Study On The Effect And Mechanism Of Recombinant Human Parathyroid Hormone Against Osteoarthritis

Objective:1.To observe the effects of recombinant human parathyroid hormone(PTH1-34)against the osteoarthritis(OA)of different stage induced by surgery in mice and explore its mechanism;2.To investigate the effects of PTH1-34 against early osteoarthritis induced by surgery in the type 1 diabetic mellitus(T1DM)mouse model.Methods:Part I:The effect of Recombinant Human Parathyroid Hormone against OA of different stages in mice1.C57BL/6J mice were randomly divided into groups,with osteoarthritis model induced by right medial meniscectomy(MM),and sham operation was performed on the opposite side of the limb.Normal saline and PTH1-34 solution were respectively intraperitoneal injected as planed,and samples of the mice were sacrificed on time;2.Micro CT scanning and 3d reconstruction was used for observing the bone mass and structure changes of mice subchondral bone;3.Paraffin embedding Routinely and preparation of 4?m paraffin sections after decalcification and dehydration of mouse specimens;4.Using HE staining to observe changes in articular chondrocytes and subchondral bone;5.The changes of articular cartilage and cartilage matrix were observed with safranin 0-fast green staining,and the histopathological score recommended by Osteoarthritis Research Society International(OARSI)was used to score the Osteoarthritis of mice;6.Immunohistochemical staining was performed to detect the expression levels of type 1 parathyroid hormone receptor(PTHR1),vascular endothelial growth factor(VEGF),osteoblast marker core-binding factor 2(RUNX2),and matrix metalloproteinase 13(mmp-13).Part II: Protective Effect of Recombinant Human Parathyroid Hormone against Early Osteoarthritis in Type 1 Diabetic mellitus Mice.1.Type 1 diabetes mellitus(T1DM)mice model: 12-week-old C57BL/6J mice were fasted(free drinking water)for 14 h,intraperitoneal injection of 1% streptozotocin(STZ)solution 4.5 ?L/g,once a day for 6 days.After one week of modeling,the fasting blood glucose of the mice was measured.The T1 DM model was successfully established once the fasting blood glucose of the mice exceeded 250 mg/dL for 3 consecutive days;2.Mice were randomly divided into groups.The mouse model of knee joint OA was constructed by operation of the destabilization of the medial meniscus(DMM)combined with partial medial meniscus resection.The mice in the intervention group were intraperitoneally injected with PTH1-34 solution.The mice were sacrificed on time;3.Micro CT scanning and 3d reconstruction was used for observing the bone mass and structure changes of mice subchondral bone;4.Routine paraffin embedding and preparation of 4 ? m paraffin sections after decalcification and dehydration of mouse specimens;5 Using HE staining to observe changes in articular chondrocytes and subchondral bone;6.The changes of articular cartilage and cartilage matrix were observed with safranin 0-fast green staining,and the histopathological score recommended by Osteoarthritis Research Society International(OARSI)was used to score the Osteoarthritis of mice.Results:Part I: PTH1-34 inhibits mouse OA progression in the early stage and promotes arthrosis destruction in mice in the late stage?1.Surgery-induced osteoarthritis in mice progresses with the course of the disease,and the amount of bone in the subchondral bone decreases in the early stage of osteoarthritis and increases in the late stage.Compared with the saline-treated group,PTH1-34 intervention immediately after surgery can increase the amount of bone and bone in the articular cartilage and inhibit the degeneration of articular cartilage;2.After 6 weeks of postoperative PTH1-34 intervention,the amount of bone under the articular cartilage of the mice was further increased,and the degeneration of the articular cartilage in this mouse was significantly aggravated;3.Immunohistochemical staining showed that the level of destruction of articular cartilage was correlated with the expression levels of vascular endothelial growth factor(VEGF),osteoblast marker core binding factor 2(RUNX2),and matrix metalloproteinase 13(MMP-13).Part II: PTH1-34 delays the decrease of subchondral bone mass and inhibits OA progression in early OA of T1 DM mice.1.Compared with healthy mice when OA was induced by surgery for 3 weeks,The subchondral bone mass of T1 DM OA mice was further decreased,and the OA score was higher;2.PTH1-34 intervention immediately after surgery improved the loss of subchondral bone mass and decreased OA score in T1 DM mice.Conclusions:1.In the early stage of OA,the amount of subchondral bone was reduced.Early use of PTH1-34 reversed the loss of subchondral bone mass and inhibited cartilage degeneration in the early stage of mice OA.The application of PTH1-34 in the late stage of OA promoted cartilage destruction.This suggests that the clinical application of PTH1-34 and related preparations for the treatment of osteoarthritis should specifically assess the benefits and harms,The application of PTH1-34 should be with caution;2.T1 DM can accelerate the loss of subchondral bone mass and promote the early progression of OA.The use of PTH1-34 can reverse the loss of subchondral bone mass in the early stage of T1 DM mice OA,and play a protective role in the early stage of T1 DM OA.