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目的:观察外源性氧化应激对重构小鼠毛囊新生能力的影响及二苯乙烯苷(TSG,一种中药何首乌活性成分)对新生毛囊的抗氧化保护。方法:用硅胶小室法重构毛囊;用不同辐照剂量(3 J/cm~2及20 J/cm~2)长波紫外线(UVA)和不同浓度(100μmol/L及500μmol/L)过氧化氢(H_2O_2)处理新鲜制备的表皮及真皮细胞,比较观察处理细胞的毛囊新生能力改变,同时用10μμmol/L TSG联合500μmol/L H_2O_2处理细胞观察其抗氧化能力;毛囊重构部位皮肤活检采用苏木精-伊红(HE)染色进行毛囊计数,免疫组化法测定皮肤标本过氧化氢酶(CAT)表达;电子自旋共振(ESR)测试TSG羟自由基清除活性。结果:3 J/cm~2UVA照射组重建毛囊密度为(304±11.79)个,20 J/cm~2 UVA为(135±11.24)个,与对照组(202±11.79)个比较,差异有统计学意义(P<0.05);500μmol/L H_2O_2组毛囊密度为(109±20.50)个,500μmol/L H_2O_2联合10μmol/L TSG处理组为(207±15.01)个。联合处理组与对照组(216±10.26)比较,差异无统计学意义(P>0.05)。免疫组化结果显示500μmol/L H_2O_2处理组CAT染色强度18.67±2.17较对照组26.07±3.74明显减低,而500μmol/L H_2O_2联合10μmol/L TSG处理组50.02±3.82较对照组增强(P<0.05);ESR结果示TSG自身缺乏羟自由基清除活性。结论:不同水平的氧化应激对重构小鼠毛囊新生能力影响不同,低水平刺激而高水平则抑制毛囊生长。TSG对小鼠重构毛囊的抗氧化保护可能是通过上调皮肤细胞(主要是真皮细胞)CAT的表达。
OBJECTIVE: To observe the effect of exogenous oxidative stress on the regeneration of mouse hair follicles and the anti-oxidative protection of TSG (a Chinese traditional medicine, Polygonum multiflorum) on the newborn hair follicles. METHODS: Hair follicles were reconstituted by silica gel chamber method. Hair follicles were treated with UVA at different doses (3 J / cm ~ 2 and 20 J / cm ~ 2) and different concentrations of hydrogen peroxide (100μmol / L and 500μmol / L) (H 2 O 2) treatment of freshly prepared epidermal and dermal cells were observed compared to the treated cells of hair follicle neocortex changes in the same time with 10μmol / L TSG combined with 500μmol / L H_2O_2 cells were observed for its antioxidant capacity; skin follicles reconstruction site biopsy using hematoxylin The hair follicles were counted by HE stain, the catalase (CAT) expression was measured by immunohistochemical method, and the hydroxyl radical scavenging activity of TSG was tested by electron spin resonance (ESR). Results: The density of reconstructed hair follicles was (304 ± 11.79) at 3 J / cm ~ 2UVA irradiation and (135 ± 11.24) at 20 J / cm ~ 2 UVA, which was statistically different from that of the control group (202 ± 11.79) The density of hair follicles was (109 ± 20.50) in 500μmol / L H 2 O 2 group and (207 ± 15.01) in 500μmol / L H 2 O 2 group and 10μmol / L TSG group. The combined treatment group and control group (216 ± 10.26), the difference was not statistically significant (P> 0.05). The results of immunohistochemistry showed that the CAT staining intensity of 500μmol / L H 2 O 2 group was significantly lower than that of control group (18.07 ± 2.17), while that of control group was 500μmol / L H 2 O 2 (50.02 ± 3.82) (P <0.05) ESR results showed that TSG itself lacked hydroxyl radical scavenging activity. CONCLUSION: Different levels of oxidative stress have different effects on the regeneration ability of reconstituted mouse hair follicles, while low levels of stimulation and high levels inhibit hair follicle growth. The anti-oxidative protection of TSG on mouse remodeling hair follicles may be through up-regulating CAT expression of skin cells (mainly dermal cells).