摘要
目的通过新生小牛与成人皮肤组织在形态学和生物力学性能方面的对比研究,寻找临床应用牛真皮基质的可能证据,并初步探索对牛真皮组织进行脱细胞处理的方法。方法收集新生小牛和成人背部全层皮肤标本,采用大体观察、HE染色、Masson三色染色、天狼猩红染色、Gomori醛复红染色、扫描电镜和透射电镜的方法,显微照相后进行形态学观察和图像分析软件的测量;采用力学生物材料试验机检测皮肤的力学性能。采用胰蛋白酶与去污剂的不同浓度和时间组合方式对牛真皮组织进行脱细胞处理,初步观察其脱细胞效果。结果新生小牛与人的皮肤比较,真皮和表皮的厚度均明显变薄,两者的比值显著减小;真皮内Ⅰ型和Ⅲ型胶原纤维分别较人真皮增多和减少,但真皮胶原纤维束的间隙率[(41.72±1.81)∶(40.66±1.40),P=0.467]和胶原纤维束粗细[(11.28±0.18)μm∶(10.88±0.66)μm,P=0.368]差异却没有统计学意义。扫描电镜观察可见新生牛真皮胶纤维束与人真皮胶原纤维束均较纤细,排列疏松,且有一定的孔隙分布;透射电镜显示新生牛真皮胶原原纤维横纹周期长度较人真皮短,但两者的胶原原纤维直径显著差异。生物力学检测示新生牛皮肤最大应力[(21.08±0.91)MPa]和杨氏模量[(82.12±1.23)MPa]均要明显高于人皮肤的最大应力[(12.76±1.60)MPa,P=0.001]和杨氏模量[(48.63±5.50)MPa,P=0.001],而最大应变却明显低于人真皮[(0.51±0.002)mm∶(0.75±0.028)mm,P=0.001]。脱细胞方法探索显示随着胰酶浓度的增加、处理时间的延长,新生牛真皮基质内细胞核成分越来越少,但对真皮胶原结构的破坏也越来越大。结论新生牛皮肤虽然在表皮与真皮厚度、真皮胶原类型比例以及生物力学上与人皮肤有明显差异,但真皮的胶原纤维束三维结构两者之间存在较大相似性。适当浓度和时间的胰蛋白酶组合可制备出空间结构和脱细胞效果较为理想的新生牛脱细胞真皮基质,为修复人体软组织缺损提供有应用前景的生物材料。
Objective To compare the dermal tissue structure,content of difference types of collagen and biomechanical properties between human and newborn bovine skin,and to find the evidence of bovine dermal matrix used in clinic,in order to explore the decellularization of newborn bovine skin.Methods The dorsal full-thickness of samples were stained by hematoxylin and eosin（HE） staining,Masson＇s trichrome staining,sirius red staining,and Gomori staining,and were examined under light microscopy.These samples were also observed by scanning electron microscopy（SEM） and transmission electron microscopy（TEM）.The relevant data of histological images were measured with image analysis software.The biomechanical properties of those samples were detected by material testing machine.And the dermal matrix was decellularized using trypsin combined detergent.Results Compared with human skin,the thickness of the dermis and epidermis were significantly thinner,and the ratio between the dermis and epidermis was significantly reduced in the newborn bovine skin.The gap rate between bundles [（41.72±1.81） vs（40.66±1.40）] and the caliber of the bundles [（11.28±0.18）μm vs（10.88±0.66）μm] were not significant in statistics（P=0.467,P=0.368）.From the observation of scanning electron microscopy（SEM） examination,the dermal collagen bundles of newborn bovine were thinner and rarefaction arranged.Compared with human dermis,the newborn bovine had longer cyclical transverse striation,and the collagenous fibrils had no significant difference by transmission electron microscope（TEM）.The maximum stress of newborn bovine[（21.08±0.91） MPa] was stronger than that of human dermis [（12.76±1.60） MPa,P=0.001],and the elastic modulus were also stronger than that of human [（82.12±1.23） MPa vs（48.63±5.50） MPa,P=0.001].The exploration of decellularization showed that according to the higher concentration of typsin and the longer of time,the nucleus were more and more less,and the destruction of dermal fiber were more and more big.Conclusion The dermis of newborn bovine and human were similar in fiber three dimensional structure.Although there have a difference in biomechanical properties,the decellularization dermal matrix can used as biomaterials for reparing the human dermis.Combination appropriate concentration and time of trypsin can be prepared an ideal bovine acellular dermal matrix.
出处
《重庆医学》
CAS
CSCD
北大核心
2012年第11期1043-1046,F0002,共5页
Chongqing medicine
基金
国家高技术研究发展计划(863计划)课题资助项目(2006AA02A121)
国家重点实验室自主研究课题资助项目(SKLZZ200904)
关键词
皮肤
组织学
生物力学
脱细胞真皮基质
skin
histology
biomechanical properties
decellularized dermal matrix
