Ethanolic extract of Persea americana Mill. (Lauraceae) seeds induced antiestrogenic effects in young female Wistar rats.

OBJECTIVES: The ethanol extract of Persea americana seeds was found to inhibit the development of estrogen-dependent conditions in female Wistar rats, suggesting the ability of its secondary metabolites to interact with estrogen receptors (ERs), either as partial agonists or as antagonists. To test this hypothesis, the abovementioned extract was assessed for its ability to mimic and/or antagonize estradiol effects. METHODS: Two experiments were conducted in ovariectomized (OVX) rats: (1) animals were treated with estradiol valerate (E2V; 1 mg/kg) or P. americana at doses of 25 and 50 mg/kg; (2) animals were treated with E2V alone (0.75 mg/kg) or in combination with P. americana at the abovementioned doses. Treatments were given orally for 3 days and animals were sacrificed for biochemical and histological analyses of the uterus and vagina. RESULTS: When administered alone, P. americana did not change the histomorphology of both organs (uterus and vagina). In combination with E2V, P. americana decreased uterine weight [30 % decrease (p<0.001) at 25 mg/kg and 24 % (p<0.01) at 50 mg/kg] and epithelium height (37 % decrease). This was associated with decreased estradiol levels (at least 86 % decrease, p<0.001) in the uterus. Similarly, vagina epithelium height decreased by at least 34 % (p<0.05) when E2V was co-administered with P. americana. CONCLUSIONS: The seed extract of P. americana contains ER antagonist secondary metabolites accounting for its ability to inhibit the development of estrogen-dependent conditions in female rats.

Ziyuglycoside II attenuated OVX mice bone loss via inflammatory responses and regulation of gut microbiota and SCFAs.

BACKGROUND AND PURPOSE: Osteoporosis (OP) is a frequent clinical problem for the elderly. Traditional Chinese Medicine (TCM) has achieved beneficial results in the treatment of OP. Ziyuglycoside II (ZGS II) is a major active compound of Sanguisorba officinalis L. that has shown anti-inflammation and antioxidation properties, but little information concerning its anti-OP potential is available. Our research aims to investigate the mechanism of ZGS II in ameliorating bone loss by inflammatory responses and regulation of gut microbiota and short chain fatty acids (SCFAs) in ovariectomized (OVX) mice. METHODS: We predicted the mode of ZGS II action on OP through network pharmacology and molecular docking, and an OVX mouse model was employed to validate its anti-OP efficacy. Then we analyzed its impact on bone microstructure, the levels of inflammatory cytokines and pain mediators in serum, inflammation in colon, intestinal barrier, gut microbiota composition and SCFAs in feces. RESULTS: Network pharmacology identified 55 intersecting targets of ZGS II related to OP. Of these, we predicted IGF1 may be the core target, which was successfully docked with ZGS II and showed excellent binding ability. Our in vivo results showed that ZGS II alleviated bone loss in OVX mice, attenuated systemic inflammation, enhanced intestinal barrier, reduced the pain threshold, modulated the abundance of gut microbiota involving norank_f__Muribaculaceae and Dubosiella, and increased the content of acetic acid and propanoic acid in SCFAs. CONCLUSIONS: Our data indicated that ZGS II attenuated bone loss in OVX mice by relieving inflammation and regulating gut microbiota and SCFAs.

Chlorogenic acid prevents ovariectomized-induced bone loss by facilitating osteoblast functions and suppressing osteoclast formation.

Osteoporosis is a usual bone disease in aging populations, principally in postmenopausal women. Anti-resorptive and anabolic drugs have been applied to prevent and cure osteoporosis and are associated to a different of adverse effects. Du-Zhong is usually applied in Traditional Chinese Medicine to strengthen bone, regulate bone metabolism, and treat osteoporosis. Chlorogenic acid is a major polyphenol in Du-Zhong. In the current study, chlorogenic acid was found to enhance osteoblast proliferation and differentiation. Chlorogenic acid also inhibits the RANKL-induced osteoclastogenesis. Notably, ovariectomy significantly decreased bone volume and mechanical properties in the ovariectomized (OVX) rats. Administration of chlorogenic acid antagonized OVX-induced bone loss. Taken together, chlorogenic acid seems to be a hopeful molecule for the development of novel anti-osteoporosis treatment.

Hyaluronan and chondroitin sulfate in chicken-vegetable bone broth delay osteoporosis progression.

Bone broth has recently gained worldwide recognition as a superfood that supplements several nutrients lacking in modern human diets; however, little is known of its efficacy on osteoporosis. Therefore, we aimed to identify the components of chicken-vegetable bone broth (CVBB) that are associated with osteoporosis prevention and verified the efficacy of these components using in vivo studies. In biochemical and cell biological experiments, CVBB was fractionated using ion exchange chromatography (IEC), and the effect of each IEC fraction on osteoclast differentiation was evaluated based on tartrate-resistant acid phosphatase (TRAP) activity, TRAP staining, and quantitative polymerase chain reaction analysis using mouse macrophage-like cells (RAW264 cell). In animal experiments, an ovariectomized (OVX) rat model was generated, followed by whole bone broth (OVX/CVBB) or IEC fraction (OVX/CVBB-Ext) administration and bone structural parameter characterization of OVX rat tibia based on micro-CT. Four CVBB fractions were obtained using IEC, and the fraction containing both hyaluronan and chondroitin sulfate (CVBB-Ext) led to the maximum inhibition of RAW264 cell differentiation. CVBB-Ext downregulated the expression of osteoclast differentiation marker genes. In animal experiments, the OVX group showed a clear decrease in bone density compared to that in the Sham operation group. The OVX/CVBB and OVX/CVBB-Ext groups showed increased bone mineral density and bone volume/tissue volume values compared to those in the OVX/control group. These results suggested that CVBB and CVBB-Ext slowed osteoporosis progression. Therefore, we conclude that hyaluronan and chondroitin sulfate in CVBB are key substances that impede osteoporosis progression. PRACTICAL APPLICATION: This study provides practical information on the effects of bone broth ingredients on osteoporosis to expand the current knowledge on the efficacy of bone broth, which is a widely consumed food. These results may help in the future development of bone broth as a dietary supplement for managing osteoporosis.

Non-targeted metabolomics strategy reveals the role of Geng-Nian-Shu in regulating ferroptosis in perimenopausal syndrome.

Ovariectomy (OVX) is usually accompanied by the occurrence of metabolic syndrome. Previous studies have shown that Geng-Nian-Shu (GNS) plays an important regulatory role in perimenopausal syndrome (PMS) rats. GNS is a traditional Chinese medicine (TCM) prescription which composed of Suanzaoren Decoction and Ganmai Dazao Decoction in "Jingui Yaolue" and Siwu Decoction in "Heji Jufang". Recently, metabolomics analysis has been used to identify slight changes in the metabolic profile and to help understand disease progression and therapeutic interventions in PMS. However, the mechanism of GNS in the treatment of PMS is still unknown. We purposed to study the metabolic characteristics of PMS by serum and fecal metabolomics, and revealed the internal mechanism of GNS regulating ferroptosis against PMS. The PMS model was established by surgical removal of 4/5 ovaries of rats. HPLC-Q-TOF/MS was used to analyze the metabolomics of rat plasma and feces to explore the potential mechanism of GNS in PMS. The expression of ferroptosis-related proteins in rat ovaries was detected by tissue Prussian blue staining, Elisa kit and Western blotting. Cluster analysis of differential metabolites in plasma and feces between the control group and the model group showed that organic acids and their derivatives, lipids and lipid molecules were mainly disturbed during PMS in rats. After GNS administration, 17 differential metabolites were adjusted, involving several major pathways, such as the tricarboxylic acid (TCA) cycle, biosynthesis of amino acids and biosynthesis of unsaturated fatty acids. Further, we found that GNS affected ferroptosis in ovarian cells by regulating endogenous substances in OVX rats. Our study provides new insights into the mechanism of OVX-induced metabolic syndrome based on non-targeted metabolomics. It provides new ideas for the development and application of GNS and the diagnosis and treatment of PMS.

Advantage of ostarine over raloxifene and their combined treatments for muscle of estrogen-deficient rats.

PURPOSE: Selective androgen (ostarine, OST) and estrogen (raloxifene, RAL) receptor modulators with improved tissue selectivity have been developed as alternatives to hormone replacement therapy. We investigated the combined effects of OST and RAL on muscle tissue in an estrogen-deficient rat model of postmenopausal conditions. METHODS: Three-month-old Sprague Dawley rats were divided into groups: (1) untreated non-ovariectomized rats (Non-OVX), (2) untreated ovariectomized rats (OVX), (3) OVX rats treated with OST, (4) OVX rats treated with RAL, (5) OVX rats treated with OST and RAL. Both compounds were administered in the diet. The average dose received was 0.6 ± 0.1 mg for OST and 11.1 ± 1.2 mg for RAL per kg body weight/day. After thirteen weeks, rat activity, muscle weight, structure, gene expression, and serum markers were analyzed. RESULTS: OST increased muscle weight, capillary ratio, insulin-like growth factor 1 (Igf-1) expression, serum phosphorus, uterine weight. RAL decreased muscle weight, capillary ratio, food intake, serum calcium and increased Igf-1 and Myostatin expression, serum follicle stimulating hormone (FSH). OST + RAL increased muscle nucleus ratio, uterine weight, serum phosphorus, FSH and luteinizing hormone and decreased body and muscle weight, serum calcium. Neither treatment changed muscle fiber size. OVX increased body and muscle weight, decreased uterine weight, serum calcium and magnesium. CONCLUSION: OST had beneficial effects on muscle in OVX rats. Side effects of OST on the uterus and serum electrolytes should be considered before using it for therapeutic purposes. RAL and RAL + OST had less effect on muscle and showed endocrinological side effects on pituitary-gonadal axis.

The bone-protective benefits of amino-conjugated calcium in an ovariectomized (OVX) rat model.

Low bone density, fragility, and microarchitectural disintegration are the symptoms of osteoporosis. An imbalance between bone growth and resorption can lead to osteoporosis. This study evaluated the effects of amino-calcium (AC) on bone protection in ovariectomized control group (NC) rats. Amino-calcium (AC) was characterized using Fourier-transform infrared spectroscopy (FT-IR), energy-dispersive X-ray spectroscopy (EDS), and nuclear magnetic resonance spectroscopy analyses (NMR). After determining the biocompatibility of amino-calcium (AC) with MC3T3-E1 cells, alkaline phosphatase staining revealed significant changes on day 7. Three of the four groups underwent ovariectomy, whereas one group received a placebo. On micro-computed tomography, in vivo, data showed increased bone volume fraction in the femoral head and shaft areas in the amino-calcium (AC) group. Hematoxylin and eosin staining showed a bone mass and architectural protection in the amino-calcium (AC) group compared with the calcium carbonate and OVX control group. RNA sequencing analysis revealed high expression of osteogenesis-related genes in MC3T3-E1 cells. RNA sequencing revealed a significant fold change in the expression of integrin-binding sialoprotein (IBSP), bone gamma-carboxyglutamate proteins 1 and 2(BGLAP1 and BGLAP2), and periostin (POSTN). The study concluded that supplementing the OVX rats with calcium enhanced bone protection.

[Adjuvant rice optimization of rice-steamed Rehmanniae Radix and its anti-osteoporosis effect].

The present study aimed to investigate the effect of various adjuvant rice on the quality of rice-steamed Rehmanniae Radix(RSRR) with Japonica rice, millet, yellow rice, black rice, and glutinous rice as raw materials, and analyze the anti-osteoporosis effect of RSRR by the optimal adjuvant rice. On the basis of the established UPLC-MS/MS method for the determination of the content of catalpol and rehmannioside D, comprehensive weighted scoring method was employed to evaluate the effect of various auxiliary rice on the quality of RSRR with the content of catalpol and rehmannioside D, character score, and taste score as indicators to optimize adjuvant rice. The osteoporosis model was induced by ovariectomy in rats. SD rats were randomly divided into a sham operation group, a model group, a positive control group, and low-dose and high-dose groups of Rehmanniae Radix, RSRR, steamed Rehmanniae Radix, and Epimedii Folium-RSRR. After treatment for 12 weeks, body weight, bone calcium content, and bone mineral density were mea-sured. The results showed that Japonica rice was selected as the optimal adjuvant due to the highest comprehensive score of RSRR steamed by Japonica rice. Rehmanniae Radix, RSRR, steamed Rehmanniae Radix, as well as Epimedii Folium-RSRR, could improve osteoporosis by increasing bone calcium content and bone mineral density. RSRR was superior to Rehmanniae Radix in improving osteo-porosis. However, there was no significant difference between RSRR and steamed Rehmanniae Radix. This study confirmed that Japo-nica rice was the optimal adjuvant rice of RSRR and verified the anti-osteoporosis effect of RSRR, which laid a foundation for further research on the pharmacological action and mechanism of RSRR.

Investigation of taurine and aqueous garlic extract diet supplementation effect on the healing of rat osteoporotic fractures.

BACKGROUND: We have evaluated the effects of taurine and aqueous garlic extract (AGE) as a dietary supplement on osteoporotic fracture (OPF) healing in the ovariectomized rat femur fracture model. METHODS: In this experimental animal study,twenty-four osteoporosis-remodeled female Wistar albino rats were randomly divided into 3 groups (n: 8) according to their supplemented diet; control, taurine, and AGE groups. Unilateral femur middiaphysis mini-open osteotomy was stabilized with Kirschner wires. Six weeks after osteotomy, the rats were sacrificed before the femurs were harvested and OPF healing was evaluated with biochemical, histologic, microcomputed-tomography, and scintigraphic methods. RESULTS: As an indicator of the antiosteoporotic effect, the calcium levels of the taurine group were significantly lower than the AGE and control groups in biochemical analyzes (p < 0.01). In histological studies, the new bone diameter and new bone volume values of the taurine group were significantly higher than the control group (p = 0.002 and p = 0.032, respectively), while higher trabecular-compact callus was observed in the taurine and AGE groups, respectively, compared to the control group. In morphological analyses, taurine and AGE groups had significantly higher bone volume/tissue volume, trabecular number, bone surface density, and lower trabecular separation than the control group (p < 0.05). The scintigraphic imaging showed a significant increase in osteoblastic activity of the taurine group compared to the control group (p = 0.005). DISCUSSION: Taurine and AGE have positive anabolic effects, respectively, on the healing of OPFs, demonstrated by biochemical, histological, morphological, and scintigraphic methods.

Ellagic acid inhibits CDK12 to increase osteoblast differentiation and alleviate osteoporosis in hindlimb-unloaded and ovariectomized mice.

BACKGROUND: Osteoporosis is a highly prevalent bone disease occurred commonly in astronauts and postmenopausal women due to mechanical unloading and estrogen deficiency, respectively. At present, there are some traditional Chinese medicine compounds for preventing and treating osteoporosis induced by simulated microgravity, but the detailed components of the traditional Chinese medicines still need to be confirmed and osteoporosis is still untreatable due to a lack of effective small-molecule natural medicine. PURPOSE: To explore the role of cyclin-dependent kinase 12 (CDK12) in osteoporosis induced by simulated microgravity and the therapeutic effect of CDK12-targeted Ellagic Acid (EA) on osteoporosis. METHODS: Our previous study has suggested that CDK12 as a potential target for treating and preventing osteoporosis. In this study, the role of CDK12 in osteoblasts and mice bone tissues was further studied under simulated microgravity. And by targeting CDK12, natural small-molecule product EA was screened out based on a large scale through the weighted set similarity (WES) method and the therapeutic effects of EA on osteoporosis was investigated in hindlimb-unloaded (HU) mouse model and ovariectomized (OVX) model. RESULTS: The results demonstrated that simulated microgravity inhibited bone formation and up-regulated the expression of CDK12. Furthermore, CDK12-siRNA or THZ531 (an inhibitor of CDK 12) promoted osteoblast differentiation, while the overexpression of CDK12 inhibited osteoblasts differentiation. And we further proved that CDK12-targeted EA showed a rescue effect on osteoblast differentiation inhibition caused by simulated microgravity. EA (50 mg·kg-1·day-1) daily intragastric administration alleviated the symptoms of osteoporosis and accompanied with the improvement of trabecular bone and cortical bone parameters with significantly overexpression of CDK12. CONCLUSION: EA efficiently improves osteoporosis by targeting CDK12, which is a suppresser of osteoblast differentiation and a novel therapeutic target for treating osteoporosis.

Protective Effects of Coumestrol on Metabolic Dysfunction and Its Estrogen Receptor-Mediated Action in Ovariectomized Mice.

Coumestrol, a phytoestrogen compound found in various plants, has been shown to act as a potent estrogen receptor (ER) agonist, with a higher binding affinity for ERß than for ERα. However, there is currently limited information regarding its beneficial effects in postmenopausal disorders and its ER-mediated mechanisms. Herein, we investigated the effects of coumestrol (subcutaneous or oral treatment) on metabolic dysfunction in ovariectomized (OVX) mice fed a high-fat diet, in comparison with the effects of 17ß-estradiol (E2) replacement. Coumestrol was administered daily at a dose of 5 mg/kg for 10 weeks. Coumestrol treatment through the subcutaneous route stimulated uterine growth in OVX mice at a level lower than that of E2. E2 and coumestrol prevented body fat accumulation, adipocyte hypertrophy, and hepatic steatosis, and enhanced voluntary physical activity. Coumestrol showed estrogen-mimetic effects in the regulation of the protein expressions involved in browning of white fat and insulin signaling, including increased hepatic expression of fibroblast growth factor 21. Importantly, the metabolic effects of coumestrol (oral administration at 10 mg/kg for 7 weeks) were mostly abolished following co-treatment with an ERß-selective antagonist but not with an ERα-selective antagonist, indicating that the metabolic actions of coumestrol in OVX mice are primarily mediated by ERß. These findings provide important insights into the beneficial effects of coumestrol as a phytoestrogen supplement for the prevention and treatment of postmenopausal symptoms.

Glycyrrhizae radix et Rhizoma-Derived Carbon Dots and Their Effect on Menopause Syndrome in Ovariectomized Mice.

With the extension of the human life span and the increasing pressure of women's work and life, menopause syndrome (MPS) refers to a problem that puzzles almost all women worldwide. Hormone replacement treatment (HRT) can effectively mitigate the symptoms but can also exert adverse effects to a certain extent. Glycyrrhizae radix et rhizome (GRR) is commonly made into a charcoal processed product, termed GRR Carbonisatas (GRRC), for use in traditional Chinese medicine (TCM). GRRC is widely used to treat MPS and other gynecological diseases. In this study, GRRC was prepared through pyrolysis. Subsequently, GRR-derived carbon dots (GRR-CDs) were purified through dialysis and characterized using transmission electron microscopy, high-resolution transmission electron microscopy, Fourier-transform infrared, ultraviolet, fluorescence, X-ray photoelectron microscopy, and high-performance liquid chromatography. The effects of GRR-CDs on MPS were examined and confirmed using ovariectomized female mice models. The GRR-CDs ranged from 1.0 to 3.0 nm in diameter and with multiple surface chemical groups, as indicated by the results. GRR-CDs can elevate the estradiol (E2) level of healthy female mice. Moreover, GRR-CDs can alleviate MPS using the typical ovariectomized mice model, as confirmed by elevating the estradiol (E2) level and reducing the degree of follicle stimulating hormone (FSH) and luteinizing hormone (LH) and raising the degree of uterine atrophy. The results of this study suggested that GRR-CDs may be a potential clinical candidate for the treatment of MPS, which also provides a possibility for nanodrugs to treat hormonal diseases.

Anti-hyperplastic effects of the Dacryodes edulis (Burseraceae) leaves aqueous extract on tamoxifen-induced endometrium hyperplasia on Wistar rat.

OBJECTIVES: Combining tamoxifen, the most common breast cancer hormonal therapy, with natural antitumor substances may prevent its hyperplastic effects on the uterine endometrium. Dacryodes edulis (DE) is traditionally recommended for the treatment of cancerous diseases. To investigate its antiproliferative properties, the present study was designed to assess the ability of the combined administration of tamoxifen with the aqueous extract of DE leaves to inhibit the trophic effect of this hormone therapy on rat uterine endometrium without compromising its non-proliferative effect on breast tissue. METHODS: Ovariectomized (OVX) female Wistar rats were simultaneously treated with tamoxifen (10 mg/kg) intraperitoneally and DE leaves (at doses of 25, 50 and 100 mg/kgBW) by gavage. Control groups received either distilled water or tamoxifen alone. Treatments lasted 37 days. The 38th day, animals were sacrificed under anesthesia (diazepam: 10 mg/kgBW and ketamine: 50 mg/kgBW). The relative uterine weight was determined and the histological analysis of the uterus and mammary gland was performed. The oxidative status of the uterus was assessed and the levels of cholesterol and estradiol were evaluated in serum and uterus. RESULTS: Tamoxifen increased uterine weight and induced endometrial hyperplasia. This effect was associated with increased uterine levels of cholesterol (164.22%; p < 0.001), estradiol (927.5%; p < 0.001) and malondiadehyde (86%; p < 0.05), but unchanged antioxidant enzymes activities. The administration of DE leaves unchanged tamoxifen-increased uterine weight but reduced uterine epithelium hypertrophy (56.4%; p < 0.01). DE also increased uterine levels of malondiadehyde and antioxidant enzymes. The levels of estradiol and cholesterol in the uterus decreased while no changes were observed in the mammary gland of animals treated with tamoxifen alone or in co-administration with DE. CONCLUSIONS: D. edulis has antiproliferative properties and could complement endocrine therapy of estrogen-dependent breast cancer.

Oral Administration of Artemisia argyi Polysaccharide Increases Estrogen Level and Maintains Blood Lipid Homeostasis in Ovariectomized Rats.

INTRODUCTION: Artemisia argyi polysaccharide (AAP) has a beneficial effect on menstruation-related symptoms and the potential regulation of lipid metabolism. It is expected to be a safe and effective ingredient for estrogen deficiency and lipid metabolic disorders. Here, we investigate the effect of AAP on body weight gain, estrogen level, and blood lipid changes in ovariectomized (OVX) rats. METHODS: Thirty-six female Wistar rats were randomly divided into six treatment groups, including a sham-operated (Sham) group, OVX group, estrogen replacement (OVX + E2) group, and AAP treatment (OVX + 125, 250, 500 mg/kg AAP) group. The body weight and feed intake were recorded every week. The level of estrogen and blood lipid was determined. The gene expressions and protein expressions of estrogen receptors (ERs), fatty acid synthetase (FAS), acetyl CoA carboxylase 2 (ACC2), and 3-hydroxy-3-methyl glutaryl coenzyme A reductase (HMGR) were determined. RESULTS: AAP treatment significantly decreased the body weight gain and average daily food intake of rats in the OVX group. Treatment with AAP significantly increased the relative weight of the uterus, plasma estrogen level, and the gene expression and protein expression of ER-α in the uterus. For blood lipids, plasma levels of triglyceride, total cholesterol, and low-density lipoprotein cholesterol were significantly reduced by AAP treatment in OVX rats. AAP treatment decreased the expression of FAS and HMGR in the liver of OVX rats. Furthermore, AAP treatment significantly increased the gene expression of ACC2, the protein expression of P-ACC2, and the ratio of P-ACC2/ACC2. CONCLUSION: In summary, AAP treatment exerts beneficial effects on body weight gain and lipid metabolism disorder induced by ovariectomy through increasing estrogen levels, inhibiting FAS, and promoting fatty acid oxidation.

Adjuvant rice optimization of rice-steamed Rehmanniae Radix and its anti-osteoporosis effect / 中国中药杂志

The present study aimed to investigate the effect of various adjuvant rice on the quality of rice-steamed Rehmanniae Radix(RSRR) with Japonica rice, millet, yellow rice, black rice, and glutinous rice as raw materials, and analyze the anti-osteoporosis effect of RSRR by the optimal adjuvant rice. On the basis of the established UPLC-MS/MS method for the determination of the content of catalpol and rehmannioside D, comprehensive weighted scoring method was employed to evaluate the effect of various auxiliary rice on the quality of RSRR with the content of catalpol and rehmannioside D, character score, and taste score as indicators to optimize adjuvant rice. The osteoporosis model was induced by ovariectomy in rats. SD rats were randomly divided into a sham operation group, a model group, a positive control group, and low-dose and high-dose groups of Rehmanniae Radix, RSRR, steamed Rehmanniae Radix, and Epimedii Folium-RSRR. After treatment for 12 weeks, body weight, bone calcium content, and bone mineral density were mea-sured. The results showed that Japonica rice was selected as the optimal adjuvant due to the highest comprehensive score of RSRR steamed by Japonica rice. Rehmanniae Radix, RSRR, steamed Rehmanniae Radix, as well as Epimedii Folium-RSRR, could improve osteoporosis by increasing bone calcium content and bone mineral density. RSRR was superior to Rehmanniae Radix in improving osteo-porosis. However, there was no significant difference between RSRR and steamed Rehmanniae Radix. This study confirmed that Japo-nica rice was the optimal adjuvant rice of RSRR and verified the anti-osteoporosis effect of RSRR, which laid a foundation for further research on the pharmacological action and mechanism of RSRR.

Prebiotics improve osteoporosis indicators in a preclinical model: systematic review with meta-analysis.

CONTEXT: Studies using experimental models have demonstrated that prebiotics are involved in antiosteoporotic mechanisms. OBJECTIVE: This study was conducted to determine the impact of supplementation with prebiotics in the basal diet of ovariectomized rats with induced osteoporosis as a preclinical model. METHODS: A comprehensive systematic search was carried out in the electronic databases PubMed, Science Direct, Web of Science, Scielo, and Google through March 2022 for studies that investigated the impact of prebiotics on bone mineral density (BMD), bone mineral content (BMC), and bone biomechanics. RESULTS: The search returned 844 complete articles, abstracts, or book chapters. After detailed screening, 8 studies met the inclusion criteria. Rats (n = 206), were randomly divided between control and treatment groups. Weighted mean differences (WMDs) with the 95%CIs were used to estimate the combined effect size. Compared with the control group, dietary intake of prebiotics significantly increased bone density in the BMD subgroups, with WMDs as follows: 0.03 g/cm3, 95%CI, 0.01-0.05, P < 0.00001, n = 46; and 0.00 g/cm2, 95%CI, 0.00-0.02, P < 0.00001, n = 81; total BMD: WMD, 0.01, 95%CI, 0.01-0.02, P < 0.00001, n = 127; bone content in BMC: WMD, 0.02 g, 95%CI, 0.00-0.04, P = 0.05, n = 107; and the 3-point-bend test: WMD, 15.20 N, 95%CI, 5.92-24.47, P = 0.00001, n = 120. CONCLUSION: Prebiotics improve indicators of osteoporosis, BMD, BMC, and bone biomechanics in ovariectomized rats. More studies are needed to increase the level of evidence. SYSTEMIC REVIEW REGISTRATION: Systematic Review Protocol for Animal Intervention Studies.

Integration of pharmacodynamics and metabolomics reveals the therapeutic effects of 6-acetylacteoside on ovariectomy-induced osteoporosis mice.

BACKGROUND: 6-acetylacteoside (6-AA) is a phenylethanoid glycoside isolated from Cistanche deserticola which had been previously proven to possess anti-osteoporotic activity previously. Currently, it is still unknown whether 6-AA plays a crucial role on the anti-osteoporotic effects of C. deserticola. PURPOSE: To elucidate the therapeutic effect and mechanism of 6-AA on osteoporosis by employing an ovariectomized mouse model in vivo and RAW264.7 cells in vitro. METHODS: Sixty female ICR mice were randomly assigned into six groups: sham-operated control group (SHAM, vehicle), ovariectomized model group (OVX, vehicle), positive group (EV, 1 mg/kg/day of estradiol valerate), low dosage (10 mg/kg/day of 6-AA), medium dosage (20 mg/kg/day of 6-AA) and high dosage (40 mg/kg/day of 6-AA) treatment groups. All substances were administered daily by intragastric gavage. After 12 weeks of intervention, trabecular bone microarchitecture was estimated and bone biomechanics were determined. Bone formation and resorption factors were determined by using the corresponding Elisa kits. The related proteins and metabolites were estimated by using western-blot and metabolomics techniques. RESULTS: OVX mice demonstrated significant atrophy of the uterine and vagina, declined biomechanical parameters such as flexural strength and maximum load, deteriorated trabecular bone microarchitecture such as decreased BMD, BMC, TMC, TMD, BVF, Tb. N, and Tb. Th and increased Tb. Sp, as well as increased bone resorption factors such as TRAP, cathepsin K, and DPD, all after 12 weeks of ovariectomy operation. Following administration of 6-AA to OVX mice, parameters related to the bone microarchitecture, bone resorption activities as well as biomechanical properties were all significantly improved. Meanwhile, the levels of NF-κB, NFATc1, RANK, RANKL and TRAF6 were significantly downregulated, while OPG, PI3K and AKT were upregulated after 6-AA intervention. This indicates that, 6-AA could prevent bone resorption by regulating the RANKL/RANK/OPG mediated NF-κB and PI3K/AKT pathways. Furthermore, 26 different metabolites corresponding to 25 metabolic pathways were identified, and 5 of which were related to the formation of osteoporosis. Interestingly, 23 abnormal metabolites were recovered after 6-AA treatment. CONCLUSION: Our results revealed the significant anti-osteoporotic effects of 6-AA on ovariectomized mice which were probably exerted via suppression of osteoclast formation and bone resorption.

Effect of Rhizoma Drynariae on differential gene expression in ovariectomized rats with osteoporosis based on transcriptome sequencing.

Osteoporosis is increasingly becoming a serious problem affecting the quality of life of the older population. Several experimental studies have shown that Chinese medicine has a definite effect on improving osteoporosis. Based on transcriptome sequencing, we analyzed the differential gene expression and mechanism of the related signaling pathways. Fifteen rats were randomly divided into an experimental group, a model group, and a sham surgery group. The rat model for menopausal osteoporosis was established using an ovariectomy method. One week after modeling, the experimental group was administered(intragastric administration)8.1 g/kg of Rhizoma drynariae, whereas the model and sham groups received 0.9% saline solution twice daily for 12 weeks. Subsequently, the rats were sacrificed, and the left femur of each group was removed for computerized tomography testing, while right femurs were used for hematoxylin and eosin staining. High-throughput RNA sequencing and functional and pathway enrichment analyses were performed. Comparing the gene expression between the experimental and model groups, 149 differential genes were identified, of which 44 were downregulated and 105 were upregulated. The criteria for statistical significance were |log2 Fold Change| > 1 and P < 0.05. Gene ontology analysis showed that the differentially expressed genes were enriched in cell component terms such as cell part and outer cell membrane part, and the genes were associated with cell process, biological regulation, metabolic processes, DNA transcription, and catalytic activity. Enrichment analysis of Kyoto Encyclopedia of Genes and Genomes pathways showed significantly enriched pathways associated with systemic lupus erythematosus, herpes simplex infection, circadian rhythm, vascular smooth muscle contraction, the AGE-RAGE signaling pathway in diabetic complications, and the TNF, Apelin, and Ras signaling pathways. Our results revealed that the Npas2, Dbp, Rt1, Arntl, Grem2, H2bc9, LOC501233, Pla2g2c, Hpgd, Pde6c, and Dner genes, and the circadian rhythm, lipid metabolism, inflammatory signaling pathway, and immune pathways may be the key targets and pathways for traditional Chinese medicine therapy of Rhizoma Drynariae in osteoporosis.

Comprehensive metabolite profiling of Phoenix rupicola pulp and seeds using UPLC-ESI-MS/MS and evaluation of their estrogenic activity in ovariectomized rat model.

Dates have been consumed since ancient times as functional foods which beside their high nutritional value possess various biological activities. Phoenix rupicola T. Anderson (Cliff date palm) produces non-conventional edible dates, however, due to low natural abundance, these dates aren't commercially important as the dates of Phoenix dactylifera L. The present study was designed to evaluate the phytochemical constituents as well as the estrogenic activity of P. rupicola dates. UPLC-ESI-MS/MS approach was used to study the metabolite profile of the 70% aqueous methanol extracts of P. rupicola dates (pulps and seeds) for the first time. A total of fifty-five metabolites were tentatively identified in both extracts, belonging to different classes, chiefly phenolic compounds viz. procyanidins, flavonoid glycosides, hydroxycinnamic acid derivatives, as well as, fatty acids, organic acids and sphingolipids. Acute toxicity studies revealed that the studied extracts were safe at oral doses up to 2 g/kg. Besides, they possessed significant (P < 0.05) estrogenic activity in ovariectomized rat model, as compared to ovariectomized (OVX) and reference standard (17ß-estradiol; OVX-E) groups. Moreover, the extracts showed significant improvement on bone metabolism, lipid profile, liver and kidney functions. In silico docking study revealed that various metabolites possessed high binding affinities to both ERs, where 2-palmitoyl glycerol (-10.28 Kcal/mol) and aminotetradecanetriol (-9.61 Kcal/mol) showed the strongest affinities to Erα and Erß, respectively. Thus, it can be concluded that P. rupicola pulp and seeds possess bioactive phytoconstituents comparable to those in P. dactylifera and can be used as a safe and efficient natural estrogen substitute in postmenopausal women.

Effects of isoflavone supplementation on endometrial thickness, endometrial hyperplasia, and cancer in ovariectomized cats.

Ovariectomy is identified as a standard treatment in different European countries. Isoflavones, as nonsteroidal compounds in plants, are common constituents of soy and soy products. Some available cat diets contain different concentrations of soy products. This study aimed to examine the effects of isoflavone supplementation on endometrial hyperplasia and endometrial thickness in ovariectomized cats. Fifteen neutered adult cats were divided into control, estradiol, and isoflavone groups (five cats per group). Subcutaneous injection of estradiol (0.5 µg) in sesame oil (100 µL) was done for 30 days in estradiol-treated cats. Isoflavone-treated cats ingested a single oral tablet of soy extract for 30 days, while the controls received subcutaneous injections of the vehicle and oral placebo for 30 days. Histopathological findings of hematoxylin and eosin-stained sections revealed a significant difference between the estradiol group and other groups in terms of hyperplastic epithelium and simple hyperplasia. Thickness of myometrium was greater in the estradiol group compared to the isoflavone and control groups. Higher concentrations of estrogen can affect the endometrium and myometrium, while 30-day ingestion of isoflavone didn't have any uterine effect.