CAJ | 학술논문

目的比较三维条件下不同代次角质形成细胞构建的表皮形态及增殖分化情况.方法采用不同代次角质形成细胞构建组织工程皮肤,通过苏木精-伊红(HE)和高碘酸-希夫(PAS)染色,观察各组组织工程皮肤的结构形态；并利用角蛋白CK1/CK10、CK5/CK14、细胞增殖核抗原Ki-67免疫组织化学染色,比较各组组织工程皮肤表皮的增殖分化能力.结果不同代次角质形成细胞构建的组织工程皮肤都有明显的表皮真皮结构.1、2代较其他代次分层情况好,2代表皮层厚度明显高于其他代次(P＜0.05).加Ⅳ型胶原组的表皮真皮间黏附的紧密程度高于未加Ⅳ型胶原组,Ⅳ型胶原对表皮的厚度影响不大(P＞0.05).Ⅳ型胶原组PAS染色阴性,说明单独加Ⅳ型胶原不能形成基底膜结构.2代角质形成细胞Ki-67增殖系数接近正常皮肤,其余各代次角质形成细胞的增殖系数逐渐减少(P＜0.05).CK1/CK10、CK5/CK14在1、2代皮肤中阳性表达明显,其余代次对于两种角蛋白的表达并不明显.结论 3代以前角质形成细胞有更好的增殖分化能力,更适合作为组织工程皮肤的种子细胞.
목적 비교삼유조건하불동대차각질형성세포구건적표피형태급증식분화정황.방법 채용불동대차각질형성세포구건조직공정피부,통과소목정-이홍(HE)화고전산-희부(PAS)염색,관찰각조조직공정피부적결구형태；병이용각단백CK1/CK10、CK5/CK14、세포증식핵항원Ki-67면역조직화학염색,비교각조조직공정피부표피적증식분화능력.결과 불동대차각질형성세포구건적조직공정피부도유명현적표피진피결구.1、2대교기타대차분층정황호,2대표피층후도명현고우기타대차(P＜0.05).가Ⅳ형효원조적표피진피간점부적긴밀정도고우미가Ⅳ형효원조,Ⅳ형효원대표피적후도영향불대(P＞0.05).Ⅳ형효원조PAS염색음성,설명단독가Ⅳ형효원불능형성기저막결구.2대각질형성세포Ki-67증식계수접근정상피부,기여각대차각질형성세포적증식계수축점감소(P＜0.05).CK1/CK10、CK5/CK14재1、2대피부중양성표체명현,기여대차대우량충각단백적표체병불명현.결론 3대이전각질형성세포유경호적증식분화능력,경괄합작위조직공정피부적충자세포.
Objective To compare the epidermal shape built by different passages of keratinocytes and its ability of proliferation and differentiation in three-dimensional conditions.Methods Different passages of keratinocytes were used to construct tissue-engineered skin.The morphology of the tissue-engineered skin was observed with HE and PAS staining,while CK1/CK10,CK5/CK14,Ki67 were detected by immunohistochemical assays.Results All the tissue-engineered skin had a significant dermoepidermal structure.The stratification of 1st and 2nd passage skins were better,and 2nd passage epidermis was thicker than that in other passages (P＜0.05).Dermoepidermal structure in collagen type Ⅳ group binded more tightly,but collagen type Ⅳ had little effect on the thickness of the epidermis (P＞0.05).In collagen type Ⅳ group PAS stain was negative,indicating type Ⅳ collagen was unable to promote the reconstruction of BM in vitro.The Ki-67 proliferation index of the 2nd keratinocyte was similar to the normal skin,the remaining passages keratinocyte proliferation gradually decreased (P＜0.05) ; the 1st and 2nd passage skins expressed CK1/CK10 and CK5/CK14.Conclusions Keratinocytes before the 3rd passage have a better ability in the proliferation and differentiation,and so they are more suitable as seed cells for tissue-engineered skin.