lower concentrations (0.1-5 micrograms/kg), TCDD caused alterations in the density of cell surface markers but not in the percentage of cells expressing a specific molecule. It is striking that the phenotypic alterations were similar to those seen in normal thymocytes undergoing spontaneous apoptosis in vitro as previously reported. Together, the current study suggests that TCDD treatment induces phenotypic changes in thymocytes that are similar to those seen in normal thymocytes undergoing apoptosis. Also, because detection of apoptosis in vivo is difficult, phenotypic alterations in the density of thymocyte surface molecules may serve as a useful biomarker for toxicity involving apoptosis.

Animals↗

PubMed1997

Fas-Fas ligand-based interactions between tumor cells and tumor-specific cytotoxic T lymphocytes: a lethal two-way street.

In the current study, we investigated the repercussions of the interaction between tumor cells (LSA) and the tumor-specific cytotoxic T lymphocyte (CTL) (PE-9) when both expressed Fas and Fas ligand (FasL). The CTL clone, PE-9, expressed high levels of Fas and FasL upon activation through the T-cell receptor (TCR). Furthermore, the activated PE-9 cells used both perforin- and FasL-based pathways to kill Fas-positive (Fas+) LSA tumor cells. Interestingly, LSA tumor cells also constitutively expressed FasL but not perforin, and killed Fas+ PE-9 CTLs and Fas+ but not Fas-negative (Fas-) activated T cells and thymocytes, as detected using the JAM test. PE-9 CTLs, cultured for 24 hours in the presence of cell lysates of FasL-bearing LSA cells but not FasL-deficient P815 cells, exhibited significant apoptosis as detected using the TUNEL method. Moreover, another FasL+ T-cell lymphoma line, EL-4, induced apoptosis in Fas+ but not in Fas- T cells in a similar fashion. The current study demonstrates for the first time that not only can the tumor-specific CTL mediate Fas-based killing of tumor cells, but FasL+ tumor cells can kill the Fas+ tumor-specific CTL. Thus, the survival of the tumor or the host may depend on which cell can accomplish this task more efficiently. The current study also suggests that FasL-based killing of CTLs by specific tumor cells may constitute a major limiting factor in successful immunotherapy.

Animals↗

PubMed1997

Hyaluronate-CD44 interactions can induce murine B-cell activation.

CD44 is a widely distributed cell surface glycoprotein whose principal ligand has been identified as hyaluronic acid (HA), a major component of the extracellular matrix (ECM). Recent studies have demonstrated that activation through CD44 leads to induction of effector function in T cells and macrophages. In the current study, we investigated whether HA or monoclonal antibodies (MoAbs) against CD44 would induce a proliferative response in mouse lymphocytes. Spleen cells from normal and nude, but not severe combined immunodeficient mice, exhibited strong proliferative responsiveness to stimulation with soluble HA or anti-CD44 MoAbs. Furthermore, purified B cells, but not T cells, were found to respond to HA. HA was unable to stimulate T cells even in the presence of antigen presenting cells (APC) and was unable to act as a costimulus in the presence of mitogenic or submitogenic concentrations of anti-CD3 MoAbs. In contrast, stimulation of B cells with HA in vitro, led to B-cell differentiation as measured by production of IgM antibodies in addition to increased expression of CD44 and decreased levels of CD45R. The fact that the B cells were responding directly to HA through its binding to CD44 and not to any contaminants or endotoxins was demonstrated by the fact that F(ab)2 fragments of anti-CD44 MoAbs or soluble CD44 fusion proteins could significantly inhibit the HA-induced proliferation of B cells. Also, HA-induced proliferation of B cells was not affected by the addition of polymixin B, and B cells from lipopolysaccharide (LPS)-unresponsive C3H/HeJ strain responded strongly to stimulation with HA. Furthermore, HA, but not chondroitin-sulfate, another major component of the ECM, induced B-cell activation. It was also noted that injection of HA intraperitoneally, triggered splenic B cell proliferation in vivo. Together, the current study demonstrates that interaction between HA and CD44 can regulate murine B-cell effector functions and that such interactions may play a critical role during normal or autoimmune responsiveness of B cells.

Animals↗

PubMed1997

Evidence for the participation of interleukin-2 (IL-2) and IL-4 in the regulation of autonomous growth and tumorigenesis of transformed cells of lymphoid origin.

In the current study, we investigated the role of interleukin-2 (IL-2) and IL-4 as autocrine growth factors responsible for autonomous growth of four murine tumor cell lines: LSA, a radiation leukemia virus-induced T-cell lymphoma; EL-4, a chemically triggered T-cell lymphoma; PE-3T, a T-cell line that underwent spontaneous transformation ex vivo; and P815, a mastocytoma. All tumor cell lines screened constitutively expressed IL-2 receptor (IL-2R) and IL-4R genes. However, only LSA and PE-3T cells expressed IL-2 and IL-4 genes constitutively, whereas EL-4 and P815 tumor cells expressed only IL-4 but not IL-2. Monoclonal antibodies (MoAbs) against IL-2, IL-4, or a combination of these, as well as MoAbs against IL-2R significantly inhibited the proliferation of LSA but not that of other tumor cell lines ex vivo. To exclude the possibility that, in other tumor cell lines, the autocrine growth factor may interact with its receptor within the cell, the ability of antisense phosphorothioate oligonucleotides to inhibit the growth of the tumor cells was tested. The antisense phosphorothioate oligonucleotides specific for IL-2, IL-4, IL-2R beta, or IL-2R gamma chains, added in culture, could markedly inhibit the growth of LSA but not that of the other tumor cell lines screened. Inasmuch as IL-2R beta and IL-2R gamma subunits also serve as a component of the receptors for IL-4, IL-7, IL-9, and IL-15, the above data suggested that such cytokine redundancy was not responsible for autonomous growth of the other tumor cell lines. Addition of exogenous IL-2 or IL-4 to the tumor cell cultures caused significant enhancement in the proliferation of PE-3T cells, whereas other cell lines were either not significantly affected or slightly inhibited from growing. Interestingly, the LSA tumor growth in nude mice was significantly inhibited after treatment of these mice with a combination of MoAbs against IL-2 and IL-4. Together, our studies show for the first time that IL-2 and IL-4 may serve as autocrine growth factors in the autonomous proliferation of tumor cells, particularly those that are retrovirally induced. Second, some tumor cell lines, despite expressing certain cytokines and their receptors constitutively, may not depend exclusively on such factors for autocrine growth.

Animals↗

PubMed1997

Evidence for the induction of apoptosis in thymocytes by 2,3,7,8-tetrachlorodibenzo-p-dioxin in vivo.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is well known for its immunotoxic effects particularly on the thymus. The exact mechanism by which TCDD induces thymic atrophy is not clear. In the current study, we investigated whether TCDD triggers apoptosis in thymocytes, when administered in vivo, by using the TdT-mediated FITC-dUTP nick end labeling method and analyzing the cell flow cytometrically. Significant apoptosis was detected at 8-12 hr after the TCDD injection but not at 24 hr or beyond, up to 120 hr of study. Furthermore, the induction of apoptosis was confirmed using the JAM test in which thymocytes from TCDD-treated mice, labeled with [3H]thymidine, exhibited increased DNA fragmentation when compared to the controls. Similar to TCDD treatment, administration of dexamethasone (5 or 100 mg/kg) into C57BL/6 mice triggered apoptosis that was only detected at 12 hr after administration of the drug and not thereafter. When thymocytes from TCDD- or dexamethasone-treated mice were cultured in vitro for 24 hr, they exhibited marked increase in apoptosis when compared to the vehicle-treated controls. However, TCDD, when added to in vitro cultures of thymocytes, failed to trigger apoptosis. Together, our studies demonstrate that TCDD can induce apoptosis in thymocytes in vivo. This can be detected only at an early stage following TCDD administration, possibly because of rapid clearance of apoptotic cells by the phagocytic cells in vivo.

Animals↗

PubMed1996

Role of Fas apoptosis and MHC genes in 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced immunotoxicity of T cells.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is well known for its immunotoxic effects particularly on the thymus as well as on T and B lymphocyte functions. Previous studies have suggested that TCDD may induce apoptosis in thymocytes although its demonstration in vivo has met with limited success. TCDD has also been shown to alter the major histocompatability complex- (MHC) encoded molecules, however, its role in immunotoxicity is not clear. In the current study, we investigated the role of Fas (CD95), an important molecule involved in the induction of apoptosis, on TCDD-mediated immunotoxicity using mice bearing homozygous lpr mutation which leads to failure of expression of Fas. When TCDD was administered orally at 0, 0.1, 1.0, or 5.0 micrograms/kg body weight for 11 days, it was found to be less toxic to the thymocytes from C57BL/6 lpr/lpr mice (Ah-responsive, Fas-) when compared to C57BL/6 +/+ mice (Ah-responsive, Fas+). Similar results were obtained when peripheral T cell responsiveness to antigenic challenge with conalbumin was studied in these mice. When mice that differed only at the MHC were compared for immunotoxic effects of TCDD, it was noted that B10.D2 (Ah-responsive, H-2d) were more sensitive to TCDD-mediated thymic atrophy and peripheral T cell dysfunction when compared to B10 mice (Ah-responsive, H-2b). In all TCDD-sensitive strains tested, the thymic atrophy was accompanied by a uniform depletion of all four subset of T cells (CD4+, CD4+CD8+, CD4-CD8-, and CD8+) and the percentage of these subsets was not altered. Furthermore, in these strains, TCDD suppressed the antigen-specific peripheral T cell responsiveness but not the responsiveness of naive resting T cells to polyclonal mitogens. Lastly, using cell-mixing experiments, it was demonstrated that TCDD directly affected the T cells responding to conalbumin but not the antigen presenting cells (APCs). Together, our studies demonstrate that although Ah locus plays the primary role, determining the toxicity of TCDD on the T cells, there are secondary factors such as expression of Fas or the MHC-phenotype which may play an important role in TCDD-mediated immunotoxicity. The role of Fas further suggests that TCDD may induce toxicity in T cells by triggering apoptosis.

Administration, Oral↗

PubMed1996

Effect of radon on the immune system: alterations in the cellularity and functions of T cells in lymphoid organs of mouse.

Exposure to radon and its progeny induces significant damage to the cells of the respiratory tract and causes lung cancer. Whether a similar exposure to radon would alter the functions of the immune system has not been previously investigated. In the current study, we investigated the effect of exposure of C57BL/6 mice to 1000 or 2500 working-level months (WLM) of radon and its progeny by inhalation, on the number and function of T lymphocytes in lymphoid organs. The control mice received uranium ore dust carrier aerosol by inhalation. Exposure to radon induced marked decrease in the total cellularity of most lymphoid organs such as thymus, peripheral lymph nodes (PLN), and lung-associated lymph nodes (LALN), when compared to the controls. The percentage of T cells increased, while that of non-T cells decreased, in all peripheral lymphoid organs at both the doses of radon. In the thymus, particularly at 2500 WLM of radon exposure, there was a marked decrease in CD4+CD8+ T cells and an increase in the immature CD4-CD8- T cells. Such alterations in both the numbers and percentages of lymphocytes and macrophages in radon-exposed mice may have resulted from the cell killing by the alpha particles as the immune cells were migrating through the lungs, or it may have been caused by altered migration of cells, inasmuch as expression of CD44, a molecule involved in migration and homing of immune cells, was significantly altered on cells found in different lymphoid organs. In the LALN, where one would predict the largest number of damaged cells to be present, there was a significant decrease in the T-cell responsiveness to mitogens while the B-cell response was not affected. Such alterations may have resulted from the direct effect of alpha-particle exposure on the migrating lymphocytes, altered percentage of lymphocytes as seen in secondary lymphoid organs, or altered expression of adhesion molecules involved in cell activation such as CD44 and CD3. Interestingly, radon exposure caused and increase in the T- and B-cell responsiveness to mitogens in the spleen and PLN. Since there is little evidence of direct radiation dose from radon in lymphoid organs, our studies demonstrating immunological alterations suggest an indirect effect of radon exposure that may have significant repercussions on the development of hypersensitivity and increased susceptibility to infections and cancer in the lung.

Administration, Inhalation↗

PubMed1995

Characterization of factors regulating successful immunotherapy using a tumor-specific cytotoxic T lymphocyte clone: role of interleukin-2, cycling pattern of lytic activity and adhesion molecules.

Adoptive immunotherapy against cancer has met with varying degrees of success, the reasons for which remain unclear. The present study characterizes factors that regulate successful immunotherapy of mice bearing a syngeneic T-cell lymphoma, designated LSA, using a tumor-specific cytotoxic T lymphocyte (CTL) clone, PE-9. Adoptive transfer of PE-9 cells afforded significant protection in normal but not in nude mice against LSA tumor. However, the PE-9 cells could protect the nude mice when injected along with normal CD4+ T cells. Administration of IL-2 along with PE-9 cells failed to enhance tumor immunotherapy. IL-2 therapy was toxic inasmuch as injection of the CTL clone PE-9 + IL-2, but not PE-9 or IL-2 alone, for 5 days into irradiated mice caused vascular leak syndrome (VLS). PE-9 cells cultured with high doses of rIL-2 in vitro also caused TCR-independent and MHC-unrestricted lysis of SV40-transformed endothelial cells. Furthermore, PE-9 cells cultured in vitro for 24-96 hr with IL-2 exhibited cycling pattern of tumor-specific cytotoxicity with maximum cytotoxicity demonstrable at 48 hr and virtually no cytolytic activity at 96 hr of culture or thereafter. The loss of cytotoxicity correlated with down-regulation of several adhesion molecule expressions on PE-9 cells, particularly the alpha beta-TCR, as well as the mRNA for TNF-alpha, IFN-gamma and perforin, although the levels of granzyme A were not altered. Interestingly, the outcome of immunotherapy by PE-9 cells depended on the cycling pattern of cytotoxicity. Our data suggest that successful immunotherapy against cancer using a CTL clone depends on several factors, such as the cycling pattern of lytic activity, density of adhesion molecules, levels of cytokines expressed and the ability of IL-2 and CTL to trigger VLS.

Animals↗