中华眼科杂志
中華眼科雜誌
중화안과잡지
Chinese Journal of Ophthalmology
2008年
9期
825-830
,共6页
房兴峰%赵靖%史伟云%谢立信
房興峰%趙靖%史偉雲%謝立信
방흥봉%조정%사위운%사립신
角膜基质%角膜%组织工程%生物相容性材料
角膜基質%角膜%組織工程%生物相容性材料
각막기질%각막%조직공정%생물상용성재료
Corneal stroma%Cornea%Tissue engineering%Biocompatible materials
目的 比较氯化钠(NaCl)-十二烷基硫酸钠(SDS)-胰蛋白酶与分散酶(Dispase)-聚乙二醇辛基苯基醚(Triton-X-100)两种角膜组织脱细胞方法的效果,探讨以脱细胞角膜基质为支架构建组织工程化角膜上皮组织的可行性.方法 实验研究.采用完全随机化设计的方法,分别用NaCl-SDS-胰蛋白酶和Dispase-Triton-X-100处理兔角膜组织,使用裂隙灯显微镜、光学显微镜及透射电镜观察经两种方法处理后的角膜基质特性和脱细胞效果.以兔角膜缘上皮细胞为种子细胞,Dispase-Triton-X-100处理的猪角膜前弹力层与基质为支架,体外重建兔角膜上皮细胞层,并进行形态学、组织病理学及免疫组织化学检测.结果 两种方法处理过的兔角膜基质大体形态相似,灰白色不透明,水肿明显,质地柔软.组织病理学和超微形态学观察显示两种方法处理的兔角膜基质胶原排列规整,NaCl-SDS-胰蛋白酶处理的角膜组织残留了部分基质细胞碎片,而Dispase-Triton-X-100处理的角膜组织未见基质细胞碎片.兔角膜缘上皮组织块接种于脱细胞猪角膜前弹力层与基质上,24 h角膜上皮细胞开始游出,3～4 d融合呈片状,7～8 d形成单细胞层.组织工程化角膜上皮细胞表达CK3.结论 Dispase-Triton-X-100处理方法的角膜组织脱细胞效果良好.以脱细胞猪角膜前弹力层与基质为支架.可在体外构建组织工程化兔角膜上皮组织.
目的 比較氯化鈉(NaCl)-十二烷基硫痠鈉(SDS)-胰蛋白酶與分散酶(Dispase)-聚乙二醇辛基苯基醚(Triton-X-100)兩種角膜組織脫細胞方法的效果,探討以脫細胞角膜基質為支架構建組織工程化角膜上皮組織的可行性.方法 實驗研究.採用完全隨機化設計的方法,分彆用NaCl-SDS-胰蛋白酶和Dispase-Triton-X-100處理兔角膜組織,使用裂隙燈顯微鏡、光學顯微鏡及透射電鏡觀察經兩種方法處理後的角膜基質特性和脫細胞效果.以兔角膜緣上皮細胞為種子細胞,Dispase-Triton-X-100處理的豬角膜前彈力層與基質為支架,體外重建兔角膜上皮細胞層,併進行形態學、組織病理學及免疫組織化學檢測.結果 兩種方法處理過的兔角膜基質大體形態相似,灰白色不透明,水腫明顯,質地柔軟.組織病理學和超微形態學觀察顯示兩種方法處理的兔角膜基質膠原排列規整,NaCl-SDS-胰蛋白酶處理的角膜組織殘留瞭部分基質細胞碎片,而Dispase-Triton-X-100處理的角膜組織未見基質細胞碎片.兔角膜緣上皮組織塊接種于脫細胞豬角膜前彈力層與基質上,24 h角膜上皮細胞開始遊齣,3～4 d融閤呈片狀,7～8 d形成單細胞層.組織工程化角膜上皮細胞錶達CK3.結論 Dispase-Triton-X-100處理方法的角膜組織脫細胞效果良好.以脫細胞豬角膜前彈力層與基質為支架.可在體外構建組織工程化兔角膜上皮組織.
목적 비교록화납(NaCl)-십이완기류산납(SDS)-이단백매여분산매(Dispase)-취을이순신기분기미(Triton-X-100)량충각막조직탈세포방법적효과,탐토이탈세포각막기질위지가구건조직공정화각막상피조직적가행성.방법 실험연구.채용완전수궤화설계적방법,분별용NaCl-SDS-이단백매화Dispase-Triton-X-100처리토각막조직,사용렬극등현미경、광학현미경급투사전경관찰경량충방법처리후적각막기질특성화탈세포효과.이토각막연상피세포위충자세포,Dispase-Triton-X-100처리적저각막전탄력층여기질위지가,체외중건토각막상피세포층,병진행형태학、조직병이학급면역조직화학검측.결과 량충방법처리과적토각막기질대체형태상사,회백색불투명,수종명현,질지유연.조직병이학화초미형태학관찰현시량충방법처리적토각막기질효원배렬규정,NaCl-SDS-이단백매처리적각막조직잔류료부분기질세포쇄편,이Dispase-Triton-X-100처리적각막조직미견기질세포쇄편.토각막연상피조직괴접충우탈세포저각막전탄력층여기질상,24 h각막상피세포개시유출,3～4 d융합정편상,7～8 d형성단세포층.조직공정화각막상피세포표체CK3.결론 Dispase-Triton-X-100처리방법적각막조직탈세포효과량호.이탈세포저각막전탄력층여기질위지가.가재체외구건조직공정화토각막상피조직.
Objective To compare two methods for preparing acellular corneal strnma and evaluate the possibility of culturing corneal epithelium on xenogeneic acellular corneal stroma.Methods Experimental study,applying completely randomized design method.Dispase followed by Triton-X-100 detergent and sodium chloride SDS detergent followed by trypsinase were apphed respectively to treat the rabbit comeea.The characteristics of corneal stroma and acellular status after treatment were examined with slit lamp,optical microscope and transmission electron microscope.The rabbit limbal cells were then cultured on the acellular porcine Bowman's membrane/stroma.Rabbit corneal epithelium lamella reconstructed in vitro and evaluated in morphology,histopathology and immunohisto-chdmistry.Results Acellolar corneal stroma prepared by two different methods is quite similar in morphology,being gray and opaque with visible edema and soft texture.Collagen fibers of the stroma were regularly in histopathology and ultrastructure.But the one prepared by the NaCl-SDS-Trypsinase metllod retained a amounts of cell debris,while there was none in the other did by Dispase-Tfiton-X-100 method.The limbal cells began to shift out at 24 hours after being inocdated on xenogeneic acellular corneal stroma,then attached on it,formed a confluent monolayer containing normal-appearing in 7 days.The tissue engineering corneal epithelium was cryosectioned and characterized immunohistochemically at 14 days after inoculation.Meanwhile,epithelium associated antigen CK3 in endochylema was stained.Conclusions Dispase-Triton-X-100 was proved better in obtaining acellular corneal stroma.It is possible to reconstruction tissue-engineered rabbit's corneal epithelium on acellular porcine corneal Bowman's membmne/stroma.
