Ginkgo biloba L. extract prevents steroid-induced necrosis of the femoral head by rescuing apoptosis and dysfunction in vascular endothelial cells via the PI3K/AKT/eNOS pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Ginkgo biloba L. extract (EGb) is one of the world's most extensively used herbal medicines. Due to the diverse pharmacological properties of EGb, it has been used in the treatment of neurological illnesses, as well as cardiovascular and cerebrovascular ailments. However, the effect and pharmacological mechanism of EGb on steroid-induced necrosis of the femoral head (SINFH) are still unclear. AIM OF THE STUDY: SINFH remains a challenging problem in orthopedics. Previous investigations have shown that EGb has the potential to reduce the occurrence of SINFH. The goal was to determine the effect and mechanism of EGb in preventing SINFH by inhibiting apoptosis and improving vascular endothelial cells (VECs) functions. MATERIALS AND METHODS: CCK-8, nitric oxide (NO) production and flow cytometry were used to determine the cell apoptosis and function. The scratch and angiogenesis tests assessed migration and tube formation. Western blot analysis detected the expressions of apoptosis-related proteins and PI3K/AKT/eNOS pathway-related proteins. Apoptosis and angiogenesis were also detected treated with the inhibitors. A mouse model of SINFH was established. Paraffin section was used to determine the necrotic pathology and apoptosis. Vessels in the femoral heads were assessed by immunofluorescence staining. RESULTS: When stimulated by methylprednisolone (MPS), cell viability, NO generation and tube formation were decreased, the apoptotic rate increased. Simultaneously, MPS decreased the expression levels of p-PI3K, p-AKT, and p-eNOS. EGb increased the expression levels of these proteins, restrained apoptosis, and restored cell functions. The addition of the inhibitors decreased anti-apoptotic effect and angiogenesis. In addition, when compared to the model mice, there were fewer empty lacunae and normal trabecular arrangement after taking different doses of EGb. The protective effect was also confirmed by the vascular quantitative analysis in vivo. CONCLUSION: This study established that EGb increased endothelial cell activity and inhibited apoptosis and function loss induced by MPS, elucidating the effect and molecular mechanism of EGb on early SINFH.

Lycium barbarum polysaccharide protects against osteonecrosis of femoral head via regulating Runx2 expression.

BACKGROUND: Osteonecrosis of femoral head (ONFH) is a pathological state caused by lack of blood supply in femoral head. This study aimed to explore the function of Lycium barbarum polysaccharide (LBP), an antioxidant agent extracted from L. barbarum, on ONFH. METHODS: Osteonecrosis rat model was generated using lipopolysaccharide (LPS) and methylprednisolone followed by examination of body weight, blood glucose, morphology, and BMSC osteoblast differentiation. The effect and underlying mechanism of LBP on the proliferation, apoptosis, and osteoblast differentiation of BMSC were determined with or without LPS or hypoxia treatment using CCK-8. Alizarin Red S staining, flow cytometry, and western blot, respectively. RESULT: LBP could protect against glucocorticoid-induced ONFH in rats, resulting in improved sparse trabecular bone, empty lacunae and bone cell coagulation. Moreover, LBP promoted the proliferation and osteoblast differentiation of bone mesenchymal-derived stem cells (BMSCs) in a dose-dependent manner. Furthermore, LBP enhanced osteoblast differentiation of BMSCs under hypoxia condition. Mechanistically, we found that LBP treatment enhanced Runx2 and ALP expression in BMSCs. LBP restored the expression of Runx2 and ALP under hypoxia, suggesting that LBP might be involved in regulating Runx2/ALP expression and contributed to osteoblast differentiation. Knockdown of Runx2 significantly inhibited BMSCs proliferation, while LBP treatment did not rescue the osteoblast differentiation ability of BMSCs with Runx2 knockdown. CONCLUSION: Our findings suggested that LBP protects against ONFH via regulating Runx2 expression, which could be utilized to treat patients suffering ONFH.

Curcumin prevents osteocyte apoptosis by inhibiting M1-type macrophage polarization in mice model of glucocorticoid-associated osteonecrosis of the femoral head.

Inflammation is a contributing factor in osteocyte apoptosis, which is strongly associated with the development of glucocorticoid-associated osteonecrosis of the femoral head (GA-ONFH). Curcumin is a naturally derived drug that regulates immunity and inhibits inflammation. This study aimed to examine the capacity of curcumin to prevent osteocyte apoptosis and GA-ONFH, while elucidating possible mechanisms of action. C57/BL6 female mice were divided into control, GA-ONFH, and curcumin-treated GA-ONFH groups. We determined the effect of curcumin on the polarization of RAW264.7 and the apoptosis of MLO-Y4 cells. We found that curcumin reduced the infiltration of M1-type macrophages in the femoral heads and alleviated systemic inflammation in GA-ONFH models. Additionally, curcumin decreased the apoptosis of osteocytes in the femoral heads and the ratio of GA-ONFH in mice. Further, in vitro curcumin intervention inhibited M1-type polarization via the Janus kinase1/2-signal transducer and activator of transcription protein1 (JAK1/2-STAT1) pathway. Taken together, this study demonstrates that curcumin is effective in preventing osteocyte apoptosis and the development of GA-ONFH in a mouse model. Curcumin prevents inflammatory-mediated apoptosis of osteocytes in part through inhibition of M1 polarization through the JAK1/2-STAT1 pathway. These findings provide novel insights as well as a potential preventive agent for GA-ONFH. This article is protected by copyright. All rights reserved.

Yougui pills exert osteoprotective effects on rabbit steroid-related osteonecrosis of the femoral head by activating ß-catenin.

OBJECTIVE: To investigate the effect and underlying mechanism of Yougui pills (YGPs) on steroid-related osteonecrosis of the femoral head (SONFH). METHODS: Male New Zealand white rabbits were divided into three groups: control group, SONFH group and YGPs group. Rabbit SONFH was induced by methylprednisolone (MPS) combined with lipopolysaccharide (LPS). At 6 weeks post induction, the femoral heads were harvested for tissue analyses, including histopathology, mechanical test of femoral heads, micro-CT, tartrate-resistant acid phosphatase (TRAP) staining, and immunohistochemistry for osteocalcin (OCN), vascular endothelial growth factor (VEGF) and ß-catenin. Protein levels of cathepsin K (CTSK), phospho-glycogen synthase kinase-3 beta (p-Ser9 GSK-3ß) and total glycogen synthase kinase-3 beta (GSK-3ß) in femoral heads were also detected. Additionally, the serum TRAP activity was measured using enzyme-linked immunosorbent assay (ELISA). Finally, the effects of YGPs treatment on osteoclast differentiation and osteoblast formation were evaluated in vitro. RESULTS: The ratio of empty lacuna was markedly lower in YGPs group than SONFH group. Micro-CT evaluation indicated that YGPs has a preventive effect on bone loss in rabbit SONFH. YGPs treatment could suppress bone resorption by reducing TRAP+ osteoclast and serum TRACP5b levels in necrotic femoral heads. Moreover, YGPs treatment could promote bone formation by up-regulating the expression of OCN, VEGF and ß-catenin, while increasing load-bearing capacity of femoral heads. Interestingly, p-Ser9 GSK-3ß downregulation, and CTSK upregulation in necrotic femoral head could be reversed by YGPs treatment, which also effectively inhibited RANKL-induced osteoclast differentiation and promoted osteoblast formation in vitro. CONCLUSION: YGPs could suppress osteoclastogenesis and promote bone formation during SONFH in rabbits by activating ß-catenin.

Pharmacological interventions for the prevention of insufficiency fractures and avascular necrosis associated with pelvic radiotherapy in adults.

BACKGROUND: Pelvic radiotherapy is a treatment delivered to an estimated 150,000 to 300,000 people annually across high-income countries. Fractures due to normal stresses on weakened bone due to radiotherapy are termed insufficiency fractures. Pelvic radiotherapy-related interruption of the blood supply to the hip is termed avascular necrosis and is another recognised complication. The reported incidences of insufficiency fractures are 2.7% to 89% and risk of developing avascular necrosis is 0.5%. These complications lead to significant morbidity in terms of pain, immobility and consequently risk of infections, pressure sores and mortality. OBJECTIVES: To assess the effects of pharmacological interventions for preventing insufficiency fractures and avascular necrosis in adults over 18 years of age undergoing pelvic radiotherapy. SEARCH METHODS: We performed electronic literature searches in the Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE, Embase and DARE to 19 April 2017. We also searched trial registries. Further relevant studies were identified through handsearching of citation lists of included studies. SELECTION CRITERIA: Randomised controlled trials (RCTs) or non RCTs with concurrent comparison groups including quasi-RCTs, cluster RCTs, prospective cohort studies and case series of 30 or more participants were screened. We included studies assessing the effect of pharmacological interventions in adults over 18 years of age undergoing radical pelvic radiotherapy as part of anticancer treatment for a primary pelvic malignancy. We excluded studies involving radiotherapy for bone metastases. We assessed use of pharmacological interventions at any stage before or during pelvic radiotherapy. Interventions included calcium or vitamin D (or both) supplementation, bisphosphonates, selective oestrogen receptor modulators, hormone replacement therapy (oestrogen or testosterone), denosumab and calcitonin. DATA COLLECTION AND ANALYSIS: Two review authors independently assessed trial quality and extracted data. We contacted study authors to obtain missing data. Data were to be pooled using the random-effects model if study comparisons were similar, otherwise results were to be reported narratively. MAIN RESULTS: We included two RCTs (1167 participants). The first RCT compared zoledronic acid with placebo in 96 men undergoing pelvic radiotherapy for non-metastatic prostate cancer.The second RCT had four treatment arms, two of which evaluated zoledronic acid plus adjuvant androgen suppression compared with androgen suppression only in 1071 men undergoing pelvic radiotherapy for non-metastatic prostate cancer.Both studies were at a moderate to high risk of bias and all evidence was judged to be of very low certainty.The studies provided no evidence on the primary outcomes of the review and provided limited data in relation to secondary outcomes, such that meta-analyses were not possible. Both studies focused on interventions to improve bone health in relation to androgen deprivation rather than radiation-related insufficiency fractures and avascular necrosis. Few fractures were described in each study and those described were not specific to insufficiency fractures secondary to radiotherapy. Both studies reported that zoledronic acid in addition to androgen deprivation and pelvic radiotherapy led to improvements in BMD; however, the changes in BMD were measured and reported differently. There was no available evidence regarding adverse effects. AUTHORS' CONCLUSIONS: The evidence relating to interventions to prevent insufficiency fractures and avascular necrosis associated with pelvic radiotherapy in adults is of very low certainty. This review highlights the need for prospective clinical trials using interventions prior to and during radiotherapy to prevent radiation-related bone morbidity, insufficiency fractures and avascular necrosis. Future trials could involve prospective assessment of bone health including BMD and bone turnover markers prior to pelvic radiotherapy. The interventions for investigation could begin as radiotherapy commences and remain ongoing for 12 to 24 months. Bone turnover markers and BMD could be used as surrogate markers for bone health in addition to radiographic imaging to report on presence of insufficiency fractures and development of avascular necrosis. Clinical assessments and patient reported outcomes would help to identify any associated adverse effects of treatment and quality of life outcomes.

Research in the precaution of recombinant human erythropoietin to steroid-induced osteonecrosis of the rat femoral head.

Objective To elucidate the effects of recombinant human erythropoietin (rHuEPO) on steroid-induced osteonecrosis of the femoral head in rats. Methods Twenty-four adult Wistar rats were randomly divided into three groups of eight rats each. The rats in the positive control group were injected with dexamethasone at 1 mg/kg twice a week for 5 weeks. The rats in the negative control group were injected with sodium chloride alone. The rats in the experimental group were injected with dexamethasone at 1 mg/kg twice a week for 5 weeks and rHuEPO (500 u/d/kg) daily for 5 weeks. The femoral head on one side was examined by hematoxylin and eosin staining, and that on the other side was examined by CD31 staining of the capillaries. Results Hematoxylin and eosin staining in the positive control group showed that the bony trabeculae had become obviously narrow and sparse with discontinuity of the integrity. The integrity of the trabeculae was better in the experimental group than positive control group. The CD31 expression was lower in the positive control group than in the other two groups. Conclusion rHuEPO can effectively prevent osteocyte apoptosis, delaying or decreasing osteonecrosis of the femoral head.

Lithium prevents rat steroid-related osteonecrosis of the femoral head by ß-catenin activation.

This study explored the use of lithium to prevent rat steroid-related osteonecrosis of the femoral head (ONFH) through the modulation of the ß-catenin pathway. ONFH was induced by methylprednisolone combined with lipopolysaccharide, and serum lipids were analyzed. ONFH was detected by hematoxylin-eosin staining. Micro-CT-based angiography and bone scanning were performed to analyze vessels and bone structure, respectively. Immunohistochemical staining for peroxisome proliferator-activated receptor gamma (PPARγ), bone morphogenetic protein-2 (BMP-2), and vascular endothelial growth factor (VEGF) was analyzed. Protein levels of phospho-glycogen synthase kinase-3ß at Tyr-216 (p-Tyr(216) GSK-3ß), total glycogen synthase kinase-3ß (GSK-3ß) and ß-catenin, as well as mRNA levels of GSK-3ß and ß-catenin in femoral heads, were assessed. The rate of empty bone lacunae in the femoral heads was lower in the lithium and control groups than in the model group. The lithium group showed preventive effects against steroid-related vessel loss by micro-CT-based angiography and VEGF staining. Lithium treatment improved hyperlipidemia and reduced PPARγ expression. Moreover, lithium improved steroid-related bone loss in micro-CT bone scans and BMP-2 staining analyses. Furthermore, local ß-catenin was reduced in steroid-related ONFH, and lithium treatment increased ß-catenin expression while reducing p-Tyr(216) GSK-3ß levels. The local ß-catenin pathway was inhibited during steroid-related ONFH. Lithium may enhance angiogenesis and stabilize osteogenic/adipogenic homeostasis during steroid-related ONFH in rats by activating the ß-catenin pathway.

[Experimental study on preventive effect of Yougui drink on femoral head necrosis in rats under micro CT].

OBJECTIVE: To explore the preventive effect of Yougui drink on femoral head necrosis in rats under micro CT. METHODS: Twenty-five SD rats were divided into steroid hormone group (group A, 10 rats ), Yougui drink group (group B,10 rats) and normal group (group C,5 rats)with random number table. Endotoxin were injected into abdominal cavity of rats in group A and B for 2 days, methylprednisolone sodium succinate were injected by gluteus for twice a week continued for 6 weeks; group B were gavaged by Yougui drink (veryday for 8 weeks; group C did not do any processing. All rats were killed on the 10th weeks,m icro CT were used to scan femoral head in vitro and preventive effect of Yougui drink (n femoral head necrosis in rats. RESULTS: There was statistical significance in BMD, BV/TV, Tb.N, Tb, Th, Thb, Sp, BS/TV and DA but no significance in SMI between group A and B. Comparison between A and C, there was significant meaning in BMD, BV/TV, Tb.N, Tb, Th, Tb, Sp, BS/TV, DA and SMI. CONCLUSION: Yougui drink on femoral head necrosis in rats under micro CT has preventive effect from BMD BV/TV, Tb.N, Tb, Th, Tb, Sp, BS/TV and DA.

[Clinical observation of jianpi bushen formula to prevent collapse of osteonecrosis of femoral head].

To observe the clinical effect of Jianpi Bushen formula to prevent the collapse of osteonecrosis of femoral head (ONFH) of type phlegm and blood stasis obstructing the collaterals. 50 cases (including 73 hips) of non-collapse ONFH (ARCO I, II, III a) were selected from the out-patient department of orthopedic in Guang'anmen Hospital attached to China Academy of Chinese Medical Science. All the cases fit for diagnostic criteria were given Jianpi Bushen formula and followed up. Staging criteria was ARCO classification. Harris score evaluated the hip function. The mean follow-up time was 4.2 years (3-5.4 years). After a mean of 4.2 years (3-5.4 years) followed-up, 12 hips collapse occurred while 61 hips not (the non-collapse rate was 83.56%). Collapse did not occurred among the all 7 hips at stage ARCO I (the non-collapse rate was 100%). Among the 49 hips at stage ARCO II, 9 hips collapse occurred while 40 hips not (the non-collapse rate was 81.63%). Among the 17 hips at stage ARCO III a, 3 hips collapse occurred while 14 hips not (the non-collapse rate was 82.35%). Kaplan-Meier analysis showed the average survival time of non-collapse was 5 years (4. 8-5.2 years). The 3 year survival rate of non-collapse was 92.5%. The 4 year survival rate was 74% and the condition tended to be stability trend. The Harris score was 71.93 +/- 11.25 before treatment and 81.63 +/- 12.16 after treatment, significantly different. These results suggest that: Jianpi Bushen formula is an effective method for treating ONFH of type phlegm and blood stasis obstructing the collaterals. It can delay or prevent the collapse of femoral head and significantly improve the hip function.

[Effect of Epimedium extract on osteoprotegerin and RANKL mRNA expressions in glucocorticoid-induced femoral head necrosis in rats].

OBJECTIVE: To investigate the effect of glucocorticoid on the expression levels of osteoprotegerin (OPG) and receptor activator of nuclear factor kappa B ligand (RANKL) mRNAs in rat femoral head and the antagonistic effect of Epimedium, and explore the mechanism of Epimedium in preventing glucocorticoid-induced femoral head necrosis. METHODS: Forty-eight adult SD rats were randomized into glucocorticoid group, Epimedium group and control group. In the former two groups, the rats received intramuscular injection of 12.5 mg prednisolone twice a week, and in Epimedium group, additional 1 ml/100 g aqueous Epimedium extract (equivalent to 0.1 g/ml of the crude drug) was administered intragastrically once daily. The control group received only intramuscular saline injection. After 4 weeks of treatment, osteonecrosis of the left femoral head was detected by HE staining, and the right femoral head was sampled for detection of OPG and RANKL mRNA expressions using real-time quantitative PCR. RESULTS: In glucocorticoid, Epimedium and control groups, the mortality rate of the rats was 12.5% (2/16), 6.25% (1/16), 0 (0/16), and femoral head necrosis occurred at a rate of 71.43% (10/14), 26.67% (4/15), and 0 (0/16), respectively. In glucocorticoid group, the expression level of OPG mRNA was significantly lower, RANKL expression significantly higher, and OPG/RANKL ratio significantly lower than those in Epimedium and control groups (P<0.05). OPG, RANKL and their ratios showed no significant differences between Epimedium group and the control group. CONCLUSION: Epimedium can prevent glucocorticoid-induced femoral head necrosis probably by antagonizing glucocorticiod-induced abnormal expressions of OPG and RANKL mRNA.

Pentosan reduces osteonecrosis of femoral head in SHRSP.

Increased oxidative stress is considered one of the main causes of steroid-induced osteonecrosis of the femoral head (ONFH). The aim of this study was to evaluate the effects of a steroid hormone and pentosan polysulfate sodium (pentosan), a heparin analog, in stroke-prone spontaneously hypertensive rats (SHRSP) as a model of ONFH. One hundred twenty-three 13-week-old male SHRSP/Izm rats were divided into four groups: a control group (group C), pentosan-administered group (group P), steroid-administered group (group S), and group administered pentosan plus steroid (group PS). Methylprednisolone acetate, as the steroid hormone, at a dose of 4 mg (15 mg/kg) was administered at 15 weeks of age. Pentosan at a dose of 3 mg/day/kg was continuously administered intraperitoneally from 13 weeks of age for 4 weeks. Rats were sacrificed at 17 weeks of age, and heart blood and both femora were collected. Triglyceride levels were significantly lower in group PS than in group S, indicating that pentosan improves lipid metabolism. The incidence of histologic ONFH was significantly lower in group P, at 14.8% (10/71 femoral heads), than in group C, at 30.4% (17/56 femoral heads), and significantly lower in group PS, at 40.8% (29/71 femoral heads), than in group S, at 91.3% (42/46 femoral heads), indicating that pentosan markedly inhibits ONFH. Immunohistochemical staining for oxidative stress showed that the stainability was significantly lower in group PS than in group S. Pentosan seems to reduce the incidence of ONFH in SHRSP by improving lipid metabolism and decreasing oxidative stress.

[Preventing steroid-induced osteonecrosis of the femoral head with Liuwei dihuang pills and molecular mechanism].

OBJECTIVE: To evaluate the effects and the molecular mechanism of Liuwei dihuang pills in preventing steroid-induced osteonecrosis of the femoral head (ONFH) so as to provide an experimental basis for preventing ONFH clinically. METHODS: Thirty-six adult Kunming mice (weighing 40-50 g, 46 g on average) were randomly divided into three groups (n=12): group A (control group), group B (model group) and group C (prevention group). In groups B and C, ONFH mice models were produced by intraperitoneal injection of horse serum at first (10 mL/kg) and a second injection of horse serum intraperitoneally (5 mL/kg) and prednisolone intramuscularly [45 mL/(kg x day), for 5 days] 2 weeks later. At the same time, the mice in group C were given Liuwei dihuang pills intragastrically [2 g/(kg x day)] and were given normal saline [10 mL/(kg x day)] in group B. In group A, mice were given normal saline intramuscularly and intragastrically as controls. The animals were sacrificed 2, 4, and 8 weeks after first treatment with prednisone, and femoral heads and livers were harvested to do histopathology analysis and apoptosis assay. RESULTS: Other mice survived throughout the experiment period except two death at 7 and 11 days after second injection of horse serum intraperitoneally in group B and one death at 24 hours after second injection of horse serum intraperitoneally in group C. The appearance and shape of the femoral head and the surface of cartilages were all normal. The histological observation showed: normal structures of liver and femoral head were seen in group A at each time point; swelling liver cells with small fat vacuole, unclear structure of hepatic cords and narrower sinus hepaticus were seen, the bone trabeculae of femoral head was thin, sparse and collapsed in some regions and the changes became more obvious with time in group B; group C had similar results to group A. The percentage of empty osteocyte lacunae was significantly higher in group B than in groups A and C (P < 0.01). The osteoprotegerin expression significantly decreased and the osteoprotegerin ligand expression significantly increased in group B when compared with groups A and C (P < 0.01). Apoptosis analysis showed that the apoptosis index in group B was significantly higher than that in groups A and C (P < 0.01). CONCLUSION: Liuwei dihuang pills can prevent steroid-induced ONFH by improving lipid metabolism, relieving bone lesion, and protecting against cell death.

Vitamin E prevents steroid-induced osteonecrosis in rabbits.

BACKGROUND AND PURPOSE: Prevention of osteonecrosis after corticosteroid administration would be important. We examined the potential of vitamin E (alpha-tocopherol) to reduce the incidence of corticosteroid-induced osteonecrosis in an animal model. METHODS: Japanese white rabbits were divided into 2 groups; the control group was fed a normal diet and the experimental group was fed alpha-tocopherol-supplemented diet in which alpha-tocopherol (600 mg/kg diet) was added to the normal diet. To induce osteonecrosis, high-dose methylprednisolone acetate (MPSL) (20 mg/kg body weight) was injected once into the right gluteus medius muscle of all rabbits. 4 weeks after the injection of MPSL, the presence or absence of osteonecrosis of bilateral femurs was examined histopathologically. The tocopherol/cholesterol ratios were calculated. The plasma levels of thiobarbituric acid-reactive substances (TBARS) were measured. RESULTS: Alpha-tocopherol-supplemented diet reduced the incidence of osteonecrosis, which developed in 14 of 20 rabbits in the control group and 5 of 21 rabbits in the experimental group (p = 0.004). The tocopherol/cholesterol ratio was higher in the experimental group than in the control group after the alpha-tocopherol administration. The plasma TBARS level was lower in the experimental group than in the control group at 4 weeks after the MPSL administration. INTERPRETATION: Our findings may offer a new approach for the prevention of corticosteroid-induced osteonecrosis.

Effect of rifampicin on the risk of steroid-induced osteonecrosis of the femoral head.

OBJECTIVE: To investigate the effects and possible mechanism of rifampicin on steroid-induced osteonecrosis of the femoral head (ONFH). METHODS: Bone marrow stromal cells (BMSC) separated from male rats were cultured in vitro without any treatment (Group mA), exposed to dexamethasone (Group mB), treated with rifampicin (Group mC), and exposed to dexamethasone and rifampicin simultaneously (Group mD) respectively (n = 5 in each group). After 7 days, P-glycoprotein (P-gp) activity and adipogenesis of the BMSC were evaluated. In an in vivo experiment, 80 rats were randomly divided into 4 groups (n= 20 in each group). Group A received intragastric saline for 5 weeks. Group B received intragastric saline for one week, followed by subcutaneous methylprednisolone and saline for 4 weeks. Group C received intragastric rifampicin for 5 weeks. Group D received intragastric rifampicin for one week, followed by subcutaneous methylprednisolone and rifampicin for 4 weeks. At the end of the experiment, all rats underwent analysis of P-gp activity of BMSC, P-gp expression in the femoral heads, MRI and histomorphometry of the femoral heads. RESULTS: In vitro, the P-gp activity of BMSC increased and lipid accumulation decreased significantly in Group mD, compared to Group mB. In vivo, P-gp activity and P-gp expression in Group D increased compared to Group B. The mean area of MRI abnormal signal, adipocytic variables and apoptotic cells in Group D decreased, mean percentage of the whole epiphysis made up by the epiphyseal ossification center and trabecular structure variables improved compared to those in Group B. The incidence of ONFH was lower in Group D (50%) than in Group B (80%). CONCLUSION: Rifampicin may decrease the risk of steroid-induced ONFH by enhancing P-gp activity, thus preventing steroid-induced BMSC adipogenesis.

Electromagnetic fields: a novel prophylaxis for steroid-induced osteonecrosis.

Establishing a means to prevent osteonecrosis after corticosteroid administration is an important theme. We asked whether pulsed electromagnetic field stimulation, a noninvasive treatment, could prevent osteonecrosis. Ninety rabbits were divided into four treatment groups: (1) exposure of 10 hours per day to electromagnetic stimulation for 1 week, followed by injection of methylprednisolone (20 mg/kg), and exposure of 10 hours per day to electromagnetism for a further 4 weeks (n = 40); (2) methylprednisolone injection only (n = 40); (3) no treatment (n = 5); and (4) exposure of 10 hours per day to electromagnetism for 5 weeks (n = 5). After 5 weeks, we harvested and histologically examined femurs bilaterally. The frequency of osteonecrosis was lower in the steroid-electromagnetism group (15/40) than in the steroid-only group (26/40). No necrotic lesions were found in the two control groups. We observed no clear effects of electromagnetism on the number, location, extent, and repair of necrotic lesions and intramedullary fat cell size in affected rabbits. Pulsed electromagnetic field stimulation reportedly augments angiogenesis factors and dilates blood vessels; these effects may lower the frequency of osteonecrosis. Exposure to pulsed electromagnetic field stimulation before corticosteroid administration could be an effective means to reduce the risk of osteonecrosis.

Preventive effects of puerarin on alcohol-induced osteonecrosis.

Alcohol can induce adipogenesis by bone marrow stromal cells and may cause osteonecrosis of the femoral head. Currently, there are no medications available to prevent alcohol-induced osteonecrosis. We hypothesized puerarin, a Chinese herbal medicine with antioxidative and antithrombotic effects, can prevent alcohol-induced adipogenesis and osteonecrosis. Both bone marrow stromal cells (in vitro) and mice (in vivo) were treated either with ethanol or with ethanol and puerarin, with an untreated group serving as a control. In the in vitro study, the number of adipocytes, contents of triglycerides, and levels of PPAR gamma mRNA expression were decreased and alkaline phosphatase activity, contents of osteocalcin, and levels of osteocalcin mRNA expression were increased in cells treated with both alcohol and puerarin, compared with cells treated with alcohol only. In the in vivo study, marrow necrosis, fat cell hypertrophy and proliferation, thinner and sparse trabeculae, diminished hematopoiesis, and increased empty osteocyte lacunae in the subchondral region of the femoral head were observed in mice treated with alcohol. However, no such changes were seen in femoral heads of mice treated with alcohol and puerarin. The data suggest puerarin can inhibit adipogenic differentiation by bone marrow stromal cells both in vitro and in vivo and prevents alcohol-induced osteonecrosis in this model.

Prevention of corticosteroid-induced osteonecrosis in rabbits by intra-bone marrow injection of autologous bone marrow cells.

OBJECTIVES: Femoral head osteonecrosis (ON) is a serious complication of steroid administration. We evaluated bone marrow transplantation (BMT) for preventing corticosteroid-induced ON. METHODS: Rabbits, injected with methylprednisolone (MPSL; 20 mg/kg), were divided into four groups: (i) MPSL alone; MPSL injection only, (ii) MPSL+needling; 2 days after MPSL injection, a hole (1.2 mm diameter) was drilled from the outer cortex 2.5 cm distal to the proximal end of the greater trochanter, (iii) MPSL+saline; 2 days after MPSL injection, 2 ml saline was injected directly into the bone marrow cavity, and (iv) MPSL+BMT; 2 days after MPSL injection, 1 x 10(7)/2 ml bone marrow cells (BMCs) were injected directly into the bone marrow cavity. Platelets, fibrinogen, prothrombin time and total cholesterol in peripheral blood were measured before and after treatment. Tissues were stained with haematoxylin and eosion and terminal deoxynucleotidyl-mediated deoxyuridine triphosphate nick-end labelling stain and immunostained for VEGF, while cell proliferation and viability of whole BMCs in the femur were analysed by cell cycle analysis and [(3)H]-thymidine uptake. RESULTS: The ON incidence in rabbits treated with MPSL alone, MPSL+needling and MPSL+saline was 72.7, 70.0 and 66.7%, respectively, while in the MPSL+BMT group, the incidence was 0%. Serological findings in the MPSL+BMT group were almost normalized. VEGF and TUNEL staining were reduced in the MPSL+BMT group compared with all other groups. There were significantly fewer BMCs in G1 phase from the MPSL+BMT group than the other groups, while uptake of [(3)H]-thymidine was significantly increased. CONCLUSION: Direct injection of autologous BMCs into femurs prevents corticosteroid-induced ON following treatment with high-dose, short-term steroids.

[Effect of gufuhuotang on hemorheology and lipid metabolism of hormonal necrosis of femoral head].

OBJECTIVE: To probe the of Gufuhuotang effect on hemorheology and lipid metabolism of hormonal necrosis of femoral head. METHOD: 32 male rabbits of New Zealand were divided at random into pathological pattern group, normal control group, low-dosage of Gufuhuotang group and high-dosage group; models of osteocyte necrosis of head of femur were made with intragluteal injection of hydroprednisone acetate; the rabbits were observed weekly and killed 6 weeks later and HE pathological section was made to observe hemorheology index and blood-liquid content, and then analysis and comparison were made. RESULT: Gufuhuotang could obviously decrease total blood low-shear viscosity, plasma viscosity, hemacocrit, serum cholesterol and triglyceride contents; under light microscope empty bone lacunas of femoral head were less, fat cells of medullary cavity were less and relatively small, and bone trabecula was sparse with no break. CONCLUSION: Gufuhuotang has a distinct preventive and curative effect on rabbits' hormonal necrosis of femoral head.