蛋清源活性肽对过氧化氢诱导的HEK293细胞谷胱甘肽抗氧化系统的影响

The Effect of Bioactive Peptides from Egg White on Glutathione Anti-oxidant System against Oxidative Stress Injury in Human Embryonic Kidney 293 Cells

研究了蛋清源活性肽WNWAD、WNW、WAD、WN、AD的体外氧自由基吸收能力(ORAC活性),以及其对过氧化氢诱导的HEK293细胞还原型谷胱甘肽(GSH)含量、氧化型谷胱甘肽(GSSG)含量、GSH/GSSG比值以及γ-谷氨酰半胱氨酸合成酶(γ-GCS)表达的影响。结果表明,浓度为5.0μmol/L的蛋清源活性肽WNWAD、WNW、WAD、WN的ORAC活性均强于Trolox,AD没有ORAC活性。过氧化氢诱导的HEK293细胞氧化损伤模型组细胞中GSH含量由(173.52±2.87)μmol/L降至(115.85±2.03)μmol/L(P<0.01),GSSG含量由(0.90±0.08)μmol/L升至(33.23±0.68)μmol/L(P<0.01);细胞中加入不同浓度的蛋清源活性肽,GSH含量与损伤组相比,显著升高(P<0.05),GSSG含量与损伤组相比明显降低(P<0.01),在0.5μmol/L WNWAD作用下GSSG含量由(33.23±0.68)μmol/L降至(0.96±0.02)μmol/L (P<0.01),接近对照组细胞中GSSG含量;GSH/GSSG比值由损伤组的3.49±0.68提至174.22±0.13(P<0.01)。蛋清源活性肽处理过的细胞中γ-GCS mRNA表达量相对于损伤组有不同程度增加,其中5.0μmol/L WN处理过的细胞中γ-GCS m RNA表达量最大,为8.53±0.06。以上研究结果表明,蛋清源活性肽能有效清除活性氧,减少GSH消耗,从而使细胞内γ-GCS含量减少。蛋清源活性肽能促进γ-GCS mRNA的表达,使GSH的合成增加,起到氧化应激抑制作用。

The aim of this study was to investigate the anti-oxidative effects of WNWAD, WNW, WAD, WN and AD against the oxidative stress induced by H2 O2 in HEK-293 cells, including ORAC, GSH and GSSG levels and the effects on gene expressions and activity of γ-GCS. Oxygen radical absorbance capacity(ORAC) results showed that 5.0μmol/L WNWAD, WAD, WNW and WN had different oxygen radical absorption capacities in vitro but AD. Then, on the base of oxidative stress model of HEK-293 cells, GSH and GSSG assay results indicated that HEK-293 cells induced by H2 O2 decreased GSH level from(173.52 ± 2.87) μmol/L to(115.85 ± 2.03) μmol/L(P<0.01), and increased GSSG level from(0.90 ± 0.08) μmol/L to(33.23 ± 0.68) μmol/L(P<0.01). Egg white peptides(except AD) can obviously increase GSH level(P<0.05) and decrease GSSG lever(P<0.01), especially 0.5 μmol/L WNWAD decreased GSSG lever in HEK-293 cells from(33.23 ± 0.68) μmol/L to(0.96 ± 0.02) μmol/L(P<0.01) and increased GSH/GSSG ratio from 3.49 ± 0.68 to 174.22 ± 0.13(P<0.01). In addition, peptides on the expression of γ-GCS mRNA result showed that the peptides improved the expression of γ-GCS. Among them, the highest expression of γ-GCS mRNA in the 5.0 μmol/L WN-treated cells was 8.53 ± 0.06. All the above results suggested that egg white peptides protected HEK-293 cells against H2 O2-induced oxidative stress by absorbing ROS to decrease the consumption of GSH and inhibiting GSH/GSSG ratio changed dramatically. Egg white peptides also can upregulate gene expressions of γ-GCS, which could make the synthesis of GSH increase to against H2 O2-induced oxidative stress.