骨髓间充质干细胞联合脱细胞真皮基质修复尿道损伤
|
摘要
背景:近年来,组织工程学的发展为尿道损伤修复提供了更多的选择。目的:探讨骨髓间充质干细胞联合脱细胞真皮基质修复尿道损伤的效果。方法:取第3代新西兰大白兔骨髓间充质干细胞,接种于脱细胞真皮基质材料表面,构建组织工程尿道。将36只新西兰大白兔随机分3组,每组12只。实验组于尿道损伤处植入骨髓间充质干细胞-脱细胞真皮基质复合物,对照组于尿道损伤处植入单纯脱细胞真皮基质材料,正常组既无损伤也不做任何处理。术后4,8,12周,尿道修复组织进行苏木精-伊红染色,术后12周,进行免疫组织化学染色和尿动力学检查。结果与结论:(1)术后4周,实验组尿道缺损区可见有少许薄层上皮再生,连续性较好;对照组修复段尿道新生黏膜尚不连续。术后8-12周,实验组修复段尿道上皮层逐渐变厚,与正常尿道上皮连续性良好,黏膜逐渐增厚,尿道黏膜光滑连续;对照组修复段尿道新生黏膜连续性较好,再生上皮层次较少;(2)术后12周,免疫组化可见实验组修复尿道标本uroplakinⅢa,CK AE1/AE3,α-SMA表达阳性;(3)实验组手术前后的最大尿道压比较差异无显著性意义;对照组术后最大尿道压高于术前,差异有显著性意义(P<0.05);(4)结果表明,骨髓间充质干细胞联合脱细胞真皮基质修复尿道损伤的效果优于单纯脱细胞真皮基质材料。
BACKGROUND: In recent years, the development of tissue engineering provides more choices for the repair of urethral injury.OBJECTIVE: To investigate the effect of bone marrow mesenchymal stem cells(BMSCs) combined with acellular dermal matrix in the repair of urethral injuries. METHODS: Passage 3 BMSCs from New Zealand white rabbits were inoculated on the acellular dermal matrix to construct tissue-engineered urethra grafts. Thirty-six New Zealand rabbits were randomized into three groups(n=12 per group). Experimental group was implanted withBMSCs-acellular dermal matrix complex at urethral injury. Control group was implanted with acellular dermal matrix material at urethral injury. Normal group had neither injury nor treatment. At postoperative 4, 8 and 12 weeks, the repaired urethral tissue was subjected to hematoxylin-eosin staining. At postoperative 12 weeks, immunohistochemical staining and urodynamic study were performed. RESULTS AND CONCLUSION: At postoperative 4 weeks, thin-layer epithelial regeneration was visible in the urethra defect area of the experimental group, and the continuity was better. The urethra mucosa of the control group was discontinuous. At postoperative 8-12 weeks, the urethral epithelial layer in the experimental group became thickened, exhibiting a good continuity with the normal urethral epithelium, thickened mucosa, and smooth and continuous urethral mucosa; the regenerated urethral mucosa of the control group exhibited good continuity, but less regenerated epithelial layers. At postoperative 12 weeks, immunohistochemical results showed the repaired urethra in the experimental group was positive for uroplakin Ⅲa, CK AE1/AE3, and α-smooth muscle actin. The maximum urethral pressure in the experimental group showed no significant difference before and after operation, while the postoperative pressure in the control group showed a significant increase(P < 0.05). Overall findings indicate that the combination of BMSCs and acellular dermal matrix has better efficacy than the acellular dermal matrix alone.
BACKGROUND: In recent years, the development of tissue engineering provides more choices for the repair of urethral injury.OBJECTIVE: To investigate the effect of bone marrow mesenchymal stem cells(BMSCs) combined with acellular dermal matrix in the repair of urethral injuries. METHODS: Passage 3 BMSCs from New Zealand white rabbits were inoculated on the acellular dermal matrix to construct tissue-engineered urethra grafts. Thirty-six New Zealand rabbits were randomized into three groups(n=12 per group). Experimental group was implanted withBMSCs-acellular dermal matrix complex at urethral injury. Control group was implanted with acellular dermal matrix material at urethral injury. Normal group had neither injury nor treatment. At postoperative 4, 8 and 12 weeks, the repaired urethral tissue was subjected to hematoxylin-eosin staining. At postoperative 12 weeks, immunohistochemical staining and urodynamic study were performed. RESULTS AND CONCLUSION: At postoperative 4 weeks, thin-layer epithelial regeneration was visible in the urethra defect area of the experimental group, and the continuity was better. The urethra mucosa of the control group was discontinuous. At postoperative 8-12 weeks, the urethral epithelial layer in the experimental group became thickened, exhibiting a good continuity with the normal urethral epithelium, thickened mucosa, and smooth and continuous urethral mucosa; the regenerated urethral mucosa of the control group exhibited good continuity, but less regenerated epithelial layers. At postoperative 12 weeks, immunohistochemical results showed the repaired urethra in the experimental group was positive for uroplakin Ⅲa, CK AE1/AE3, and α-smooth muscle actin. The maximum urethral pressure in the experimental group showed no significant difference before and after operation, while the postoperative pressure in the control group showed a significant increase(P < 0.05). Overall findings indicate that the combination of BMSCs and acellular dermal matrix has better efficacy than the acellular dermal matrix alone.
引文
[1]Hong YK,Choi KH,Lee YT,et al.Predictors for success of internal urethrotomy in patients with urethral contracture following perineal repair of pelvic fracture urethral injuries.Injury.2017;48(5):1035-1039.
[2]唐勇,李养群,赵穆欣,等.口腔黏膜卷管游离移植一次性重建远段尿道[J].中华整形外科杂志,2017,33(1):34-36.
[3]江志勇,李学德,何庆鑫,等.下唇黏膜或加颊黏膜与生殖器带蒂皮瓣联合修复复杂性长段前尿道狭窄[J].中国男科学杂志,2015,29(6):44-47.
[4]何越,田智泉,郜香黎,等.人羊膜脱细胞基质填充材料的生物相容性及动物有效性研究[J].中国美容整形外科杂志,2016,27(6):381-384.
[5]南文滨,徐志浩,王永学,等.脱细胞胶原基质双层材料与骨髓间充质干细胞的生物相容性研究[J].国际生物医学工程杂志,2016,39(1):16-19.
[6]Hon HH,Mubang RN,Wernick BD,et al.Acellular Dermal Matrix Versus Inferior Deepithelialized Flap Breast Reconstruction:Equivalent Outcomes,with Increased Cost.Plast Reconstr Surg Glob Open.2017;5(6):e1382.
[7]Qayyum AA,Kaur KP,Mathiasen AB,et al.Influence of patient related factors on number of mesenchymal stromal cells reached after in vitro culture expansion for clinical treatment.Scand J Clin Lab Invest.2017;77(7):541-548.
[8]Zhang X,Bendeck MP,Simmons CA,et al.Deriving vascular smooth muscle cells from mesenchymal stromal cells:Evolving differentiation strategies and current understanding of their mechanisms.Biomaterials.2017;145:9-22.
[9]Shudo Y,Goldstone AB,Cohen JE,et al.Layered smooth muscle cell-endothelial progenitor cell sheets derived from the bone marrow augment postinfarction ventricular function.J Thorac Cardiovasc Surg.2017;154(3):955-963.
[10]吴梦青,朱榕嘉,刘开彦.失血性贫血小鼠骨髓间充质干细胞向成骨成脂分化的变化[J].中国实验血液学杂志,2014,22(1):177-182.
[11]丁浩.大鼠血管内皮细胞对大鼠骨髓间充质干细胞成脂诱导的影响研究[D].泸州:西南医科大学,2015.
[12]赵刚,刘微微,高伟玮,等.不同组织来源间充质干细胞体外成骨分化能力的比较研究[J].中国骨质疏松杂志,2017,23(5):561-566,584.
[13]唐宇星,赵庆,杨中萌,等.音猬因子修饰纳米晶胶原基骨复合骨髓间充质干细胞修复股骨缺损[J].中国组织工程研究,2017,21(14):2180-2185.
[14]陈能,邵云峰,刘傥,等.带部分松质骨小牛皮质骨复合骨髓间充质干细胞植入兔体内成骨及骨形态发生蛋白2的表达[J].中国组织工程研究,2017,21(17):2684-2689.
[15]Ishihara K,Nakayama K,Akieda S,et al.Simultaneous regeneration of full-thickness cartilage and subchondral bone defects in vivo using a three-dimensional scaffold-free autologous construct derived from high-density bone marrow-derived mesenchymal stem cells.J Orthop Surg Res.2014;9:98.
[16]余瑞新,杜远立.骨髓间充质干细胞向软骨分化的研究进展[J].中国医药导刊,2012,14(1):94,103.
[17]Lemcke H,Gaebel R,Skorska A,et al.Mechanisms of stem cell based cardiac repair-gap junctional signaling promotes the cardiac lineage specification of mesenchymal stem cells.Sci Rep.2017;7(1):9755.
[18]邓宁波,曾元清,韩腾龙,等.h HO-1联合GATA-4修饰大鼠骨髓间充质干细胞抗凋亡及向心肌分化的实验研究[J].解放军医学杂志,2017,42(4):314-319.
[19]韩效林,李敬东,王婉玲,等.抗坏血酸促进骨髓间充质干细胞的上皮转分化[J].中国组织工程研究,2017,21(1):13-17.
[20]Di G,Du X,Qi X,et al.Mesenchymal Stem Cells Promote Diabetic Corneal Epithelial Wound Healing Through TSG-6-Dependent Stem Cell Activation and Macrophage Switch.Invest Ophthalmol Vis Sci.2017;58(10):4344-4354.
[21]Rowart P,Erpicum P,Krzesinski JM,et al.Mesenchymal Stromal Cells Accelerate Epithelial Tight Junction Assembly via the AMP-Activated Protein Kinase Pathway,Independently of Liver Kinase B1.Stem Cells Int.2017;2017:9717353.
[22]郭海林.组织工程尿道重建从基础到临床应用的反思[J].现代泌尿外科杂志,2016,21(2):152-155,158.
[23]杜小文,陈浩浩,刘庆,等.内纵外螺旋双层结构管状尿道支架复合体修复尿道缺损的可行性及其血管化方法[J].中华泌尿外科杂志,2017,38(1):59-65.
[24]林阳彦,王沫,杨勇,等.尿道上皮细胞/片状多孔丝素蛋白支架复合物修复兔尿道缺损的实验研究[J].中国男科学杂志,2016,30(3):14-18.
[25]Kajbafzadeh AM,Abbasioun R,Sabetkish S,et al.Future Prospects for Human Tissue Engineered Urethra Transplantation:Decellularization and RecellularizationBased Urethra Regeneration.Ann Biomed Eng.2017;45(7):1795-1806.
[26]Sim?es IN,Vale P,Soker S,et al.Acellular Urethra Bioscaffold:Decellularization of Whole Urethras for Tissue Engineering Applications.Sci Rep.2017;7:41934.
[27]蔡炳,李科,脱颖,王德娟.诱导多能干细胞向尿道上皮细胞分化体系的建立[J].中华腔镜泌尿外科杂志:电子版,2016,10(3):40-43.
[28]李泸平.组织工程化尿道构建及尿道缺损修复的实验性研究[D].郑州:郑州大学,2016.
[2]唐勇,李养群,赵穆欣,等.口腔黏膜卷管游离移植一次性重建远段尿道[J].中华整形外科杂志,2017,33(1):34-36.
[3]江志勇,李学德,何庆鑫,等.下唇黏膜或加颊黏膜与生殖器带蒂皮瓣联合修复复杂性长段前尿道狭窄[J].中国男科学杂志,2015,29(6):44-47.
[4]何越,田智泉,郜香黎,等.人羊膜脱细胞基质填充材料的生物相容性及动物有效性研究[J].中国美容整形外科杂志,2016,27(6):381-384.
[5]南文滨,徐志浩,王永学,等.脱细胞胶原基质双层材料与骨髓间充质干细胞的生物相容性研究[J].国际生物医学工程杂志,2016,39(1):16-19.
[6]Hon HH,Mubang RN,Wernick BD,et al.Acellular Dermal Matrix Versus Inferior Deepithelialized Flap Breast Reconstruction:Equivalent Outcomes,with Increased Cost.Plast Reconstr Surg Glob Open.2017;5(6):e1382.
[7]Qayyum AA,Kaur KP,Mathiasen AB,et al.Influence of patient related factors on number of mesenchymal stromal cells reached after in vitro culture expansion for clinical treatment.Scand J Clin Lab Invest.2017;77(7):541-548.
[8]Zhang X,Bendeck MP,Simmons CA,et al.Deriving vascular smooth muscle cells from mesenchymal stromal cells:Evolving differentiation strategies and current understanding of their mechanisms.Biomaterials.2017;145:9-22.
[9]Shudo Y,Goldstone AB,Cohen JE,et al.Layered smooth muscle cell-endothelial progenitor cell sheets derived from the bone marrow augment postinfarction ventricular function.J Thorac Cardiovasc Surg.2017;154(3):955-963.
[10]吴梦青,朱榕嘉,刘开彦.失血性贫血小鼠骨髓间充质干细胞向成骨成脂分化的变化[J].中国实验血液学杂志,2014,22(1):177-182.
[11]丁浩.大鼠血管内皮细胞对大鼠骨髓间充质干细胞成脂诱导的影响研究[D].泸州:西南医科大学,2015.
[12]赵刚,刘微微,高伟玮,等.不同组织来源间充质干细胞体外成骨分化能力的比较研究[J].中国骨质疏松杂志,2017,23(5):561-566,584.
[13]唐宇星,赵庆,杨中萌,等.音猬因子修饰纳米晶胶原基骨复合骨髓间充质干细胞修复股骨缺损[J].中国组织工程研究,2017,21(14):2180-2185.
[14]陈能,邵云峰,刘傥,等.带部分松质骨小牛皮质骨复合骨髓间充质干细胞植入兔体内成骨及骨形态发生蛋白2的表达[J].中国组织工程研究,2017,21(17):2684-2689.
[15]Ishihara K,Nakayama K,Akieda S,et al.Simultaneous regeneration of full-thickness cartilage and subchondral bone defects in vivo using a three-dimensional scaffold-free autologous construct derived from high-density bone marrow-derived mesenchymal stem cells.J Orthop Surg Res.2014;9:98.
[16]余瑞新,杜远立.骨髓间充质干细胞向软骨分化的研究进展[J].中国医药导刊,2012,14(1):94,103.
[17]Lemcke H,Gaebel R,Skorska A,et al.Mechanisms of stem cell based cardiac repair-gap junctional signaling promotes the cardiac lineage specification of mesenchymal stem cells.Sci Rep.2017;7(1):9755.
[18]邓宁波,曾元清,韩腾龙,等.h HO-1联合GATA-4修饰大鼠骨髓间充质干细胞抗凋亡及向心肌分化的实验研究[J].解放军医学杂志,2017,42(4):314-319.
[19]韩效林,李敬东,王婉玲,等.抗坏血酸促进骨髓间充质干细胞的上皮转分化[J].中国组织工程研究,2017,21(1):13-17.
[20]Di G,Du X,Qi X,et al.Mesenchymal Stem Cells Promote Diabetic Corneal Epithelial Wound Healing Through TSG-6-Dependent Stem Cell Activation and Macrophage Switch.Invest Ophthalmol Vis Sci.2017;58(10):4344-4354.
[21]Rowart P,Erpicum P,Krzesinski JM,et al.Mesenchymal Stromal Cells Accelerate Epithelial Tight Junction Assembly via the AMP-Activated Protein Kinase Pathway,Independently of Liver Kinase B1.Stem Cells Int.2017;2017:9717353.
[22]郭海林.组织工程尿道重建从基础到临床应用的反思[J].现代泌尿外科杂志,2016,21(2):152-155,158.
[23]杜小文,陈浩浩,刘庆,等.内纵外螺旋双层结构管状尿道支架复合体修复尿道缺损的可行性及其血管化方法[J].中华泌尿外科杂志,2017,38(1):59-65.
[24]林阳彦,王沫,杨勇,等.尿道上皮细胞/片状多孔丝素蛋白支架复合物修复兔尿道缺损的实验研究[J].中国男科学杂志,2016,30(3):14-18.
[25]Kajbafzadeh AM,Abbasioun R,Sabetkish S,et al.Future Prospects for Human Tissue Engineered Urethra Transplantation:Decellularization and RecellularizationBased Urethra Regeneration.Ann Biomed Eng.2017;45(7):1795-1806.
[26]Sim?es IN,Vale P,Soker S,et al.Acellular Urethra Bioscaffold:Decellularization of Whole Urethras for Tissue Engineering Applications.Sci Rep.2017;7:41934.
[27]蔡炳,李科,脱颖,王德娟.诱导多能干细胞向尿道上皮细胞分化体系的建立[J].中华腔镜泌尿外科杂志:电子版,2016,10(3):40-43.
[28]李泸平.组织工程化尿道构建及尿道缺损修复的实验性研究[D].郑州:郑州大学,2016.