Antiosteoporotic activity and constituents of Podocarpium podocarpum

Our study aimed to investigate the antiosteoporotic properties of the ethanol extract of Podocarpium podocarpum (DC.) Yang et Huang (PE) in ovariectomized (OVX) rats and to characterize the active constituents. As a result, PE significantly inhibited the increased urinary Ca excretion and activity of bone resorption markers including tartrate-resistant acid phosphatase (TRAP), deoxypyridinoline crosslinks and cathepsin K in OVX rats, whereas exhibited little effects on the body, uterus and vagina weight. Detailed micro-CT analysis showed that PE notably enhanced bone quality, with increased bone mineral content (BMC), bone volume fraction (BVF), connectivity density (CD), tissue mineral content (TMC), tissue mineral density (TMD) and trabecular number (Tb. N), and decreased trabecular separation (Tb. Sp), in OVX animal. Those findings implied that PE had notable antiosteoporotic effect, especially effective in preventing bone resorption, with little side-effects on reproductive tissue. Further chemical investigation led to the isolation of 17 flavonoids, most of which showed significantly stimulatory effect on osteoblastic proliferation, ALP activity and mineralized nodes formation as well as inhibitory effect on osteoclastic TRAP activity in osteoblastic and osteoclastic cells. Our results indicated that PE, with abundant flavonoids, had remarkable antiosteoporotic activity and therefore can be a promising candidate for the treatment of postmenopausal osteoporosis induced by estrogen deficiency through herbal remedy.     

Panax notoginseng saponins suppress radiation-induced osteoporosis by regulating bone formation and resorption

BackgroundWhile radiation-based therapies are effective for treating numerous malignancies, such treatments can also induce osteoporosis.PurposeWe assessed the antiosteoporotic properties of total saponins extracted from the leaves of Panax notoginseng (LPNS) in a mouse model of radiation-induced osteoporosis and in vitro.Study design/methodsThe bone mineral densities, the marker of bone formation and resorption, and inflammatory factors were measured in vivo. Cell proliferation and differentiation were detected in vitro.ResultsThe results showed that bone mineral densities in irradiated mice administered LPNS were significantly increased compared to those in irradiated mice which had not received LPNS. LPNS attenuated the inflammation caused by irradiation, and significantly increased blood serum AKP activity, the mRNA levels of RUNX2 and osteoprotegerin, and the numbers of CFU-Fs formed by bone marrow cells collected from irradiated mice. In contrast, LPNS decreased the numbers of osteoclast precursor cells (CD117⁺/RANKL⁺ cells and CD71⁺/CD115⁺ cells) and the mRNA levels of TRAP and ATP6i. These results suggest that LPNS functions as a negative regulator of bone resorption. In vitro assays showed that LPNS promoted the differentiation of bone marrow mesenchymal stem cells and mononuclear cells into osteoblasts and osteoclasts, respectively, but had no effect on osteoclast activation.ConclusionThese results demonstrate that LPNS has significant antiosteoporotic activity, which may warrant further investigations concerning its therapeutic effects in treating radiation-induced osteoporosis.     

Icariin ameliorates estrogen-deficiency induced bone loss by enhancing IGF-I signaling via its crosstalk with non-genomic ERα signaling

BackgroundRapid, non-genomic estrogen receptor (ER) signaling plays an integral role in mediating the tissue selective properties of ER modulators. Icariin, a bone bioactive flavonoid, has been reported to selectively activate non-genomic ERα signaling in in vitro and in vivo studies.PurposeThe mechanisms underlying the estrogen-like bone protective effects of icariin are not fully understood, especially those that are related to insulin-like growth factor I (IGF-1) signaling. The bone protective effects of icariin were investigated in female mature ovariectomized (OVX) rats and the signaling of IGF-IR- ERα cross-talk was determined in osteoblastic cells.Study design and methodsIcariin at 3 different dosages (50, 500 and 3000 ppm) were orally administrated to rats for 3 months through daily intake of phytoestrogen-free animal diets containing icariin. Bone marrow stromal cells (BMSCs) and osteoclast precursors from femurs were harvested for experiments and RNA-sequencing. The interactions between IGF-IR and non-genomic ERα signaling were examined in pre-osteoblastic MC3T3-E1 cells and mature osteoblasts differentiated from BMSCs.ResultsOur results show that chronic administration of icariin to OVX rats significantly protected them against bone loss at the long bone and lumbar spine without inducing any uterotrophic effects. Ex vivo studies using BMSCs and osteoclast precursors confirmed the stimulatory effects of icariin on osteoblastogenesis and its inhibitory effects on osteoclastogenesis, respectively. RNA-sequencing analysis of mRNA from BMSCs revealed that icariin at 500 ppm significantly altered IGF-1 signaling as well as PI3K-Akt pathways. Our results demonstrated for the first time the rapid induction of interactions between IGF-IR and ERα as well as IGF-IR signaling and the downstream Akt phosphorylation by icariin in MC3T3-E1 cells. The activation of ERα and Akt phosphorylation by icariin in MC3T3-E1 cells and the osteogenic effects of icariin on ALP activity in mature osteoblasts were shown to be IGF-IR-dependent.ConclusionOur findings reveal that icariin activates both ERα and Akt via enhancing rapid induction of IGF-1 signaling in osteoblastic cells for osteogenesis and might be regarded as a novel pathway-selective phytoestrogen for management of postmenopausal osteoporosis.     

Kirenol inhibits RANKL-induced osteoclastogenesis and prevents ovariectomized-induced osteoporosis via suppressing the Ca2+-NFATc1 and Cav-1 signaling pathways

BackgroundOsteoporosis is a threat to aged people who have excessive osteoclast activation and bone resorption, subsequently causing fracture and even disability. Inhibiting osteoclast differentiation and absorptive functions has become an efficient approach to treat osteoporosis, but osteoclast-targeting inhibitors available clinically remain rare. Kirenol (Kir), a bioactive diterpenoid derived from an antirheumatic Chinese herbal medicine Herba Siegesbeckiae, can treat collagen-induced arthritis in vivo and promote osteoblast differentiation in vitro, while the effects of Kir on osteoclasts are still unclear.PurposeWe explore the role of Kir on RANKL-induced osteoclastogenesis in vitro and bone loss in vivo.MethodsThe in vitro effects of Kir on osteoclast differentiation, bone resorption and the underlying mechanisms were evaluated with bone marrow-derived macrophages (BMMs). In vivo experiments were performed using an ovariectomy (OVX)-induced osteoporosis model.ResultsWe found that Kir remarkably inhibited osteoclast generation and bone resorption in vitro. Mechanistically, Kir significantly inhibited F-actinring formation and repressed RANKL-induced NF-κB p65 activation and p-p38, p-ERK and c-Fos expression. Moreover, Kir inhibited both the expression and nuclear translocation of NFATc1. Ca²⁺ oscillation and caveolin-1 (Cav-1) were also reduced by Kir during osteoclastogenesis in vitro. Consistent with these findings, 2–10 mg/kg Kir attenuated OVX-induced osteoporosis in vivo as evidenced by decreased osteoclast numbers and downregulated Cav-1 and NFATc1 expression.ConclusionsKir suppresses osteoclastogenesis and the Cav-1/NFATc1 signaling pathway both in vitro and in vivo and protects against OVX-induced osteoporosis. Our findings reveal Kir as a potential safe oral treatment for osteoporosis.     

METTL3‐m6 A methylase regulates the osteogenic potential of bone marrow mesenchymal stem cells in osteoporotic rats via the Wnt signalling pathway

ObjectivesBone marrow mesenchymal stem cells (BMSCs) hold a high osteogenic differentiation potential, but the mechanisms that control the osteogenic ability of BMSCs from osteoporosis (OP‐BMSCs) need further research. The purpose of this experiment is to discuss the osteogenic effect of Mettl3 on OP‐BMSCs and explore new therapeutic target that can enhance the bone formation ability of OP‐BMSCs.Materials and MethodsThe bilateral ovariectomy (OVX) method was used to establish the SD rat OP model. Dot blots were used to reveal the different methylation levels of BMSCs and OP‐BMSCs. Lentiviral‐mediated overexpression of Mettl3 was applied in OP‐BMSCs. QPCR and WB detected the molecular changes of osteogenic‐related factors and Wnt signalling pathway in vitro experiment. The staining of calcium nodules and alkaline phosphatase detected the osteogenic ability of OP‐BMSCs. Micro‐CT and histological examination evaluated the osteogenesis of Mettl3 in OP rats in vivo.ResultsThe OP rat model was successfully established by OVX. Methylation levels and osteogenic potential of OP‐BMSCs were decreased in OP‐BMSCs. In vitro experiment, overexpression of Mettl3 could upregulate the osteogenic‐related factors and activate the Wnt signalling pathway in OP‐BMSCs. However, osteogenesis of OP‐BMSCs was weakened by treatment with the canonical Wnt inhibitor Dickkopf‐1. Micro‐CT showed that the Mettl3(+) group had an increased amount of new bone formation at 8 weeks. Moreover, the results of histological staining were the same as the micro‐CT results.ConclusionsTaken together, the methylation levels and osteogenic potential of OP‐BMSCs were decreased in OP‐BMSCs. In vitro and in vivo studies, overexpression of Mettl3 could partially rescue the decreased bone formation ability of OP‐BMSCs by the canonical Wnt signalling pathway. Therefore, Mettl3 may be a key targeted gene for bone generation and therapy of bone defects in OP patients.

In this study, the osteoporosis rat model was successfully established by OVX. OP‐BMSCs were successfully isolated and cultured from the femur of OP rat. Lentiviral‐mediated overexpression of Mettl3 could partially rescue the impaired osteogenic ability of OP‐BMSCs by activating the canonical Wnt signalling pathway in vitro and in vivo .     

The Improved Biological Performance of a Novel Low Elastic Modulus Implant

Background

The mismatch of the elastic modulus between implants and bone tissue can lead to stress shielding, bone resorption and poor osseointegration. Compared with normal bone tissue, this problem is much more serious in osteoporosis. The purpose of this study was designed to find out whether the novel Ti-24Nb-4Zr-7.9Sn (TNZS) implant with low elastic modulus and high strength was suitable for biomedical material, especially in osteoporosis.

Methodology

In vitro study, the viability and Alkaline phosphatase (ALP) activity of osteoblasts on the TNZS and Ti-6V-4V (TAV) were observed. In vivo study, 30 adult female New Zealand rabbits were selected and divided randomly into two groups: sham-operation (SHAM, n = 6) and ovariectomised in combination with methylprednisolone treatment (OVX+MP, n = 24). Two implants were then placed in the tibia of each OVX + MP group rabbit, one in each side (left: TAV; right: TNZS). The OVX + MP group rabbits were sacrificed at 4 and 12 weeks after the implantation. The osteoporotic bone responses to the TNZS and TAV implants were evaluated by pull-out test, Micro-CT analyses and histological observation.

Principal Findings

Compared with the TAV group, the TNZS group showed a significant increase (P<0.05) in cell viability and ALP activity, new bone formation and pull-out force.

Conclusions

The novel TNZS implants show good biological performance both in vitro and in vivo, which suggests that the alloys are suitable for biomedical applications, especially in osteoporosis.     

The effect of dehydroepiandrosterone administration on intestinal calcium absorption in ovariectomized female rats

[Purpose] Dehydroepiandrosterone (DHEA) administration reportedly recovers osteoporosis, a bone disorder associated with bone deficiency in postmenopausal women. However, the physiological mechanism of DHEA in osteoporosis remains elusive, especially in terms of intestinal calcium absorption. Therefore, we investigated the effect of DHEA administration on calcium absorption in ovariectomized (OVX) female rats using an estrogen receptor antagonist.[Methods] Female Sprague-Dawley rats (n=23, 6 weeks old) were randomized into three groups: OVX control group (OC, n=7), OVX with DHEA treatment group (OD, n=8), and OVX with DHEA inhibitor group (ODI, n=8) for 8 weeks.[Results] Intestinal calcium accumulation, as well as the rate of absorption, demonstrated no significant differences during the experimental period among investigated groups. The bone mineral density (BMD) of the tibia at the proximal metaphysis was higher in the OD group than that in the OC group (p<0.05); however, BMD of the ODI group showed no significant difference from investigated groups. Furthermore, the BMD of the tibia at the diaphysis did not significantly differ among these groups.[Conclusion] We revealed that DHEA administration does not involve intestinal Ca absorption, although this treatment improves BMD levels in OVX rats. These observations indicate that the effect of DHEA on the bone in postmenopausal women is solely due to its influence on bone metabolism and not intestinal calcium absorption.     

Antiosteoporotic activity of icariin in ovariectomized rats

Icariin was evaluated for its antiosteoporotic activity in an ovariectomized rat model of osteoporosis. The rats were divided into sham and OVX groups. The OVX rats were then subdivided into five groups treated with water, nylestriol (1 mg/kg body weight, weekly, orally) or icariin (ICA) (5, 25, and 125 mg/kg body weight, daily, orally) for 12 weeks. In OVX rats, the increases of body weight, serum BGP and ALP were significantly decreased by ICA treatment. In OVX rats, atrophy of uterus and descent of BMD were suppressed by treatment with ICA. In addition, ICA (125 mg/kg body weight) completely corrected the decreased serum concentration of Calcium, Phosphorus, and E₂ observed in OVX rats. ICA (125 mg/kg body weight) increased biomechanical strength significantly in comparison to the sham group. Histological results also showed its protective action through promotion of bone formation. The findings, assessed on the basis of biochemical, bone mineral density, biomechanical, and histopathological parameters, showed that ICA has a definite antiosteoporotic effect, similar to estrogen, especially effective for prevention bone fracture induced by estrogen deficiency.     

Adipose-derived stem cells alleviate osteoporosis by enchancing osteogenesis and inhibiting adipogenesis in a rabbit model

Background aimsOsteoporosis (OP) is characterized by a reduction in bone quality, which is associated with inadequacies in bone marrow mesenchymal stromal cells (BMSCs). As an alternative cell source to BMSCs, adipose-derived stem cells (ASCs) have been investigated for bone repair because of their osteogenic potential and self-renewal capability. Nevertheless, whether autologous ASCs can be used to promote bone regeneration under osteoporotic conditions has not been elucidated.MethodsThe OP rabbit model was established by means of bilateral ovariectomy (OVX). Both BMSCs and ASCs were harvested from OVX rabbits and expanded in vitro. The effects of osteogenic-induced ASCs on the in vitro adipogenic and osteogenic capabilities of BMSCs were evaluated. Autologous ASCs were then encapsulated by calcium alginate gel and transplanted into the distal femurs of OVX rabbits (n = 12). Hydrogel without loading cells was injected into the contralateral femurs as a control. Animals were killed for investigation at 12 weeks after transplantation.ResultsOsteogenic-induced ASCs were able to promote osteogenesis and inhibit adipogenesis of osteoporotic BMSCs through activation of the bone morphogenetic protein 2/bone morphogenetic protein receptor type IB signal pathway. Local bone mineral density began to increase at 8 weeks after ASC transplantation (P < 0.05). At 12 weeks, micro–computed tomography and histological evaluation revealed more new bone formation in the cell-treated femurs than in the control group (P < 0.05).ConclusionsThis study demonstrated that ASCs could stimulate proliferation and osteogenic differentiation of BMSCs in vitro and enhance bone regeneration in vivo, which suggests that autologous osteogenic-induced ASCs might be useful to alleviate OP temporally.     

Deficiency of Adiponectin Protects against Ovariectomy-Induced Osteoporosis in Mice

Adipokine adiponectin (APN) has been recently reported to play a role in regulating bone mineral density (BMD). To explore the mechanism by which APN affects BMD, we investigated BMD and biomechanical strength properties of the femur and vertebra in sham-operated (Sham) and ovariectomized (OVX) APN knockout (KO) mice as compared to their operated wild-type (WT) littermates. The results show that APN deficiency has no effect on BMD but induces increased ALP activity and osteoclast cell number. While OVX indeed leads to significant bone loss in both femora and vertebras of WT mice with comparable osteogenic activity and a significant increase in osteoclast cell number when compared to that of sham control. However, no differences in BMD, ALP activity and osteoclast cell number were found between Sham and OVX mice deficient for APN. Further studies using bone marrow derived mesenchymal stem cells (MSCs) demonstrate an enhanced osteogenic differentiation and extracellular matrix calcification in APN KO mice. The possible mechanism for APN deletion induced acceleration of osteogenesis could involve increased proliferation of MSCs and higher expression of Runx2 and Osterix genes. These findings indicate that APN deficiency can protect against OVX-induced osteoporosis in mice, suggesting a potential role of APN in regulating the balance of bone formation and bone resorption, especially in the development of post-menopausal osteoporosis.     

Osteogenic effects of flavonoid aglycones from an osteoprotective fraction of Drynaria fortunei—An in vitro efficacy study

Drynaria fortunei (Kunze) J. Sm. is a traditional Chinese herb used for the treatment of osteoporosis and other bone metabolic disorders. Previous studies demonstrated that “small polar active fraction in Drynaria fortunei (SDF)”exerted osteoprotective effects in ovariectomized (OVX) mice. This study aims to investigate the constituents in SDF and systemically evaluate their osteogenic effects in vitro. Five flavonoid aglycones, naringenin, kurarinone, kushennol F, xanthogalenol, and sophoraflavanone G were identified in SDF. All the compounds did not show effects on proliferation of osteoblastic UMR 106 cells at the concentrations of 0.1-1000 nM, but significantly increased the ALP activity of the cells at most of the concentrations from 10 nm to 1000 nM. Xanthogalenol at the concentration of 100 nM significantly increased concentration of acid-solubilized calcium. ICI 182,780, antagonist of estrogen receptor (ER), diminished the effect of kushennol F on ALP activity and the effect of xanthogalenol on acid-solubilized calcium. In conclusion, flavonoid aglycones in SDF could promote differentiation and mineralization of osteoblastic UMR 106 cells in vitro, which was explained by activation of ER signaling pathway. This study provides scientific evidences for the conduction of in vivo experiments to confirm potential effects of flavonoid aglycones on preventing OVX-induced osteoporosis.     

The effects of luteolin on osteoclast differentiation, function in vitro and ovariectomy-induced bone loss

Flavonoids, a group of polyphenolic compounds abundant in plants, are known to prevent bone loss in ovariectomized (OVX) animal models. Inhibition of osteoclast differentiation and bone resorption is considered as an effective therapeutic approach in the treatment of postmenopausal bone loss. Luteolin, a plant flavonoid, has potent anti-inflammatory properties both in vivo and vitro. In this study, we found that luteolin markedly decreased the differentiation of both bone marrow mononuclear cells and Raw264.7 cells into osteoclasts. Luteolin also inhibited the bone resorptive activity of differentiated osteoclasts. We further investigated the effects of luteolin on ovariectomy-induced bone loss using micro-computed tomography, biomechanical tests and serum markers assay for bone remodeling. Oral administration of luteolin (5 and 20 mg/kg per day) to OVX mice caused significant increase in bone mineral density and bone mineral content of trabecular and cortical bones in the femur as compared to those of OVX controls, and prevented decreases of bone strength indexes induced by OVX surgery. Serum biochemical markers assays revealed that luteolin prevents OVX-induced increases in bone turnover. These data strongly suggest that luteolin has the potential for prevention of bone loss in postmenopausal osteoporosis by reducing both osteoclast differentiation and function.     

In Vivo Screening for Anti-Osteoporotic Fraction from Extract of Herbal Formula Xianlinggubao in Ovariectomized Mice

MethodsThe extract of XLGB formula was separated into three fractions using chromatography, i.e., XLGB-A, XLGB-B and XLGB-C. They were administrated to 4-month old ovariectomized (OVX) mice for 6 weeks to determine which bioactive fraction(s) were more effective for preventing OVX-induced bone loss evaluated by microCT, biomechanical testing and biochemical markers. The main peaks of the key fraction were identified using reference compounds isolated from the fraction. In addition, the effects of the composite compounds in XLGB-B on osteoblasts’ proliferation and mineralization were evaluated in UMR 106 cells.ResultsXLGB-B with a yield of 13.0% from herbal Fufang XLGB was identified as the most potential one among the three fractions for prevention of OVX-induced bone loss confirmed with bone mass, bone microarchitecture, bone strength and bone turnover markers. Nine compounds in HPLC fingerprint were identified in the XLGB-B fraction, including phenylpropanoids from Herba Epimedii, terpenes from Radix Dipsaci and coumarins from Fructus Psoraleae. In addition, the identified compounds effectively promoted proliferation and/or mineralization of osteoblast-like UMR 106 cells in vitro.ConclusionXLGB-B with defined phytochemical structures was screened as the key fraction that demonstrated preventive effects on OVX-induced bone loss in mice. The present study laid down a foundation towards a new generation of herbal Fufang characterized with “less herbal materials for achieving equal treatment efficacy” in development strategy of TCM for prevention of OVX-induced osteoporosis.     

Dual effects of isoflavonoids from Pueraria lobata roots on estrogenic activity and anti-proliferation of MCF-7 human breast carcinoma cells

Pueraria lobata root (PLR), well known as Kudzu root, has recently become commercially available in Western dietary supplements for menopausal symptoms. The scientific basis for its action has been attributed to the action of phytoestrogens. This study aimed to investigate the estrogen-like activity of isoflavonoids isolated from P. lobata root and their safety with respect to their effect on breast cancer cell proliferation. In an E-screen assay, crude MeOH extract of PLR significantly increased the proliferation of MCF-7 cells in a concentration-dependent manner. Among the four fractions obtained by solvent fractionation of MeOH extract, the n-BuOH fraction had significant estrogen-like activities at all concentrations tested. Phytochemical analysis of the n-BuOH fraction led to the isolation of 10 isoflavones (1–10), among which genistein (10) had significant estrogen-like activities at all concentrations tested. These activities were significantly enhanced by treatment with genistein and 17β-estradiol compared with 17β-estradiol alone, and this effect was mediated by decreased expression of estrogen receptor (ER)α and phospho-ERα in MCF-7 cells. In a cell cytotoxicity assay, genistein (10) exhibited significant cytotoxicity in both ER-positive MCF-7 and ER-negative MDA-MB-231 breast cancer cells. This cytotoxicity was characterized by the induction of apoptotic cells stained with annexin V conjugated with Alexa Fluor 488 and involved activation of mitochondria-independent and -dependent apoptosis pathways in MCF-7 cells. Our results demonstrated that genistein (10) has estrogen-like effects dependent on ER pathway activation and anti-proliferative effects mediated by the apoptosis pathway rather than the ER pathway in MCF-7 breast cancer cells.     

Pharmacologic stem cell based intervention as a new approach to osteoporosis treatment in rodents

Background

Osteoporosis is the most prevalent skeletal disorder, characterized by a low bone mineral density (BMD) and bone structural deterioration, leading to bone fragility fractures. Accelerated bone resorption by osteoclasts has been established as a principal mechanism in osteoporosis. However, recent experimental evidences suggest that inappropriate apoptosis of osteoblasts/osteocytes accounts for, at least in part, the imbalance in bone remodeling as occurs in osteoporosis. The aim of this study is to examine whether aspirin, which has been reported as an effective drug improving bone mineral density in human epidemiology studies, regulates the balance between bone resorption and bone formation at stem cell levels.

Methods and Findings

We found that T cell-mediated bone marrow mesenchymal stem cell (BMMSC) impairment plays a crucial role in ovariectomized-induced osteoporosis. Ex vivo mechanistic studies revealed that T cell-mediated BMMSC impairment was mainly attributed to the apoptosis of BMMSCs via the Fas/Fas ligand pathway. To explore potential of using pharmacologic stem cell based intervention as an approach for osteoporosis treatment, we selected ovariectomy (OVX)-induced ostoeporosis mouse model to examine feasibility and mechanism of aspirin-mediated therapy for osteoporosis. We found that aspirin can inhibit T cell activation and Fas ligand induced BMMSC apoptosis in vitro. Further, we revealed that aspirin increases osteogenesis of BMMSCs by aiming at telomerase activity and inhibits osteoclast activity in OVX mice, leading to ameliorating bone density.

Conclusion

Our findings have revealed a novel osteoporosis mechanism in which activated T cells induce BMMSC apoptosis via Fas/Fas ligand pathway and suggested that pharmacologic stem cell based intervention by aspirin may be a new alternative in osteoporosis treatment including activated osteoblasts and inhibited osteoclasts.     

Gut microbiota modulates osteoclast glutathione synthesis and mitochondrial biogenesis in mice subjected to ovariectomy

ObjectivesOsteoporosis is a common bone disease in the elderly mainly regulated by osteoblasts (OBs) and osteoclasts (OCs). The gut microbiota has been recognized as an important factor in many physiological and pathological processes in the host. Thus, we hypothesize that the gut microbiota is necessary for postmenopausal osteoporosis and that germ‐free (GF) mice are protected from osteoporosis.Material and MethodsOsteoporosis models were established by performing ovariectomy (OVX) in mice. Bone mass was measured by micro‐CT, and gut microbiota were assessed by 16s rDNA sequencing. Reactive oxygen species (ROS) were detected by dihydroethidium (DHE) staining in vivo and 2’,7''‐dichlorodihydrofluorescein diacetate (DCFH‐DA) staining in vitro.ResultsFirmicutes and Bacteroidetes in the intestine are pivotal in OC differentiation, and the Firmicutes/Bacteroidetes ratio (F/B ratio) is a specific indicator of osteoporosis. Furthermore, we found that Firmicutes and Bacteroidetes affect the de novo synthesis of glutathione (GSH) by regulating its key enzyme glutamate–cysteine ligase catalytic subunit (Gclc) and inhibiting mitochondrial biogenesis and ROS accumulation via the cAMP response element‐binding (CREB) pathway. In addition, supplementing OVX mice with the probiotic Lactobacillus salivarius LI01 from the Firmicutes phylum prevented osteoporosis.ConclusionsOur results reveal that GSH plays a vital role in OVX‐induced bone loss, and probiotics that affect GSH metabolism are potential therapeutic targets for overcoming osteoporosis.     

Beneficial effects of losartan on vascular injury induced by advanced glycosylation end products and their receptors in spontaneous hypertension rats

The purpose of this study was to examine the effects of ENA Actimineral Resource A (ENA-A), seaweed origin alkaline water, on postmenopausal osteoporosis in ovariectomized (OVX) rats. The 12-week old Wistar rats were divided randomly into 4 groups: ovariectomized (OVX), OVX plus 0.5% ENA-A, OVX plus 5% ENA-A and OVX plus 10% ENA-A. A histopathological analysis indicated that ENA-A could prevent OVX-induced bone loss by increasing femur trabecular bone area in a dose-dependent manner. ENA-A significantly (p < 0.05) increased serum estradiol levels, decreased serum osteocalcin activity and suppressed serum pyridinoline (PYD) levels. The in vitro effects of ENA-A were also studied using MC3T3-E1 cells. ENA-A significantly stimulated cell proliferation and increased both ALP activity and calcium deposition in a dose-dependent manner. These results suggest that the treatment of ovariectomized rats with ENA-A not only prevents bone resorption but also appears to maintain the cancellous bone structure of postmeopausal osteoporosis.     

Antiosteoporotic flavonoids from Podocarpium podocarpum

A novel bi-isoflavonoid, podocarnone (1), together with five known flavonoids, namely genistein (2), afzelin (3), astragalin (4), luteolin (5) and pratensein (6), were isolated from the whole plants of Podocarpium podocarpum (DC.) Yang and Huang for the first time. The structure of podocarnone (1) was elucidated by detailed spectroscopic analyses, including 1D and 2D NMR techniques. All the isolated compounds were evaluated for their proliferative effects on osteoblasts derived from neonatal rat calvaria and inhibitory effects on multinucleated osteoclasts from rat marrow cells so as to explore the antiosteoporotic activity of these components. Podocarnone (1) exhibited potent stimulatory effects on osteoblastic proliferation and ALP (alkaline phosphatase) activity, and significantly inhibited the activity of osteoclastic TRAP (tartrate-resistant acid phosphatase) in the low concentration range of 10^?12–10^?14 mol/L, which were equivalent to the activity of genistein (2) in the concentration range of 10^?7–10^?9 mol/L. The other five known flavonoids also showed varied degrees of antiosteoporotic activities, and structure–activity relationship analysis revealed the number of phenolic rings contained in these structures maybe responsible for the antiosteoporosis property.