再生多孔丝素膜在创面的降解及对免疫细胞的影响

The Degradation on Trauma and Effect on Immune Cells of Regenerated Porous Silk Fibroin Membrane

目的研究再生多孔丝素膜在创伤局部应用后的降解吸收及对免疫细胞的影响。方法取SD大鼠,切除背部皮肤建立创面,面积为2cm×2cm。将125I标记的再生多孔丝素膜覆盖在创伤面,再将切除皮肤的表皮层盖在再生多孔丝素膜上进行缝合。在术后3h和3、6、13、20、27、34、41、48、55d,采用单光子发射计算机断层成像术(SPECT)进行显像,示踪至同位素信号消失。处死大鼠,取创面皮肤经HE染色后对其中的炎症细胞进行分析。脾脏细胞经双色免疫荧光及流式细胞术检测CD3+CD25+/CD3+T细胞的比率。结果大鼠种植再生多孔丝素膜55d,SPECT显像的同位素信号基本消失。创面皮肤下HE染色观察到少量的炎症细胞,未观察到残留丝素成分。实验组脾脏中CD3+CD25+/CD3+T细胞的百分率为(7.34±1.85)%,对照组为(7.18±0.37)%,两者无明显差异(P>0.05)。结论再生多孔丝素膜创面局部应用可在2个月内完全降解吸收,对免疫细胞的激发作用较小,是一种较为理想的创面修复组织工程材料。

Objective To investigate the degradation and absorption of regenerated porous silk fibroin membrane after local application on trauma and its effect on immune cells.Methods The SD rats were selected to create wound surface with an area of 2 cm × 2 cm by removing its skin of back.Then,the wound surface was covered with 125I-regenerated porous silk fibroin membrane and the epidermis was layed on the regenerated porous silk fibroin membrane.At last the wound was sutured.After operation,single photon emission computed tomography（SPECT） was used for imaging at 3 h and 3,6,13,20,27,34,41,48,55 d and it stopped when the isotope signal disappeared.At the end of the imaging,the rats were killed to obtain the skin of wound surface.Inflammatory cells was analysed through HE staining of the skin and two-color immunofluorescence and flow cytometry technology were used to detect the ratio of splenic CD3 ＋ CD25 ＋ /CD3 ＋ T cells.Results After transplanted porous silk fibroin membrane was kept on for 55 days the rats,the isotope signal disappeared.Little inflammatory cells were observed and there was no residual regenerated porous silk fibroin membrane on wound surface observed by HE staining of the skin.At the experimental group,the percentage of splenic CD3 ＋ CD25 ＋ /CD3 ＋ T cells was（7.34 ± 1.85） % while this result was（7.18 ± 0.37） % in the control group.There existed no significant difference between the two groups（P 0.05）.Conclusion Regenerated porous silk fibroin membrane can be completely degraded and absorbed within 2 months with local application on the wound and its ability of stimulating immune cells is weak.Regenerated porous silk fibroin membrane is an ideal wound repair tissue engineering materials.