Study On The Effect Of Morinda Officinalis On Proliferation Of Osteoblasts In Traumatic Osteonecrosis Of Femoral Head Based On Wnt/β-catenin Pathway

Objective:1.Taking the rabbit model of traumatic osteonecrosis of the femoral head(TONFH)as the research object,the effect of Morinda officinalis How on traumatic femoral head necrosis was observed.2.Taking hypoxia-induced rat osteoblasts as the research object,the mechanism of the effect of Morinda officinalis How-containing serum Wnt/β-catenin pathway on osteoblast proliferation was discussed.Methods: 55 male adult healthy New Zealand white rabbits were chosen in this study,weighing from 2kg to 4kg.The rabbits were managed in accordance with SPF level standards.According to the random allocation table,48 rabbits after successful modeling were randomly assigned to the model control group,Morinda officinalis lowdose group,Morinda officinalis middle-dose group,Morinda officinalis high-dose group,12 animals in each group.they were given Chinese medicine Morinda officinalis low(0.35 g crude drug amount / kg),medium(0.7g crude drug amount / kg),high dose(1.4g crude drug amount / kg)and normal saline for intragastric administration intervention respectively.Observation and detection at 4 and 8 weeks after intragastric administration were conducted: The general conditions of rabbit were observed,such as weight change,activity,mental status,diet;X-ray images were token and the morphology of the femoral head,sclerosis or cystic change of the femoral head and bone density were observed and recorded.Cell experiment part: The rat osteoblasts were token for culturation.After hypoxia treatment,different dosage of Morinda officinalis-containing serum were added into the cells.The cells underwent cck-8cytoactive detection to find the most suitable concentration of drug-containing serum for intervention.The cells after hypoxia treatment were randomly divided into model control group,Morinda officinalis low-dose group,Morinda officinalis middle-dose group,Morinda officinalis high-dose group.The cells of each group were taken to be seeded on the plate,and added these cells into serum-free DMEM culture media and corresponding dosage of Morinda officinalis-containing serum was added to culture.Western blot was used to detect β-catenin and GSK-3β protein expression levels of the cells until36h’s culturation.Results:1.Chinese medicine Morinda officinalis has obvious curative effect on traumatic femoral head necrosis.Under the intervention,Statistical analysis and comparison of various indicators in each group have overall statistical significance.2.Microscopic observation of osteoblast morphology: The third-generation osteoblasts are partly fusiform,partly round or polygonal,with extended tentacles visible;they are arranged in a net shape,and the arrangement is relatively tight.Abundant cytoplasm can be seen inside the cell,and clear nuclei and nucleoli can be seen.After hypoxia treatment,the osteoblasts can be seen to be significantly narrower,sparsely arranged,and some cells have apoptosis.The remaining surviving cells can be seen clear nuclei and nucleoli.Microscopic observation of cell morphology after drug-containing serum intervention: the model group mostly s howed a long spindle shape,and the low-,medium-,and high-dose groups mostly showed a round spindle shape;The cells in the model group are sparsely arranged,and it can be seen that some cells have apoptosis,which is similar to the cell morphology afte r hypoxia treatment;The cells intervened by the drug-containing serum are arranged in a network and are arranged more closely.3.The best intervention concentration of osteoblasts is containing 10% Morinda officinalis serum suggested by the cck-8 detection method.And Morinda officinalis high-dose group has the best effect on promoting osteoblast proliferation.4.Western blot results: β-catenin protein expression: When drugcontaining serum interfered with osteoblasts for 36 h,the protein content of the remaining three groups was significantly higher than that of the model group(P<0.05),Morinda officinalis high-dose group and Morinda officinalis medium-dose group were significantly higher than the low-dose group(P<0.05),and the Morinda officinalis high-dose group was significantly higher than the Morinda officinalis medium-dose group(P<0.05).GSk-3β protein expression: When the serum of medicine interfered with osteoblasts for 36 h,the expression levels of Morinda officinalis low-dose group,Morinda officinalis medium-dose group,Morinda officinalis high-dose group were all significantly decreased compared with the control group(P<0.05);There is no significant difference between Morinda officinalis high,medium and low dose groups(P>0.05).Conclusion: Morinda officinalis has a good effect on the treatment of traumatic femoral head necrosis.It can improve the trabecular bone fracture and disorder of femoral head necrosis in a dose-dependent manner.The mechanism may be related with Morinda officinalis regulating the Wnt/β-catenin pathway to promote the proliferation of osteoblasts,inhibit their apoptosis,and delay the necrosis of femoral head.