阴道植入物对成纤维细胞增殖和蛋白合成的影响

Effect of transvaginal mesh on proliferation and collagen synthesis of fibroblasts

目的：探讨不同网片对盆底功能障碍性疾病患者阴道前壁成纤维细胞增殖、胶原蛋白合成的影响。方法：选取2013年3～5月在该院妇产科,行盆底重建术的患者5例为研究对象。术中切取阴道前壁组织,组织块培养法于体外行成纤维细胞原代培养。用抗波形蛋白抗体和抗角蛋白抗体进行免疫组化法鉴定成纤维细胞。将培养至第4～6代成纤维细胞分为4组,设加入Prolift网片为Prolift组、加入Avaulta网片为Avaulta组、加入TVT-O网片为TVT-O组及不加网片为正常对照组。采用四甲基偶氮唑蓝比色法（MTT法）检测4组培养2、4、7天时的细胞增殖情况,采用Western bolt法检测细胞与网片共培养4天时成纤维细胞分泌Ⅰ、Ⅲ型胶原蛋白情况。结果：1网片与细胞共培养2天时,正常对照组、Prolift组、Avaulta组和TVT-O组吸光值分别为（0.790±0.023）、（0.655±0.017）、（0.644±0.022）和（0.538±0.067）;培养4天时,分别为（0.843±0.014）、（0.716±0.045）、（0.673±0.025）和（0.283±0.008）;培养7天时,分别为（2.040±0.035）、（1.201±0.126）、（1.109±0.087）和（0.196±0.017）。正常对照组各时期细胞吸光度值分别与其他3组比较,差异均有统计学意义（P〈0.05）。Prolift组与Avaulta组各时期之间的吸光度值分别比较,差异无统计学意义（P〉0.05）。培养2天时,Prolift组和Avaulta组分别与TVT-O组比较,差异均无统计学意义（P〉0.05）;培养4天和7天时各组比较,差异均有统计学意义（P〈0.05）;2网片与细胞共培养4天时,4组Ⅰ型胶原蛋白表达量分别为（7.801±0.253）、（5.421±0.191）、（5.274±0.195）和（4.041±0.154）;Ⅲ型胶原分别为（1.062±0.104）、（0.870±0.051）、（0.865±0.047）和（0.651±0.073）。正常对照组的Ⅰ、Ⅲ型胶原蛋白分别与其他3组进行比较,差异均有统计学意义（P〈0.05）,其中Prolift组和Avaulta组分别比较,差异无统计学意义（P〉0.05）,但Prolift组、Avaulta组分别与TVT-O组进行比较,差异均有统计学意义（P〈0.05）。结论：网片孔径越大,对成纤维细胞增殖、胶原蛋白合成影响越小。

Objective： To explore the effects of different meshes on proliferation and collagen synthesis of fibroblasts in anterior va- ginal wall of patients with pelvic floor dysfunction. Methods： Five patients with pelvic floor dysfunction undergoing pelvic floor reconstruc- tion surgery in department of gynecology and obstetrics of the hospital from March to May in 2013 were selected as study objects. Anterior va- ginal wall tissue samples were obtained during operation, then tissue block culture method was used to conduct primary culture of fibroblasts in vitro, fibroblasts were identified by immunohistochemical staining with anti - vimentin antibody and anti - keratin antibody. The fibroblasts from the fourth generation to the sixth generation were divided into four groups ： Prolift group （ treated with Prolift mesh）, Avaulta group （ treated with Avanlta mesh）, TVT - O group （ treated with TVT - O mesh） and normal control group （ without mesh） . MTI＇ method was used to detect cell proliferation on the second, the fourth and the seventh day in the four groups; Western bolt was used to detect type I , Iil collagens secreted by fibroblasts on the fourth day of coculture of fibroblasts and mesh. Results： On the second day of coculture of fibroblasts and mesh, light absozption values in normal control group, Prolift group, Avaulta group and TVT - O group were （ 0. 790 ± 0. 023 ）, （0. 655 ±0. 017）, （0. 644 ±0. 022） and （0. 538 ±0. 067）, respectively; on the fourth day of eoculture of fibroblasts and mesh, light ab- sorption values in normal control group, Prolift group, Avaulta group and TVT- O group were （0. 843 ± 0. 014）, （0. 716 ± 0. 045 ）, （0. 673 ± 0. 025 ） and （ 0. 283 ± 0. 008 ）, respectively ; on the seventh day of cocuhure of fibroblasts and mesh, light absorption values in normal control group, Prolift group, Avauha group and TVT - 0 group were （ 2. 040 ± 0. 035 ）, （ 1. 201 ± 0. 126 ）, （ 1. 109 ± 0. 087 ） and （0. 196 ± 0. 017 ） , respectively. There was statistically significant difference between normal control group and the other three groups （P 〈 0. 05 ） . There was no statistically significant difference between Prolifl group and Avauha group （ P 〉 0. 05 ） . On the second day of eocul- ture of fibroblasts and mesh, there was no statistically significant difference between Prolift group, Avanha group and TVT - O group, respec- tively （ P 〉 0. 05 ） ; on the fourth day and the seventh day, there was statistically significant difference （ P 〈 0. 05 ） . On the fourth day of cocuhure of fibroblasts and mesh, the expression levels of type I collagen in the four groups were （7. 801 ± 0. 253 ） , （5. 421± 0. 191 ）, （5. 274 ±0. 195） and （4. 041 ±0. 154）, respectively; the expression levels of type m collagen in the four groups were （ 1. 062±0. 104）, （0. 870 ± 0. 051 ） , （0. 865 ± 0. 047 ） and （ 0. 651 - 0. 073 ） , respectively. There were statistically significant differences in the expression levels of type I and 11[ collagens between normal control group mad the other three groups （ P 〈 0. 05 ） . There was no statistically significant difference between Prolift group and Avaulta group （ P 〉 0. 05 ）, but there was statistically significant difference between Prolift group,Avaulta group and TVT - O group, respectively （ P 〈 0.05） . Conclusion： The greater the mesh aperture is, the smaller the impact on proliferation and collagen synthesis of fibroblasts is.