Caspase-3 gene knockout defines cell lineage specificity for programmed cell death signaling in the ovary.
Caspase-3 gene knockout defines cell lineage specificity for programmed cell death signaling in the ovary.
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DOI：
10.1210/endo.142.6.8078
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发表时间：
2001-06
期刊：
影响因子：
4.8
通讯作者：
T. Matikainen;G. Perez;T. Zheng;T. Kluzak;B. Rueda;R. Flavell;J. Tilly
中科院分区：
文献类型：
--
作者：
T. Matikainen;G. Perez;T. Zheng;T. Kluzak;B. Rueda;R. Flavell;J. Tilly
以前的研究已经提出参与caspase-3，一个下游执行器酶共同的程序性细胞死亡（PCD）的许多范例，在介导的卵巢生殖细胞和体细胞的凋亡。在此，我们使用caspase-3基因敲除小鼠直接测试这种蛋白酶在卵母细胞和/或颗粒细胞死亡中的功能需求。使用在体内和体外的方法，我们确定，卵母细胞死亡开始作为一个结果，无论是发展线索或病理性损伤是不受caspase-3的情况下。然而，在小鼠和人类卵巢退化的窦状卵泡的颗粒细胞表现出强烈的免疫反应，使用的抗体对裂解（激活）形式的胱天蛋白酶-3。此外，caspase-3突变型雌性小鼠具有异常闭锁卵泡，其中含有不能通过凋亡消除的颗粒细胞，如通过DNA切割的TUNEL（末端脱氧核苷酸转移酶介导的脱氧-UTP缺口末端标记）分析和核形态的4 '，6-二脒基-2-苯基吲哚染色（固缩）所证实的。这些体内结果得到了野生型和caspase-3缺陷型窦卵泡或分离颗粒细胞体外培养结果的支持。与野生型颗粒细胞中血清饥饿诱导的凋亡发生相反，缺乏激素支持的caspase-3缺失颗粒细胞为TUNEL阴性，通过4 '，6-二脒基-2-苯基吲哚染色显示染色质浓缩减弱，并显示核小体间DNA裂解延迟。这样的离体研究结果强调存在的细胞自主（颗粒细胞内在）缺陷，导致凋亡执行caspase-3缺陷。我们的结论是，半胱天冬酶-3是功能上所需的颗粒细胞凋亡卵泡闭锁，但该酶在女性的生殖细胞凋亡。
Previous studies have proposed the involvement of caspase-3, a downstream executioner enzyme common to many paradigms of programmed cell death (PCD), in mediating the apoptosis of both germ and somatic cells in the ovary. Herein we used caspase-3 gene knockout mice to directly test for the functional requirement of this protease in oocyte and/or granulosa cell demise. Using both in vivo and in vitro approaches, we determined that oocyte death initiated as a result of either developmental cues or pathological insults was unaffected by the absence of caspase-3. However, granulosa cells of degenerating antral follicles in both mouse and human ovaries showed a strong immunoreaction using an antibody raised against the cleaved (activated) form of caspase-3. Furthermore, caspase-3 mutant female mice possessed aberrant atretic follicles containing granulosa cells that failed to be eliminated by apoptosis, as confirmed by TUNEL (terminal deoxynucleotidyl transferase-mediated deoxy-UTP nick end labeling) analysis of DNA cleavage and 4',6-diamidino-2-phenylindole staining of nuclear morphology (pyknosis). These in vivo results were supported by findings from in vitro cultures of wild-type and caspase-3-deficient antral follicles or isolated granulosa cells. Contrasting the serum starvation-induced occurrence of apoptosis in wild-type granulosa cells, caspase-3-null granulosa cells deprived of hormonal support were TUNEL-negative, showed attenuated chromatin condensation by 4',6-diamidino-2-phenylindole staining and exhibited delayed internucleosomal DNA cleavage. Such ex vivo findings underscore the existence of a cell autonomous (granulosa cell intrinsic) defect in apoptosis execution resulting from caspase-3 deficiency. We conclude that caspase-3 is functionally required for granulosa cell apoptosis during follicular atresia, but that the enzyme is dispensable for germ cell apoptosis in the female.