新型激光微孔化猪脱细胞真皮基质的制备及应用

Preparation and application of novel laser micropore porcine acellular dermal matrix

目的介绍一种新型微孔化猪脱细胞真皮基质并验证其安全性、可行性。方法取新鲜健康小白猪断层真皮,采用高渗盐-胰蛋白酶法脱细胞,经超声波充分震荡洗涤,获得猪脱细胞真皮基质,经特定激光微孔技术,在脱细胞真皮基质上贯穿打孔即获得新型激光微孔化猪脱细胞真皮基质。将24只SD大鼠均切除背部全层皮肤至深筋膜,造成大小2.0cm×2.0cm创面,随机分为实验组和对照组,每组各12只,同步I期行自体中厚皮片与真皮材料复合移植于大鼠全层皮肤缺失创面,实验组采用移植真皮材料为激光微孔化猪脱细胞真皮基质(LPADM),对照组采用移植真皮材料为普通的猪皮脱细胞真皮基质(ADM),组织学检查、电镜观察两组真皮材料的物理性状、结构完整性、细胞残留及移植后皮片活性、血管化和移植后动物安全性变化。结果制备的真皮基质材料LPADM和ADM均呈瓷白色,有光泽,柔软而有弹性;组织学检查未见上皮细胞、内皮细胞残留,结构完整性好;电镜检查胶原纤维排列整齐,保持较好的结构完整性。实验组术后7dLPADM组织学检查显示移植真皮血管化充分,术后14d复合移植皮片均存活;对照组术后7d移植皮片色暗淡,多有起泡,并逐渐失活坏死,ADM组织学检查未见新生血管形成,术后14d复合移植皮片干性坏死。两组实验动物均无异常死亡或感染,亦未见皮肤过敏反应,体重无减轻。结论以特定激光微孔化等技术制备的新型猪脱细胞真皮基质动物移植实验中血管化充分、复合皮片愈合良好,获得满意的实验效果。

Objective To prepare a new type of micropore porcine acellular dermal matrix with the aid of laser（PLADM）, and validate the safety and practicability of the PLADM. Methods LPADM was prepared by a specific laser micropore technology with the punch worked in cycles and suspended in the midair on the harvested split-thickness dermis of the white healthy pigs followed by the procedures of high-sodium-SDS acellular treatment,digestion of trypsin ,immersion in glutaric dialdehyde solution and fully concussive washing by ultrasonic for the cross-linking treatment. Chose 24 SD rats, of which 12 cases were LPADM grafted with the split-thickness autograft （LPADM group） synchronous on the Rats full-thickness skin defects wound, the others were ADM grafted with the split-thickness autograft （ADM group）. All the dermal material adopt the accellular and cross-linking treatment used for transplantation of LPADM and ADM in two groups,the physical properties were evaluated before further experiments,The activity, immunogenicity and vascularization of the LPADM was histologically observed and tested at regular time points after surgery by histological examination and transmission electron microscopy. Results The well-prepared micropore LPADM was characterized by its porcelain white color, shiny,softness and good elasticity;Histological examination showed that the LPADM was totally devoid of epidermis and cellular components;transmission electron microscopy examination showed alignment of collagen fiber, maintain the structural integrity finely.All the SD rats survived after treatment with both transplanting methods,no infection or skin allergy ,as well as weight loss was found after surgery. Histological examination showed that the neovascularization of the grafted LPADM was adequate. Also there was little damage of the wounds and slight scars were found during the process of healing. Histological examination showed the transplantation dermal vascularization fully of LPADM experimental group in 7d postoperation, composite transplantation dermatome all live in 14d postoperation;transplantation dermatome of the control group looked dim, blister, inactivation and necrosis gradually, histological examination of ADM did not see new blood vessels forming and the composite transplantation dermatome mummification necrosis in 14d postoperation. Conclusion The new type of laser micropore porcine acellular dermis matrix（LPADM） prepared by the specific laser micropore technology shows vascularization fully and good healing of composite skin,had a good effect on the wounds of the animal transplantation experiments.