| Part 1 Effects of oral blood-activating and bone formation decoction and impacting bone graft treating pre-col lapse stage ONFHObjectiveTo observe the clinical curative effects and security of oral blood-activating and bone formation decoction and impacting bone graft with nonvascular fibular allograft treating pre-collapse stage kidney deficiency and blood stasis type of ONFH,and discuss its mechanism of action.Methods62 patients(80 hips)between February 2013 and October 2016 in Linyi People ' s Hospital with osteonecrosis of the femoral head according to the patients ' willing divided into treatment group and control group,60 cases(78 hips)completed treatment and follow-up.The treatment group(25 cases of male,female 4 cases,aged(39.7±6.9),the course of the disease(4.6±1.6)months)took Chinese herbal medicine blood-activating and bone formation decoction and impacting bone graft.The control group(26 males,5 females,age(38.7±6.7),course(4.8±1.7)months)treated by impacting bone graft only.The general data between the two groups had no statistical significance(P>0.05).The outcome was evaluated both clinically by Harris hip score(HHS)and radiologically by X-rays or CT,imaging examination to observe morphological changes.Recorded the cases of femoral head collapse,and monitored serum low density lipoprotein(LDL),Triglyceride(TG),serum bone gla protein(BGP).ResultThe two groups of postoperative incision were primary healing,no early complications and were received more than 2 year follow-up,the treatment group 4 hips femoral head collapsed,and the control group 12 hips collapsed,the difference was statistically significant(P=0.033).After the 6,12,24 months'treatment,the HHS of the treatment group and control group were higher than that before the treatment(P<0.01),while the treatment group was better than that of the control group,the difference was statistically significant(F=11.334,P<0.01),processing method and time factors interaction effect(F=6.097,P<0.01).The two groups after treatment serum bone gla protein had increased,the treatment group compared with control group difference was statistically significant(F=11.528,P<0.01);processing method and time factors interaction effect(F=13.702,P<0.01).Two groups of treatment after 6,12 months the level of serum bone gla protein was higher than before treatment(P<0.05);12 months after treatment compared with 6 months after treatment,although increased,but there were no statistically significant difference(P>0.05)in the two groups.Two groups of serum triglyceride and triglyceride before treatment there was no statistical significance(P>0.05)difference comparison,6 and 12 months after treatment group was lower than that before treatment(P<0.05),the control group with quite(P>0.05)before treatment,the treatment group compared with control group difference was statistically significant(P<0.05).Different time had time effect(P<0.05),the time factors and processing factors had interaction effect(P<0.01).ConclusionTwo methods for the early stage of the collapse ONFH can obtain good clinical curative effect.Compared with impacting bone graft and nonvascular fibular allograft only,taking Chinese herbal medicine blood-activating and bone formation decoction and impacting bone graft and nonvascular fibular allograft effectively improve clinical symptoms of the hips,decrease the rate of the collapse of the femoral head,promote the bone repair.These effects may be related to effective reduce serum TG and LDL levels or increase the serum level of BGP.Part 2 Effects of oral blood-activating and bone formation decoction and extracorporeal shock wave treatment for early eldly kidney deficiency and blood stasis type of ONFHObjectiveTo observe the clinical curative effects and security of oral blood-activating and bone formation decoction and extracorporeal shock wave treatment for early elderly kidney deficiency and blood stasis type of ONFH,and discuss its mechanism of action.MethodsUsing prospective controlled study,56 patients(96 hips)between June 2014 and December 2016 in Linyi People ' s Hospital outpatient or hospitalization with early eldly osteonecrosis of the femoral head according to the patients ' willing divided into treatment group and control group.50 cases(86 hips)completed treatment and follow-up.The treatment group(23 cases of male,1 case of female,aged 60 to 68,63.4 years on average,the course of the disease from 1 to 8 months,an average of 3.6 months.ARCO I stage 6 cases(10 hips),ARCO ? stage 18 cases(30 hips)),took Chinese herbal medicine blood-activating and bone formation decoction,daily 1 agent,orally one dose daily,and extracorporeal shock wave,once a day.The control group(25 cases of male,1 case of female,aged 60 to 68,64.0 years on average,the course of the disease from 1 to 10 months,an average of 3.7 months)ARCO I stage 7 cases(12 hips)),ARCOII stage 19 cases(34 hips),treated by ESWT only.The general data between the two groups had no statistical significance(P>0.05).Compared.with each other in such parameters as scores of Harris,Visual analogue score,low density lipoprotein(LDL),total cholesterol(TC)bone gla protein(BGP),insulin like growth factor-1(IGF-1)and security index before and after treatment.Finally,we evaluated the curative effect of Chinese medical syndrome after treatment.Resut:Four cases of treatment group and two cases of control group duo to lack of efficacy or failure to adhere to the drug or extracorporeal shock wave treatment falls off or eliminate.Complete treatment and follow-up of the 50 cases(86 hips)in treatment group and control group,with a minimum of 12 months follow-up.The treatment group had 90.0%curative effect of Chinese medical syndrome,significant higher than that of the control group 70%(P<0.05).After the 6 and 12 months' treatment,the HHS of the treatment group and control group were higher than before the treatment,while the treatment group was better than that of the control group,the difference was statistically significant(P=26.863,P<0.01),processing method and time factors interaction effect(F=17.046,P<0.01).The two groups after treatment serum BGP had increased,the treatment group compared with control group difference was statistically significant(F=7.561,P<0.01);Two groups of treatment and 6 months after,the level of serum BGP was higher than that before treatment(P<0.05);6 months after treatment compared with that after treatment,although increased,but there were no statistically significant difference(P>0.05)in the two groups.Two groups of serum TG?LDL and IGF-1 before treatment were no statistical significance(P>0.05)difference comparison.After treatment and 6 months after,the treatment group was lower than that before treatment(P<0.01),the control group was quite(P>0.05)before treatment,the difference of the treatment group compared with control group was statistically significant(P<0.01).The time factors and processing factors had interaction effect(P<0.01).ConclusionTaking Chinese herbal medicine blood-activating and bone formation decoction and extracorporeal shock wave treatment than extracorporeal shock wave treatment only can effectively improve clinical symptoms for early elderly kidney deficiency and blood stasis type of ONFH.These effects may be related to effective regulation of metabolic balance of lipid and bone.Part 3 Blood-activating and bone formation decoction medicated serum of dexamethasone induced influence the differentiation of bone marrow mesenchymal stem cellsObjectiveInterfering with dexamethasone induced BMSCs by blood-activating and bone formation decoction medicated serum in vitro experiment,to observe its effect on osteogenesis and adipogenic differentiation of BMSCs and the Wnt/?-catenin signaling pathway related to osteogenesis and adipogenic differentiation factor expression,and to discusses the mechanism of action of kidney and blood stasis type of ONFH with blood-activating and bone formation decoction.Methods:In vitro experiment,obtained the BMSCs of clean SD rats,and isolated,cultured,and identified the cells.Prepared of the blood-activating and bone formation decoction medicated serum,and observed the proliferation of BMSCs intervened with different concentrations of drug-containing serum.Induced the BMSCs adipogenic differentiation with DEX,and interfered with different concentrations of blood-activating and bone formation decoction.To observe the gross morphology of BMSCs by oil red 0 and alizarin red staining,and grasp osteogenesis and adipogenic differentiation of BMSCs by detecting triglyceride and alkaline phosphatase in BMSCs of each group after the intervention.Through observing the Wnt/?-catenin signaling pathway related to osteogenesis and adipogenic differentiation factor expression with the methods of RT-PCR and Western Blotting,to explore the mechanism of action of blood-activating and bone formation decoction in the treatment of kidney and blood stasis type of ONFH in molecular and cellular level.ResultsCCK8 experimental results showed that the proliferation ability of BMSCs with different concentrations of medicated serum intervention were stronger than that of the blank serum group(P<0.05).In the adipogenic differentiation tests,comparison of oil red 0 staining,the blank control groups did not see fat particles appeared in the experiments,the model groups and blank serum groups were visible large red dye,the concentration of blood-activating and bone formation decoction medicated serum lipochondrion were less than that of the model group and the blank serum group,the fat particles of the middle dose groups were significantly higher than of the low dose groups and the high dose groups,TG detection situation:blood-activating and bone formation decoction medicated serum groups to compare with model groups were significantly lower(P<0.01),the middle dose groups were lower than the low dose groups(P<0.05)and the high dose groups(P>0.05).RT-PCR results showed that different concentrations of medicated serum inhibit DKK1 and Sfrpl gene expression(P<0.05),the middle dose groups were the lowest,and the expression of Sfrpl the middle dose groups was significantly lower than the low dose groups and the high dose groups(P<0.01),the expression of DKK1 the middle dose groups was significantly lower than the low dose groups(P<0.01)and was no significant difference than the high dose groups(P>0.05).Western Blotting results showed that DKK1 and Sfrpl protein expression of the different concentrations of medicated serum were significantly weakened than the model groups(P<0.01),the expression of the middle dose groups were the weakest,and the expression significantly lower than the low dose groups and the high dose groups(P<0.01).In osteogenesis differentiation tests:comparison of alizarin red staining,calcium nodules appeared in the blank control groups,model groups and blank serum groups,different concentrations of medicated serum groups can be seen obviously calcium nodules,the calcium nodules in equivalent dose groups were more than that in the low dose groups and the high dose groups.ALP detection situation:blood-activating and bone formation decoction medicated serum groups to compare with model groups were significantly higher(P<0.01).Among them,the equivalent dose groups were significantly higher than the the low dose groups(P<0.01)and no significant difference than the high dose groups(P>0.05).RT-PCR results showed that different concentrations of medicated serum enhanced p-catenin and Wnt3a gene expression(P<0.01),and the expression of the middle dose groups was significantly higher than the low dose groups and the high dose groups(P<0.01).Western Blotting results show that ?-catenin and Wnt3a gene expression were significantly higher(P<0.01).The middle dose groups were significantly higher than that of the low dose groups and the high dose groups(P<0.01).ConclusionDifferent concentrations of blood-activating and bone formation decoction medicated serum raised ?-catenin and Wnt3a level,decreased DKK1 and Sfrpl,promoted the Wnt/?-catenin signaling pathway opening and inhibite adipogenic differentiation and promote osteogenesis differentiation through Wnt/?-catenin signaling pathway consequently. |
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