枸杞多糖粗提物对紫外线诱导HaCaT细胞清除活性氧的影响

Scavenging effect of crude polysaccharides extracted from Lycium barbarum on reactive oxygen species in ultraviolet radiation-induced HaCaT cells

目的 探讨枸杞多糖粗提物对紫外线诱导的HaCaT细胞清除活性氧（ROS）的影响及可能机制。方法 将HaCaT细胞分为空白组、枸杞多糖组、UVA组、UVB组、UVA ＋ 枸杞多糖组、UVB ＋ 枸杞多糖组。用噻唑蓝法检测细胞增殖活性；分光光度计检测枸杞多糖粗提物对UVA和UVB吸收情况；用DCFH-DA荧光探针检测细胞内ROS水平；酶生化法测定胞质超氧化物歧化酶（SOD）、谷胱甘肽过氧化物酶（GSH-Px）活性及乳酸脱氢酶（LDH）漏出量。结果 0、100、200、300、400、500、600、1 500、2 000 mg/L枸杞多糖粗提物对HaCaT细胞增殖活性无明显影响。枸杞多糖粗提物对280 ~ 400 nm紫外线透光率较大；与空白组比较，UVA和UVB组 LDH漏出量、ROS水平显著升高，胞内SOD及GSH-Px活力降低，差异均有统计学意义（P 〈 0.001或0.05）。照射前加枸杞多糖粗提物（UVA ＋ 枸杞多糖组、UVB ＋ 枸杞多糖组）可明显升高细胞内SOD、GSH-Px活性，减少LDH释放，降低ROS水平，与UVA或UVB组比较，差异均有统计学意义（P 〈 0.05）。结论 枸杞多糖粗提物不具有遮光剂的作用，但可有效清除 ROS，降低 LDH漏出率，抑制紫外线致 HaCaT细胞光损伤，可能与其增强抗氧化酶活性有关。

Objective To evaluate the scavenging effect of crude polysaccharides extracted from Lycium barbarum （LBP） on reactive oxygen species in ultraviolet radiation-induced HaCaT cells, and to explore its possible mechanism. Methods Cultured immortalized human keratinocyte HaCaT cells were divided into 6 groups： blank control group receiving no treatment, LBP group treated with crude LBP alone, ultraviolet A （UVA） group treated with UVA radiation alone, ultraviolet B （UVB） group treated with UVB radiation alone, UVA ＋ LBP group treated with crude LBP for 24 hours followed by UVA radiation, and UVB ＋ LBP group treated with crude LBP for 24 hours followed by UVB radiation. MTT colorimetry was performed to evaluate the cellular proliferative activity, UV spectrophotometric method to measure the UVA and UVB absorption of crude LBP, dichlorodihydrofluorescein diacetate （DCFH-DA） fluorescent probe assay to detect the level of ROS, enzymatic-biochemical method to estimate the activities of superoxide dismutase （SOD） and glutathione peroxidase （GSH-Px）, as well as to detect the leakage of lactate dehydrogenase （LDH）. Results Crude LBP at different concentrations of 0, 100, 200, 300, 400, 500, 600, 1 500, 2 000 mg/L had no obvious effects on the proliferative activity of HaCaT cells. Crude LBP had a high transmittance of ultraviolet rays at 280 - 400 nm. Compared with the blank control group, the UVA group and UVB group both showed significantly higher LDH leakage and ROS level, lower activities of SOD and GSH-Px （P 〈 0.001 or 0.05）. Pretreatment with crude LBP before the ultraviolet radiation could significantly increase the activities of SOD and GSH-Px, decrease the LDH leakage and ROS level in the UVA ＋ LBP group and UVB ＋ LBP group compared with the UVA group or UVB group （P 〈 0.05）. Conclusion Crude LBP have no effect of sunscreening agents, but can effectively scavenge ROS, decrease LDH leakage, inhibit ultraviolet radiation-induced photodamage in HaCaT cells, which may be associated with the enhancement of antioxidant enzyme activity.