应用组织工程方法构建瘢痕疙瘩模型

摘要/Abstract

摘要： 目的应用组织工程方法构建人瘢痕疙瘩动物模型,并探讨采用这一模型进行瘢痕疙瘩临床和实验室研究的可行性。方法对人体瘢痕疙瘩组织中成纤维细胞进行原代培养,将体外培养第6~8代的成纤维细胞,接种到培养液预湿的聚乳酸-乙醇酸共聚物(PLGA)支架,形成体外复合体,将复合体转移至旋转式细胞培养系统容器内培养;1周后种植在20只雌性裸鼠皮下,同体两侧对照,第4、8、16、24周取材,对获得的瘢痕疙瘩组织进行组织学评价。结果术后裸鼠全部成活。复合体移植24w后,在裸鼠皮下形成的瘢痕疙瘩保持了其原有的胶原形态,在电镜下可观察到植入物内同时存在纤维细胞和成纤维细胞,成纤维细胞内可见丰富的粗面内质网,细胞特性保持不变。结论 PLGA与瘢痕疙瘩成纤维细胞具有较好的亲和性,PLGA-瘢痕疙瘩成纤维细胞复合体在裸鼠体内可形成瘢痕疙瘩,值得进一步开发研制。

Abstract: Objective To construct animal model of keloid by using tissue engineering and to explore its potential clinical significance. Method Fibroblasts from keloid were isolated and cultured.After being cultured to generation six or eight,fibroblasts were inoculated into scaffolds constructed from copolymers of polylactic acid (PLA) and polyglycolic acid (PGA),and then cultured in rotatory cell culture system for 1 week prior to subcutaneous transplantation into 20 athymic mice.After 4,8,16,24 weeks,the specimens were cut out and examined histologically. Results All the mice survived after surgery.The collagen patterns of all the keloids were remained after 24 weeks.A lot of fibroblasts and their secreted collagen were observed under light microscope.Enlarged rough endoplasmic reticula inside the fibroblasts were revealed by transmission electron microscopy.The fibroblasts in specimens remained the capability of synthesizing and secreting collagen. Conclusions There is a good affinity between PLA /PGA and fibroblasts.Complex of PLA /PGA and fibroblasts could be cultured into keloids in athymic mice.

中图分类号:

R-33

汪海滨, 罗盛康. 应用组织工程方法构建瘢痕疙瘩模型[J]. 南方医科大学学报, 2005, 25(07): 815-819,832.

WANG Hai-bin, LUO Sheng-kang. Construction of animal models of keloid by tissue engineering[J]. Journal of Southern Medical University, 2005, 25(07): 815-819,832.

参考文献

[1] 鲁峰,高建华,黎小间.病理性瘢痕和正常皮肤中成纤维细胞间缝隙连接通讯的差异[J].解放军医学杂志,2000,15(2):150-2.Lu F, Gao JH, Li XJ. Differences between the gap junctional communication function of fibrolast derived from pathological scars and normal skin[J]. Med J Chin PLA, 2000, 15(2): 150-2.
[2] Luo S,Benathan M,Raffoul W,et al. Abnormal balance between proliferation and apoptotic cell death in fibroblasts derived from keloid lesions [J]. Plast Reconstr Surg, 2001, 107( 1 ): 87-96.
[3] Zoppi RA,Ciontant S,Duek EAR, et al. Porous poly (L-lactide)films obtained by immersion precipitation process: morphology,Phase separation and culture of VERO cell [J]. Polymer, 1999, 40:3275-89.
[4] Peretti GM, Randolph MA, Villa MT, et al. Cell-based tissueengineered allogeneic implant for cartilage repair [J]. Tissue Eng,2000, 6(5): 567-76.
[5] Shea LD, Wang D, Franceshi RT, et al. Engineered bone development from a pre-osteoblast cell line on three-dimensional scaffolds[J]. Tissue Eng, 2000, 6(6): 605-17.
[6] Liu K, Yang Y,Mansbrige J. Comparison of the stress response to cryopreservation in monolayer and three-dimensional human fibroblast cultures: stress proteins, MAP kinases, and growth factor gene expression [J]. Tissue Eng, 2000, 6(5): 539-54.
[7] Sjiki T, Iwata H, Peak HJ, et al. Transmission electron microscopic study ofhepatocytes in bioartificial liver[J]. Tissue Eng, 2000, 6(6):627-640.
[8] Kim BS,Mooney DJ. Development of biocompatible synthetic extracellular matrices for tissue engineering [J]. Trends Biotechnol,1998, 16: 224-30.
[9] 郑磊,王前,裴国献,等.可降解聚合物在骨组织工程中的应用进展[J].中国修复重建外科杂志,2000,14(3):175-80.Zheng L, Wang Q, Pei GX, et al. Development of biodegradable polymer scaffolds for bone tissue engineering [J]. Chin J Rep Reconstr Surg, 2000, 14(3): 175-80.
[10] Mikos AG, Thorsen A J, Czerwonka LA,et al. Preparation and characterization ofpoly (L-lactic acid) foams[J]. Polymer, 1994, 35: 1068-77.
[11] Puelacher WC, Wisser J, Vacanti CA, et al. Temporomandibular joint disc replacement made by tissue engineered growth of cartilage[J]. J Oral Maxillofac Surg, 1994, 52(11): 1172-7.
[12] 刘彦春,王炜,曹谊林,等.卵磷脂、多聚赖氨酸和PLA包埋PGA与软骨细胞体外培养的实验研究[J].实用美容整形外科杂志,1997,8:225-7.Liu YC, Wang W, Cao YL, et al. The culture of chondrocyte seeding onto PLA conated PGA, lecithin and poly-L-lysine [J]. Prac Orthop Surg J, 1997, 8: 225-7.
[13] Shasti VP, Martin I, Langer R. Macroporous polymer foams by hydrocarbon templating [J]. Proc Natl Acad Sci USA, 2000, 97(5):1970-5.
[14] Vunjak Novakovic G, Martin I, Obradovic B, et al. Bioreactor cultivation conditions modulate the composition and mechanical properties of tissue-engineering cartilage[J]. J Orthop Res, 1999, 17(1): 130-8.
[15] Granet C, Laroche N, Vico L, et al. Rotating-wall vessels, promising bioreactors for osteoblsatic cell culture: comparison with other 3D conditions[J]. Med Biol Eng, 1998, 36(4): 513-9.
[16] Kim BS, Putnam AJ, Kulik TJ, et al. Optimizing seeding and culture methods to engineer smooth muscle tissue on biodegradable polymer matrices [J]. Biotechnol Biol Eng, 1998, 57(1): 46-54.
[17] 熊猛,艾玉峰,王潘勇,等.采用组织工程方法体外构建血管模型的初步实验研究[J].中国修复重建外科杂志,2001,15(2):109-12Xiong M, Ai YF, Wang PY, et al. Reconstruction of tissue engineered vascular model in vitro [J]. Chin J Reparat Reconstr Surg,2001, 15(2): 109-12.