Bhave Vishakha S, Donthamsetty Shashikiran, Latendresse John R, Muskhelishvili Levan, Mehendale Harihara M
Department of Toxicology, College of Pharmacy, University of Louisiana at Monroe, 700 University Avenue, Monroe, LA 71209, USA.
Toxicol Appl Pharmacol. 2008 Apr 15;228(2):225-38. doi: 10.1016/j.taap.2007.12.023. Epub 2008 Jan 3.
Previous studies have shown that injury initiated by toxicants progresses even after most of the toxicant is eliminated from the body. One mechanism of progression of injury is the extracellular appearance of hydrolytic enzymes following leakage or upon cell lyses. Under normal conditions, after exposure to low to moderate doses of toxicants, secretory phospholipase A(2) (sPLA(2)) and other hydrolytic enzymes are known to appear in the extracellular spaces in order to cleanup the post-necrotic debris in tissues. We tested the hypothesis that sPLA(2) contributes to progression of toxicant-initiated liver injury because of hydrolysis of membrane phospholipids of hepatocytes in the perinecrotic areas in the absence of sufficient cyclooxygenase-2 (COX-2). Male Sprague-Dawley rats were administered either a moderately hepatotoxic dose (MD, 2 ml CCl(4)/kg, ip) or a highly hepatotoxic dose (HD, 3 ml CCl(4)/kg, ip) of CCl(4). After MD, liver sPLA(2) and COX-2 were co-localized in the necrotic and perinecrotic areas and their activities in plasma and liver increased before decreasing in tandem with liver injury (ALT and histopathology) leading to 100% survival. In contrast, after the HD, high extracellular and hepatic sPLA(2) activities were accompanied by minimal COX-2 activity and localization in the liver throughout the time course. This led to progression of liver injury and 70% mortality. These data suggested a destructive role of sPLA(2) in the absence of sufficient COX-2. Time- and dose-dependent destruction of hepatocytes by sPLA(2) in isolated hepatocyte incubations confirmed the destructive ability of sPLA(2) when present extracellularly, suggesting its ability to spread injury in vivo. These findings suggest that sPLA(2), secreted for cleanup of necrotic debris upon initiation of hepatic necrosis, requires the co-presence of sufficiently induced COX-2 activity to prevent the run-away destructive action of sPLA(2) in the absence of the tissue protective mechanisms afforded by COX-2 induction.
先前的研究表明，即使大部分毒物已从体内清除，由毒物引发的损伤仍会继续发展。损伤进展的一种机制是水解酶在泄漏或细胞裂解后出现在细胞外。在正常情况下，暴露于低至中等剂量的毒物后，分泌型磷脂酶A(2)（sPLA(2)）和其他水解酶会出现在细胞外间隙，以清除组织中的坏死碎片。我们检验了这样一个假设：由于在缺乏足够的环氧化酶-2（COX-2）的情况下，sPLA(2)会水解坏死周围区域肝细胞的膜磷脂，从而导致毒物引发的肝损伤进展。给雄性Sprague-Dawley大鼠腹腔注射中等肝毒性剂量（MD，2 ml CCl(4)/kg）或高肝毒性剂量（HD，3 ml CCl(4)/kg）的CCl(4)。给予MD剂量后，肝脏中的sPLA(2)和COX-2在坏死和坏死周围区域共定位，它们在血浆和肝脏中的活性在与肝损伤（ALT和组织病理学）同步下降之前升高，最终大鼠存活率为100%。相比之下，给予HD剂量后，在整个时间进程中，肝脏中细胞外和肝内的sPLA(2)活性都很高，而COX-2活性和定位却很低。这导致肝损伤进展，死亡率达70%。这些数据表明，在缺乏足够COX-2的情况下，sPLA(2)具有破坏作用。在分离的肝细胞培养中，sPLA(2)对肝细胞的时间和剂量依赖性破坏证实了其在细胞外存在时的破坏能力，表明它能够在体内传播损伤。这些发现表明，在肝坏死开始时分泌以清除坏死碎片的sPLA(2)，需要足够诱导的COX-2活性同时存在，以防止在没有COX-2诱导所提供的组织保护机制的情况下，sPLA(2)产生失控的破坏作用。
