A Research on the Induction of Apoptosis in Lymphocytes by Malaria Infection

(1)伯氏疟原虫（Plasmodium bergbei，P.b.）诱导小鼠淋巴细胞凋亡我们分离并收获了三组小鼠的外周血淋巴细胞，1）20只感染P.b.NK65强毒株的中度寄生虫血症小鼠（D-4），20只感染P.b.NK65强毒株的高度寄生虫血症小鼠（D-5），10只未感染的对照小鼠。将来自各组的淋巴细胞分别与微珠缀合的抗MLA L3 T4（CD 4）辅助T细胞和抗MLA ly 2（CD 8a）抑制T细胞反应，并通过磁性细胞分离器进行分选。从每个标记组的淋巴细胞中提取DNA，并在琼脂糖凝胶中电泳DNA，显示来自高度寄生小鼠的CD 8a阳性T细胞的DNA片段化。(2)恶性疟原虫（P.f.）诱导人淋巴细胞凋亡我们分离并收获了6例感染P.f.的患者的外周血淋巴细胞。这些淋巴细胞与抗Fas单克隆抗体（小鼠）反应，然后与FITC标记的抗小鼠lgs反应。荧光显微镜显示，约1%的外周血淋巴细胞显示阳性荧光，这表明疟疾感染诱导了异常的细胞死亡程序。(3)凋亡淋巴细胞的电镜观察用抗Fas单克隆抗体（鼠）处理外周血淋巴细胞，再用微球标记的抗鼠IgG（鼠）孵育，然后用磁性细胞分离器分离。电子显微镜下可见单个核细胞核碎裂或核内染色质分布异常，并有微珠附着。

(1) Plasmodium bergbei (P.b.) induces apoptosis in mouse lymphocytesWe isolated and harvested peripheral lymphocytes from three groups of mice, 1) 20 mice infected with virulent strain of P.b.NK65 showing moderate level of parasitemia (D-4), 20 mice infected with the same strain of P.b.showing high parasitemia (D-5), and 10 control mice with no infection. Lymphocytes from each group were reacted with microbeads-conjugated anti-MLA L3T4 (CD4) helper T-cells and anti-MLA ly2 (CD8a) suppressor T cells respectively, and subjected to sorting through the Magnetic Cell Separator. Extraction of the DNAs from the each labeled group of lymphocytes and electrophoresis of the DNAs in agarose gels revealed the fragmentation of DNAs from CD8a positive T cells of highly parasitized mice.(2) Plasmodium falciparum (P.f.) induces apoptosis in human lymphocytesWe isolated and harvested peripheral lymphocytes from 6 patients contracting P.f.infection. Those lymphocytes were reacted with anti-Fas monoclonal antibody (mouse) and then with FITC-conjugated anti mouse lgs. Fluorescent microscopy revealed that about 1% of the peripheral lymphocytes showed positive fluorescence, suggesting the induction of an abnormal program of cell death by malaria infection.(3) Electron micrography of apoptotic lymphocytesPeripheral lymphocytes reacted with anti-Fas monoclonal antibody (mouse) was then incubated with microbeads-conjugated anti mouse lgs (rat), and subjected to the Magnetic Cell Separator. Electron microgram of the harvested cells revealed fragmentation of the nucleus or abnormal distribution of the chromatin in the nucleus in mononuclear cells, with the attachment of microbeads.