HIV慢性感染者CD8~+T淋巴细胞及其各亚群细胞增生、消亡情况的观察

目的探讨HIV-1慢性感染者(病程>12个月)CD8+T淋巴细胞及其各亚群:纯真CD8+T细胞(TN)、中心记忆CD8+T细胞(TCM)、效应记忆CD8+T细胞(TEM)和效应记忆CD8+T细胞RA(TEMRA)的增生、消亡情况。方法观察23例未经抗病毒治疗、病程大于12个月的HIV-1感染者以及16例健康对照者,通过流式细胞仪分析,根据CD45RO,CD27表达情况,CD8+T细胞分为TN(CD45RO-CD27+)、TCM(CD45RO+CD27+)、TEM(CD45RO+CD27-)和TEMRA(CD45RO-CD27-)。以Ki-67抗原的表达代表细胞的增生,AnnexinV检测阳性代表细胞的消亡,比较HIV-1慢性感染者和健康者CD8+T细胞及其各亚群增生消亡的情况;并把HIV-1慢性感染者按CD4+T细胞计数200个/mm3以及350个/mm3为观察点分成3组,比较3组间的CD8+细胞及其各亚群增生消亡的情况。结果健康组(n=16)CD8+细胞各亚群检出率为TN34.92%±12.68%、TCM25.44%±10.36%、TEM14.64%±10.58%和TEMRA25.00%±12.59%;与之相比HIV-1慢性感染组(n=23)检出率为TN17.14%±8.03%(t=5.368,P=0.000)、TCM31.40%±14.02%(t=-1.448,P=0.156)、TEM20.17%±13.17%(t=-1.393,P=0.172)和TEMRA31.48%±15.16%(t=-1.405,P=0.168)。CD8+T细胞增生率为:健康组0.15%±0.09%,感染组1.33%±0.90%(z=-4.655,P=0.000);健康组消亡率为8.74%±4.73%,感染组24.08%±13.72%(z=-4.169,P=0.000)。HIV-1慢性感染者按CD4+T细胞计数分成CD4≤200个/mm3组(A组,n=5)、CD4计数200～350个/mm3组(B组,n=12)和CD4≥350个/mm3组(C组,n=6),3组间各亚群组成比中TN所占比例在A组明显减少,而CD8+T细胞各亚群的增生率、消亡率在3组间差异没有统计学意义。结论①与健康对照组相比,HIV-1慢性感染者CD8+T细胞及其各亚群消亡率增加,增生率也相应大幅增加,细胞进入高速更新的状态;②HIV-1慢性感染者CD8+TN亚群的检出率明显下降,并在CD4≤200个/mm3组降至最低,CD8+TN资源随病情进展不断耗竭。

Investigation on Proliferation and Death of CD8+ T-cell and Its Subsets in Chronic HIV-1 Infected Persons

Objective To observe the changes associated with proliferation and death of CD8⁺ T cell and its subsets including naive CD8⁺ T-cell (T_N), central memory CD8⁺ T-cell (T_CM),effection memory CD8⁺ T-cell (T_EM) and effection memory CD8⁺ T-cell RA (T_EMRA) in chronic HIV-1 infected patients and healthy controls. Same observations were also carried out according to different CD4⁺ T-cell count in chronic HIV-1 infected patients. Methods Sixteen healthy controls and 23 untreated HIV-1 infected patients whose disease course was over 12 months were enrolled in observation. Flow cytometry tests were carried out after routine staining. CD8⁺ T-cells were divided into four subsets based on CD45RO and CD27 expression. They were T_N(CD45RO-CD27+), T_CM (CD45RO+CD27+), T_EM (CD45RO+CD27-) and T_EMRA(CD45RO- CD27-)respectively. Cell proliferation was studied by measuring expression of the Ki-67 antigen. Cell death was studied by annexin V staining. Cell proliferation and death ratio were compared between HIV-1 chronic infection group and healthy group. Same investigations were conducted among three HIV infected groups which were divided according to CD4⁺ T-cell count at 200/mm³ and 350/mm³ levels. Results The distribution of CD8⁺ T-cell subsets in normal control(n=16) were T_N 34.92%±12.68, T_CM 25.44%±10.36%, T_EM 14.64%±10.58%, and T_EMRA 25.00%±12.59%,respectively. The counterparts in HIV-1 infected group(n=23) were T_N 17.14%±8.03%(t=5.368, P=0.000), T_CM 31.40%±14.02%(t=-1.448, P=0.156), T_EM 20.17%±13.17%(t=-1.393, P=0.172), and T_EMRA 31.48%±15.16%(t=-1.405, P=0.168) respectively. The ratio of proliferating cells in CD8⁺ T-cells were 0.15%±0.09% in healthy control and 1.33%±0.90% in infection group(z=-4.655, P=0.000). The ratio of dying cells in CD8⁺ T-cells were 8.74%±4.73% in healthy control and 24.08%±13.72% in infection group(z=-4.169, P<0.001). There was a statistically significant difference in T_N subset among CD4≤200/mm³ group(n=5), CD4 200~350/mm³ group(n=12) and CD4>350/mm³ group(n=6). But there was no significant difference in proliferation and death ratios among these three groups. Conclusion 1 Compared with healthy control, death and proliferation ratios in CD8⁺ T-cells and its subsets increased dramatically. CD8⁺ T-cells were shifted into high flow-ratio cell cycling. 2 The composite ratio of CD8⁺ T_N subset was dramatically down regulated especially in the CD4<200/mm³ group. The resource of CD8⁺ T-cells T_N subset was exhausted following disease progress.