TGF-β_1对膀胱逼尿肌Ⅲ型胶原表达的调节作用

The effect of transforming growth factor β_1 on the expression of collagen type Ⅲ in the bladder detrusor

目的 探讨转化生长因子β(TGF -β1 )与膀胱逼尿肌胶原表达的关系,以及TGF -β1 单克隆抗体对膀胱纤维化的抑制作用。方法 在严格无菌操作下取大鼠膀胱逼尿肌,剪碎、离心后加入适量含有浓度为 15%小牛血清、2.0g/LNaHCO3、100U/ml青霉素、0.1mg/ml链霉素的培养液。细胞在 37℃、含体积百分数为 5%的CO2 温箱中培养。培养细胞达到 80%融合时,加入胰蛋白酶,在相同的培养液中培养。分别加入浓度为0ng/ml、5ng/ml、10ng/ml的TGF -β1 和TGF -β1 单克隆抗体共同孵育24h,收集细胞进行Ⅲ型胶原原位杂交。结果 TGF- β1 浓度越高,视野中阳性染色细胞数目越多,光密度值越大,TGF- β1 浓度为 0ng/ml、5ng/ml、10ng/ml时,光密度值分别为 0.425±0.062、0.536±0.096、0.642±0.087;TGF- β1 单克隆抗体浓度越高,视野中阳性染色细胞数目越少,光密度值越小,TGF- β1 单克隆抗体浓度为 0ng/ml、5ng/ml、10ng/ml时,光密度值分别为 0.425±0.062、0.317±0.072、0.221±0.074。结论 TGF- β1 可促进膀胱逼尿肌中胶原的沉积,TGF- β1 单克隆抗体可抑制膀胱逼尿肌中胶原的形成。

Objective To explore the relationship between transforming growth factorβ 1(TGF-β 1) and the expression of detrusor collagen.Methods Detrusor smooth muscle of rats was cut into small pieces under sterile conditions. Suspensions were cultured in medium with 15% fetal calf serum, 2.0g/L NaHCO 3, 100U/ml penicillin and 0.1mg/ml streptomycin. Cells were grown at 37℃ in an incubator with a humidified atmosphere of 5% CO 2 in air. When cultures reached a confluence of about 80%, cells were suspended by incubating them with trypsin. They were maintained and expanded with the same culture medium. TGF-β 1 of 0ng/ml, 5ng/ml, 10ng/ml and TGF-β 1 antibody of 0ng/ml, 5ng/ml, 10ng/ml were added to medium for 24 hours, respectively. Cells were collected for in-situ hybridization to demonstrate the expression of type Ⅲ collagen.Results The positive staining for type Ⅲ collagen was significantly increased with the concentration of TGF-β 1 increase. When TGF-β 1 concentration was increased from 0ng/ml to 10ng/ml, the light density value was increased from 0.425 to 0.642. However, the positive staining for type Ⅲ collagen was decreased with TGF-β 1 antibody concentration increase. The light density values were decreased from 0.425 to 0.221 when concentration of TGF-β 1 antibody was increased from 0ng/ml to 10ng/ml.Conclusions TGF-β 1 has an effect of accelerating deposition of collagen in bladder detrusor.