HIV-1 gp120/V3-derived epitopes promote activation-induced cell death to superantigen-stimulated CD4+/CD45RO+ T cells.

The third hypervirable (V3) domain of the HIV-1 envelope glycoprotein gp120 has been implicated in HIV pathogenesis via co-receptor usage of chemokine receptors CCR5 and CXCR4. As the protagonist cell populations in the asymptomatic phase of HIV-1 infection are infected macrophages and effector/memory (CD45RO+) CD4+ T cells that express CCR5, we established an in vitro model using human primary monocyte-derived macrophages and lymphocytes to investigate the role of V3 in affecting antigen presentation. We used staphylococcal enterotoxin A (SEA) as a superantigen at a low concentration of 1ng/ml, to activate naïve CD4+ T cells. Exposure of cells to SEA and lipoV3-liposomes increased the percentage of CD4+/CD45RO+/CCR5+ T cell population as compared to cells treated with SEA and plain liposomes. A consequent decrease of the percentage of CD4+/CD45RO+/CXCR4+ subset was observed. The V3-mediated activation was competitively inhibited by soluble V3-derived peptides with higher cationic charge. V3 enhanced also apoptosis as demonstrated by flow cytometry and intracellular calcium ion assays. These results reinforce the postulation that V3 alters the antigen presentation function itself, independent of specific antigens, thus leading to an enhanced activation-induced cell death (AICD) of responding T cells.

CX3CR1 receptor is up-regulated in monocytes of coronary artery diseased patients: impact of pre-inflammatory stimuli and renin-angiotensin system modulators.

Fractalkine/CX3CR1 pathway is considered a major modulator of atherosclerosis. In the present study, expression of CX3CR1 on PBMCs/monocytes of healthy individuals and coronary artery diseased patients was initially assessed by flow cytometry. Effects of pre-inflammatory cytokines interferon (INF)-gamma and tumor necrosis factor (TNF)-alpha on expression of CX3CR1 and a single representative of each major chemokine family (CCR5 and CXCR4) were further assessed in three cell models: THP-1 monocytes, Jurkat T lymphocytes and primary monocytes isolated from healthy donors. Finally, effects of angiotensin-converting enzyme (ACE) inhibitors captopril, lisinopril and angiotensin receptor blocker (ARB) losartan on chemokine receptor expression were evaluated in the same cell models either in a naive or stimulated state. INF-gamma significantly affected the chemokine receptor phenotype of THP-1 cells by increasing the rate of CX3CR1-positive cells. Pre-treatment with the ACE inhibitors, captopril and lisinopril, and the ARB, losartan, did not influence these effects. Captopril and lisinopril similarly had no effect on either stimulated or naive primary monocytes. Yet, a small but repeatable increase in CX3CR1 expression after treatment with losartan was noted. Nevertheless, the latter observation did not retain statistical significance after applying the Bonferroni correction. In conclusion, our data did not indicate any significant effect of the ACE inhibitors on the chemokine receptor phenotype of monocytes.

In vitro cytopathic effects of mycotoxin T-2 on human peripheral blood T lymphocytes.

The trichothecene mycotoxin T-2 is reported to exhibit immunotoxic activity. The potential presence of T-2 in foods renders it as public health hazard and its toxicity needs to be better understood. We investigated the in vitro effects of T-2 at sub-toxic (0.1 ng/ml) and toxic (10 ng/ml) levels on freshly isolated human peripheral blood lymphocytes (PBLs). We observed no direct influence on untreated PBLs. The toxic dose of T-2, however, totally inhibited phytohemagglutinin-induced T lymphocyte proliferation and caused early apoptosis that peaked after 8h of exposure. Both major T lymphocyte subsets (CD4+ and CD8+) were affected as they appeared to show a positive response to T-2 at 8h followed by their sharp reduction after 96 h. Further investigation on the naïve (CD45RA+) and memory (CD45RO+) subpopulations confirmed these observations and indicated that T-2 affected equally all the subpopulations studied, although PHA preferentially stimulated CD45RO+ T lymphocytes. Sub-toxic T-2 appeared to exhibit co stimulatory properties to PHA-stimulated cells. These results support the hypothesis that T-2 affects the activation-induced cell death mechanism of T lymphocytes.

Opposite effect of angiotensin receptor blockade on CXCL8 production and CXCR1/2 expression of angiotensin II-treated THP-1 monocytes.

Interleukin-8 (IL-8) or CXCL8 is a potent chemotactic factor that is involved in atherogenesis. IL-8 mediates its pre-inflammatory effects through interaction with CXCR1 and CXCR2. In the present study, we investigated the effects of angiotensin II (Ang II) on IL-8 synthesis and CXCR1/CXCR2 expression of THP-1 monocytes. IL-8 was measured in the culture medium using ELISA. Expression of chemokine receptors CXCR1 and CXCR2 was evaluated by flow cytometry. Results demonstrated that the addition of Ang II increased IL-8 production in the THP-1 monocytes. The Ang II type 1 receptor blocker (ARB) losartan significantly blocked Ang II-induced IL-8 production. Notably, losartan blocked LPS-induced IL-8 production by THP-1 monocytes and produced a small but statistically significant reduction of baseline IL-8 production of naïve THP-1 cells. Losartan also produced a statistically significant increase of fluorescence intensity of naïve CXCR1- and CXCR2-positive THP-1 monocytes, probably as a negative feedback effect secondary to IL-8 downregulation. In conclusion, we demonstrated that Ang II increased IL-8 production by THP-1 monocytes. Losartan significantly suppressed the latter effect, suggesting an AT-1 mediated pathway. Moreover, losartan suppressed the IL-8 production of naïve THP-1 monocytes and LPS-treated THP-1 monocytes, suggesting a broader spectrum of pleiotropic effects. Extrapolating this in vitro observation to in vivo pathways, we propose Ang II-induced IL-8 production by monocytes as another pre-atherogenic potential of Ang II that can be effectively blocked by the AT1 receptor blockade.

Evaluation of specific immune responses to BoNT/A and tetanus toxoid in patients undergoing treatment for neurologic disorders.

Botulinum neurotoxins (BoNTs) are used in the treatment of many neurological disorders. The primary structure of BoNTs shows a high degree of homology with the tetanus neurotoxin, the toxoid of which is used as a vaccine. Because of the potential cross-reactivity between these toxins, we investigated the effects of Botulinum neurotoxin A (BoNT/A) and tetanus toxoid on peripheral blood mononuclear cells (PBMC) and the corresponding serum antibody levels, in twenty patients who had been treated with BoNT/A. We observed very low PBMC immunostimulation by BoNT/A at the tested dose (15 units/ml), as demonstrated by the low lymphocyte proliferation, and the absence of detectable antibodies cross-reacting with tetanus. However, exposure of PBMC from tetanus-sensitized patients to both neurotoxins showed that BoNT/A exerted a co stimulatory effect on tetanus-stimulated cells. Interestingly, in flow cytometry analysis, BoNT/A seemed to also alter the ratio of naïve (CD45RA) : memory/effector (CD45RO) T lymphocyte subsets, in favour of CD45RO. These preliminary data give a new insight on the potential immune crossreactivity between the two antigens. In view of the wide use of both neurotoxins, these immunotoxic effects merit a more detailed investigation.