Influence Of HB-EGF Overexpression In Prrx1~+ Bone Marrow Mesenchymal Stromal Cells On Skeletal Development

Heparin-binding EGF-like growth factor（HB-EGF）is a member of the epidermal growth factor（EGF）superfamily.After HB-EGF binds to the EGF receptor（EGFR）or ERBB4,it causes receptor dimerization and tyrosine phosphorylation which triggers a series of signaling cascades,including extracellular signal-regulated protein kinases（ERK）,mitogen-activated protein kinase（JNK）and phosphatidylinositol 3-kinase（PI3K）pathways.These pathways participate in many physiological and pathological processes such as tumorigenesis and skeletal development.Derived from the mesoderm,mesenchymal stromal cells（MSC）are multipotent cells that express CD73,CD90,CD105.MSC also show plastic adherent properties under normal culture conditions and can differentiate into a variety of cell types including osteoblasts,chondrocytes and adipocytes in vitro.It has been reported that HB-EGF is important for osteogenic and chondrogenic differentiation.HB-EGF inhibits the expression of ALP and OCN,suggesting that HB-EGF may negatively regulate osteogenic differentiation.In addition,the expression of HB-EGF is increased in articular chondrocytes in both osteoarthritis patients and rodent surgical osteoarthritis model.This increased HB-EGF may lead to imbalance of cartilage degradation-formation in osteoarthritis.With the lack of effective markers to label mesenchymal cell,there is little research about the influence of HB-EGF on skeletal development in vivo.Recent studies have shown that Prrx1 can be used as a genetic marker for mesenchymal stromal cells,so we generated a mouse model of HB-EGF overexpression in Prrx1⁺BM-MSC using Cre/loxP system to explore the effect of HB-EGF on skeletal development.In this study,Prrx1 is expressed in the articular cartilage,growth plate,trabecular bone and periosteum of adult mice.We found that the femurs of Prrx1-Cre;ROSA26^{tm1（HB-EGF）}mice were shorter,with significantly decrease in bone mass,defective mineralization in cortical bone and abnormal cartilage morphology and structure.While the skeleton of the mutant mice was normal at p1 without any change in the size of bone and the morphology of the knee joints.Further investigation showed that the proliferation of BM-MSC derived from Prrx1-Cre;ROSA26^{tm1（HB-EGF）}mice was enhanced in vitro,while their osteogenic and chondrogenic differentiation were inhibited and the expression of related genes was down-regulated.In conclusion,overexpression of HB-EGF in Prrx1⁺MSC may influence the skeletal development in mice,resulting in decreased bone mass and osteoarthritis-like phenotypes,by inhibiting osteogenic and chondrogenic differentiation of BM-MSC.This study suggests that HB-EGF plays an important role in regulating the multilineage differentiation of BM-MSC.These findings have advanced our understanding of osteoarthritis pathogenesis,which may contribute to future clinical research and treatment.