Rat Model Of Osteoporosis Research Articles

Network Pharmacological Analysis and Animal Experimental Study on Osteoporosis Treatment with GuBen-ZengGu Granules.

We explored the molecular pathway and material basis of GuBen-ZengGu granules (GBZGG) in treating osteoporosis using network pharmacology and animal experiments. The effective active components and potential targets of GBZGG were obtained from the TCMSP database and BATMAN-TCM database. Disease-related genes were obtained from GeneCard, NCBI, and DisGeNET. Next, a protein interaction network was established using the STRING database, and core genes were screened using the MCODE module. Cytoscape 3.8.0 was used to construct the network of component-disease-pathway-target, and KEGG pathway enrichment analyses were performed using the clusterProfiler R package to predict the mechanism of GBZGG in treating osteoporosis. An osteoporosis rat model was established by ovarian excision (OVX), and the partial results of network pharmacology were experimentally verified. Pharmacodynamic results showed that GBZGG increased bone mineral density (BMD) and significantly improved the indexes of femur microstructure in model rats. The network pharmacology results showed that quercetin, luteolin, stigmasterol, angelicin, kaempferol, bakuchiol, bakuchiol, 7-O-methylisomucronulatum, isorhamnetin, formononetin, and beta-sitosterol are the major components of GBZGG, with MAPK1, AKT1, JUN, HSP90AA1, RELA, MAPK14, ESR1, RXRA, FOS, MAPK8, NCOA1, MYC, and IL-6 as its core targets for treating osteoporosis. Biological effects could be exerted by regulating the signaling pathways of fluid shear stress and the signaling pathways of atherosclerosis, advanced glycation end products (AGE-RAGE) of diabetic complications, prostate cancer, interleukin (IL-17), tumor necrosis factor (TNF), hepatitis B, mitogen-activated protein kinase (MAPK), etc. The results of animal experiments showed that GBZGG could reduce the serum levels of IL-6 and TNF-α, increase the expression of bone morphogenetic protein-2 (BMP-2) and runt-related transcription factor 2 (RUNX2) protein, and inhibit the activity of extracellular-regulated protein kinases (ERK1/2) and phosphorylation ERK1/2 (p-ERK1/2) protein. GBZGG reduces the expression of ERK1/2 and p-ERK1/2 proteins and mRNAs through the inhibitory effects on IL-6 and TNF-α and negatively regulates the MAPK/ERK signaling pathway. The osteoporosis model showed that it effectively improved the loss of bone mass and destruction of bone microstructure in rats and maintained a positive balance for bone metabolism.

Role of 17-β Estradiol and Ramipril in OPG/RANKL Pathway in a Rat Model of Post-Menopausal Osteoporosis

Role of 17-β Estradiol and Ramipril in OPG/RANKL Pathway in a Rat Model of Post-Menopausal Osteoporosis

Effect of different forms of selenium in osteoporosis rat model induced by retinoic acid

AbstractOsteoporosis is a typical physiological disease, the main symptoms of which are brittle fracture, bone pain and easily deformed. As an individual ages, the prevalence of osteoporosis increases year by year. In the present study, selenium with antioxidant, immunomodulatory and anti-tumor effects was used to prevent osteoporosis induced by retinoic acid. The serum calcium contents in the selenium-treated groups (sodium selenite and selenomethionine) were significantly higher (P<0.05) than those in the model group in both the prevention and treatment studies. After prevention, glutamic-oxalacetic transaminase transaminase (GOT), glutamate transaminase (GPT), alkaline phosphatase (ALP) and tartrate-resistant acid phosphatase (TRACP) levels were significantly (P<0.05) decreased. In the treatment study, the serum calcium and phosphorus contents of the rats increased after selenium treatment. There was no significant change (P>0.05) in the activity of GOT and GPT. The content of ALP decreased obviously and the TRACP enzyme activity increased. Overall, these results showed that different forms of selenium compounds have great potential in preventing and treating osteoporosis.

Pilose antler (Cervus elaphus Linnaeus) polysaccharide and polypeptide extract inhibits bone resorption in high turnover type osteoporosis by stimulating the MAKP and MMP-9 signaling pathways

Ethnopharmacological relevancePilose antler is a traditional Chinese medicine used to improve kidney function, strengthen tendons and bones, and prolong life, among other uses. It is widely employed in the treatment of osteoporosis. However, the molecular mechanisms underlying the treatment of high turnover osteoporosis are not fully understood. Aim of the studyThe present study aimed to investigate the molecular mechanism underlying pilose antler polysaccharide and polypeptide extracts in inhibiting bone resorption in high turnover osteoporosis, and compare the effects of the two components alone and in combination to explore whether they could produce synergistic enhancement effects. Materials and methodsThe quantitative and qualitative characteristics of pilose antler polysaccharide and polypeptide extracts were detected by UV–visible spectrophotometry and high-performance liquid chromatography. A rat model of retinoic acid-induced osteoporosis was used to evaluate the inhibitory effect of the extracts on bone resorption. Enzyme-linked immunosorbent assay (ELISA) was used to detect the activity of factors related to high turnover type osteoporosis in rat serum. Western blotting was used to detect the expression of proteins related to the MAKP and MMP-9 signaling pathways in rat femurs. Fluorescence quantitative PCR was used to detect the transcription levels of genes related to the MAKP and MMP-9 signaling pathways in rat femur tissues. Hematoxylin and eosin staining were used to observe the osteoprotective effects of pilose antler polysaccharides and polypeptides. ResultsThe yield of pilose antler polysaccharides was 8.3%, and was mainly composed of mannose, glucosamine hydrochloride, glucuronic acid, Galacturonic acid, Galactose hydrochloride, glucose, and galactose. The yield of the polypeptides was 26.2%, and eighty percent of the molecular weight of the antler polypeptides was 1.6 kDa-7kD, among which, the molecular weight of 7kD peptide accounted for 52% of the total. Both polysaccharides and peptides could reduce the activities of TRACP, OCN, ERK1, JNK, and MMP-9 in rat serum and reduce both the protein expression and gene transcription levels of ERK1, JNK, and MMP-9 in rat femur tissue with significant differences compared with the model group. Both extracts exerted significant protective effects on rat femur tissue. The effect of pilose antler polypeptides alone was better than that of polysaccharides either alone or in combination. ConclusionsPilose antler polysaccharides, polypeptides, and their mixtures could inhibit the occurrence of bone resorption of high turnover osteoporosis by stimulating the MAKP and MMP-9 signaling pathways to reduce the expression of the ERK1, JNK, and MMP-9 genes and proteins, and could help alleviate bone loss caused by retinoic acid. Pilose antler polypeptides had a stronger effect on inhibiting bone resorption. The combination of the two components did not show synergistic enhancement effect, and the polysaccharide tended to moderate the inhibitory enhancement effect of the polypeptide.

Integrated proteomics and metabolomics analysis of lumbar in a rat model of osteoporosis treated with Gushukang capsules

BackgroundGushukang (GSK) capsules are a Chinese patented medicine that is widely used in clinics for the treatment of osteoporosis (OP). Animal experiments have revealed that the bone mineral density of osteoporotic rats increase after treatment with GSK capsules. However, the specific mechanism and target of GSK in the treatment of osteoporosis are unclear. Further studies are needed.MethodsMetabolomics (GC/MS) and proteomics (TMT-LC-MC/MC) with bioinformatics (KEGG pathway enrichment), correlation analysis (Pearson correlation matrix), and joint pathway analysis (MetaboAnalyst) were employed to determine the underlying mechanisms of GSK. The differential expression proteins were verified by WB experiment.ResultsThe regulation of proteins, i.e., Cant1, Gstz1, Aldh3b1, Bid, and Slc1a3, in the common metabolic pathway of differential proteins and metabolites between GSK/OP and OP/SHAM was corrected in the GSK group. The regulation of 12 metabolites (tyramine, thymidine, deoxycytidine, cytosine, L-Aspartate, etc.) were differential in the common enrichment metabolic pathway between GSK /OP and OP/SHAM. Differential proteins and metabolites jointly regulate 11 metabolic pathways, such as purine metabolism, pyrimidine metabolism, histidine metabolism, beta-alanine metabolism, and so on.ConclusionGSK may protect bone metabolism in osteoporotic rats by affecting nucleotide metabolism, amino acid metabolism, and the immune system.

Gut microbiota and fecal metabolic signatures in rat models of disuse-induced osteoporosis.

Assessing the correlation between gut microbiota (GM) and bone homeostasis has increasingly attracted research interest. Meanwhile, GM dysbiosis has been found to be associated with abnormal bone metabolism. However, the function of GM in disuse-induced osteoporosis (DIO) remains poorly understood. In our research, we evaluated the characteristics of GM and fecal metabolomics to explore their potential correlations with DIO pathogenesis. DIO rat models and controls (CON) underwent micro-CT, histological analyses, and three-point bending tests; subsequently, bone microstructures and strength were observed. ELISAs were applied for the measurement of the biochemical markers of bone turnover while GM abundance was observed using 16S rDNA sequencing. Metabolomic analyses were used to analyze alterations fecal metabolites. The potential correlations between GM, metabolites, and bone loss were then assessed. In the DIO group, the abundance of GM was significantly altered compared to that in the CON group. Moreover, DIO significantly altered fecal metabolites. More specifically, an abnormally active pathway associated with bile acid metabolism, as well as differential bacterial genera related to bone/tissue volume (BV/TV), were identified. Lithocholic acid, which is the main secondary bile acid produced by intestinal bacteria, was then found to have a relationship with multiple differential bacterial genera. Alterations in the intestinal flora and metabolites in feces, therefore, may be responsible for DIO-induced bone loss. The results indicated that changes in the abundance of GM abundance and fecal metabolites were correlated with DIO-induced bone loss, which might provide new insights into the DIO pathogenesis. The detailed regulatory role of GM and metabolites in DIO-induced bone loss needs to be explored further.

Effect of Desmodium renifolium on ovariectomized rat model of osteoporosis via BMP-2/Smads signaling pathway

This study aimed to investigate the effect of Desmodium renifolium(Linn.) Schindl on ovariectomized rat model of osteoporosis and explore the underlying mechanism. Rats were randomized into a sham group, a model group, a Xianlin Gubao group, and low-, medium-, and high-dose D. renifolium groups. Bilateral oophorectomy was performed in other groups except sham group(removal of bilateral peri-ovarian adipose tissue). The sham group and model group were administrated with equal volume of 0.5% CMC-Na, Xianlin Gubao group with Xianlin Gubao, and D. renifolium groups with different concentrations of alcoholic extract of D. renifolium once a day for 14 weeks. The body weight of rats were recorded during the experiment. The levels of estradiol(E_2), 1,25-dihydroxyvitamin D_3 [1,25(OH)_2D_3], calcium(Ca), and phosphorus(P) in serum were determined after the administration. The femur microstructure was analyzed via micro-CT. RT-PCR and Western blot were employed to respectively determine the mRNA and protein levels of bone morphogenetic protein 2(BMP-2), Runt-related transcription factor 2(Runx2), and osterix(OSX) in the tibia of rats. The results indicated that D. renifolium significantly inhibited the weight growth, improved the uterus appearance, elevated the levels of E_2, Ca, P, and 1,25(OH)_2D_3 in serum, increased the number and reduced the fracture of bone trabeculae, ameliorated the bone microstructure parameters, and up-regulated the mRNA and protein levels of BMP-2, Runx2, and OSX. All the results demonstrated that D. renifolium had significant protective effect on the ovariectomized rat model of osteoporosis by regulating the BMP-2/Smads signaling pathway.

Eucommia Ulmoides Extracts Exert Protective Effect on Osteoporosis in Rats

Our study proposed to probe the therapeutic effect of Eucommia ulmoides extracts (EUE) on osteoporosis in rats and the potential mechanism. Castrated osteoporosis model was constructed in rats, followed by orally administrated with 50, 100, and 200 mg/kg EUE per day for one and half months, with sham animals as the negative control. The pathological conditions of femur were detected by HE staining, while the apoptotic state was evaluated by TUNEL. ELISA was utilized to measure the serum level of IL-6, calcium, and phosphorus. Significantly damaged bone trabecular structure and aggravated apoptosis were observed in castrated osteoporosis model rats, which were markedly alleviated by the administration of EUE. Furthermore, compared to control, the serum level of phosphorus was greatly declined and the IL-6 level was dramatically increased in castrated osteoporosis model rats, which were reversed by the administration of EUE. Taken together, EUE improved the pathological state of the femur tissue in osteoporosis rats by regulating the levels of phosphorus and IL-6 in serum and inhibited the apoptosis of the femur tissue cells, which effectively alleviated the osteoporosis induced by estrogen deficiency.

Overall adjustment acupuncture improves osteoporosis and exerts an endocrine-modulating effect in ovariectomized rats.

Acupuncture is a widely practiced, convenient, and safe treatment modality within complementary and integrative medicine. Increasing studies have revealed the efficacy of acupuncture for the treatment of osteoporosis in both human and non-human subjects. The aim of the present study was to assess the improvement of osteoporosis after overall adjustment acupuncture (OA) as well as its endocrine-modulating effect in an ovariectomized rat model. In total, 32 female Sprague-Dawley (SD) rats were randomly divided into the sham, model, ovariectomy+estrogen (OVX+E), and OVX+OA (OVX+A) groups with eight rats in each group. The postmenopausal osteoporosis (PMOP) rat model was induced by bilateral ovariectomy. At 12 weeks after surgery, rats in the OVX+E group received estradiol (0.2 mg/kg/i.g./qod) for 12 weeks, and rats in the OVX+A group were treated with acupuncture at Zusanli (ST36), Shenshu (BL23), and Dazhu (BL11) points (qod) for 12 weeks. At the end of the treatment, all rats were sacrificed, and the body weight, uterus index, bone mineral density (BMD), bone mineral content (BMC), bone trabeculae structural parameters, femoral biomechanical properties, femoral histomorphology, and several hormone levels were examined. In OVX rats, OA abrogated the body weight gain and improved osteoporosis in terms of BMD, BMC, bone trabeculae structural parameters, bone strength, and bone tissue histomorphology. Moreover, OA modulated the serum levels of estradiol, corticotropin releasing hormone (CRH), adrenocorticotropic hormone (ACTH), and corticosterone (CORT). OA improves osteoporosis and exerts an endocrine-modulating effect in ovariectomized rats.

SDH5 down-regulation mitigates the damage of osteoporosis via inhibiting the MyD88/NF-κB signaling pathway

Background Osteoporosis has become a serious public health problem especially in postmenopausal women. This work aims to assess both the function and mechanism of SDH5 in osteoporosis. Methods The animal model of osteoporosis in sprague-dawley rats was established by ultilizing Ovariectomy (OVX). The trabecular bone morphometry had been determined by micro-CT, and tibia injury of rats was detected through HE and alcian blue staining. Meanwhile, the levels of oxidative stress factors, including MDA, CAT, GSH-Px, and SOD, were detected by ELISA. The proliferation and apoptosis of osteoblasts isolated from OVX-induced rats were found out by CCK-8 and flow cytometry respectively. The expression of SDH5, Osterix (Osx), Type I collagen (CoL1A1), osteocalcin (OC), SOD1, SOD2, p-MyD88/MyD88, and p-NF-κB p65/NF-κB p65 was assessed by Western blot. The effect and mechanism of SDH5 knockdown on osteoporosis were verified by lipopolysaccharide (LPS) treatment. Results In the osteoporosis rat model, the expression of SDH5 had an up-regulated tend. A higher bone mineral density (BMD) value was found in the SDH5 knockdown group. SDH5 inhibition ameliorated bone loss, mitigated bone histopathological injury, alleviated oxidative stress, and elevated osteogenic marker protein expression in vivo and in vitro. SDH5 down-regulation also promoted the proliferation and restrained apoptosis of osteoblasts extracted from OVX-induced rats. Furthermore, we found that the underlying mechanism was associated with the inhibition of the MyD88/NF-κB pathway. Conclusion Down-regulation of SDH5 mitigates the damage of osteoporosis both in vivo and in vitro via inhibiting the MyD88/NF-κB signaling activation.

Therapeutic Effects of Strontium Ranelate and Risedronate on a Glucocorticoid-Induced Osteoporosis Rat Model: A Comparative Histological and Morphometric study

Therapeutic Effects of Strontium Ranelate and Risedronate on a Glucocorticoid-Induced Osteoporosis Rat Model: A Comparative Histological and Morphometric study

Casticin promotes osteogenic differentiation of bone marrow stromal cells and improves osteoporosis in rats by regulating nuclear factor-κB/mitogen-activated protein kinase.

Osteoporosis has influenced millions of people, especially postmenopausal women, which has become a big burden to the whole world. Although the diverse roles of casticin (CAS) on different diseases were identified, whether it was implicated with osteoporosis was unknown. A rat model of osteoporosis was established through dexamethasone (DEX) treatment and a cell model reflecting the osteogenic and osteoclast induction was constructed in bone marrow stromal cells (BMSCs). The calcification at the late stage of induction was measured via Alizarin Red S staining. Western blot was applied to evaluate the levels of proteins. Hematoxylin and eosin staining revealed that the number of bone trabecular in DEX-induced osteoporosis rats was decreased, while increased doses of CAS treatment elevated the number of bone trabecular. CAS treatment alleviated DEX-induced osteoporosis in rats. Moreover, we found that CAS inhibited the nuclear factor-κB/mitogen-activated protein kinase (NF-κB/MAPK) pathway. In addition, CAS promoted osteogenic differentiation of BMSCs and reduced osteoclastogenesis of bone marrow monocytes. Finally, CAS was observed to retard the receptor activator of NFκ-B ligand-induced NF-κB/MAPK pathway. CAS promoted osteogenic differentiation of BMSCs and improved osteoporosis in rats by regulating the NF-κB/MAPK pathway. This might shed a light into using CAS as a drug treating osteoporosis in the future.

Effect of the Pulsed Electromagnetic Field Treatment in a Rat Model of Senile Osteoporosis In Vivo.

This study assessed the effects of pulsed electromagnetic fields (PEMF) in a rat model of senile osteoporosis and the underlying molecular events. 24-month-old male Sprague-Dawley (SD) rats were randomly divided into control and PEMF groups (n = 8 per group) using a random digit table, while 3-month-old male SD rats were set as the young-age control group. Rats in the PEMF group were treated by PEMF for 40 min/day for 5 days/week. Bone mineral density/microarchitecture, level of serum bone-specific alkaline phosphatase (BALP), tartrate-resistant acid phosphatase 5b (TRACP5b), and Wnt/β-catenin signaling genes in rat bone marrow cells were then analyzed. The 12-week PEMF intervention showed a significant effect on inhibition of age-induced bone density loss and deterioration of trabecular bone structures in the PEMF group rats versus control rats, that is, the treatment enhanced bone mineral density of the proximal femoral metaphysis and the fifth lumbar (L5) vertebral body and improved the proximal tibia and L4 vertebral body parameters using bone histomorphometry analysis. Furthermore, the BALP level in the bones was significantly increased, but the TRACP5b level was reduced in the PEMF group of rats versus control rats. PEMF also dramatically upregulated expression of Wnt3a, LRP5, β-catenin, and Runx2 but downregulated PPAR-γ expression in the aged rats. The results demonstrated that PEMF could prevent bone loss and architectural deterioration due to the improvement of bone marrow mesenchymal stromal cell differentiation and proliferation abilities and activating the Wnt signaling pathway. Future clinical studies are needed to validate these findings. © 2022 Bioelectromagnetics Society.

Tricin protects rats from ovariectomized-induced osteoporosis by enhancing Wnt/β-catenin pathway

Purpose: To investigate the effects of tricin on ovariectomized-induced osteoporosis, and unravel theunderling mechanism of action.Methods: An osteoporosis rat model was established by conducting ovariectomy (OVX). Changes in the microstructure of the trabecular bone were visualized using Hematoxylin and eosin (H&E) staining, and a three-point bending test was employed to assess the biomechanical stability of the femurs, after the administration of tricin (20 and 40 mg/kg). Subsequently, bone marrow mesenchymal stem cells (BMSCs) were isolated and treated with tricin (7 and 15 μM). Alizarin red staining was performed to assess mineralization, and Runt-related transcription factor 2 (RUNX2); osteocalcin (OCN) and collagen type I alpha 1 (Col1a1) were quantified using western blot analysis. The Wnt/β-catenin pathway related proteins, i.e., Wnt3a, β-catenin, glycogen synthase kinase-3 β (GSK-3 β) were determined.Results: Ovariectomy induced thinner and discontinuous trabecular bone, with increased marrow cavities, while application of tricin significantly improved the density and regularity meshwork, but reduced marrow cavities. Tricin also enhanced biomechanical competence as seen in the upregulated maximum load, stiffness, young modulus and maximum stress compared with OVX group (p < 0.01). Furthermore, tricin increased calcification in BMSCs, and significantly upregulated the expressions of RUNX2, OCN and COL1A1 when compared with OVX group (p < 0.01). It promoted Wnt/β-catenin signaling by enhancing Wnt3a and β-catenin, while inhibiting GSK3β expression, compared with OVX group (p < 0.05 or p < 0.01).Conclusion: Tricin exerts protective effects against OVX-induced osteoporosis by enhancing Wnt/β-catenin pathway. Thus, tricin is a potential therapeutic agent for the management of osteoporosis.

Mechanism of Bushen Huoxue Recipe on Improving Arterial Calcification in Rats with Osteoporosis Based on OPG/RANKL Signaling Pathway

<bold>Objective</bold> To observe the effect of Bushen Huoxue Recipe on thoracic aortic calcification and explore the possible mechanism of improving arterial calcification by Bushen Huoxue Recipe based on osteoprotegerin / receptor activator for nuclear factor-κB ligand (OPG/RANKL) signaling pathway in rat model of osteoporosis. <bold>Methods</bold> A total of 32 seven-month-old male SD rats were randomly divided into control group, sham operation group, osteoporosis group and Bushen Huoxue group, with eight rats in each group. The control group did not receive any intervention, the sham operation group exposed but did not remove the testicles, and the other two groups removed bilateral testicles to establish the model of osteoporosis. The intervention began at the 16th week after operation. The Bushen Huoxue group was given 55 g/(kg·d) Bushen Huoxue Recipe by gavage, and the other three groups were given the same amount of normal saline, once a day, lasting for eight weeks. Dual-energy X-ray absorptiometry was used to measure the bone mass density (BMD) of femoral shaft of right hindlimb; alizarin red staining was used to detect integrated optical density (IOD) of calcification deposition on the vascular wall of thoracic aorta, and the calcification degree of thoracic aorta was observed; ELISA method was used to detect serum levels of PⅠNP, BGP and CTX-Ⅰ; immunohistochemical staining was used to detect the expression of OPG and RANKL proteins in thoracic aorta. <bold>Results</bold> 1) BMD and IOD value of thoracic aorta calcification deposition: compared with the control group, there were no significant changes of BMD and IOD value in thoracic aorta calcification deposition of the sham operation group (<italic>P</italic>&gt;0.05), while the BMD of the osteoporosis group decreased significantly, and the IOD value of thoracic aorta calcification deposition increased significantly (<italic>P</italic>&lt;0.05). Compared with the osteoporosis group, the BMD of the Bushen Huoxue group increased significantly, and IOD value of thoracic aorta calcification deposition decreased significantly, the difference was statistically significant (<italic>P</italic>&lt;0.05). 2) The level of PⅠNP, BGP and CTX-Ⅰ in the serum: compared with the control group, the level of PⅠNP and BGP of the osteoporosis group decreased significantly, the level of CTX-Ⅰ, a bone resorption marker increased significantly, the difference was statistically significant (<italic>P</italic>&lt;0.05). Compared with the osteoporosis group, the level of PⅠNP and BGP in the Bushen Huoxue group increased significantly (<italic>P</italic>&lt;0.05), and there was no significant difference in CTX-Ⅰ level (<italic>P</italic>&gt;0.05). 3) The expression level of OPG and RANKL protein in thoracic aorta: compared with the control group, the expression level of OPG in the osteoporosis group decreased significantly while the expression level of RANKL increased significantly, the difference was statistically significant (<italic>P</italic>&lt;0.05); compared with the osteoporosis group, the expression level of OPG protein in thoracic aorta of the Bushen Huoxue group increased significantly, and the expression level of RANKL protein decreased significantly, the difference was statistically significant (<italic>P</italic>&lt;0.05). <bold>Conclusion</bold> Bushen Huoxue Recipe can improve BMD of osteoporosis rats, promote bone formation, and improve the condition of artery calcification induced by osteoporosis. The possible mechanism may be related to regulating the expression of OPG/RANKL signaling pathway through up-regulation of OPG expression and down-regulation of RANKL expression in thoracic aorta.

Suppression of high bone remodelling by E'Jiao in ovariectomised rats.

The current prevention options for postmenopausal osteoporosis are very limited. E'Jiao is a collagen-rich traditional Chinese medicine with the potential to prevent osteoporosis but more comprehensive investigations are lacking. This study aimed to investigate the skeletal protective effects of E'Jiao in a rat model of osteoporosis caused by ovariectomy. Female Sprague Dawley rats (n=42) were randomly assigned into baseline, sham, ovariectomised (OVX) control, OVX-treated with low-dose (0.26g/kg), medium dose (0.53g/kg) and high dose E'Jiao (1.06g/kg), as well as calcium carbonate (1% w/v) groups. Daily treatment through oral gavage was initiated 7 days after OVX. The rats were euthanised after eight weeks of treatment. Bone mineral density and content were measured at baseline, 1 and 2 months after treatment. Blood was collected for the measurement of bone remodelling markers. Femur and tibial bones were collected for histomorphometry and biomechanical strength analysis. Untreated OVX rats showed high bone remodelling marked by the increased bone formation and bone resorption markers, as well as increased mineralising surface/bone surface ratio. In addition, osteoclast surface and single-labelled surface were increased while mineral apposition rate was reduced in the untreated OVX rats. These changes were antagonised by E'Jiao at all doses. However, the structural, cellular and biomechanical parameters were not affected by ovariectomy and treatment. In conclusion, E'Jiao prevented high bone remodelling during oestrogen deficiency but a long-term study will be required to establish its effects on structural and biomechanical changes due to oestrogen deficiency.

Improvement of Osteoporosis in Rats With Hind-Limb Unloading Treated With Pulsed Electromagnetic Field and Whole-Body Vibration.

Physical factors have been used to address disuse osteoporosis, but their effects and mechanism remain unclear. The purpose of this study was to determine the effects of pulsed electromagnetic field (PEMF) and whole-body vibration (WBV) on disuse osteoporosis to increase knowledge about treating osteoporosis. A disuse osteoporosis rat model was developed by hind-limb unloading (HU) for 6weeks. Forty 4-month-old female Sprague-Dawley rats were divided into 5 groups and given the following interventions: HU, HU treated with PEMF (HUP), HU treated with WBV (HUW), HU treated with both PEMF and WBV (HUPW), and no intervention (controls). After 8weeks of intervention, measurements were taken. HU induced a decrease in bone mineral density (BMD), whereas HUP, HUW, and HUPW increased it. Moreover, the bone resorption markers tartrate-resistant acid phosphatase (TRAP) and C-terminal peptide of type 1 collagen in the HU group significantly increased, whereas the osteogenesis markers osteocalcin and N-terminal propeptide of type 1 procollagen significantly decreased. The markers osteocalcin and N-terminal propeptide of type 1 procollagen significantly increased, but TRAP and C-terminal peptide of type 1 collagen significantly decreased in the HUPW, HUP, and HUW groups compared with the HU group. In particular, HUPW effectively increased osteocalcin and decreased TRAP compared with HUP and WBV. Microcomputed tomography analysis of the femur indicated that HUPW improved trabecular number, bone volume over total volume, bone surface over bone volume, trabecular separation, and the structure model index compared with HUP and that it improved bone surface over bone volume, trabecular separation, and structure model index compared with HUW. The HUPW group showed a significant increase in maximum load compared with the HUW group and a significant increase in elastic modulus compared with the HUP group. PEMF, WBV, and their combination all attenuated bone resorption and enhanced osteogenesis. WBV and the combination of treatments have great potential to improve osteogenesis compared with PEMF. In addition, HUPW significantly attenuated bone resorption compared with HUW and HUP. The results of this study indicated that HUPW could effectively improve disuse osteoporosis compared with HUP, given that trabecular number and bone volume over total volume are associated with disuse osteoporosis. Moreover, BMD recovered well with HUP, HUW, and HUPW but the bone structure-especially mechanical performance-did not, indicating that osteoporosis should be evaluated with BMD and mechanical performance, not with BMD in isolation.

Discovery of potential active ingredients of Er-Zhi-Wan, a famous traditional Chinese formulation, in model rat serum for treating osteoporosis with kidney-yin deficiency by UPLC-Q/TOF-MS and molecular docking

Er-Zhi-Wan (EZW), a classical traditional Chinese formulation, has attracted more and more attention. This study was carried out to analyze the constituents of EZW absorbed into blood and find out the potential active ingredients for treating osteoporosis (OP) with kidney-yin deficiency (KYD). The rat model of OP with KYD was achieved by ovariectomies and using the mixture of thyroxine and reserpine. Then ultra-high performance liquid chromatography coupled with a quadrupole time-of-flight mass spectrometer (UPLC-Q/TOF-MS) combined with statistical analysis was used to analyze the constituents of EZW absorbed into blood and differential components between the normal and OP with KYD rats. Finally, the components identified in OP with KYD rats were docked with targets of OP with KYD found in online databases. The results of molecular docking were adopted to find the potential active ingredients and further verified in vitro experiment. A total of 21 prototype compounds and 69 metabolites were identified in serum. Among them, 63 components in model rats and 50 components in normal rats were summarized, respectively. Most of the identified metabolites in serum of model rats were produced by hydrolysis, oxidation or glucuronidation, while in serum of normal rats were produced by hydrolysis, oxidation and methylation. According to the results of molecular docking, specnuezhenide, salidroside, tyrosol, echinacoside and verbascoside could be classified as potential active ingredients. The activity of salidroside and a metabolite was verified by pharmacodynamics analysis. In summary, UPLC-Q/TOF-MS system was combined with molecular docking to search the potential active ingredients from model rats of OP with KYD, which provided a new idea for the research on the pharmacodynamic material basis of other traditional medicine. Moreover, the result of this study lays the foundation for further study regarding the mechanism of EZW in treating OP with KYD.

Dietary Effects of Nanopowder Eggshells on Mineral Contents, Bone Turnover Biomarkers, and Regulators of Bone Resorption in Healthy Rats and Ovariectomy-Induced Osteoporosis Rat Model.

Postmenopausal osteoporosis is a critical issue for female health worldwide. This current study was designed to evaluate the role of nanopowder eggshell (NPES) in healthy and ovariectomy-induced osteoporosis rats. Fifty-six female rats were divided into healthy rats (35) and ovariectomized rats (21). The healthy rats were subdivided into five groups (G1-G5) and received one of the following treatments: saline, 20 or 40mg/kg of calcium carbonate, and 20 or 40mg/kg of NPES. The 21 ovariectomized rats were divided into three groups (G6-G8) and received either saline, 40mg/kg of calcium carbonate, or 40mg/kg of NPES. Biochemical and histopathological assessments of bone formation and resorption were performed. Biomarkers of bone formation (calcium and osteocalcin (OCN)) and calcium content in left femur ashes were significantly higher in healthy rats given 40-mg/kg NPES than in healthy control rats and healthy rats given 40-mg/kg calcium carbonate. The ovariectomized groups had significantly lower levels of vitamin D3, OCN, and osteoprotegerin (OPG) than the healthy control. Alanine transaminase (ALT), alkaline phosphatase (ALP), and receptor activator of nuclear factor-κB ligand (RANKL) were significantly increased in the ovariectomized group than in the healthy control group. Treatment with NPES and calcium carbonate reduced liver enzymes in ovariectomized rats. NPES treatment significantly increased Vit D3, OCN, OPG, and bone ash mineral content (calcium, magnesium, zinc, and phosphorus) in ovariectomized rats. NPES also increased femur cortical thickness, osteoblast number, and collagen fiber. The current study suggests that NPES can modulate bone turnover biomarkers and increase bone trace elements. Moreover, NPES alleviates bone resorption in ovariectomy-induced osteoporosis.

MiR-152-5p suppresses osteogenic differentiation of mandible mesenchymal stem cells by regulating ATG14-mediated autophagy

BackgroundOsteoporosis affects the mandible resulting in bone loss. Though impairments are not life threatening, they affect a person's quality-of-life particularly vulnerable elderly. MicroRNAs (miRNAs) are novel regulatory factors that play an important role in regulating bone metabolism. Autophagy is evolutionarily conserved intracellular self-degradation process and is vital in the maintenance of both miRNA and bone homeostasis. However, the role of autophagy in the pathogenesis of miRNA regulating osteoporosis remains unclear.MethodsIn the study, we established a rat osteoporosis model induced by ovariectomy (OVX) and isolated mesenchymal stem cells from mandible (MMSCs-M). Several miRNAs were identified to regulate osteoporosis in some studies. qRT-PCR was applied to examine the expression of miRNA, autophagy and osteogenic differentiation-related genes. Western blotting assays were performed to detect the expression of autophagy and osteogenic differentiation proteins. Immunofluorescence and transmission electron microscope were used to verify the autophagy activity. Transfecting technology was used to enhance or suppress the expression of miR-152-5p which enable us to observe the relationship between miR-152-5p, autophagy and osteogenic differentiation. Additionally, the measurement of reactive oxygen species was used to investigate the mechanism of autophagy affecting osteogenic differentiation.ResultsWe found an upregulated expression of miR-152-5p in MMSCs-M in OVX group. Downregulated autophagy-related gene, proteins and autophagosome were detected in vitro of OVX group compared with sham group. Moreover, downregulation of miR-152-5p promoted osteogenic differentiation of MMSCs-M as well as enhanced autophagy-related proteins in OVX group. Conversely, overexpression of miR-152-5p showed opposite effect in sham group. Meanwhile, we found Atg14 (autophagy-related protein homolog 14) was identified to be a direct target of miR-152-5p theoretically and functionally. In other words, we confirmed inhibition of miR-152-5p promoted the osteogenic differentiation via promoting ATG14-mediated autophagy. Furthermore, miR-152-5p/ATG14-mediated autophagy regulated osteogenic differentiation by reducing the endogenous ROS accumulation and maintaining cellular redox homeostasis.ConclusionOur data suggest that miR-152-5p is the first identified to regulate osteogenic differentiation by directly targeting autophagy-related protein ATG14 and regulating oxidative stress and therapeutic inhibition of miR-152-5p may be an efficient anabolic strategy for osteoporosis.