课题背景：课题为贵州省省长基金（黔省专合字[2007]70号）和贵州省科技厅基金（黔科合J字[2007]2106号）重点资助项目。目前阴茎白膜修复材料会导致瘢痕收缩、硬件形成、弯曲复发等不足。膀胱黏膜无细胞基质是膀胱黏膜下层脱细胞后获得的无细胞而富含胶元的生物材料，已用于修复动物膀胱、尿道的研究。课题拟自制兔膀胱无细胞基质，构建新西兰兔阴茎白膜缺损膀胱无细胞基质移植物覆盖模型，以寻找理想的白膜修复材料。

应用要点：实验应用膀胱细胞外基质重建新西兰兔阴茎白膜，结果表明膀胱无细胞基质移植物修复阴茎白膜后，有较好的组织相容性，未出现明显的炎症反应和纤维化，有较好的研究前景。

偏倚或不足：实验样本量偏小，实验对象为小型动物，今后在增大样本量，逐渐完善其代谢、毒理作用以及临床前动物实验的前提下，可考虑进一步申请伦理学等临床相关审核批准，进行人体白膜重建，以期真正达到临床应用标准。

摘要

背景：阴茎弯曲的矫正需用材料替代缺损的白膜或增大病变处白膜，目前的白膜替代材料没有达到理想的效果，国外学者开始考虑用脱细胞的细胞外基质重建阴茎白膜。
目的：应用膀胱无细胞基质移植物重建新西兰兔阴茎白膜，动态观察其重建效果。
设计、时间及地点：随机对照动物实验，于2005-12/2007-06在四川大学华西实验动物中心、四川大学组织工程实验室及贵阳医学院组织工程实验室完成。
材料：选用健康封闭新西兰兔50只，均为雄性，体质量2.6~3.0 kg，10只新西兰兔用于膀胱无细胞移植物的制备。
方法：将剩余40只兔按随机数字表法分为对照组和膀胱无细胞基质移植物组（n=20），在阴茎背侧切除白膜造成10 mm×5 mm缺损，分别采用白膜原位缝合及膀胱黏膜无细胞基质修复。
主要观察指标：分别于术后2，6，12和24周，每个时间点取5只动物进行阴茎海绵体内快速注射生理盐水诱导阴茎勃起，观察阴茎弯曲情况；然后处死动物术区取材，进行苏木精-伊红、Masson和Stirus染色，检测炎性标志因子诱导型一氧化氮合成酶和促纤维化因子转化生长因子β1的表达；应用图像采集系统测量胶原纤维的面积。
结果：①术后6周时弯曲发生率最高，以后逐渐减少，到24周只有对照组发生弯曲1例。②术后2周时，Ⅰ型和Ⅲ型胶原纤维共同存在，比例相当；以后Ⅰ型胶原纤维逐渐增加(P < 0.05)，Ⅲ型胶原纤维逐渐减少(P < 0.05)；到24周以Ⅰ型胶原纤维为主，Ⅲ型胶原纤维已不明显。③诱导型一氧化氮合成酶和转化生长因子在术后2周时表达最强，6周时表达明显下降。
结论：膀胱黏膜无细胞基质移植物修复新西兰兔阴茎白膜显示了良好的组织相容性，未出现明显的炎症反应和纤维化，是较为理想的阴茎白膜修复材料。
关键词：移植物；阴茎；膀胱；生物相容性材料；疾病模型，动物

孙发，杨宇如，魏强，卢一平，李虹，韩平，宋超，石家齐，谷江．膀胱无细胞基质移植物重建兔阴茎白膜[J].中国组织工程研究与临床康复，2008;12(18):3441-3445 [www.zglckf.com/zglckf/ejournal/upfiles/08-18/18k-3441(ps).pdf]

Bladder acellular matrix grafts for reconstructing tunica albuginea in rabbits

Abstract

BACKGROUND：Penile curvature can be corrected with the materials to substitute the tunica albuginea defect or enlarge the affected tunica albuginea. At present, no ideal substitute material has been found, so the scholars overseas consider reconstructing tunica albuginea with the acellular extracellular matrix.
OBJECTIVE: To conduct a dynamic observation of the reconstruction of tunica albuginea in New Zealand rabbits with bladder acellular matrix grafts.
DESIGN, TIME AND SETTING: From December 2005 to June 2007, a randomized controlled animal experiment was carried out in the West China Experimental Animal Center and the Tissue Engineering Laboratory of Sichuan University, as well as the Tissue Engineering Laboratory of Guiyang Medical College.
MATERIALS: Fifty New Zealand male rabbits were adopted, weighing 2.6-3.0 kg, and 10 of them were used for the preparation of bladder acellular grafts.
METHODS: The other 40 rabbits were at random divided into control group and bladder acellular matrix graft group, with 20 animals in each group. A defect area of 10 mm × 5 mm was induced in those rabbits, and repaired with orthotopic suture of tunica albuginea or bladder acellular matrix grafts, respectively.
MAIN OUTCOME MEASUERS: At postoperative weeks 2, 6, 12 and 24, an penile erection was induced by a rapid intracavernosal injection with saline into 5 rabbits at each time point, to observe penile curvature; Then rabbits were sacrificed, followed by hematoxylin-eosin, Masson and sirius red stain. Protein expression of the inflammatory markers inducible nitric oxide synthase (iNOS) and transforming growth factor-β1 (TGF-β1) was evaluated; Image collection system was used to measure the area of collagen fiber.
RESULTS: The incidence of penile deviation was the highest at 6 weeks and then reduced gradually, and only 1 case of slight penile deviation was observed in control group at 24 weeks. Collagen Ⅰ and Ⅲ occupied equally the grafting site 2 weeks postoperatively; After that, the collage Ⅰ increased evidently (P < 0.05) and collagen Ⅲ decreased gradually (P < 0.05). Collagen Ⅰ occupied mostly the grafting site at 24 weeks, while collagen Ⅲ was not observed. Expression of iNOS and TGF-β1 was the highest at 2 weeks and precipitously decreased at 6 weeks.
CONCLUSION: The bladder acellular matrix grafts demonstrate good histocompatibility in the repair of tunica albuginea in New Zealand rabbits, does not stimulate significant inflammatory response or cause fibrosis, so it is an ideal material for the tunica albuginea repair.

Sun F, Yang YR, Wei Q, Lu YP, Li H, Han P, Song C, Shi JQ, Gu J. Bladder acellular matrix grafts for reconstructing tunica albuginea in rabbits.Zhongguo Zuzhi Gongcheng Yanjiu yu Linchuang Kangfu 2008;12(18):3441-3445(China)
[www.zglckf.com/zglckf/ejournal/upfiles/08-18/18k-3441(ps).pdf]