The Construction Of DMP1 Knockout Rabbit Model Of Hypophosphatemic Rickets

Hypophosphatemic rickets(HR)is a bone disease caused by loss of the loss of phosphates in the kidney and decrease in serum phosphorus levels,leading to bone mineralization abnormalities.Autosomal recessive hypohosphatemic rickets(ARHR)is a kind of HR.Its typical clinical manifestations are rickets and osteomalacia,accompanied by short stature,abnormality in bone and tooth development,elevated serum FGF23 levels,and decreased serum phosphorus levels.The main pathogenic gene of ARHR is DMP1(Dentin Matrix Protein 1)gene,located on 4q21-22 with 7 exons,encoding an acidic non-collagenous extracellular matrix protein which highly expresses in bones and teeth.It is an important regulator in the formation and mineralization of bone and tooth.DMP1 regulates homeostasis of phosphatase enzymes in vivo through fibroblast growth factor 23(FGF23),a protein with endocrine function released from osteocytes and osteoblasts.In addition,DMP1 accelerates phosphate metabolism,leading to a decrease in serum phosphorus levels.Therefore,the study on DMP1 will help us increase the understanding of bone mineralization process and phosphorus transportation mechanism in the kidney,finally reveal the pathogenesis of ARHR.With the development of science and technology,human beings show an urgent desire to have a deep understanding of the diseases.Researchers have found that most of the common and rare diseases are related to mutations in various genes.However,due to the complexity of clinical manifestations,the pathogenesis of the diseases is not clear,which leads to the lack of effective prevention and treatment means.Therefore,establishing an animal model that could simulate the symptoms of human diseases is useful for the studies on the pathogenesis of the diseases and the screen of the treatment.Nowadays,researchers have successfully constructed a mouse ARHR model.Human ARHR can’t be fully simulated since the histological characteristics of bone and the pathological changes of arthritic bones in mice are different from those of humans.However,as a classic model organism,rabbits share more physiological,anatomical,and genetic characteristics with humans,and require lower feeding costs and shorter gestational cycles.Therefore,rabbits are a more reliable and ideal model organism for better simulating human diseases.In this study,we successfully constructed a rabbit hypophosphatemic rickets model by DMP1 gene knockout rabbit by using CRISPR/Cas9 system.We have successfully obtained DMP1 knockout rabbits,and no off-target mutations were detected at these potential sites.We analyzed the phenotypes of DMP1 knockout rabbits,the results showed that abnormal expression of serum,elevated serum FGF23,decreased serum phosphorus;teeth,lower jaw,vertebrae and long-bone mineralized incompletely and reduced bone density;typical symptoms of rickets with rachitic rosary in ribs;long-bone is abnormal with cells mutated,resulting in shorter and thicker long-bones,which were observed in DMP1 knockout rabbits.These phenotypes were consistent with the clinical manifestations of human ARHR patients.This novel rabbit model may help reveal the pathogenesis of hypophosphatemic rickets and provide an ideal animal model for drug screening and clinical treatment of hypophosphatemic rickets.