常规制备家兔血清对人脐静脉内皮细胞-304的影响

目的：探讨中药血清药理学研究中常规制备的正常家兔血清浓度变化对培养体系中细胞的影响。方法：常规制备无污染、无溶血正常家兔血清，通过细胞形态学、台盼蓝染色活细胞计数、细胞毒四唑盐（MTT）比色试验、乳酸脱氢酶（LDH）及血管紧张素转换酶（ACE）等指标，观察5％、10％、20％、40％、60％和80％家兔血清对人脐静脉内皮细胞（VEC）－304的影响。结果：5％和10％两浓度组各指标均无变化，对VEC－304没有损伤作用。40％、60％和80％3个浓度组家兔血清浓度对细胞均有不同程度的损伤，而且随着浓度的增加，呈现出损伤加重的趋势，而20％的浓度从细胞形态、台盼蓝染色活细胞计数、MTT及ACE方面没有表现出明显的损伤作用，只有LDH含量差异有显著性（P＜0．01）。结论：常规制备的家兔血清在无污染和无溶血的情况下，其对细胞的作用浓度是有一定限制的，当达到40％或更高浓度时有较为明显的损伤。所以，在中药血清药理学研究中，应进行常规制备正常血清及含药血清的细胞毒实验，以尽量减少血清本身带给实验的影响。

Effect of Conventionally Prepared Rabbit Serum on Human Umbilical Vein Endothelial Cell-304

OBJECTIVE: To study the effect of conventionally prepared rabbit serum, usually used in the field of serum pharmacology of TCM, in various concentration on cultured system of human umbilical vein endothelial cell (VEC)-304 cell line. METHODS: Prepared conventionally from healthy rabbit, the effect of uncontaminated and nonhemolytic serum of six concentrations (5%, 10%, 20%, 40%, 60% and 80%) on VEC-304 cells was observed using parameters such as cell morphologic examination, trypan blue exclusion test, methyl thiazolyl tetrazolium (MTT) test, lactate dehydrogenase (LDH) and angiotensin-converting release test. RESULTS: Serum of 5% and 10% concentration showed no effect in all the above-mentioned tests. Serum of 20% concentration also showed no effect in the tests except for LDH releasing (P < 0.01). But when the concentration of serum reached up to 40% or above, distinct cytotoxicity and serious injury appeared in a dose-dependent manner. CONCLUSION: High concentration of conventionally prepared rabbit serum could injure the incubated cell. Therefore, in order to lower the background effect of serum as far as possible in TCM serum pharmacological study, the cytotoxicity of rabbit serum should be tested first of all.