In vitro apoptosis and expression of apoptosis-related molecules in lymphocytes from patients with systemic lupus erythematosus and other autoimmune diseases
In vitro apoptosis and expression of apoptosis-related molecules in lymphocytes from patients with systemic lupus erythematosus and other autoimmune diseases
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DOI：
10.1002/art.1780400216
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发表时间：
1997-02-01
影响因子：
--
通讯作者：
Kalden, JR
中科院分区：
文献类型：
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作者：
Lorenz, HM;Grunke, M;Kalden, JR
Objective.分析系统性红斑狼疮（SLE）患者外周血单个核细胞（PBMC）凋亡的相关因素，并与正常人及其他自身免疫性疾病患者PBMC进行比较。分离来自正常健康供体或患有SLE、混合性结缔组织病（MCTD）、类风湿性关节炎（RA）或各种血管炎的患者的PBMC。使用碘化丙啶染色定量通过不同信号传导途径活化后的凋亡百分比。检测Fas/APO-1、bcl-2蛋白表达及bcl-2、bcl-x（L）、bar、巴克、Fas/APO-1、Fas配体（Fas-L）、c-myc、mad、max mRNA表达。我们证实了以前的研究结果，SLE PBMC中凋亡细胞的数量增加，与正常供体细胞体外孵育后。用CD 28单克隆抗体+佛波酯PMA（CD 28 MAW PMA）、葡萄球菌肠毒素B（SEE）或植物血凝素（PHA）激活PBMC后，SLE细胞中凋亡细胞的百分比不变（SEE）或减少（CD 28 MAB/PMA，PHA），正常供体与SLE细胞之间的差异不明显。在mRNA水平上，SLE相关基因产物的表达在SLE细胞和正常供体细胞之间没有差异。与正常供者T淋巴细胞相比，SLE患者T淋巴细胞Fas/APO-1蛋白表达增加，而bcl-2蛋白表达在培养3天后上调。细胞活化进一步增加bcl-2蛋白水平，消除正常供体和SLE患者之间的差异。在Ri细胞中，细胞凋亡的百分比与正常供体PBMC中的细胞凋亡的百分比相似，而使用来自其他自身免疫性疾病（MCTD、韦格纳肉芽肿、大动脉炎、结节性多动脉炎）患者的细胞的结果与使用SLE PBMC发现的结果相当。在培养液中加入生长因子如白细胞介素-2（IL-2）、IL-4或IL-15可降低正常供体和SLE细胞的体外凋亡百分比。基于这些数据，我们得出结论，加速体外细胞凋亡和增加Fas/APO-1和bcl-2蛋白表达的SLE是非特异性的疾病，并可能至少部分地解释了在体内增加PBMC的激活水平，从患者，SLE，MCTD，或自身免疫性血管炎结合体外孵育条件下的“非炎症"和生长因子的撤退。
Objective. To analyze factors related to apoptosis in systemic lupus erythematosus (SLE) peripheral blood mononuclear cells (PBMC) and to compare the findings in SLE PBMC with those in normal donor PBMC or PBMC from patients with other autoimmune diseases.Methods. PBMC from normal healthy donors or patients with SLE, mixed connective tissue disease (MCTD), rheumatoid arthritis (RA), or various vasculitides were isolated. The percentage of apoptosis after activation through different signaling pathways was quantified using propidium iodide staining. Protein expression of Fas/APO-1 or bcl-2, and messenger RNA (mRNA) expression of bcl-2, bcl-x(L), bar, bak, Fas/APO-1, Fas ligand (Fas-L), c-myc, mad, or max were determined.Results. We confirmed previous findings of increased numbers of apoptotic cells in SLE PBMC compared with normal donor cells after in vitro incubation. After activation of PBMC with CD28 monoclonal antibody plus phorbol myristate acetate (CD28 MAW PMA), staphylococcal enterotoxin B (SEE), or phytohemagglutinin (PHA), the percentage of apoptotic cells was unchanged (SEE) or diminished (CD28 MAb/PMA, PHA) in SLE cells, and the difference between normal donor and SLE cells was less pronounced. On the mRNA level, expression of apoptosis-related gene products did not differ between SLE cells and normal donor cells. Expression of Fas/APO-1 protein was increased in freshly isolated SLE T lymphocytes compared with normal donor T lymphocytes, whereas bcl-2 protein was up-regulated after a 3-day culture period. Cellular activation further increased bcl-2 protein levels, eliminating differences between normal donors and SLE patients. In Ri cells, the percentage of apoptosis was similar to that in normal donor PBMC, whereas results using cells from patients with other autoimmune diseases (MCTD, Wegener's granulomatosis, Takayasu arteritis, polyarteritis nodosa) were comparable with those found using SLE PBMC. Addition of growth factors such as interleukin-2 (IL-2), IL-4, or IL-15 to culture medium decreased the percentage of in vitro apoptosis in both normal donor and SLE cells.Conclusion. Based on these data, we conclude that accelerated in vitro apoptosis and increased Fas/APO-1 and bcl-2 protein expression in SLE are nonspecific for the disease, and might be explained at least in part by the increased in vivo activation levels of PBMC from patients,vith SLE, MCTD, or autoimmune vasculitides combined with in vitro incubation under ''noninflammatory'' conditions and growth factor withdrawal.