Study On The Efficacy Of Different Administration Routes Of Placental Mesenchymal Stem Cell Preparations In The Treatment Of OPF In Tree Shrews

Objective:To explore the efficacy of placental mesenchymal stem cell preparations in the treatment of osteoporotic fractures in tree shrews,and the effect of different administration routes on the curative effect of osteoporotic fractures in tree shrews.Methods:The female tree shrews were randomly divided into an operation group and a sham-operated group.The operation group was used to establish an osteoporosis(OP)tree shrew model.The operation group simulated postmenopausal osteoporosis by removing both ovaries and uterus,while the sham-operated group only removed the large omentum equal to the size of the uterus and ovaries.The model was formed within 6 months.The successful model was confirmed by the DXA test.OP tree shrews were treated with tibial fractures of the right hind limbs to establish an osteoporotic fracture(OPF)tree shrew model.Twenty-four OPF tree shrews were randomly divided into 4 groups,6 in each group.PMSCs were resuspended in saline and the cell concentration was adjusted to 1×106/mL.In group A,PMSCs were injected with 1 mL in the tail vein.PMSCs in group B were injected with 0.5 mL each in the wound and tail vein.PMSCs in group C were injected with 0.5 mL of PMSCs in the abdominal cavity and tail vein,group D was injected with the same amount of normal saline in the tail vein.From the 3rd day after the fracture,injections were given once a week for 3 consecutive weeks,for a total of 3 injections.After the first treatment for 10 weeks,the body weight,bone mineral density(BMD),three-point bone biomechanics of each group of tree shrews were measured,and the pathological changes of bone tissue were observed by HE staining.ELISA kit to detect the serum estrogen,bone alkaline phosphatase(BALP),osteocalcin(OCN)and tartrate resistant acid phosphatase(TRAP)expression levels of each group of tree shrews.And Real-time fluorescent quantitative PCR(RT-qPCR)to detect the mRNA expression level of callus related indicators Bone Morphogenetic Protein-2(BMP2),Vascular Endothelial Growth Factor(VEGF),Osteoprotective Protein(OPG)and Nuclear factor kappa B receptor activator ligand(RANKL).Results:Comparing the four groups of tree shrews,the weight of group A was significantly reduced compared with group B,C and D,but only the difference between group D and group A was statistically significant(P<0.05).The BMD of group A was significantly increased compared with groups C and D,and the BMD of group B was significantly increased compared with group D,and the differences were statistically significant(P<0.05).In the detection of femoral three-point bone biomechanics,the maximum load of group A was significantly higher than that of groups B,C,and D,and compared with groups C and D,group A has significantly higher structural rigidity than group D.The energy absorption of groups A and B was significantly higher than that of group D,and the difference was statistically significant(P<0.05 or P<0.01).The results of HE staining showed that the basic structure of the trabecular bone of the tree shrews of group D was incomplete,broken,and the reticular structure disappeared,and the pathological changes of the tree shrews of group B and C were improved compared with group D,but the trabecular bone still remained Obvious fracture phenomenon.The pathological changes of tree shrews in group A have been significantly improved,and the trabecular bone structure is relatively complete and arranged in an orderly manner.The serological test results showed that in the detection of estrogen and BALP levels,group A was significantly higher than that of groups B,C,and D,and group B was higher than that of group D.In the detection of OCN levels,group A was higher than that of groups B and D.In the detection of TRAP level,group A was lower than the three groups B,C,and D to different degrees,and the differences were statistically significant(P<0.05 or P<0.01).In addition,the mRNA levels of OPG and BMP2 in groups A,B,and C increased to varying degrees compared with group D,and group B showed the highest level.And the mRNA levels of RANKL in groups A,B,and C were lower than those in group D to different degrees,among which group B showed the lowest level,and the differences were statistically significant(P<0.01).The levels of VEGF in groups A,B and C increased compared with group D,but the difference was not significant(P>0.05).Conclusion:PMSCs transplantation can increase the OPF tree shrew model BALP,OCN,OPG and other bone formation indicators,reduce TRAP and other bone resorption indicators,improve tree shrew weight,BMD,estrogen and other related indicators.PMSCs may regulate the process of bone formation and bone resorption,and Regulate paracrine action and play a therapeutic role together.Full-dose tail vein injection of PMSCs has the most significant improvement in the systemic bone strength of tree shrew OPF model,which may be related to the direct entry of stem cells into the systemic circulation through the tail vein to exert systemic effects.Secondly,tail vein combined wound injection has better curative effect,among which the improvement of bone callus is more significant,which may be related to the direct contact of PMSCs to the fracture site.The tail vein combined with abdominal cavity injection is less effective.