葡萄糖浓度波动对人肝L02细胞损伤的实验研究

Experimental study on the damage of glucose fluctuation on cultured human L02 hepatocytes

研究了葡萄糖浓度波动对于体外培养的人肝实质L02细胞的影响以及可能的机制.利用人肝实质细胞株L02进行传代培养.实验共分为4组:正常组(N)、持续高糖组(HG)、波动组(GF)和渗透压对照组(OP).各组细胞培养72 h后,测定培养液中谷丙转氨酶(ALT)、谷草转氨酶(AST)和乳酸脱氢酶(LDH)活力,肝细胞内糖原含量,谷胱甘肽(GSH)、丙二醛(MDA)、超氧化酶歧化酶(SOD)、Na^+K^+-ATP酶和Ca^(2+)Mg^(2+)-ATP酶的活力,胞内游离钙离子([Ca^(2+)]_i)浓度以及细胞膜的流动性.高糖组和波动组细胞培养液中ALT,AST和LDH活力上升,细胞内GSH,SOD,Na^+K^+ATP酶和Ca^(2+)Mg^(2+)-ATP酶活性均下降,MDA和糖原含量上升,细胞膜流动性下降;高糖组和波动组与正常组比较均差异显著(p<0.001),波动组较高糖组也有显著差异(p<0.01).葡萄糖浓度波动能够导致细胞膜通透性的改变和细胞内酶的严重泄漏,同时对肝细胞有明显的氧化损伤和毒性作用,而且比单纯的高糖对肝细胞的损害更为明显和严重.

The goal of this study is to investigate the effects of glucose fluctuation on cultured human L02 hepatocytes in vitro and discuss its possible mechanism. Cultured cells were randomly divided into 4 groups： normal （N）, high glucose （HG）, glucose fluctuation （GF） and osmotic pressure （OP）. After 72 hours, alanine transferase （ALT）, aspartate amino transferase （AST） and lactate dehydrogenase （LDH） in the medium were measured. In the cells, glycogen, gluta- thione （GSH）, superoxide dismutase （SOD）, ma/ondialdehyde （MDA）, Na＋ K＋ -ATP and Ca2＋Mg2＋ -ATP enzyme activities were detected. In addition, cell membrane fluidity was also detected. Compared with the N group, the activities of ALT, AST and LDH in the HG and GF groups increased markedly, while the activities of GSH, SOD, Na＋ K＋ ATP and Ca2＋ Mg2＋-ATP enzymes were decreased significantly. The contents of glycogen and MDA in the cells of HG and GF groups were increased obviously, while cell membrane fluidity was decreased markedly （all p〈0. 01）. There are also significantly statistical differences between the HG and GF groups（all p〈0.01 ）. Our results demonstrate that glucose fluctuation can change the permeability of cell membrane, lead to the leakage of enzymes and cause marked oxidative damage and cell toxicity in human L02 hepatocytes. Moreover, the deleterious effects of GF on the L02 cells are significantly higher than that of the HG group.