Glucocorticoid-induced Osteonecrosis Of The Femoral Head Research Articles

Enhanced bone defect repairing effects in glucocorticoid-induced osteonecrosis of the femoral head using a porous nano-lithium-hydroxyapatite/gelatin microsphere/erythropoietin composite scaffold.

Glucocorticoid-induced osteonecrosis of the femoral head (GIONFH) is a common debilitating disease that occurs in young and middle-aged adults. To treat early GIONFH, core decompression and bone graft are regarded as effective measures. However, the ideal bone graft should possess bioactivity as well as biomechanical properties. The most commonly used bone graft materials are currently unsatisfactory. In this study, we fabricated a composited scaffold using lithium (Li) to activate the Wnt signal pathway and erythrogenin (EPO) to upregulate the HIF-1/VEGF pathway to improve the osteogenic and angiogenic effects of the scaffold. We obtained the porous gelatin/nano-lithium-hydroxyapatite/gelatin microsphere/rhEPO (Li-nHA/GMs/rhEPO) composited scaffold and assessed its mechanical properties, release properties, and in vitro bioactivity. Then, we implanted the scaffold into the femoral heads of GIONFH rabbits after core decompression surgery and evaluated the osteogenic and angiogenic abilities of the scaffold in vivo as well as its bone defect repair efficacy. As the results show, the Li-nHA/GM/rhEPO scaffold possessed good mechanical compression strength and enabled continuous release of Li and rhEPO. Moreover, the scaffold improved the viability of glucocorticoid-treated BMMSCs and vascular endothelial cells and increased the expression of osteogenic and angiogenic factors. In the in vivo study, the composited scaffold improved new bone formation and exerted effects on repairing femoral head defects in GIONFH rabbits. Additionally, the osteogenic and angiogenic factors were increased along with the activation of factors in the Wnt signal pathway and the HIF-1/VEGF pathway. In conclusion, the Li-nHA/GM/rhEPO scaffold can upregulate the Wnt and HIF-1/VEGF pathways at same time and has effects on improving osteogenesis and angiogenesis, which benefits the repair of GIONFH.

Icariin Protects against Glucocorticoid-Induced Osteonecrosis of the Femoral Head in Rats

Background/Aims: Glucocorticoid (GC)-related osteonecrosis of the femoral head (ONFH) is a common complication following administration of steroids to treat many diseases. Our previous study demonstrated that icariin (ICA) might have a beneficial effect on the bone marrow mesenchymal stem cells (BMSCs) of patients with steroid-associated osteonecrosis. In this study, we investigated the underlying mechanisms of ICA associated with the potential enhancement of osteogenesis and anti-adipogenesis in GC-related ONFH. Methods: In vitro cell proliferation was evaluated by CCK-8 assay. Alizarin red S and alkaline phosphatase (ALP) activity were used to measure osteogenic differentiation, while adipogenic differentiation was revealed by oil red O staining and TG content assay. The expression level of osteogenesis-associated genes and PPARγ was evaluated by RT-qPCR, western blotting and immunofluorescence. A total of 30 female SD rats were randomly separated into three groups: a control group, a methylprednisolone (MPS) group and a MPS + ICA group. Serum ALP and TG (triglyceride), micro-CT scanning, histological and immunohistochemical analyses were performed in the animal model. Results: In the in vitro study, ICA promoted proliferation, improved osteogenic differentiation and suppressed adipogenic differentiation of BMSCs treated with MPS. The group treated with MPS and 10^-6 M ICA expressed higher levels of Runx2, ALP, bone morphogenetic protein (BMP) 2, and OC and lower expression of PPARγ than the MPS group. In the in vivo study, ICA prevented bone loss in a rat model of GC-related ONFH as shown by micro-CT scanning, histological and immunohistochemical analyses. Conclusions: ICA is an effective compound for promoting bone repair and preventing or delaying the progression of GC-associated ONFH in rats. This effect can be explained by its ability to improve the balance between adipogenesis and osteogenesis, indicating that ICA is an effective candidate for management of GC-associated ONFH.

The Pathophysiological Sequence of Glucocorticoid-Induced Osteonecrosis of the Femoral Head in Male Mice.

In search of the sequence of pathogenic events leading to glucocorticoid-induced osteonecrosis, we determined the molecular, biomechanical, cellular, and vascular changes in the femur of C57BL/6 mice receiving prednisolone for 14, 28, or 42 days. The femoral head, but not the distal femur, of mice treated for 14 days showed a decrease in the expression of the hypoxia-inducible factor (Hif)-1α and vascular endothelial growth factor (VEGF), the number of osteoblasts, and bone formation rate and strength and showed an increase in osteoclasts. These changes were accompanied by conversion of the normal dendritic vasculature to pools of edema as detected by magnetic resonance imaging, providing robust diagnostic evidence of early osteonecrosis. At that time point, there were no detectable changes in bone density, cortical or cancellous bone architecture, midshaft or distal cancellous bone, or osteocyte apoptosis. In mice treated for 28 days, femoral head cancellous density, cortical width, and trabecular thickness decreased, and by 42 days the femoral heads had full-depth cortical penetrations and cancellous tissue osteonecrosis. These results indicate that the femoral head is a particularly sensitive anatomical site to the adverse effects of glucocorticoid excess on bone and that decreases of Hif-1α and VEGF expression, bone vascularity, and strength precede the loss of bone mass and microarchitectural deterioration, thus rendering the femoral head vulnerable to collapse.

Parathyroid Hormone (PTH) Induces Autophagy to Protect Osteocyte Cell Survival from Dexamethasone Damage.

BackgroundGlucocorticoids (GC) have direct adverse effects on osteocytes, the most abundant bone cell type, and play an important role in osteonecrosis of the femoral head (ONFH). Teriparatide has been reported to be an effective treatment for ONFH. However, the underlying mechanism is unclear.Material/MethodsAn osteocyte cell line, MLO-Y4, was used under various doses of dexamethasone (Dex) with or without rhPTH (1–34). Cell viability, autophagy, and apoptosis markers and osteocyte characteristic mRNAs were investigated to better understand this phenomenon.ResultsInduction of apoptosis by Dex was increased in a time- and dose-dependent manner in MLO-Y4 cells. Autophagy markers (LC3-II and Beclin-1) were increased at the low dose of Dex (10−7 or 10−6 M) and decreased at the high dose (10−5 M). In MOL-Y4 cells, rhPTH (1–34) was shown to be protective against Dex-induced apoptosis. The upregulation of LC3-II and Beclin-1 and decreased level of Caspase-3 was observed in the rhPTH (1–34)-treated group compared with the Dex-only-treated group. Furthermore, the changes induced by Dex in osteocytes, such as increased SOST, RANKL, and DMP-1 mRNA level and decreased Destrin mRNA level, were reversed by rhPTH (1–34). A similar result was found in osteocyte-specific proteins sclerostin expression encoded by SOST mRNA, which acted as a bone formation inhibitor.ConclusionsThe self-activation of autophagy may be a protective mechanism against apoptosis induced by Dex. The protection effect of rhPTH (1–34) for GC-induced ONFH thus results, at least in part, from enhanced autophagy.

Regulatory effect of microRNA-34a on osteogenesis and angiogenesis in glucocorticoid-induced osteonecrosis of the femoral head.

Glucocorticoid-induced osteonecrosis of the femoral head (GIOFH) is a common and devastating orthopedic disease, and its underlying mechanism remains unclear. The aim of this study was to determine the role of microRNA-34a (mir-34a) in GIOFH. C57 mouse mesenchymal stem cells (mMSCs) and human umbilical vein endothelial cells (HUVECs) were cultured with dexamethasone (Dex). A total of 48 adult rats were treated with glucocorticoids, and after the onset of GIOFH, each femoral head was removed. Mir-34a mimics, an inhibitor and over-expressing lentivirus were used in vitro and in vivo, respectively. Real-time PCR, immunohistochemistry, ELISA, cell proliferation assays, osteoblastic differentiation, and endothelial activity assays were employed to evaluate the effect of mir-34a on mMSCs, osteoblasts, and vascular endothelial cells in glucocorticoid-treated mice. We found that Dex inhibited mMSC proliferation and osteoblastic differentiation, as well as the viability and activity of endothelial cells. Dex also caused osteonecrosis and decreased new vessel formation in vivo. Mir-34a alleviated the inhibitory effects of Dex on mMSCs and osteoblasts, while facilitating its inhibitory effects on endothelial cells. Mir-34a is an important regulator in osteogenesis and angiogenesis, and it might be useful as a therapeutic target for GIOFH. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:417-424, 2018.

Substrain-specific differences in bone parameters, alpha-2-macroglobulin circulating levels, and osteonecrosis incidence in a rat model.

Osteonecrosis of the femoral head (ONFH) is a potentially devastating complication that occurs in up to 40% of young adults receiving chronic glucocorticoid (GC) therapy. Through a validated GC therapy rat model, we have previously shown that Wistar Kyoto (WK) rats exhibit a genetic susceptibility to GC-induced ONFH compared to Sasco Fischer (F344) rats. We have undertaken this study in order to investigate differences between these two strains for their bone parameters, alpha-2-macroglobulin (A2M) circulating levels and incidence of GC-induced osteonecrosis of the femoral head. WK and F344 rats were treated either with 1.5 mg/kg/day of prednisone or placebo for 6 months. Blood was taken every month. The femoral heads were harvested for histological examination to detect ONFH and analyzed with micro-computed tomography. After 3 months of GC-therapy, plasma A2M was elevated in treated rats only. GC-treated WK rats exhibited histological evidence of early ONFH through higher rates of cellular apoptosis and empty osteocyte lacunae in the subchondral bone compared to placebos and to F344 rats. Furthermore, micro-CT analysis exhibited femoral head collapse only in GC-treated WK rats. Interestingly, GC-treated F344 rats exhibited significant micro-CT changes, but such changes were less concentrated in the articular region and were accompanied histologically with increased marrow fat. These µCT and histological findings suggest that elevated A2M serum level is not predictive and suitable as an indicative biomarker for early GC-induced ONFH in rodents. Elevated A2M levels observed during GC treatment suggests that it plays role in the host reparative response to GC-associated effects. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1183-1194, 2017.

Role of coagulopathy in glucocorticoid-induced osteonecrosis of the femoral head.

The two major theories of glucocorticoid (GC)-induced osteonecrosis of the femoral head (ONFH) are apoptosis and ischaemia. The traditional theory implicates ischaemia as the main aetiological factor because the final common pathway of ONFH is interruption of blood supply to the bone. The most common causes of interruption of blood supply include fat embolism and coagulation disorders. GCs can directly or indirectly lead to coagulation disorders, producing a hypercoagulable state, followed by poor blood flow, ischaemia, and eventually ONFH. This review summarizes the existing knowledge on coagulation disorders in the context of GC-induced ONFH, including hypofibrinolysis and thrombophilia, endothelial cell dysfunction and damage, endothelial cell apoptosis, lipid metabolism, platelet activation, and the effect of anticoagulant treatment.

Vitamin K2 Ameliorates Damage of Blood Vessels by Glucocorticoid: a Potential Mechanism for Its Protective Effects in Glucocorticoid-induced Osteonecrosis of the Femoral Head in a Rat Model

Glucocorticoid has been reported to decrease blood vessel number and harm the blood supply in the femoral head, which is recognized to be an important mechanism of glucocorticoid-induced osteonecrosis of the femoral head (ONFH). To prevent glucocorticoid-induced ONFH, medication that promotes both bone formation and angiogenesis would be ideal. Vitamin K2 has been revealed to play an important role in bone metabolism; however, few studies have focused on the effect of Vitamin K2 on new vascular formation. Thus, this study aimed to investigate whether Vitamin K2 promoted new blood vessel formation in the presence of glucocorticoids, both in vitro and in vivo. The effect of Vitamin K2 on viability, migration, in vitro tube formation, and VEGF, vWF, CD31, KDR, Flt and PDGFB in EAhy926 incubated with or without dexamethasone were elucidated. VEGF, TGF-β and BMP-2, angiogenesis-related proteins secreted by osteoblasts, were also detected in the osteoblast-like cell line of MG63. In addition, blood vessels of the femoral head in rats administered with or without methylprednisolone and Vitamin K2 were evaluated using angiography and CD31 staining. In vitro studies showed that Vitamin K2 significantly protected endothelial cells from dexamethasone-induced apoptosis, promoted endothelial cell migration and in vitro tube formation. Angiogenesis-related proteins both in EAhy926 and MG63 were also upregulated by Vitamin K2 when cotreated with dexamethasone. In vivo studies showed enhanced blood vessel volume and CD31-positive staining cells in rats cotreated with VK2 and methylprednisolone compared to rats treated with methylprednisolone only. Collectively, Vitamin K2 has the ability to promote angiogenesis in vitro and to ameliorate vessels of the femoral head in glucocorticoid-treated rats in vivo, indicating that Vitamin K2 is a promising drug that may be used to prevent steroid-induced ONFH.

Exosomes from Human Synovial-Derived Mesenchymal Stem Cells Prevent Glucocorticoid-Induced Osteonecrosis of the Femoral Head in the Rat.

Osteonecrosis of the femoral head (ONFH) represents a debilitating complication following glucocorticoid (GC)-based therapy. Synovial-derived mesenchymal stem cells (SMSCs) can exert protective effect in the animal model of GC-induced ONFH by inducing cell proliferation and preventing cell apoptosis. Recent studies indicate the transplanted cells exert therapeutic effects primarily via a paracrine mechanism and exosomes are an important paracrine factor that can be directly used as therapeutic agents for tissue engineering. Herein, we provided the first demonstration that the early treatment of exosomes secreted by human synovial-derived mesenchymal stem cells (SMSC-Exos) could prevent GC-induced ONFH in the rat model. Using a series of in vitro functional assays, we found that SMSC-Exos could be internalized into bone marrow derived stromal cells (BMSCs) and enhance their proliferation and have anti-apoptotic abilities. Finally, SMSC-Exos may be promising for preventing GC-induced ONFH.

Vitamin K2 Prevents Glucocorticoid-induced Osteonecrosis of the Femoral Head in Rats.

Glucocorticoid medication is one of the most common causes of atraumatic osteonecrosis of the femoral head (ONFH), and vitamin K2 (VK2) has been shown to play an important and beneficial role in bone metabolism. In this study, we hypothesized that VK2 could decrease the incidence of glucocorticoid-induced ONFH in a rat model. Using in vitro studies, we investigated how bone marrow-derived stem cells in the presence of methylprednisolone proliferate and differentiate, specifically examining osteogenic-related proteins, including Runx2, alkaline phosphatase and osteocalcin. Using in vivo studies, we established glucocorticoid-induced ONFH in rats and investigated the preventive effect of VK2. We employed micro-CT scanning, angiography of the femoral head, and histological and immunohistochemical analyses, which demonstrated that VK2 yielded beneficial effects for subchondral bone trabecula. In conclusion, VK2 is an effective antagonist for glucocorticoid on osteogenic progenitors. The underlying mechanisms include acceleration of BMSC propagation and promotion of bone formation-associated protein expression, which combine and contribute to the prevention of glucocorticoid-induced ONFH in rats.

Association of ABCB1/MDR1 polymorphisms in patients with glucocorticoid-induced osteonecrosis of the femoral head: Evidence for a meta-analysis

PurposeGlucocorticoid-induced osteonecrosis of the femoral head (GC-induced ONFH) is a rebarbative disease affecting people from all ages, especially young adults, and often leads to severe joint pain and limitations on physical activity. Numerous studies have reported that ABCB1 polymorphisms are associated with GC-induced ONFH, but the results are inconclusive, partially because the sample size of published studies is relatively small. Therefore, we performed a meta-analysis including seven case–control studies to estimate such association. MethodsPublished literature from Medline, Embase, and CNKI were searched for eligible publications. Pooled odds ratio (OR) together with their 95% confidence (CI) was calculated using a fixed effect model or random effect model. The meta-analysis was performed in accordance to PRISMA Statement Criteria. ResultsThe ABCB1 3435T allele reduces the GC-induced ONFH risk based on the evidence from the co-dominant model (CT vs. CC, OR=0.73, 95% CI: 0.53–1.00; TT vs. CC, OR=0.43, 95% CI: 0.26–0.69), dominant model (CT+TT vs. CC, OR=0.64, 95% CI: 0.48–0.87), allele contract model, (T vs. C, OR=0.68, 95% CI: 0.54–0.84), and recessive model (TT vs. CC+CT, OR=0.52, 95% CI: 0.34–0.81). Similarly, the ABCB1 2677T/A allele reduce the GC-induced ONFH risk based on the evidence from the co-dominant model (GT/A vs. GG, OR=0.66, 95% CI: 0.45–0.96; T/AT/A vs. GG, OR=0.52, 95% CI: 0.34–0.82), dominant model (GT/A+T/AT/A vs. GG, OR=0.61, 95% CI: 0.43–0.87), and allele contract model (T/A vs. G, OR=0.73, 95% CI: 0.58–0.90). ConclusionsThe meta-analysis revealed that 3435T allele and ABCB1 2677T/A allele may decrease the risks of GC-induced ONFH.

Administration of erythropoietin exerts protective effects against glucocorticoid-induced osteonecrosis of the femoral head in rats.

Accumulating evidence has indicated that erythropoietin (EPO) plays a role in anti-apoptosis and tissue protection in a number of human diseases. The present study was implemented to evaluate these anti-apoptotic and tissue-protective effects in glucocorticoid-induced osteonecrosis in rats. Osteonecrosis was induced by low-dose lipopolysaccharide and subsequent high-dose methylprednisolone pulse. Rats in the preventive group were treated with 500 U/kg/day recombinant human EPO (rhuEPO) for 1 week. Hematological and histomorphometric methods were then used to determine the effects of the administration of rhuEPO. An analysis of trabecular bone architecture was performed to evaluate bone mass change in the osteonecrosis zone. Terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay was performed to determine the apoptotic index of osteoblasts and osteocytes. Immunoblot analysis was performed to assess the expression of caspase-3 and vascular endothelial growth factor (VEGF) in the femoral head. Treatment with rhuEPO greatly improved the histological performance. Additionally, the incidence of osteonecrosis markedly decreased in the rats in the rhuEPO-treated group (22.2%) compared with the control group (66.7%). Furthermore, the expression of caspase-3 markedly decreased in the rhuEPO-treated group. Consistently, the apoptosis of osteoblasts and osteocytes, as determined by TUNEL assays, was inhibited following the administration of rhuEPO. By contrast, the expression of VEGF increased in the osteonecrosis zone in the rats treated with rhuEPO. The results from the present study demonstrate that EPO exerts prominent protective effects against glucocorticoid-induced osteonecrosis of the femoral head in rats by inhibiting the apoptosis of osteoblasts and osteocytes and increasing the expression of VEGF.

Pravastatin enchances the osteogenic process to the glucocorticoid-induced osteonecrosis of the femoral head

Glucocorticoid (GC)-induced osteonecrosis of the femoral head (ONFH) is a disabling bone disease and it affects a lot of patients in the world widely. To investigate the effect of pravastatin on the early stage of GC-induced ONFH, white rabbits were injected twice with lipopolysaccharide (LPS) at 10 ug/kg each and three times with methyprednisolone (MPS) at 20 mg/kg each to the constructedONFH model. The model groups were randomly divided into groups B and C. Group C was treated with pravastatin from the 5th week. Six rabbits in each group were tested; the blood lipids of 4, 8, 12 and 16 weeks. The rabbits were sacrificed at the 8, 12 and 16 weeks after the blood test. Both histopathologic and molecular studies were performed. The blood lipids levels of cholesterol (CHOL), triglyceride (TG) and high-density lipoprotein (HDL) changed significantly in the model group. After the continuously treatment of pravastatin, the blood lipids levels recovered significantly in group C were compared with those of group B. Necrosis was reduced and new bone repair increased; the repair of ONFH was enhanced in group C. The results suggested that, pravastatin was useful to treat the GC-induced ONFH in the early stage. Key words: Bone morphogenetic protein-2 (BMP2), core binding factor a1 (Cbfa1), glucocorticoid, osteonecrosis of the femoral head, pravastatin, vascular endothelial growth factor (VEGF).