Intra-articular Injection Of Mg²⁺ Promotes Microfracture-mediated Cartilage Repair

Objective（s）:To investigate the effect of the combination of MFX and Mg²⁺on osteochonage repair and the optimal injection concentration of Mg²⁺.Explore the possible molecular pathways after injection of Mg²⁺to provide experimental basis for its clinical application.Methods:Twenty-four New Zealand white rabbits were randomly divided into4 groups（12 knees each）:microfracture group（MFX group）and MFX combined with3 different concentrations of Mg²⁺treatment（0.05 mol/L,0.5 mol/L,5 mol/L）.MFX surgery was performed after the reconstruction of the osteochondral defect（diameter5 mm,depth 2 mm）in the trochlear sulcus of both knees.Mg²⁺is injected into both knee joint cavities immediately after surgery,2 weeks and 4 weeks after surgery.The rabbits were sacrificed 6 weeks and 12 weeks after surgery,and the distribution and changes of type I collagen（COL-I.）and type II collagen（COL-II.）were detected by general observation,the International Society for Cartilage Repair（ICRS）score,histological staining（HE staining,crocus O-fixed green staining and Alisine blue staining）,and immunohistochemical detection of type I collagen（COL-I.）and type II collagen（COL-II.）.Micro-CT imaging was performed 6 weeks after surgery to observe the healing of subchondral bone,and reverse transcription quantitative polymerase chain reaction（RT-q PCR）was used to detect the expression of m RNA of SIRT1,BMP-2,SOX-9 and HIF-1α.Results:All 24 rabbits were successfully operated,and the postoperative incisions were healed without infection and complications.Six weeks after surgery,Micro-CT and general view showed that the healing of the 0.5mol/L group was better than that of other groups,and the ICRS scores of the 0.5mol/L group and the MFX group were statistically significant（P<0.01）,the HE stain,Alisin blue staining and CroséO staining showed deeper positive staining in the 0.5mol/L group,and the O’Driscoll score showed that the 0.5mol/L group was statistically significant with the MFX group（P<0.01）.Type I collagen staining showed that the positive area ratio of MFX group was better than that of different Mg²⁺concentration groups,which was statistically significant（P<0.01）,while type II collagen staining showed that 0.5mol/L was better than other groups,but not statistically significant compared with MFX group（P=0.1006）.Gross view and ICRS score showed that the healing of the 0.5mol/L group was better than that of the other groups 12 weeks after microfracture surgery,which was statistically significant with the MFX group（P<0.01）.HE staining,Alisine blue staining and crocus O staining showed that the regenerated cartilage in the 0.5mol/L group was similar to natural cartilage,and the O’Driscoll score showed that there was a significant difference between the 0.5mol/L group and the MFX group（P<0.01）;Type I collagen staining showed that the positive area ratio of MFX was higher than that of different concentrations of Mg²⁺concentration group,and there was a significant significance with 0.5mol/L（P<0.01）;Type II.collagen 0.5mol/The L-positive area ratio was significantly higher than that of other groups,and was statistically significant with the MFX group（P<0.01）.RT-q PCR results showed that after injection of Mg²⁺,the m RNA levels of SIRT1/BMP-2/SOX-9 and HIF-1αwere upregulated compared with the MFX group.Conclusion（s）:MFX combined with Mg²⁺treatment has a positive effect on osteochondrocartilage repair.Among them,when the injection concentration of Mg²⁺is 0.5 mol/L,it is most effective for enhancing microfracture-mediated osteochondrocartilage repair.At the same time,intra-articular injection of Mg²⁺promoted the synthesis of type II collagen and the formation of glycosaminoglycans by upregulating the m RNA expression levels of SIRT1-BMP-2/SOX-9 and HIF-1α.