| HIV infection induces a progressive deterioration of the immune system and ultimately leads to onset of acquired immune deficiency syndrome (AIDS). Highly active antiretroviral therapy (HAART) has been used for clinical treatment, and it partly restores the immune system in those individuals infected with HIV. In light of this incomplete success, with technological developments, extensive efforts have been made for immunological recovery. Enhancing HIV-specific CD8+T cell immune responses is one of the most important mechanisms and gradually become the focus of research. At present, a number of new therapies and strategies are entering testing or clinical stages, including the use of various cytokines.Interleukine21(IL-21) bears special relevance to HIV/SIV/SHIV infection. IL-21is relatively recently discovered and has been shown to exert significant immune enhancing and regulatory functions. It has been demonstrated that CD4+T cells are the main producers of this cytokine. It is noteworthy that these cells are also the primary targets of HIV/SIV/SHIV in infected individuals and non-human primates. During acute HIV infection, virus-specific CD8+T cells are preferentially expanded to control viral replication. Many scholars pay much attention to the ability of IL-21to regulate CD8+T cell responses. At the same time, many cytokines are coordinately induced following HIV infection as part of the well described "cytokine storm" of acute infection. In the clinic, it is too difficult to know the accurate time of HIV infection in human subjects and to study acute HIV infection. Hence, little is known about the dynamic changes in serum/plasma IL-21production during acute HIV/SIV/SHIV infection, but SHIV/Chinese origin rhesus model has been created successfully through SHIV infection and make it find the method of resolving the questions. The immunological mechanisms of IL-21regulating SHIV-specific CD8+T cells also remain to be elucidated.Although some cytokines play a positive role in defense against HIV/SIV/SHIV, promoting virus replication in CD4+T cells of virus-infected individuals is the major obstacle to successful clinical therapy and vaccine adjuvants development. It is necessary to assess the effect of IL-21administration on CD4+T cells. This thesis has finished with three parts:The objective of the first part was to study dynamic changes in the levels of plasma IL-21in rhesus macaques (RMs) following SHIV infection. We investigated the dynamics of IL-21and a few immunological indicators after rectal infection with SHIV-1157ipd3N4and further analyzed the potential association exists between either of those parameters in acute SHIV infection. All peripheral blood samples from22RMs infected with SHIV were collected accordingly. Absolute CD4+and CD8+T cells was quantified, real-time fluorescence quantitative RT-PCR assay was used to determine virus loads for SHIV RNA, and the concentrations of plasma IL-21and IFN-y were determined by ELISA. This is the first result that the levels of plasma IL-21during SHIV infection were significantly higher than basal ones. Moreover, the levels of plasma IL-21were correlated positively with the concentrations of plasma IFN-y. Similarly, they were correlated positively with the absolute numbers of CD4+T cells in these RMs, but not with CD8T cells. In conclusion, IL-21has potential role in the immunopathogenesis of HIV/SI V/SHIV, and might be an important factor in immune construction.In vivo, the significantly elevated IL-21in acute SHIV infection bound with IL-21R and may raise a number of biological effects. In the second part, we next examined IL-21influences on immune regulation of HIV-specific CD8+T cells from SHIV-infected RMs and explored the mechanisms in vitro. CD8+T cells from RMs infected with SHIV were stimulated with IL-21and the expression of IL-21R, IFN-y, perforin, Granzyme B and CD107a of CD8+T cells was assayed, respectively. The effects of IL-21on killing activity, proliferation and apoptosis level of CD8+T cells were also measured. Moreover, the changes of STAT3and STAT5activation in CD8+T cells were determined. We show that, IL-21(10ng/mL) was found to obviously increase IFN-y level (P<0.05) in the supernatant of CD8+T cells compared with the control group. The expression of IFN-7mRNA was significantly enhanced by real time PCR, and IFN-γ which requires de novo synthes is following activation peaked at4h post stimulation. The released increases of perform, Granzyme B were mediated by IL-21from activated CD8+T cells. Moreover, percentage of CD107a+CD8+T cells was higher than those of the control. Compared with the medium control, the expression of IL-21R on the surface of CD8+T cells was significantly increased by laser scanning confocal microscopy, real time PCR and FC. In addition, the expressions of pSTAT3was significantly up-regulated (P<0.05), while had no effect on pSTAT5. Thus, our studies reveal that IL-21administration increases CD8+T cell-mediated immune response and the mechanism of the immune regulation may be through the JAK/pSTAT3signal pathway.The above multicenter studies indicated that IL-21enhances function of CD8+T cells and specific anti-HIV immune responses. Hence it is necessary to evaluate the effect of IL-21administration on CD4+T cells. In the third part, CD4+T cells from RMs infected with SHIV were analyzed for CD69, Ki67and P27Ag expression, respectively. Lymphocyte subsets were also analyzed in the presence of rhIL-21. Here we showed that, the expression percentages of CD4+T cells in response to IL-21(0.28±0.047)%was equal with medium control (0.25±0.003)%, which were significantly lower than that of IL-2(2.03±0.97)%(P<0.05). IL-21did not change the proportions of the T cell subsets. The expressions of Ki67in CD4+T cells in response to IL-21[(1.35±0.18)%] was higher than those of medium control [(0.68±0.07)%], but with no statistical significance. The expressions of peripheral blood CD4+T cells Ki67were greatly higher in the presence of rh IL-2than those in the controls (P<0.01). Moreover, the expression rates of P27of CD4+T cells was (0.56±0.06)%,(0.85±0.11)%,(0.89±0.09)%,(0.88±0.19)%and (1.03±0.25)%in response to5,10,50,100and200μg/mL IL-21stimulation, respectively, was higher than those of medium control (0.47±0.04)%, but with no statistical significance. In addition, P27expression on CD4+T cells after20U/mL and40U/mL IL-2stimulation was much higher than that of medium control. In conclusion, IL-21administration did not alter CD4+T cell immune activation and viral replication. The results showed that IL-21was safer than IL-2.To sum up, in light of an increased production of circulating IL-21in SHIV-infected RMs, and it exerts its biological effects on SHIV-specific CD8+T cells with minimal stimulation of HIV-1replication, and IL-21has an acceptable safety profile, being well tolerated at low doses, these data suggest that IL-21may be considered as a anti-AIDS drug and a therapeutic agent to boost anti-HIV immune responses in HIV-infected AIDS patients. |
PDF Full Text Request