In situ localization of procollagen gene expression on cryosections of undecalcified fracture callus

Objective:To analyze the expression of procollagen gene in fracture callus,and to search for the technique of in situ hybridization for undecalcified skeletal tissue.Methods:In situ hybridization of procollagen gene expression was performed on the undecalcified cryosections of rat fracture callus at 7,14,and 28d.Results:The hybridization signals achieved were clear and easy to be localized with high specificity.On the 7th day,the expressions of pro α1(Ⅲ) in fibroblasts and some chondrocyte-like cells were dominant;and at the end of second week high expression of type-Ⅱ procollagen mRNA was observed in chondrocytes.At the end of fourth week,the cartilaginous callus was almost all replaced by woven bone tissue,and some type-I procollagen mRNA positive osteoblasts and hypertrophic chondrocytes were found scattering in the woven bone and remnants of cartilaginous callus.Conclusions:The modified method employed in this study is easier,quicker,and more sensitive with high specificity than the conventional technique for a situ hybridization of procollagen gene expression of decalcified rat fracture callus.The phenomenon of shared phenotype expression,which was demonstrated among cells engaged in fracture healing,indicates an important approach to reveal the mechanism of the origin,differentiation,and orientation of cells.