EXPRESSION OF BCL 2 IN SIV INFECTED THYMIC TISSUE

BCL 2 在 SIV 感染的胸腺组织中的表达

• 批准号: 6453762
• 负责人: M ROSENZWEIG
• 金额: $ 11.11万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-05-01 至 2002-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6453762
• 关键词: AIDS; animal tissue; communicable diseases; immunology; inflammation; lymphatic system; virus

Cell death due to apoptosis plays a critical role in the development of T cells in the thymus Several intracellular molecular mechanisms have been identified that regulate apoptosis in thymocytes, including bcl-2, bcl-x, bax, R-ras and fas Retroviral infection of the thymus with HIV-1 or SIV results in selective thymocyte depletion, and apoptosis has been shown to contribute to this increase in cell death The precise mechanism whereby lentiviruses induce apoptosis remains controversial Using a novel system of in vitro T cell differentiation, we developed a multiparameter flow cytometry assay to determine the role of fas, fas-ligand and bcl-2 in apoptosis of thymocytes as a result of SIV infection We detected a 3-fold increase of apoptosis (detected by Tdt staining) in in vitro cultures 48 post infection with SIV239 Furthermore, the increase in Tdt+ cells is accompanied by a decrease in bcl-2 (2- 3-fold) expression and an increase in surface fas expressio n (4-5 fo ld) as determined by flow cytometry Fas ligand was found to be increased in single positive thymocytes in vivo, but not in the in vitro culture system This data clearly demonstrates that both the bcl-2 and fas pathways are involved in SIV induced apoptosis of thymocytes and that lentiviral infection leads to thymocyte apoptosis Using FACS we demonstrated that SIV infection results in an initial 15-20% decrease of surface fas expression in CD4+CD8+ and CD4+CD8- cells, and not CD4-CD8+ cells However, by day 7 post-infection, we observed a profound reversal of this phenomenon, with a dramatic increase (6 fold) in fas expression in CD4+CD8+ and CD4+CD8- cells, as compared to uninfected cultures These studies have been extended to examine the effects of acute SIV-infection in vivo, with respect to alterations in the thymus Data from these studies demonstrated that the surface levels of fas on thymocytes from normal neonates is generally low (1-3% of all cells), and does not demonstrate any specific phenotypic distribution After SIV infection the level of surface fas changes modestly, increasing to a maximum of 8% of thymocytes of d14 post-infection This was not accompanied by any significant alteration in surface expression of fas-ligand This observation was in contrast to the fluctuations in bcl-2 levels Uninfected neonatal thymocytes express large amounts of bcl-2, consistent with the observations of others However, rapidly during the course of SIV infection the number of bcl-2 positive cells decreases, as well as the amount of bcl-2 per cell as detected by flow cytometry The lowest levels of bcl-2 occur coincident with the largest amount of apoptosis, detected by Tdt staining By day 21 of acute SIV infection, the levels of virus begin to decline and this is associated with the resumption of normal thymic function Specifically, by d21, the number of apoptotic cells is drastically reduced, and the number of bcl-2 cells begins to approach baseline levels The level of bcl-2 on a per cell basis recovers only by day 50 post-infection Based on these observations, it would appear that dysregulation of the bcl-2 pathway is the predominant mechanism associated with the apoptosis that accompanies retroviral infection of the thymus REFERENCES Wykrzykowska JJ, Rosenzweig M, Veazey RS, Simon MA, Halvorsen K, Desrosiers RC, Johnson RP, Lackner AA Early regeneration of thymic progenitors in rhesus macaques infected with simian immunodeficiency virus J Exp Med 1998, 187:1767-1778 Gardner JP, Rosenzweig M*, Marks DF, Harper D, Gaynor K, Fallon RJ, Wall DA, Johnson RP, Scadden DT T lymphopoietic capacity of cord blood-derived CD34+ progenitor cells Exp Hematol 1998; 26:991-999

细胞凋亡引起的细胞死亡在细胞凋亡中起着关键作用。 胸腺中T细胞的发育 已经确定了调节胸腺细胞凋亡的机制， 包括bcl-2、bcl-x、bax、R-ras和fas逆转录病毒感染， 胸腺感染HIV-1或SIV导致选择性胸腺细胞耗竭， 细胞凋亡已被证明有助于这种细胞死亡的增加 慢病毒诱导细胞凋亡的确切机制仍然存在 使用一种新的体外T细胞分化系统， 我们开发了一种多参数流式细胞术测定， Fas、Fas配体和bcl-2在胸腺细胞凋亡中的作用 SIV感染的结果我们检测到细胞凋亡增加了3倍 （通过Tdt染色检测） SIV 239此外，Tdt+细胞的增加伴随着 bcl-2表达减少（2- 3倍）， 流式细胞术测定Fas表达（4-5倍）Fas配体 发现在体内单个阳性胸腺细胞中， 不在体外培养系统中。该数据清楚地表明， bcl-2和fas通路均参与SIV诱导的细胞凋亡 慢病毒感染导致胸腺细胞 我们用流式细胞仪证实SIV感染导致细胞凋亡， CD 4 + CD 8+中表面Fas表达最初降低15-20%， CD 4 + CD 8-细胞，而不是CD 4-CD 8+细胞。 感染后，我们观察到这种现象的深刻逆转， CD 4 + CD 8+细胞中Fas表达显著增加（6倍）， CD 4 + CD 8-细胞，与未感染的培养物相比 已经扩展到检查体内急性SIV感染的影响， 这些研究的数据 结果表明，正常人胸腺细胞表面Fas的水平 新生儿通常较低（占所有细胞的1-3%）， SIV感染后表现出任何特定的表型分布 表面脂肪酸水平变化不大，最大增加到 8%的胸腺细胞感染后d14，这是不伴随任何 Fas-配体表面表达的显著改变 观察结果与bcl-2水平的波动相反 未感染的新生胸腺细胞表达大量bcl-2， 与其他人的观察结果一致，然而， SIV感染过程中bcl-2阳性细胞减少， 以及通过流式细胞术检测的每个细胞的bcl-2的量 最低水平的bcl-2与最大量的 细胞凋亡，通过Tdt染色检测到急性SIV感染的第21天， 病毒水平开始下降，这与 恢复正常胸腺功能具体而言，到d21， 凋亡细胞的数量急剧减少，bcl-2的数量 细胞开始接近基线水平。 细胞基础仅在感染后50天恢复 通过观察，似乎bcl-2通路的失调 是与细胞凋亡相关的主要机制， 伴随胸腺逆转录病毒感染 JJ，Rosenzweig M，Vezey RS，Simon MA，Halvorsen K，Desrosiers RC， 约翰逊RP，Lackner AA小鼠胸腺祖细胞的早期再生 猕猴感染猴免疫缺陷病毒J Exp Med 1998，187：1767-1778 Gardner JP，Rosenzweig M*，Marks DF，哈珀D， Gaynor K，Fallon RJ，Wall DA，约翰逊RP，Scadden DT T淋巴细胞生成 脐血来源的CD 34+祖细胞的能力 1998; 26：991-999