Femoral Head Necrosis In Rats Research Articles

Effect of Genetically Engineered Drugs on Aseptic Femoral Head Necrosis in Rats

Effect of Genetically Engineered Drugs on Aseptic Femoral Head Necrosis in Rats

Extracellular vesicles from human umbilical cord mesenchymal stem cells prevent steroid-induced avascular necrosis of the femoral head via the PI3K/AKT pathway.

Extracellular vesicles (EVs) secreted by human umbilical cord mesenchymal stem cells (hucMSC) have excellent therapeutic potential for many diseases. The aim of this study was to define the role of hucMSC-EVs in the prevention and treatment of steroid-induced avascular necrosis of the femoral head (SANFH). After establishing the SANFH rat model, the effects of hucMSC-EVs were assessed by measuring the microstructure of the femoral head using HE staining, micro-computed tomography (micro-CT), and TUNEL staining. The administration of hucMSC-EVs caused a significant reduction to glucocorticoids (GCs)-induced osteoblast apoptosis and empty lacuna of the femoral head, while effectively improving the microstructure. HucMSC-EVs rescued the deactivation of the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) signaling pathway induced by GCs, and reversed the proliferation and migration of osteoblasts inhibited by GCs. In addition, hucMSC-EVs attenuated the inhibitory effects of GCs on rat osteoblast osteogenesis, angiogenesis of endothelial cells, and prevented osteoblast apoptosis. However, the promoting effects of hucMSC-EVs were abolished following the blockade of PI3K/AKT on osteoblasts. hucMSC-EVs were found to prevent glucocorticoid-induced femoral head necrosis in rats through the PI3K/AKT pathway.

Expression Dynamics of Bone Homeostasis Genes in the Development of Aseptic Femoral Head Necrosis in Rats

Expression Dynamics of Bone Homeostasis Genes in the Development of Aseptic Femoral Head Necrosis in Rats

A novel model of traumatic femoral head necrosis in rats developed by microsurgical technique

BackgroundClinical angiography and vascular microperfusion confirmed that the femoral head retains blood supply after a collum femur fracture. However, no animal model accurately mimics this clinical situation. This study was performed to establish a rat model with retained viability of the femoral head and partial vasculature deprivation-induced traumatic caput femoris necrosis by surgery.MethodsThirty rats were randomly divided into three groups (n = 10 per group): normal group, sham-operated group (Control), and ischemic osteonecrosis group. The femoral head of the normal group of rats underwent a gross anatomy study and microangiography to identify femoral head blood supply. Microsurgical techniques were used to cauterize the anterior-superior retinacular vessels to induce osteonecrosis. Hematoxylin and Eosin (H&E) staining were used for femoral head histologic assessment. Morphologic assessments of the deformity in and trabecular bone parameters of the femoral head epiphysis were performed using micro-CT.ResultsThe blood supply of the femoral head in rats primarily came from the anterior-superior, inferior, and posterior retinacular arteries. However, anterior-superior retinacular vasculature deprivation alone was sufficient in inducing femoral head osteonecrosis. H&E showed bone cell loss in nuclear staining, disorganized marrow, and trabecular structure. The bone volume (BV) decreased by 13% and 22% in the ischemic group after 5 and 10 weeks, respectively. The mean trabecular thickness (Tb.Th) decreased from 0.09 to 0.06 mm after 10 weeks. The trabecular spacing (Tb.Sp) increased from 0.03 to 0.05 mm after 5 weeks, and the epiphyseal height-to-diameter (H/D) ratio decreased.ConclusionsWe developed an original and highly selective rat model that embodied femoral head traumatic osteonecrosis induced by surgical anterior-superior retinacular vasculature deprivation.

The effect of Moringa oleifera polysaccharides on the regulation of glucocorticoid-induced femoral head necrosis: In vitro and in vivo

One of the common challenges in using glucocorticoid in the long term is the development of femoral head necrosis. To address this challenge, the use of glucocorticoid suppressors like plant polysaccharides has been considered. In this study, Moringa oleifera polysaccharide was isolated through hot water–ethanol precipitation method and purified by DEAE-Sepharose fast flow column. Then, they were characterized by FT-IR, NMR, methylation, and chromatography assays. The polysaccharide biocompatibility was investigated by MTT assay and its effect on osteoblasts was evaluated by controlling gene expression. Also, the effect of polysaccharide on dexamethasone-induced femoral head necrosis in rats was assessed by hydroxyproline, hexosamine and morphometric parameters. The results show that 2 Da molecular weight polysaccharide is mainly composed of Rha, Ara, Fru, Xyl, Man and Gal in the molar ratio of 1.7:2.1:3.4:5.9:5.8:1.3. Meanwhile, MTT results on osteoblasts cells showed polysaccharide biocompatibility, while significantly reducing the negative effects of glucocorticoid. Likewise, polysaccharide significantly reduced the levels of apoptosis and intracellular ROS of glucocorticoid-induced femoral necrosis. Moreover, the results of gene expression indicated a decrease in the expression of TNF-α and IL-6 genes using polysaccharide, which is very effective in preventing apoptotic activity. Also, Polysaccharide increased bone density, bone volume per tissue volume, trabecular thickness, and the hexosamine to hydroxyproline ratio in the rat serum in the presence of glucocorticoids, which are very effective in the process of femoral head necrosis. Furthermore, polysaccharide significantly increases the OCN, RUNX2 and COL-1 genes expression in cartilage tissue, which is in line with the result of morphometric parameters. Overall, this study suggests that the use of polysaccharide could result in the treatment of femoral head necrosis.

Glucocorticoids induce femoral head necrosis in rats through the ROS/JNK/c-Jun pathway.

Osteonecrosis of the femoral head (ONFH) is a common clinical disease with a high disability rate. Apoptosis of osteoblasts caused by high-dose short-term or low-dose long-term glucocorticoid (GC) administration is the biological basis of steroid-induced avascular necrosis of the femoral head (SANFH). The pathogenesis of SANFH has not yet been fully elucidated, and there is currently a lack of effective clinical treatments. Here, we investigated the role of the reactive oxygen species (ROS)/JNK/c-Jun signaling pathway in SANFH. Dexamethasone (Dex) was used to induce apoptosis in osteoblasts, and this resulted in a significant increase in levels of p-JNK, p-c-Jun, Bax, caspase-3, caspase-9, cytochrome C, Beclin-1, and LC3, and a decrease in levels of P62 and Bcl-2. In addition, intracellular ROS levels were increased and mitochondrial membrane potential was decreased. Administration of 3-MA, an autophagy inhibitor, attenuated Dex-mediated changes in autophagy and apoptosis. A rat model of ONFH exhibited severe bone trabecular hollow bone pits along with a significant increase in femoral head cell apoptosis compared with the control group. Additionally, micro-CT analysis showed that both bone tissue content and femoral head integrity were significantly reduced in the ONFH group. Furthermore, 3-MA treatment decreased the effect of Dex on GC-induced ONFH and osteoblast apoptosis in rats and could counteract microstructure destruction due to femoral head necrosis. In summary, our data suggest that GC can induce osteoblast apoptosis and autophagy through the ROS/JNK/c-Jun signaling pathway, which contributes to ONFH.

Effects of erythropoietin for precaution of steroid-induced femoral head necrosis in rats

BackgroundSteroids such as glucocorticoid have been widely used for their excellent anti-inflammatory, anti-immune, and anti-shock properties. However, the long-term use in high doses has been found to cause necrosis of femoral head and other serious adverse reactions. Thus, it is of great importance to safely use these medications on patients without inducing bone necrosis.MethodsIn this preclinical study, we examined the effects of erythropoietin (EPO) to attenuate the induction of steroid-induced femoral bone necrosis using rats to build up the in-vivo models. Rats were randomly divided into three groups: negative control group (group A), disease group (group B), and EPO group (group C). 20 mg/kg methylprednisolone was administrated into group B and group C for 6 weeks with two intramuscular injections per week per rat. Group C was further given daily intraperitoneal injections of rHuEPO during this period. Group A received only injection of saline at the same schedule. 12 weeks after the initial drug administration, the rats’ femoral tissues were harvested for HE staining, immunohistochemistry studies for PECAM-1(also CD31) expression and Western Blotting for VEGF expression.ResultsHistology studies showed that compared with the disease group, EPO group had significant improvement and bone morphology being much closer to the negative control group. Immunohistochemical studies revealed that EPO group had statistically much more expression of PECAM-1 than the other groups did. Western Blot demonstrated that the EPO group had significantly higher VEGF expression than the disease group.ConclusionResults suggested that simultaneous injection of EPO could partially prevent steroid-induced ANFH.

NEL-like type 1 gene transfection for prevention of traumatic femoral head necrosis in rats

Objective To determine the effect of NEL-like type 1 gene (NELL-1) transfection in vivo in the repair of traumatic femoral head necrosis. Methods Twenty-four SD rats were randomly divided into three groups (8 rats per group) according to the lottery method, ie, sham group (served as normal control), NELL-1 treatment group (injected NELL-1 gene by recombinant adenovirus vectors around the hip one week after osteonecrosis model induced surgically) and placebo group (given an equal volume of saline solution at the same time after the induction of osteonecrosis). Femurs were taken from the animals 5 weeks after surgery. Gross observation was performed for morphology changes, X-ray assessment for femoral head height and length ratio (H/L), Micro-CT measure for bone parameters of femoral head including total volume (TV), bone volume (BV), total mineralized content (TMC), trabecular thickness (Tb.Th) and trabecular space (Tb.SP), and histological study for osteocytes, osteoblasts and osteoclasts. Results Preserved femoral head shape was noted in NELL-1 treatment group compared to the obvious flattening of the femoral head in placebo group. No heterotopic osteogenesis was observed in any group. Femoral head H/L ratio for 0.753 2±0.040 2 in NELL-1 treatment group was higher than 0.598 4±0.037 0 in placebo group (P 0.05), but all were increased compared to placebo group (P 0.05). Most osteocytes were alive in NELL-1 treatment group. More active osteoblasts and osteoclasts were noted in NELL-1 treatment group than those in placebo group. Conclusion NELL-1 gene transfection can preserve femoral head shape and bone content, promote osteoblast activity and neovascularization and hence is an effective treatment for rat traumatic osteonecrosis. However, the activity of osteoclasts is stimulated simultaneously. Key words: Osteonecrosis; Femoral head; Genes; Transfection

Preventive effect of erythropoietin on femoral head necrosis in rats

Objective To explore the preventive effect of recombinant human erythropoietin erythropoietin(EPO)on femoral head necrosis in rats. Methods Sixty male SD rats were randomly into three groups:blank group, control group, and experimental group.The control group was subdivided into two subgroups:subgroup A and subgroup B.The rats in subgroup A were injected with prednisone acetate 50 mg/kg in the root of the gluteal muscle on every Monday and Friday, once per day for 4 weeks, and those in subgroup B were injected with the same volume of sodium chloride injection of recombinant human erythropoietin erythropoietin(rh EPO,5 000 U/kg)three h before the modeling. Results The empty lacunae rate of hematoxylin and eosin(HE)stained sections in the experimental group, control group and blank group was(14.7±1.78)%,(23.21±2.34)%, and (7.45±1.23)% respectively; At 1st, 2nd and 4th, blood triglyceride content was(1.25±0.32), (1.28±0.26)and(1.27±0.4) mmo/L in blank group,(2.17±0.23),(2.48±0.41)and(1.88± 0.38) mmo/L in control group,and(1.42±0.34),(1.47±0.26)and(1.50±0.49)mmo/L in experimental group; the total cholesterol content was(1.62±0.13),(1.62±0.14)and(1.62±0.19) mmo/L in blank group,(2.41±0.18),(2.52±0.24)and(2.13±0.37)mmo/L in control group, and(1.64±0.18),(1.69±0.24)and(1.66±0.31)mmo/L in experimental group.Histologically, the femoral head surface was more smooth in experimental group than control group.In experimental group, most of the trabecular bones arranged densely and regularly,and shrink,sparse,and fracture were seen in less regions. Conclusion EPO can reduce the empty lacunae rate,increase the microvessels in femoral head blood supply, reduce blood lipid levels,and prevent the progress of femoral head necrosis. Key words: Femoral head necrosis; Erythropoietin

Osteoblast differentiation and mature gene regulation inside femoral head with steroid-induced osteonecrosis in response to mechanical stress stimulation

Objective To study dynamic changes of gene expressions and protein synthesis of Runx2 (runt-related transcription factor-2), Osterix, osteocalcin and AJ18 inside the femoral head with steroid-induced osteonecrosis after mechanical stress stimulation in rats. Methods A total of 50 Wistar rats (half male in sex) weighing 250-270 g (mean 260 g) were involved in this study and randomly divided into experimental group (40 rats) and normal group (10 rats). The rats in experimental group were injected with dexamethasone (20 mg/kg) via bilateral gluteus maximus alternatively once a week and then trained on laboratory animal treadmill twice weekly to make rat model of femoral head necrosis. After identifying the successfully induced model by Hematoxylin and eosin stain, glucocorticoid injection was ceased and the experimental group was randomly divided into model control group, 4 weeks, 6 weeks, 8 weeks groups after hormone training stopped. Then, total RNA and total protein were extracted from femoral head for detecting dynamic changes of genes expressions and proteins synthesis of Runx2, Osterix, osteocalcin and A J18 after mechanical stress stimulation inside the femoral head with steroid-induced osteonecrosis by means of real-time quantitative PCR and Western blot assay. Results In 4 weeks, 6 weeks, 8 weeks groups after hormone training stopped, the gene expressions and proteins synthesis of Runx2, Osterix and osteocalein were reduced more significantly compared with model control group, mBNA expression values of Runx2, Osterix and osteoealcin were 0. 1809, 0. 1639, 0. 1374 and 0. 4219, 0. 3026, 0.2652 and 0. 2857, 0.2027, 0. 1583 times of those in model control group. The expressions of Runx2, Osterix and osteocalcin showed a downward trend with time. The mBNA expression and protein synthesis of AJ18 at 4th, 6th and 8th weeks after hormone training stopped were 2.6391,4. 2718 and 5. 3165 times of model control group. Conclusions In addition to hormonal factors, inappropriate mechanical stress inhibits expressions and proteins synthesis of Runx2, Osterix and osteocalcin, while the expression and protein synthesis of AJ18 are upgraded in early steroid-induced femoral head necrosis in rats. Key words: Femur head necrosis; Dexamethasome; Gone expression regulation; Rats; Wistar