Depyrogenation of digestive enzymes reduces lipopolysaccharide tolerance in isolated cardiac myocytes.
Depyrogenation of digestive enzymes reduces lipopolysaccharide tolerance in isolated cardiac myocytes.
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消化酶的去热原降低了离体心肌细胞的脂多糖耐受性。
DOI：
10.1006/jmcc.1997.0432
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发表时间：
1997
期刊：
影响因子：
--
通讯作者：
Bayna,E
中科院分区：
文献类型：
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作者：
Lew,WY;Lee,M;Yasuda,S;Bayna,E
分离的肌细胞可用于检查物质如脂多糖（LPS）和细胞因子的直接心脏效应。然而，用于标准细胞分离程序的消化酶被几百ng/ml LPS污染。我们用一系列Triton X-114和多粘菌素B洗涤液去除消化酶的热原，以去除99.7-99.9%的LPS。这将LPS污染水平从100-300 ng/ml降低到0.15-0.70 ng/ml，同时保持从新西兰白色兔的左心室分离的良好质量的细胞。我们评估了是否短暂暴露于LPS污染物水平，如标准细胞分离过程中发生的，诱导LPS耐受。将心肌细胞（用去热原酶分离）预暴露于100 ng/ml LPS 1小时，洗涤，然后暴露于100 ng/ml LPS的激发剂量。LPS的挑战剂量诱导的时间依赖性减少细胞缩短超过6小时，在没有预暴露的心肌细胞，但不是在预暴露于较早剂量的LPS的心肌细胞。我们研究了与去热原酶相比，用未经处理的酶分离的肌细胞是否产生LPS耐受性。在用未经处理的酶分离的心肌细胞中，暴露于1000-10 000 ng/ml LPS 6 h后，细胞缩短显著减少。在用去热原酶分离的心肌细胞中，仅需要5-50 ng/ml LPS就可诱导相当的心脏抑制。我们的结论是，短暂暴露于LPS污染物水平，这发生在标准的细胞分离程序，诱导低反应性或耐受随后剂量的LPS在分离的心肌细胞。
The isolated myocyte is useful for examining the direct cardiac effects of substances such as lipopolysaccharide (LPS) and cytokines. However, the digestive enzymes used for standard cell isolation procedures are contaminated by several hundred ng/ml LPS. We depyrogenated the digestive enzymes with a series of Triton X-114 and Polymyxin B washes to remove 99.7–99.9% of the LPS. This lowered LPS contamination levels from 100–300 ng/ml to 0.15–0.70 ng/ml, while maintaining good quality cell isolations from the left ventricle of New Zealand white rabbits. We evaluated whether brief exposure to LPS contaminant levels, as occur during standard cell isolations, induces LPS tolerance. Cardiac myocytes (isolated with depyrogenated enzymes) were pre-exposed to 100 ng/ml LPS for 1 h, washed, then exposed to a challenge dose with 100 ng/ml LPS. The LPS challenge dose induced a time-dependent decrease in cell shortening over 6 h in myocytes without pre-exposure, but not in myocytes pre-exposed to an earlier dose of LPS. We examined whether LPS tolerance develops in myocytes isolated with untreated enzymes, compared with depyrogenated enzymes. In myocytes isolated with untreated enzymes, there was a significant decrease in cell shortening after 6 h exposure to 1000–10 000 ng/ml LPS. In myocytes isolated with depyrogenated enzymes, it required only 5–50 ng/ml LPS to induce a comparable cardiac depression. We conclude that brief exposure to LPS contaminant levels, which occur with standard cell isolation procedures, induces a hyporesponsiveness or tolerance to subsequent doses of LPS in isolated cardiac myocytes.