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CO 2点阵激光Fusion融合模式对小鼠皮肤增生性瘢痕的影响及其机制

Effect of CO 2 fractional laser Fusion fusion mode on hyperplastic scarring in mouse skin and its mechanism

摘要目的:探讨CO 2点阵激光Fusion融合模式对小鼠皮肤增生性瘢痕(HS)的影响及其机制。 方法:将50只雄性C57BL/6J小鼠采用随机数字表法分成空白对照组(空白涂膜剂基质0.1 ml)、模型组(空白涂膜剂基质0.1 ml)、10 mJ激光治疗组(10 mJ组)、20 mJ激光治疗组(20 mJ组)和阳性对照复方醋酸地塞米松乳膏(DXM,0.1 ml)组,每组10只。空白对照组皮下注射生理盐水,其余各组小鼠均于背部皮肤注射博来霉素(BLM)(1 mg/ml)制作HS模型,3周后采用温哥华瘢痕量表(VSS)评估皮肤瘢痕增生情况。治疗4周后处死小鼠,取瘢痕全层皮肤组织,苏木精-伊红(HE)染色、马松(Masson)染色法观察病理形态,免疫荧光染色检测血小板内皮细胞黏附分子(CD31)、转化生长因子β1(TGF-β1)表达,蛋白质印迹法(Western blot)检测α-平滑肌肌动蛋白(α-SMA)、纤连蛋白(FN)、Ⅰ型胶原蛋白(Col Ⅰ)表达。组间数据比较采用单因素方差分析,最小显著性差异法(LSD)或Tamhane’s T2法进行两两比较。结果:模型组、10 mJ组、20 mJ组和DXM组VSS总评分高于空白对照组[(7.900±0.800)、(8.200±0.600)、(8.000±0.700)、(7.900±0.700)分比(0.000±0.000)分, t=31.227、43.218、36.140、35.689, P均<0.05],差异均有统计学意义。治疗4周后,空白对照组小鼠背部皮肤组织中表皮细胞排列整齐,层次清晰;模型组有较厚的表皮增生;10 mJ组有少量毛囊生出;20 mJ组毛囊排列较整齐;DXM组基本无表皮增生。空白对照组、10 mJ组、20 mJ组、DXM组的CD31、TGF-β1染色强度以及α-SMA、FN、Col Ⅰ蛋白相对表达量均明显低于模型组,且20 mJ组低于10 mJ组[α-SMA(3.840±0.410)、(2.560±0.260)、(2.180±0.250)、(1.330±0.150)比(0.980±0.120), t=21.171、17.448、13.684、5.762;FN(3.880±0.420)、(2.920±0.300)、(2.460±0.260)、(1.180±0.140)比(1.020±0.140), t=20.429、18.149、15.421、2.556;Col Ⅰ(3.360±0.340)、(2.480±0.260)、(2.020±0.220)、(1.360±0.180)比(1.000±0.160), t=19.861、15.330、11.857、4.727, P均<0.05],差异均有统计学意义。 结论:CO 2点阵激光Fusion融合模式可减轻小鼠皮肤瘢痕增生,其机制可能与抑制瘢痕组织中CD31、TGF-β1和α-SMA、FN、Col Ⅰ蛋白表达有关。

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abstractsObjective:To investigate the effect of CO 2 fractional laser Fusion fusion mode on skin proliferative scar (HS) in mice and its mechanism. Methods:Totally, 50 male C57BL/6J mice were divided into blank control group (blank coating agent substrate 0.1 ml), model group (blank coating agent substrate 0.1 ml), 10 mJ laser treatment group (10 mJ group), 20 mJ laser treatment group (20 mJ group) and positive control compound dexamethasone acetate cream (DXM, 0.1 ml) using the random number table method, 10 mice in each group. The blank control group was injected subcutaneously with the same volume of saline, and the rest of the mice in each group were injected with BLM (1 mg/ml) in the dorsal skin to make the HS model, and the skin scar proliferation was assessed by the VSS after 3 weeks. After 4 weeks of treatment, the mice were executed, and the whole skin tissue of the scar was taken, and the pathological morphology was observed by hematoxylin and eosin (HE) staining and Masson staining, and the expression of CD31 and TGF-β1 was detected by immunofluorescence staining, and the expression of α-smooth muscle actin (α-SMA), fibronectin (FN) and collagen Ⅰ (Col Ⅰ) was detected by Western blotting. Data were compared between groups using one-way ANOVA, LSD method or Tamhane’s T2 method for two-way comparison.Results:The model group, 10 mJ group, 20 mJ group, and DXM group had higher skin scar scores in total VSS scores than the blank control group [(7.900±0.800), (8.200±0.600), (8.000±0.700), (7.900±0.700) scores vs. (0.000±0.000) scores, t=31.227, 43.218, 36.140, 35.689, P<0.05], all differences were statistically significant. After 4 weeks of treatment, the epidermal cells in the back skin tissue of mice in the blank control group were neatly arranged and clearly layered; the model group had thicker epidermal hyperplasia; the 10 mJ group had a small amount of hair follicles; the 20 mJ group had more neatly arranged hair follicles; the DXM group basically had no epidermal hyperplasia. The intensity of CD31 and TGF-β1 staining in the blank control group, 10 mJ group, 20 mJ group and DXM group were all significantly lower than that in the model group, and the intensity of CD31 and TGF-β1 staining in the 20 mJ group was lower than that in the 10 mJ group. The staining intensity of CD31 and TGF-β1 and the relative expression of α-SMA, FN and Col Ⅰ proteins in the blank control group, 10 mJ group, 20 mJ group and DXM group were significantly lower than those in the model group, and those in the 20 mJ group were lower than the 10 mJ group [α-SMA: (3.840±0.410), (2.560±0.260), (2.180±0.250), (1.330±0.150) vs. (0.980±0.120), t=21.171, 17.448, 13.684, 5.762; FN: (3.880±0.420), (2.920±0.300), (2.460±0.260), (1.180±0.140) vs. (1.020±0.140), t=20.429, 18.149, 15.421, 2.556; Col Ⅰ: (3.360±0.340), (2.480±0.260), (2.020±0.220), (1.360±0.180) vs. (1.000±0.160), t=19.861, 15.330, 11.857, 4.727, all P< 0.05]. Conclusion:CO 2 fractional laser Fusion fusion mode can reduce skin scar proliferation in mice which may be related to the inhibition of CD31, TGF-β1 and α-SMA, FN, Col Ⅰ protein expression in scar tissue.

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