Chaitanya Ganta Vijay, Babu Phanithi Prakash
Department of Animal Sciences, School of Life Sciences, University of Hyderabad, Hyderabad, Andhra Pradesh, India.
Cell Mol Neurobiol. 2009 Jun;29(4):563-73. doi: 10.1007/s10571-009-9348-8. Epub 2009 Feb 19.
Poly (ADP-ribose) polymerase (PARP) is a nuclear repair enzyme whose role is widely depicted in various physiological and pathological processes. In the present study, we wanted to check the status of PARP and the role of various cell death proteases involved in apoptotic and non-apoptotic forms of cell death during transient focal cerebral ischemia in rat model. The activation of these proteases can result in the production of PARP fragments which can be treated as specific signature fragments to the particular protease involved in the pathology and hence the type of cell death.
In the ischemic samples, we observed activation of calpain, cathepsin-b, caspase-3, and granzyme-b which were known to act on and cleave PARP to produce specific signature fragments by Western blot and immunohistochemical analysis. Cresyl violet staining showed the presence of apoptotic and necrotic cell deaths. Further we observed interaction of AIF and gra-b with PARP in double immunofluorescence and co-immunoprecipitation experiments.
Activation of calpains, cathepsin-b, caspase-3, and granzyme-b correlated with either apoptotic or necrotic cell deaths in cresyl violet staining. The appearance of PARP signature fragments gives a clear idea on the involvement of particular protease in the pathology. Appearance of signature fragments like 89- and 50-kDa indicates the involvement of apoptotic and necrotic cell death in the pathology. Further interaction of AIF and gra-b with PARP also indicates the involvement of non-apoptotic modes of cell death during the pathology of focal cerebral ischemia.
聚（ADP - 核糖）聚合酶（PARP）是一种核修复酶，其作用在各种生理和病理过程中被广泛描述。在本研究中，我们想检测PARP的状态以及在大鼠短暂性局灶性脑缺血期间参与凋亡和非凋亡形式细胞死亡的各种细胞死亡蛋白酶的作用。这些蛋白酶的激活可导致PARP片段的产生，这些片段可被视为参与病理过程的特定蛋白酶以及细胞死亡类型的特异性标志性片段。
在缺血样本中，通过蛋白质印迹和免疫组织化学分析，我们观察到钙蛋白酶、组织蛋白酶 - b、半胱天冬酶 - 3和颗粒酶 - b的激活，这些酶已知作用于PARP并将其切割以产生特异性标志性片段。甲酚紫染色显示存在凋亡和坏死性细胞死亡。此外，在双重免疫荧光和免疫共沉淀实验中，我们观察到凋亡诱导因子（AIF）和颗粒酶 - b与PARP的相互作用。
钙蛋白酶、组织蛋白酶 - b、半胱天冬酶 - 3和颗粒酶 - b的激活与甲酚紫染色中的凋亡或坏死性细胞死亡相关。PARP标志性片段的出现清楚地表明了特定蛋白酶在病理过程中的参与情况。89 kDa和50 kDa等标志性片段的出现表明凋亡和坏死性细胞死亡参与了病理过程。此外，AIF和颗粒酶 - b与PARP的相互作用也表明在局灶性脑缺血病理过程中存在非凋亡性细胞死亡模式。
