虾青素通过TGF-β1介导的Smads/ERK通路抑制肺纤维化

Astaxanthin inhibits lung fibrosis via down-regulation of TGF-β1-mediated Smads/ERK signaling pathway

目的研究虾青素(astaxanthin,AX)对博莱霉素(bleomycin,BLM)所致SD大鼠肺纤维化的干预作用及机制。方法 SD雄性大鼠32只,体质量(200±20)g,采用随机数字表法分为Sham组(假手术组)、AX组、BLM组和AX+BLM组。经气道注入5 mg/kg的BLM(BLM组和AX+BLM组)或生理盐水(Sham组)后,每天灌服虾青素(2 mg/kg)油溶液(AX组和AX+BLM组)或植物油(BLM组)连续14 d,最后一次给药24 h后取材。检测肺系数,肺组织进行HE染色及Masson染色,显微镜下观察大鼠肺部炎症及纤维化程度;碱水解法检测羟脯氨酸含量;免疫组化法观察肺组织α-SMA蛋白表达情况;酶联免疫夹心法(ELISA)检测血清中TGF-β1水平;Western blot法检测Smad2/3及ERK1/2蛋白。结果与Sham组及AX组相比,BLM组肺损伤严重,有明显的肺纤维化病灶形成;与BLM组比较,AX+BLM组肺炎症程度及纤维化程度明显减轻(P<0.01),肺组织胶原蛋白沉积减少,羟脯氨酸含量明显降低(P<0.01),α-SMA蛋白表达减少,TGF-β1水平也明显降低(P<0.05),同时Smad2/3蛋白及ERK1/2蛋白磷酸化水平明显下降。结论虾青素能明显抑制BLM诱导的肺纤维化,其作用机制可能与降低TGF-β1水平,从而下调Smads/ERK通路有关。

Objective To determine the effect of astaxanthin （AX） on bleomycin （BLM） -induced lung fibrosis in SD rats and explore the underlying mechanism. Methods Thirty-two healthy male SD rats （weighing 200 - 220 g） were randomly divided into 4 groups, that is, Sham, BLM, AX, and AX ＋ BLM groups. After intratracheal administration of BLM （5 mg/kg） or normal saline （Sham）, the rats were orally administered with AX （2 mg/kg） oil solution or vegetable oil daily for the AX group and AX ＋ BLM group or BLM group respectively for 14 consecutive days. All rats were sacrificed in 24 h after the last administration and the lung tissues were harvested. Lung pathological changes were evaluated by lung coefficient calculation, hematoxylin-eosin staining, Masson trichrome staining and hydroxyproline assay. The expression of α-SMA in the lung tissue was determined by immunohistochemical assay, and the serum level of TGF-β1 was measured by enzyme linked immunosorbent assay （ELISA）. The expression of Smad2/3 and ERK1/2 in the lung tissues were determined by Western blotting. Results Compared with the Sham and AX groups, the BLM group showed severe lung injury, with obvious formation of lung fibrosis. But the AX ＋ BLM group had notably alleviated lung inflammation and fibrosis （ P 〈 0.01 ） , reduced collagen deposition, decreased hydroxyproline content （ P 〈 0.01 ）, lower expression of ot-SMA, declined level of TGF-β1 （ P 〈 0.05 ）, and decreased expression levels of Smad2/3 and p-ERK1/2. Conclusion AX exerts obvious inhibitory induced lung fibrosis, which may be related with its decreasing the TGF-β1 level and thus Smads/ERK signaling pathway. effect on BLM- down-regulating