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M J Tremblay

Publications and source records attributed to M J Tremblay.

30 records · Page 2Linked to original sources
PubMed1999

Protective effect of a thermoreversible gel against the toxicity of nonoxynol-9.

BACKGROUND AND OBJECTIVES: One major problem associated with the use of nonoxynol-9 is that it can induce local inflammation and ulceration of the vaginal and cervical mucosa that might favor the entry of pathogens. With the aim of developing a gel formulation that could be effective in preventing sexually transmitted infections, the authors have evaluated the capacity of a polyoxypropylene/polyoxyethylene polymer to reduce or eliminate the toxicity of nonoxynol-9. STUDY DESIGN: The cytotoxicity of nonoxynol-9 alone or incorporated into the gel was investigated in human cervical and colon epithelial cells and after daily intravaginal application for 2 weeks in rabbits. RESULTS: In vitro experiments showed that nonoxynol-9 was highly toxic to human cervical and colon epithelial cells in a dose-dependent manner. However, the incorporation of the spermicide into the gel markedly reduced its toxicity under the same experimental conditions. In vivo studies showed that in animals treated with nonoxynol-9, the spermicide was very toxic to the vaginal and cervical mucosa as evidenced by the presence of bleeding, irritation, epithelial disruption, necrosis, the accumulation of leukocytes in the submucosa, and the loss of integrity of the epithelial cells. Of prime importance, the incorporation of nonoxynol-9 into the gel markedly reduced the toxicity of this potent spermicide/microbicide. CONCLUSION: The gel formulation could be used as an interesting approach to eliminate the toxicity of potent spermicides/microbicides such as nonoxynol-9.

Animals↗
PubMed1998

Prostaglandin E2 Up-regulates HIV-1 long terminal repeat-driven gene activity in T cells via NF-kappaB-dependent and -independent signaling pathways.

Replication of human immunodeficiency virus type-1 (HIV-1) is highly dependent on the state of activation of the infected cells and is modulated by interactions between viral and host cellular factors. Prostaglandin E2 (PGE2), a pleiotropic immunomodulatory molecule, is observed at elevated levels during HIV-1 infection as well as during the course of other pathogenic infections. In 1G5, a Jurkat-derived T cell line stably transfected with a luciferase gene driven by HIV-1 long terminal repeat (LTR), we found that PGE2 markedly enhanced HIV-1 LTR-mediated reporter gene activity. Experiments have been conducted to identify second messengers involved in this PGE2-dependent up-regulating effect on the regulatory element of HIV-1. In this study, we present evidence indicating that signal transduction pathways induced by PGE2 necessitate the participation of cyclic AMP, protein kinase A, and Ca2+. Experiments conducted with different HIV-1 LTR-based vectors suggested that PGE2-mediated activation effect on HIV-1 transcription was transduced via both NF-kappaB-dependent and -independent signaling pathways. The involvement of NF-kappaB in the PGE2-dependent activating effect on HIV-1 transcription was further confirmed using a kappaB-regulated luciferase encoding vector and by electrophoretic mobility shift assays. Results from Northern blot and flow cytometric analyses, as well as the use of a selective antagonist indicated that PGE2 modulation of HIV-1 LTR-driven reporter gene activity in studied T lymphoid cells is transduced via the EP4 receptor subtype. These results suggest that secretion of PGE2 by macrophages in response to infection or inflammatory activators could induce signaling events resulting in activation of proviral DNA present into T cells latently infected with HIV-1.

Calcium↗
PubMed1998

Prostaglandin E2 induces resistance to human immunodeficiency virus-1 infection in monocyte-derived macrophages: downregulation of CCR5 expression by cyclic adenosine monophosphate.

The chemokine receptor CCR5 can function as a coreceptor for human immunodeficiency virus-1 (HIV-1) entry into CD4(+) T cells and macrophages, especially during the early stages of HIV-1 infection. The regulation of CCR5 expression may affect not only leukocyte migration, but also infectivity by HIV-1 and, therefore, acquired immunodeficiency syndrome (AIDS) pathogenesis. We report here that agents which increase intracellular concentrations of cyclic adenosine monophosphate (cAMP) rapidly downregulate CCR5 gene expression, with consequent loss of CCR5 expression and function in monocytes/macrophages. Chemotaxis and intracellular Ca2+ mobilization in monocytes pretreated with prostaglandin E2 or dibutyryl-cAMP for 24 hours were significantly reduced in response to the CCR5 ligand, MIP-1beta. Moreover, HIV-1 entry into monocyte-derived macrophages pretreated with dibutyryl-cAMP or prostaglandin E2 was markedly decreased. Our findings suggest that resistance to HIV-1 can be induced by agents which increase cellular levels of cAMP and that this may suggest additional therapeutic strategies to limit infection by HIV-1.

Cyclic AMP↗
PubMed1998

Modulation of interferon-gamma-induced macrophage activation by phosphotyrosine phosphatases inhibition. Effect on murine Leishmaniasis progression.

Phagocyte functions are markedly inhibited after infection with the intracellular protozoan parasite Leishmania. This situation strongly favors the installation and propagation of this pathogen within its mammalian host. Previous findings by us and others have established that alteration of several signaling pathways (protein kinase C-, Ca2+- and protein-tyrosine kinases-dependent signaling events) were directly responsible for Leishmania-induced macrophage (MO) dysfunctions. Here we report that modulation of phosphotyrosine-dependent events with a protein tyrosine phosphatases (PTP) inhibitor, the peroxovanadium (pV) compound bpV(phen) (potassium bisperoxo(1,10-phenanthroline)oxovanadate(Vi)), can control host-pathogen interactions by different mechanisms. We observed that the inhibition of parasite PTP resulted in an arrest of proliferation and death of the latter in coincidence with cyclin-dependent kinase (CDK1) tyrosine 15 phosphorylation. Moreover the treatment of MO with bpV(phen) resulted in an increased sensitivity to interferon-gamma stimulation, which was reflected by enhanced nitric oxide (NO) production. This enhanced IFN-gamma-induced NO generation was accompanied by a marked increase of inducible nitric oxide synthase (iNOS) mRNA gene and protein expression. Finally we have verified the in vivo potency of bpV(phen) over a 6-week period of daily administration of a sub-toxic dose. The results revealed its effectiveness in controlling the progression of visceral and cutaneous leishmaniasis. Therefore PTP inhibition of Leishmania and MO by the pV compound bpV(phen) can differentially affect these eukaryotic cells. This strongly suggests that PTP plays an important role in the progression of Leishmania infection and pathogenesis. The apparent potency of pV compounds along with their relatively simple and versatile structure render them attractive pharmacological agents for the management of parasitic infections.

Animals↗
PubMed1998

The lipophosphoglycan of Leishmania donovani up-regulates HIV-1 transcription in T cells through the nuclear factor-kappaB elements.

We have recently demonstrated that the parasite Leishmania donovani and its surface molecule, lipophosphoglycan (LPG), can activate HIV-1 replication in monocytoid cells. Our present interest was to determine whether LPG could also up-regulate HIV-1 transcription in T cells. Using a CD4-positive human lymphoid T cell line (1G5) containing a stably integrated HIV-1 long terminal repeat (LTR)-luciferase construct, we found that LPG is a potent inducer of HIV-1 LTR activity. Treatment of 1G5 cells with signaling antagonists revealed that protein tyrosine kinase- and protein kinase A-dependent pathways were actively participating in the LPG-induced enhancement of HIV-1 LTR-driven activity. Transfection of Jurkat E6.1 cells with plasmids containing wild-type and nuclear factor-kappaB (NF-kappaB)-mutated HIV-1 LTR-luciferase constructs has suggested a role for NF-kappaB binding sites in the LPG-mediated induction of HIV-1 LTR activity. An LPG-induced binding factor specific to the NF-kappaB consensus sequences could be observed using electrophoretic mobility shift assay. Finally, transfection experiments performed with a vector containing HIV-1 kappaB binding sites only showed similar LPG-mediated induction, which was abrogated by sodium salicylate, a known NF-kappaB inhibitor. We thus demonstrate that the LPG-mediated induction of HIV-1 LTR activity in T cells involves several second messengers culminating in activation of HIV-1 LTR-driven transcription via NF-kappaB-binding consensus sequences. In conclusion, these results reinforce the idea that L. donovani is a putative cofactor in HIV-1 pathogenesis.

Animals↗
PubMed1998

Mycobacterium tuberculosis mannose-capped lipoarabinomannan can induce NF-kappaB-dependent activation of human immunodeficiency virus type 1 long terminal repeat in T cells.

Tuberculosis has emerged as an epidemic, extended by the large number of individuals infected with human immunodeficiency virus type 1 (HIV-1). The major goal of this study was to determine whether the mycobacterial cell wall component mannose-capped lipoarabinomannan (ManLAM) of Mycobacterium tuberculosis (M. tuberculosis) could activate transcription of HIV-1 in T cells with the use of an in vitro cell culture system. These experiments are of prime importance considering that CD4-expressing T lymphocytes represent the major virus reservoir in the peripheral blood of infected individuals. Using the 1G5 cell line harbouring the luciferase reporter gene under the control of the HIV-1 LTR, it was first found that culture protein filtrates (CFP) from M. tuberculosis or purified ManLAM could activate HIV-1 LTR-dependent gene expression unlike similarly prepared CFP extracts devoid of ManLAM. The implication of protein tyrosine kinase(s), protein kinase A and/or protein kinase C was highlighted by the abrogation of the ManLAM-mediated activation of HIV-1 LTR-driven gene expression using herbimycin A and H7. It was also determined, using electrophoresis mobility shift assays, that M. tuberculosis ManLAM led to the nuclear translocation of the transcription factor NF-kappaB. M. tuberculosis ManLAM resulted in clear induction of the luciferase gene placed under the control of the wild-type, but not the kappaB-mutated, HIV-1 LTR region. Finally, the ManLAM-mediated activation of HIV-1 LTR transcription was found to be independent of the autocrine or paracrine action of endogenous TNF-alpha. The results suggest that M. tuberculosis can upregulate HIV-1 expression in T cells and could thus have the potential to influence the pathogenesis of HIV-1 infection.

Binding Sites↗
PubMed1998

Level of ICAM-1 surface expression on virus producer cells influences both the amount of virion-bound host ICAM-1 and human immunodeficiency virus type 1 infectivity.

Using virions harvested from 293T cells stably expressing either low or high levels of surface ICAM-1, we determined that the number of virus-embedded host ICAM-1 proteins is positively influenced by the expression level of ICAM-1 on virus producer cells. Moreover, the increase in virion-bound host cell membrane ICAM-1 led to a concomitant enhancement of virus infectivity when a T-cell-tropic strain of human immunodeficiency virus type 1 (HIV-1) was used. The phenomenon was also seen when primary human cells were infected with virions pseudotyped with the envelope protein from a macrophage-tropic HIV-1 isolate, thus ruling out any envelope-specific effect. We also observed that target cells treated with NKI-L16, an anti-LFA-1 antibody known to increase the affinity of LFA-1 for ICAM-1, were markedly more susceptible to infection with HIV-1 particles bearing on their surfaces large numbers of host-derived ICAM-1 proteins. Given that cellular activation of leukocytes is known to modify the conformational state of LFA-1 and induce ICAM-1 surface expression, it is tempting to speculate that activation of virus-infected cells will lead to the production of HIV-1 particles bearing more host ICAM-1 on their surfaces and that such progeny virions will preferentially infect and replicate more efficiently in activated cells which are prevalent in lymphoid organs.

Cell Line↗
PubMed1998

T cells expressing activated LFA-1 are more susceptible to infection with human immunodeficiency virus type 1 particles bearing host-encoded ICAM-1.

The incorporation of host-derived proteins in nascent human immunodeficiency virus type 1 (HIV-1) particles is a well-established phenomenon. We recently demonstrated that the physical presence of host-encoded ICAM-1 glycoproteins on HIV-1 leads to a significant increase in virus infectivity in an ICAM-1/LFA-1-dependent fashion (J.-F. Fortin, R. Cantin, G. Lamontagne, and M. Tremblay, J. Virol. 71:3588-3596, 1997). We show here that conversion of LFA-1 to high affinity for ICAM-1 with the use of anti-LFA-1 antibodies (clones NKI-L16 and MEM83) markedly enhances the susceptibility of different target T-lymphoid cell lines, as well as of primary peripheral blood mononuclear cells, to infection by ICAM-1-bearing HIV-1 particles (6- to 95-fold). It is known that T-cell receptor (TCR) cross-linking induces a transient increase in LFA-1 affinity for ICAM-1. Treatment of peripheral blood mononuclear cells with anti-TCR antibodies (clone OKT3) resulted in a transient increase in susceptibility to infection by ICAM-1-positive virions that parallels the previously reported kinetics of the LFA-1/ICAM-1 adhesion mechanism. Our results led us to postulate that the strong interaction taking place between virally incorporated ICAM-1 and cell surface-activated LFA-1 markedly enhances the efficiency of virus binding and entry, thus favoring greater infection by ICAM-1-bearing HIV-1 particles. In view of the knowledge that primary HIV-1 isolates harbor host-derived ICAM-1 on their surfaces, these results provide new information about the role of host-derived ICAM-1 in the life cycle of HIV-1 and how it could positively modulate the dynamics of the viral infection, mainly in cellular compartments, such as the lymphoid tissues, where the level of cellular activation is high and where the probability of encountering a T cell expressing the activated LFA-1 form is also elevated.

Antibodies↗
PubMed1998

Modulation of human immunodeficiency virus type 1-induced syncytium formation by the conformational state of LFA-1 determined by a new luciferase-based syncytium quantitative assay.

The ICAM-1/LFA-1 interaction has been clearly demonstrated to play an active role in syncytium formation induced by human immunodeficiency virus type 1 (HIV-1). Since it is known that a high-affinity state of LFA-1 for ICAM-1 can be induced through conformational change, such a high-affinity state may also contribute to the process of syncytium formation. In this study, we have investigated the involvement of the conformational status of LFA-1 in HIV-1-dependent syncytium formation by using the anti-LFA-1 antibody NKI-L16, which is known to activate the high-affinity state. Initial visual observations by light microscopy indeed suggested that the addition of the NKI-L16 antibody led to bigger and more numerous syncytia when different cell lines were tested. To further analyze this NKI-L16-dependent increment of syncytium formation in a quantitative assay, a new luciferase-based assay was developed by using a T-cell line containing an HIV-1 long terminal repeat (LTR)-driven luciferase construct (1G5) in coincubation with an HIV-1-positive cell line (J1.1). Upon fusion, the viral Tat protein could diffuse to the 1G5 cells, leading to a transcriptional increase of the HIV-1 LTR-driven luciferase gene. Initial evaluation of this assay showed a good correlation between the level of syncytium formation determined by microscopic observation and the level of measured luciferase activity. In addition, this assay showed a greater induction of enzymatic activity correlating with syncytium formation in comparison to a similar incubation with the HeLa-CD4-LTR-beta-gal indicator cell line. By using this test, NKI-L16 treatment of 1G5/J1.1 cells led to a three- to sevenfold increase in HIV-1 LTR-driven luciferase activity. The syncytium-dependent luciferase activity in NKI-L16-treated cells could be blocked by classical syncytium inhibitors such as soluble CD4, anti-CD4, and anti-gp120 antibodies. Inhibition could also be observed with specific blocking agents for the chemokine receptor CXCR4, as well as with soluble ICAM-1, anti-LFA-1, anti-ICAM-1, and anti-ICAM-2 blocking antibodies, indicating the requirement for the LFA-1/ICAM interaction. Treatment of peripheral blood mononuclear cells with NKI-L16 resulted in a higher level of syncytium formation in the presence of the cell line J1.1. Conversely, when PBMCs were infected with two different syncytium-inducing HIV-1 primary isolates, coincubation with NKI-L16-pretreated 1G5 cells led to higher levels of luciferase activity for both virus isolates. Our results therefore show for the first time a direct role for the LFA-1 high-affinity state in virus-mediated syncytium formation. Based on the demonstration that an increase in ICAM-1 binding is induced by T-cell activation, these data suggest an in vivo involvement of the high-affinity state of LFA-1 in HIV-1-induced syncytium formation. Moreover, syncytia might preferentially occur in lymph nodes, since this microenvironment harbors a high proportion of activated T cells.

Antibodies↗
PubMedDate not supplied

Human herpesvirus 8 viral FLICE-inhibitory protein inhibits Fas-mediated apoptosis through binding and prevention of procaspase-8 maturation.

Viral FLICE-inhibitory proteins (v-FLIPs) encoded by several herpesviruses and poxviruses share the ability to inhibit apoptosis after engagement of death receptors. In the current article, we provide insights into the mechanisms by which the v-FLIP of human herpesvirus 8 (HHV-8) (also referred to as Kaposi's sarcoma-associated virus) protects cells from apoptosis after Fas-induced signaling. Using v-FLIP expression vectors, our results clearly show that HHV-8 v-FLIP reduces the cleavage of procaspase-8 into its active p18 and p10 protease subunits upon Fas-induced cell death. These results were confirmed by lower caspase-8 and caspase-3 protease activities in extracts of HeLa cells expressing HHV-8 v-FLIP. Coimmunoprecipitation studies further indicate that HHV-8 v-FLIP physically interacts with procaspase-8, but not with Fas-associated protein with death domain in the cellular cytoplasm. These results suggest that binding of HHV-8 v-FLIP to procaspase-8 affects the recruitment and the activation of the latter at the death-induced signaling complex, resulting in diminished apoptotic cascade initiation. Because cellular FLIP was recently reported to modulate promoter containing NF-kappaB motifs and that both HHV-8 and human immunodeficiency virus type 1 (HWV-1) can infect monocytes, we studied the effects of v-FLIP on HIV-1 gene expression. Cotransfection experiments indicated that v-FLIP expression is associated with activation of HIV long terminal repeats: events that were strictly dependent on the presence of NF-kappaB consensus elements. In conclusion, HHV-8 v-FLIP can possibly contribute to the pathogenesis of both HHV-8 and HIV-1 through impaired Fas-dependent killing of infected cells by cytotoxic T cells and through activation of HIV gene expression.

Amino Acid Sequence↗