HIV-1感染者特异CTL反应的研究进展

艾滋病病毒I型(HIV-1)感染可以同时活化宿主的T细胞免疫与体液免疫。被活化的HIV-1特异性细胞毒性T淋巴细胞(CTL),可通过释放穿孔素和颗粒酶或细胞因子与趋化因子等途径,来直接杀伤被感染的靶细胞,或控制病毒进入靶细胞及其复制,在与HIV-1斗争中发挥着极其重要的作用。随着研究的深入,人们对机体抵御HIV-1感染的CTL细胞免疫反应的认识也逐渐加深,已经从单一功能与单一相关因素水平进展为多功能及多因素水平的分析,从而锁定真正具有免疫保护作用的CTL反应功能群,解析控制病毒血症的主要机制,以期为新型HIV疫苗的设计以及效果评价提供科学依据。文章主要就具有免疫保护作用的CTL反应的特征及CTL反应与HIV疾病进展关系的研究等最新进展加以综述。     

Decreased HIV-specific CD4 T cell proliferation in long-term HIV-infected individuals on antiretroviral therapy

The effect of highly active antiretroviral therapy (HAART) on HIV-specific CD4 T cell proliferation in long-term HIV-infected individuals was studied. Subjects receiving treatment for over a year were compared with individuals not receiving therapy. The absolute number of HIV-specific memory CD4 T cells proliferating in response to HIV antigen was substantially higher in untreated subjects than in those on HAART. A decrease in HIV-specific memory CD4 T cells could explain the rebound in HIV replication after the termination of HAART.     

Mycobacterial T cell responses in HIV-infected patients with advanced immunosuppression

Antimycobacterial T cell reactivity at different stages of HIV infection was investigated. Subjects were screened with purified protein derivative (PPD), early secreted antigenic target (ESAT)-6, and culture filtrate protein (CFP)-10 antigens for interferon (IFN)-gamma-producing effector T cell responses by direct ex vivo enzyme-linked immunospot (ELISpot) assay. The proportion of responders to PPD tuberculin decreased with a reduction in CD4 T cell count, whereas the proportion of responder to ESAT-6 and CFP-10 did not. The main sources of IFN-gamma secretion were CD4 cells, and the relative responses to ESAT-6 and CFP-10 significantly increased in HIV-infected patients with decreasing CD4 cell count. This may reflect early signs of reactivation, reinfection, or a restricted, inefficient immune response to Mycobacterium tuberculosis.     

Suppression of HIV-specific T cell activity by lymph node CD25+ regulatory T cells from HIV-infected individuals

CD25(+) CD4(+) FoxP3(+) regulatory T (Treg) cells isolated from the peripheral blood of asymptomatic HIV-infected individuals have been demonstrated to significantly suppress HIV-specific immune responses in vitro. CD25(+) Treg cell suppressor activity in the peripheral blood seems to diminish with progression of HIV disease, and it has been suggested that loss of Treg cells contributes to aberrant immune activation and disease progression. However, phenotypic studies suggest that Treg cells may migrate to, and be maintained or even expanded in, tissue sites of HIV replication. Currently, it is not known whether tissue-associated Treg cells maintain suppressive activity in the context of HIV infection, particularly in individuals with advanced disease. The present study demonstrates that CD25(+) Treg cells isolated from lymph nodes and peripheral blood of HIV(+) subjects, even those with high viral loads and/or low CD4(+) T cell counts, maintain potent suppressive activity against HIV-specific cytolytic T cell function. This activity was better in lymph node as compared with peripheral blood, particularly in patients with high levels of plasma viremia. In addition, the expression of certain CD25(+) Treg-associated markers on CD4(+) T cells isolated from lymph nodes differed significantly from those on CD4(+) T cell subsets isolated from the peripheral blood. These data suggest that CD25(+) Treg cell-mediated suppression of HIV-specific responses continues throughout the course of HIV disease and, because of their particularly potent suppression of HIV-specific CTL activity in lymphoid tissue, may considerably impact the ability to control HIV replication in vivo.     

Enhanced HIV-specific immune responses in chronically HIV-infected patients receiving didanosine plus hydroxyurea

BACKGROUND: The role of hydroxyurea (HU) in the treatment of HIV infection remains controversial. HU potentiates didanosine (ddI) antiviral activity and might exert immunomodulatory effects. PATIENTS AND METHODS: Immunologic parameters were examined in HIV-infected patients enrolled in a simplification trial in which ddI-HU was provided to subjects who had been on complete virus suppression under highly active antiretroviral therapy (HAART) for longer than 6 months. A total of 84 of these patients showed plasma viraemia repeatedly below 5000 HIV-RNA copies/ml, and were the main study population. A group of 22 patients who continued on HAART and another of 22 drug-naive HIV-positive individuals were taken as controls. RESULTS: At 12 months, the levels of naive and memory T-cell subsets were similar in patients on ddI-HU and under HAART, whereas immune activation tended to be lower in the former group. The frequency of HIV-specific CD8+ T cells (CTL) directed against 125 peptides derived from Gag, Pol, Env, Nef and HIV regulatory proteins was similar among patients on ddI-HU and untreated controls, and significantly higher than in patients under HAART. This higher CTL response in patients on ddI-HU was seen even when considering only subjects with undetectable viral load. HIV-specific CD4+ T-cell responses were absent in almost all patients on HAART, whereas they were present in up to 19% of patients on ddI-HU. CONCLUSION: Treatment with ddI-HU provides higher levels of HIV-specific CD8+ and CD4+ T-cell responses than standard triple drug regimens. Thus, hydroxyurea might exert a beneficial immunomodulatory effect in HIV infection.     

HIV subtypes induce distinct profiles of HIV-specific CD8(+) T cell responses

CD8(+) T cells play an important role in controlling HIV infection and qualitative differences in HIV-specific CD8(+) responses may determine the degree of immune control. We studied 56 HIV-infected, ARV-naive Ugandans and examined the role of subtypes in modulating their HIV-specific T cell responses. Gag-specific responses were readily detectable in our study population. Interestingly, we found significantly decreased Gag-specific cytolysis (as measured by CD107 expression) in subtype D (n = 21) compared to subtype A (n = 35) HIV infection. Sequence analyses within identified epitopes suggest patterns of conservation that are subtype specific. We conclude that HIV subtypes may promote distinct profiles of T cells responses and immune control.     

Profile of T cell immune responses in HIV-infected children from Uganda

Human immunodeficiency virus (HIV) immunopathogenesis in children remains poorly understood. We assessed T cell immune activation in antiretroviral therapy-naive children in Uganda (n=154). Increased CD4+ and CD8+ T cell activation strongly correlated with decreased CD4+ T cell percentage. Interestingly, no correlation between plasma HIV RNA level and T cell activation was observed after controlling for CD4+ T cell count. In addition, the presence of Gag-specific CD4+ T helper responses was associated with increased HIV-specific CD8+ T cells. Understanding the balance between immune activation and T cell immunity in HIV-infected children may provide further insights into the mechanisms leading to effective immune control.     

Distinct patterns of HIV-specific memory T lymphocytes in HIV-exposed uninfected individuals and in HIV-infected patients

BACKGROUND: Repeated exposure to HIV is not always associated with infection and multiple cohorts of HIV-exposed but seronegative individuals (ESN) have been described. HIV-specific CD4 and CD8 T lymphocytes are detected both in HIV patients and in ESN; we verified whether different patterns of HIV-specific memory T lymphocytes would be detected in individuals in whom exposure to HIV results or does not result in infection. METHODS: Gag-specific T cells were analysed in 15 ESN, 14 HIV patients, and 15 healthy controls using extensive flow cytometry analysis. RESULTS: Data confirmed that gag-specific T lymphocytes are present in ESN. Gag-specific T cells mainly secrete interleukin-2 in ESN and interferon-gamma in HIV patients. In addition the CD4/CD8 and the memory/naive ratios are altered, central memory (45RA-/CCR7+) CD4 and CD8 T lymphocytes are more abundant, and terminally differentiated (45RA+/CCR7- and 27-/28-) CD8 T lymphocytes are augmented in ESN individuals. CONCLUSIONS: Exposure to HIV occurs in high risk seronegative individuals; the observation that naive cells and CM are skewed in ESN indicate that this exposure is robust enough to modulate the CM/EM ratio. The increase in late effectors and in natural killer cells seen in ESN suggests a role for these cells in preventing actual infection.     

HIV感染者行腹腔镜手术后主要淋巴细胞亚群的变化

目的 检测艾滋病病毒（HIV）阳性者行腹腔镜手术前后主要淋巴细胞亚群等的变化，并与HIV阴性者对照比较，探讨腹腔镜手术对HIV阳性者主要细胞免疫功能的影响。方法 12例胆囊疾病的病人分二组：观察组（HIV阳性）和对照组（HIV阴性）各6人。分别用同样方法做腹腔镜胆囊切除术（LC）。二组于手术前1天（POD-1），手术后3天（POD3）、7天（POD7）检测血常规，白蛋白，CD3、CD4、CD8及其百分比和CD4／CD8。H1v阳性者检测H1v-RNA。结果 观察组手术前1天至手术后1月HIV-RNA〈50拷贝／ml。二组间比较，观察组手术前后CD4值显著低于对照组（P〈0．05）。观察组内比较，POD3的CD4显著低于POD-1和POD7（P〈0．05）。对照组手术前后CD4变化无显著性差异（P〉0．05）。观察组手术前后CD4／CD8也显著低于对照组（P〈0．01）。二组内比较手术前后该比值无显著性差异（P〉0．05）。二组手术后恢复好，无并发症。随访观察（≤20个月）无艾滋病相关症状。结论 LC可能对HIV阳性者主要细胞免疫功能有短暂轻度抑制。总体上HIV阳性者行腹腔镜手术是安全的。术前应常规检测CD4和CD4／CD8，将CD4作为手术风险主要评估指标，术前CD4≥200是确保手术安全性的基本条件。     

Poliovirus vaccination responses in HIV-infected patients: correlation with T4 cell counts

HIV infection is known to impair both cellular and humoral immunity. The antibody levels to poliovirus were measured in 17 HIV-infected and 3 HIV-seronegative homosexual men before and after vaccination with enhanced inactivated poliovirus vaccine (eIPV). The subjects were tested for neutralizing antibodies to poliovirus types 1, 2, and 3; the number of peripheral blood T4 cells and lymphocyte responses after stimulation with phytohemagglutinin, concanavalin A, and pokeweed mitogen were measured. Before vaccination, all subjects demonstrated neutralizing antibodies to the three types of poliovirus. After eIPV administration, a rise in anti-poliovirus antibody titer was noted in subjects whose T4 cell count was greater than 200/ml, whereas the group with low T4 cell counts failed to respond. The response to eIPV immunization correlated with the number of circulating T4 cells and the mitogen-induced lymphocyte stimulation index.     

Thalidomide stimulates T cell responses and interleukin 12 production in HIV-infected patients.

We performed a placebo-controlled study to evaluate the effects of immunomodulatory treatment with thalidomide on HIV levels, TNF-alpha levels, and immune status of 31 HIV-infected individuals, after temporary suppression of viral replication with antiretroviral drugs. Treatment with a combination of zidovudine and lamivudine (ZDV/LMV) for 14 days resulted in a median decline in plasma viremia of 1.94 log10 RNA equivalents/ml. After discontinuation of ZDV/LMV, thalidomide therapy (200 mg/day for 4 weeks) did not retard the prompt return of HIV titers to the pretreatment levels, and had no effect on plasma levels of TNF-alpha. In contrast, thalidomide treatment resulted in significant immune stimulation. We observed increased levels of plasma soluble IL-2 receptor, soluble CD8 antigen, and IL-12 (p < 0.01 for all parameters), as well as increased cutaneous delayed-type hypersensitivity reactions to recall antigens (p < 0.01) in thalidomide-treated patients. These changes were associated with a median increase in HIV titer of 0.2 log10 RNA equivalents/ml in the thalidomide-treated group (p < 0.05), which resolved after stopping the drug. Further studies were performed in vitro to elucidate the mechanism of thalidomide-induced immune stimulation. When purified T cells from HIV-infected individuals were stimulated by immobilized anti-CD3 in the presence of thalidomide, a costimulatory effect of the drug was observed, resulting in increased production of IL-2 and IFN-gamma, and increased T cell-proliferative responses. Further experiments showed that thalidomide increased IL-12 production by antigen-presenting cells in a T cell-dependent manner. Our findings suggest a potential application for thalidomide as a novel immune adjuvant in HIV disease.     

Age-related impairment of T cell proliferative responses related to the decline of CD28+ T cell subsets

The impairment of phytohaemagglutinin-triggered lymphocyte proliferation represents a prominent immunologic abnormality in elderly individuals. To assess whether the reduced function is related to a CD28/B7 signalling deficiency, purified T lymphocytes and antigen presenting cells (APCs) were analyzed for their phenotypic profile and/or functional capacities. T cell responses to immobilized OKT3 monoclonal antibodies (mAb) or a combination of anti-CD2 mAb and phorbol esters were unaffected in old subjects when compared to the younger counterpart. In contrast, CD28 costimulation in the presence of OKT3 or anti-CD2 mAb, gave rise to significantly diminished T cell proliferative responses. These findings correlated with a marked decline of CD28(+) T cell frequency, which mainly involved the CD4(-)CD45RO(-) cell subset. The defect in CD28 expression could not be reversed by T cell stimulation, as a comparable increase in CD28 levels occurred in both ;aged' and ;young' T cells after in vitro activation. Moreover, the elderly group did not exhibit a reduction of interleukin (IL)-2 synthesis, as assessed at 24 h of culture, regardless of the stimulant used. Finally, B7.2 (CD86) expression by ;aged' CD14(+) APCs was unaffected in both resting and interferon-gamma activated cells. These results suggest that an intrinsic defect in CD28 expression might in part account for the age-related decline of T cell proliferative responses.     

Age-related impairment of T cell proliferative responses related to the decline of CD28+ T cell subsets

The impairment of phytohaemagglutinin-triggered lymphocyte proliferation represents a prominent immunologic abnormality in elderly individuals. To assess whether the reduced function is related to a CD28/B7 signalling deficiency, purified T lymphocytes and antigen presenting cells (APCs) were analyzed for their phenotypic profile and/or functional capacities. T cell responses to immobilized OKT3 monoclonal antibodies (mAb) or a combination of anti-CD2 mAb and phorbol esters were unaffected in old subjects when compared to the younger counterpart. In contrast, CD28 costimulation in the presence of OKT3 or anti-CD2 mAb, gave rise to significantly diminished T cell proliferative responses. These findings correlated with a marked decline of CD28⁺ T cell frequency, which mainly involved the CD4⁻CD45RO⁻ cell subset. The defect in CD28 expression could not be reversed by T cell stimulation, as a comparable increase in CD28 levels occurred in both `aged' and `young' T cells after in vitro activation. Moreover, the elderly group did not exhibit a reduction of interleukin (IL)-2 synthesis, as assessed at 24 h of culture, regardless of the stimulant used. Finally, B7.2 (CD86) expression by `aged' CD14⁺ APCs was unaffected in both resting and interferon-γ activated cells. These results suggest that an intrinsic defect in CD28 expression might in part account for the age-related decline of T cell proliferative responses.     

Respiratory syncytial virus-specific CD8+ memory T cell responses in elderly persons

BACKGROUND: We investigated respiratory syncytial virus (RSV)-specific CD8(+) memory T cell responses in healthy control participants (n=31) and in patients with chronic obstructive pulmonary disease (COPD) (n=9), with respect to frequency, memory phenotype, and proliferative requirements. METHODS: The properties of RSV-specific CD8(+) T cells were analyzed by use of RSV tetramers. The proliferative requirements of RSV-specific CD8(+) T cells were analyzed by culture of peripheral-blood mononuclear cells with RSV peptide in combination with distinct cytokines. RESULTS: RSV-specific CD8(+) memory T cells showed a high level of expression of CD27 and interleukin-7R alpha and a low level of expression of CCR7. In the healthy participants, the frequency of RSV tetramer(+) CD8(+) T cells was significantly lower than the frequency of influenza virus A (FLU) tetramer(+) CD8(+) T cells (P=.0001). In contrast to FLU tetramer(+) CD8(+) T cells, we could detect RSV tetramer(+) CD8(+) T cells in the subgroup of elderly healthy participants (age, > or =55 years) and in the patients with COPD only after in vitro expansion. Expanded RSV-specific T cells produced interferon- gamma and granzyme B. CONCLUSION: We provide evidence that a pool of functional RSV-specific CD8(+) memory T cells persists in the peripheral blood of healthy individuals and patients with COPD. Low numbers of RSV-specific memory T cells in the elderly and in patients with COPD may explain the increased susceptibility to RSV infection in these populations.     

Analysis of B cell and T cell proliferative responses induced by monomorphic and pleomorphic Trypanosoma brucei parasites in mice

Cells collected from C57B1/6 mice infected with monomorphic and pleomorphic clones of Trypanosoma brucei parasites (ILTat 1.4 and GUTat 3.1) were analysed for the incorporation of 125I-Iododeoxyuridine into DNA of total splenic lymphocytes and of B and T lymphocytes isolated on a fluorescence activated cell sorter. The monomorphic T. brucei ILTat 1.4 parasites triggered delayed and low splenic DNA synthetic responses in comparison to those arising in mice infected with the pleomorphic T. brucei GUTat 3.1 organisms. Mice infected with both parasite clones mounted splenic DNA synthetic responses similar to those arising in animals infected with the pleomorphic organisms alone and similar responses were induced by lethally irradiated T. bruceiGUTat 3.1 and T. brucei ILTat 1.4 parasites. In mice infected with the pleomorphic parasites, DNA synthesis was first detected in the T cell population and B cell DNA synthetic responses were detected between 1 and 2 days later. In contrast only T cell DNA synthetic responses were detected after infection with the monomorphic T. brucei ILTat 1.4 parasites. It is suggested that the previously reported failure of monomorphic T. brucei parasites to trigger antibody production in infected mice is a result of their inability to stimulate B lymphocytes.     

Therapeutic vaccination with p24-VLP and zidovudine augments HIV-specific cytotoxic T lymphocyte activity in asymptomatic HIV-infected individuals

This study evaluates the impact of therapeutic vaccination with p24-VLP and zidovudine on the induction or maintenance of HIV-specific cytotoxic lymphocyte activity in a cohort of asymptomatic patients with CD4 counts greater than 400 cells/microl. In a dummy, randomized, phase II clinical trial of the therapeutic vaccine, participants were randomized to one of three arms for 6 months: p24-VLP (500 microg) in alum monthly plus zidovudine 200 mg tds, alum adjuvant plus zidovudine, or p24-VLP plus placebo. Subjects were studied for a total of 52 weeks from baseline. Monitoring included viral load, CD4 and CD8 counts, markers of immune activation, delayed-type hypersensitivity (DTH) skin testing, and cytotoxic T lymphocyte (CTL) measurement. The nine subjects who received p24-VLP and zidovudine had an augmentation and/or broadening of their CTL response compared with baseline (p = 0.004). The eight subjects receiving p24-VLP and seven subjects receiving zidovudine did not have a statistically significant increase or broadening of CTL activity. The augmentation of the CTL response in the subjects who received p24-VLP and zidovudine was not associated with a decline in viral load or an increase in CD8 counts. This study suggests that HIV-specific CTL activity can be augmented in HIV-infected individuals receiving p24-VLP and zidovudine, supporting the hypothesis of therapeutic vaccination in the presence of antiretroviral therapy.