Effect of Tempeh and Daidzein on Calcium Status, Calcium Transporters, and Bone Metabolism Biomarkers in Ovariectomized Rats.

Menopause marks a critical life stage characterized by hormonal changes that significantly impact bone health, leading to a heightened susceptibility to bone fractures. This research seeks to elucidate the impact of daidzein and tempeh on calcium status, calcium transporters, and bone metabolism in an ovariectomized rat model. Forty female Wistar rats, aged 3 months, participated in a two-phase experiment. The initial phase involved inducing a calcium deficit, while the second phase comprised dietary interventions across five groups: Sham (S) and Ovariectomy (O) with a standard diet, O with bisphosphonate (OB), O with pure daidzein (OD), and O with tempeh (OT). Multiple parameters, encompassing calcium levels, calcium transporters, bone histopathology, and serum bone metabolism markers, were evaluated. The findings revealed that the OT group showcased heightened levels of bone turnover markers, such as pyridinoline, C-telopeptide of type I collagen, bone alkaline phosphatase, and procollagen type I N-terminal propeptide, in contrast to S and O groups, with statistical significance (p < 0.05). Histopathologically, both the OD and OT groups exhibited effects akin to the OB group, indicating a decrease in the surface area occupied by adipocytes in the femoral bone structure, although statistically non-equivalent, supporting the directionally similar trends. Although TRPV5 and TRPV6 mRNA expression levels in the jejunum and duodenum did not display statistically significant differences (p > 0.05), the OD and OT groups exhibited increased expression compared to the O group. We hypothesized that obtained results may be related to the effect of isoflavones on estrogen pathways because of their structurally similar to endogenous estrogen and weak estrogenic properties. In conclusion, the daily consumption of pure daidzein and tempeh could potentially improve and reinstate calcium status, calcium transport, and bone metabolism in ovariectomized rats. Additionally, isoflavone products demonstrate effects similar to bisphosphonate drugs on these parameters in ovariectomized rats.

Nodakenin Ameliorates Ovariectomy-Induced Bone Loss by Regulating Gut Microbiota.

Disordered gut microbiota (GM) structure and function may contribute to osteoporosis (OP). Nodakenin has been shown to ameliorate osteoporosis; however, its anti-osteoporotic mechanism is unknown. This study aimed to further reveal the mechanism of the anti-osteoporotic action of nodakenin from the perspective of the microbiome and metabolome. An osteoporosis model was induced in mice through ovariectomy (OVX), with bone mass and microstructure assessed using µCT. Subsequently, ELISA and histologic examination were used to detect biochemical indicators of bone conversion and intestinal morphology. Using metabolomics and 16S rRNA sequencing, it was possible to determine the composition and abundance of the gut microbiota in feces. The results revealed that nodakenin treatment improved the bone microstructure and serum levels of bone turnover markers, and increased the intestinal mucosal integrity. 16S rRNA sequencing analysis revealed that nodakenin treatment decreased the relative abundance of Firmicutes and Patescibacteria, as well as the F/B ratio, and elevated the relative abundance of Bacteroidetes in OVX mice. In addition, nodakenin enhanced the relative abundance of Muribaculaceae and Allobaculum, among others, at the genus level. Moreover, metabolomics analysis revealed that nodakenin treatment significantly altered the changes in 113 metabolites, including calcitriol. A correlation analysis revealed substantial associations between various gut microbiota taxa and both the osteoporosis phenotype and metabolites. In summary, nodakenin treatment alleviated OVX-induced osteoporosis by modulating the gut microbiota and intestinal barrier.

Ozone therapy improves early stages of osseointegration in ovariectomized rats.

OBJECTIVE: the aim of this study was to analyze the influence of ozone therapy (OZN) on peri-implant bone repair in critical bones by installing osseointegrated implants in the tibia of ovariectomized rats. METHODOLOGY: ovariectomy was performed on 30 Wistar rats, aged six months (Rattus novergicus), and, after 90 days, osseointegrated implants were installed in each tibial metaphysis. The study groups were divided into the animals that received intraperitoneal ozone at a concentration of 700 mcg/kg - OZ Group (n=15) - and a control group that received an intraperitoneal saline solution and, for this reason, was named the SAL group (n=15). The applications for both groups occurred during the immediate post-operative period on the 2nd, 4th, 6th, 8th, 10th, and 12th day post-surgery. At various stages (14, 42, and 60 days), the animals were euthanized, and tests were performed on their tibiae. These tests include histomorphometric and immunohistochemical analyses, computerized microtomography, sampling in light-cured resin for calcified sections, and confocal microscopy. The obtained data were then analyzed using One-way ANOVA and the Shapiro-Wilk, Kruskal-Wallis, and student t-tests (P<0.05). RESULTS: our findings indicate that the OZ group (3.26±0.20 mm) showed better cellular organization and bone neoformation at 14 days (SAL group, 0.90±1.42 mm) (P=0.001). Immunohistochemistry revealed that osteocalcin labeling was moderate in the OZ group and mild in the SAL group at 14 and 42 days post-surgery. The data from the analysis of calcified tissues (microtomography, histometric, and bone dynamism analysis) at 60 days showed no statistically significant differences between the groups (P=0.32). CONCLUSION: it was concluded that ozone therapy anticipated the initial phases of the peri-implant bone repair process.

Inhibition of NF-κB and ERK signaling pathways in osteoclasts and M1 macrophage polarization: Mechanistic insights into the anti-osteoporotic effects of Pseudolaric acid B.

Osteoporosis, characterized by bone metabolism disruption leading to gradual bone loss and increased fracture susceptibility, is linked to the excessive activation of osteoclasts. Pseudolaric acid B (PAB), identified as an NF-κB signaling inhibitor crucial for osteoclastogenesis, is explored here for its protective effects in osteoporosis. Noncytotoxic PAB's impact on osteoclast differentiation was assessed through cell viability and osteoclast formation assays, with subsequent testing of osteoclast function via bone resorption assays. Quantitative real-time polymerase chain reaction evaluated PAB's genetic-level impact on osteoclastogenesis. Network pharmacology, western blot, and luciferase reporter gene assays were employed to elucidate PAB's regulatory mechanism. In an in vivo model of osteoporosis induced by ovariectomy (OVX) in mice, micro-CT, H&E staining, and TRAP staining facilitated histomorphometry analysis, while flow cytometry verified macrophage polarization. PAB demonstrated inhibitory effects on osteoclast formation and bone resorption in BMM and RAW264.7 cells, suppressing osteoclast-specific genes. Bioinformatic analysis, western blot, and luciferase assay results indicated PAB's inhibition of IκBα phosphorylation in the NF-κB signaling pathway and ERK in MAPKs, elucidating its mechanism. In vivo experiments confirmed PAB's attenuation of osteoporosis by reducing osteoclast formation in OVX mice. PAB further facilitated macrophage conversion from M1 to M2 and suppressed IL-1ß, TNF-α, and IL-6 synthesis. In conclusion, PAB prevents osteoporosis by inhibiting RANKL-induced osteoclastogenesis through NF-κB and ERK signaling pathway suppression, coupled with macrophage polarization. These findings indicate the potential therapeutic role of PAB in osteoporosis.

Evaluation of the analgesic efficacy of grapiprant compared with robenacoxib in cats undergoing elective ovariohysterectomy in a prospective, randomized, masked, non-inferiority clinical trial.

OBJECTIVES: The main objective of this study was to compare the postoperative analgesic effects of grapiprant with those of robenacoxib in cats undergoing ovariohysterectomy (OVH). METHODS: In total, 37 female cats (age range 4 months-10 years, weighing ⩾2.5 kg) were enrolled in a prospective, randomized, masked, non-inferiority (NI) clinical trial. Cats received oral robenacoxib (1 mg/kg) or grapiprant (2 mg/kg) 2 h before OVH. Analgesia was assessed via the Feline Grimace Scale (FGS), the Glasgow Composite Measure Pain Scale-Feline (CMPS-F), von Frey monofilaments (vFFs) and pressure algometry (ALG) 2 h before treatment administration, at extubation, and 2, 4, 6, 8, 18 and 24 hours after extubation. Hydromorphone (<8 h postoperatively) or buprenorphine (>18 h postoperatively) were administered to cats with scores of ⩾5/20 on CMPS-F and/or ⩾4/10 on FGS. NI margins for CMPS-F and vFFs were set at 3 and -0.2, respectively. A mixed-effect ANOVA was used for FGS scores (P <0.05). Data are reported as mean ± SEM. RESULTS: The data from 33 cats were analyzed. The upper limit of the 95% confidence interval (CI) (0.35) was less than the NI margin of 3 for CMPS-F, and the lower limit of the 95% CI (0.055) was greater than the NI margin of -0.2 for vFFs, indicating NI of grapiprant. The FGS scores were greater than baseline at extubation for both treatments (1.65 ± 0.63; P = 0.001); however, there was no difference between treatments. There was no difference between treatments, nor treatment by time interaction, for vFFs (P <0.001). The CMPS-F scores for both treatments were higher at extubation but returned to baseline after 4 h (P <0.001). For ALG, there was no difference in treatment or treatment by time interaction. The robenacoxib group had lower pressure readings at extubation and 6 h compared with baseline. CONCLUSIONS AND RELEVANCE: These results indicate that grapiprant was non-inferior to robenacoxib for mitigating postsurgical pain in cats after OVH performed via ventral celiotomy. The impact of grapiprant for analgesia in OVH via the flank is unknown.

Comparison of analgesic efficacy of tramadol, morphine and methadone in cats undergoing ovariohysterectomy.

OBJECTIVES: The aim of this study was to compare the analgesic efficacy and the effect on physiological variables and behavior of the use of tramadol, methadone and morphine as preoperative analgesia in healthy cats undergoing elective ovariohysterectomy. METHODS: Cats undergoing ovariohysterectomy were randomly assigned to receive one of the following premedication treatments intramuscularly: methadone (0.2 mg/kg; n = 10); morphine (0.2 mg/kg; n = 10); or tramadol (3 mg/kg; n = 10). Induction of anesthesia was done with propofol, and maintenance of anesthesia was done with isoflurane. Intraoperative heart rate, arterial blood pressure, respiratory rate, end-tidal isoflurane concentration and frequency of rescue analgesia (fentanyl 2.5 µg/kg) were compared between groups. Postoperative analgesia was assessed using the UNESP-Botucatu Multidimensional Composite Pain Scale, and perioperative serum glucose, cortisol concentrations and postoperative rescue analgesia were evaluated. RESULTS: Intraoperative rescue analgesia was required in 76.5% of cats at some time during surgery, and 27% of cats required postoperative rescue analgesia up to 6 h after extubation. There were no significant differences between groups with respect to intraoperative and postoperative rescue analgesia, pain scale scores and end-tidal isoflurane concentrations. In the immediate postoperative period, after extubation, most of the patients presented with hypothermia; however, 1-6 h postoperatively, hyperthermia was observed in most of the patients, and was most common in the tramadol group. CONCLUSIONS AND CLINICAL RELEVANCE: Under the conditions of this study, methadone, morphine and tramadol produced satisfactory postoperative analgesia in most of the cats undergoing ovariohysterectomy, and the effects lasted up to 6 h postoperatively. Intraoperative analgesia was not sufficient in most cases. Significant cardiovascular or respiratory effects contraindicating the use of these drugs were not found. Postanesthetic hyperthermia occurred with all opioids studied and was more frequent in the tramadol group.

Ethanol extract of Cyathulae Radix inhibits osteoclast differentiation and bone loss.

Cyathulae Radix, a traditional Chinese medicine and a common vegetable, boasts a history spanning millennia. It enhances bone density, boosts metabolism, and effectively alleviates osteoporosis-induced pain. Despite its historical use, the molecular mechanisms behind Cyathulae Radix's impact on osteoporosis remain unexplored. In this study, we investigated the effects and mechanisms of Cyathulae Radix ethanol extract (CEE) in inhibiting osteoporosis and osteoclastogenesis. Eight-week-old female mice underwent ovariectomy and were treated with CEE for eight weeks. Micro-computed tomography (micro-CT) assessed histomorphometric parameters, bone tissue staining observed distal femur histomorphology, and three-point bending tests evaluated tibia mechanical properties. Enzyme-linked immunosorbent assay (ELISA) measured serum estradiol (E2), receptor activator for nuclear factor B ligand (RANKL), and osteoprotegerin (OPG) levels. Osteoclastogenesis-related markers were analyzed via Western blotting (WB) and quantitative real-time polymerase chain reaction (qRT-PCR). Additionally, CEE effects on RANKL-induced osteoclast formation and bone resorption were investigated in vitro using tartrate-resistant acid phosphatase (TRAP) staining, qRT-PCR, and WB assay. Compared with the ovariectomy (OVX) group, CEE treatment enhanced trabecular bone density, maximal load-bearing capacity, and various histomorphometric parameters. Serum E2 and OPG levels significantly increased, while Receptor activator of nuclear factor-κB (RANK) decreased in the CEE group. CEE downregulated matrix metallopeptidase 9 (MMP-9), Cathepsin K (CTSK), and TRAP gene and protein expression. In bone marrow macrophages (BMMs), CEE reduced mature osteoclasts, bone resorption pit areas, and MMP-9, CTSK, and TRAP expression during osteoclast differentiation. Compared with DMSO treatment, CEE markedly inhibited RANK, TNF receptor associated factor 6 (TRAF6), Proto-oncogene c-Fos (c-Fos), Nuclear factor of activated T-cells cytoplasmic 1 (NFATc1) expressions, and Extracellular regulated protein kinases (ERK), c-Jun N-terminal kinase (JNK), NF-kappa B-p65 (p65) phosphorylation in osteoclasts. In conclusion, CEE significantly inhibits OVX-induced osteoporosis and RANKL-induced osteoclastogenesis, potentially through modulating the Estrogen Receptor (ER)/RANK/NFATc1 signaling pathway.

[Effects of periodontitis on bone and tryptophan metabolism of gut microbiota in estrogen-deficient mice].

Objective: To study the effects of periodontitis on bone and tryptophan metabolism of gut microbiota in the context of estrogen deficiency. Methods: Thirty-two female C57BL6/J mice were randomly divided into four groups based on table of random numbers (n=8 in each group): Sham group, in which mice were given sham surgery; Sham_Lig group, in which mice were given sham surgery and were induced to periodontitis by ligating the bilateral maxillary second molars with 5-0 silk threads at the fourth week; Ovx group, in which mice were given bilateral ovariectomy; Ovx_Lig group, in which mice were given bilateral ovariectomy and were induced to periodontitis at the fourth week. After 8 weeks of ligation, the mice of 4 groups were euthanized for collecting the samples of femur, tibia, mandible and skull. Those samples were scanned by micro-CT to measure the bone mineral density (BMD), bone volume versus total volume ratio (BV/TV), trabecular number (Tb.N), trabecular thickness (Tb.Th) and trabecular spacing (Tb.Sp). The cecum contents of 4 groups of mice were collected for gut microbiota 16S rRNA gene sequencing. The tryptophan and its metabolites in intestinal tracts were detected by liquid chromatography-mass spectrometry. Pearson correlation analysis was performed to analyze the correlation between the abundance of gut microbiota and the content of tryptophan and its metabolites. Results: Femur BMD [(82.23±3.97) mg/cm3], BV/TV [(9.25±1.37)%] and Tb.Th [(70.95±5.70) µm] in Ovx_Lig group were significantly lower than Ovx group [(96.30±3.76) mg/cm3 (P=0.004); (14.45±1.55)% (P=0.022) and (87.58±8.02) µm (P<0.001), respectively]. The ß-diversity analysis of gut microbiota based on Bray-Curtis distance showed that samples of Ovx_Lig group and Ovx group were obviously grouped. Linear discriminant analysis effect size (LEfSe) showed that Alistipes was the representative genus in Ovx_Lig group. The relative abundance of Alistipes in Ovx_Lig group [(0.42±0.14)%] were significantly higher than that in Ovx group [(0.17±0.05)%] (t=4.45, P<0.001). Tryptophan metabolism analysis showed that the content of kynurenic acid [(531.12±158.60) ng/g] in Ovx_Lig group were significantly higher than that in Ovx group [(400.42±57.96) ng/g] (t=2.19, P=0.046). And the content of indole-3-carbaldehyde [(383.37±144.06) ng/g] in Ovx_Lig group were significantly lower than Ovx group [(701.72±141.93) ng/g] (t=4.45, P<0.001). Correlation analysis showed that relative abundance of Alistipes was positively correlated with kynurenic acid (r=0.32, P=0.088), while negatively correlated with indole-3-carbaldehyde (r=-0.32, P=0.088). Conclusions: Periodontitis can induce bone destruction of femur in estrogen-deficient mice, the mechanism of which may be related to Alistipes in gut and the tryptophan metabolites kynurenic acid and indole-3-carbaldehyde.

Osteoblastgenic and Osteogenic Effects of KY-273 with CDK8/19 Inhibitory Activity in Bone Marrow Mesenchymal Stem Cells and Female Rats.

Osteoporosis is caused by imbalance between osteogenesis and bone resorption, thus, osteogenic drugs and resorption inhibitors are used for treatment of osteoporosis. The present study examined the effects of (R)-4-(1-hydroxyethyl)-3-{4-[2-(tetrahydropyran-4-yloxy)ethoxy]phenoxy}benzamide (KY-273), a diphenyl ether derivative, on CDK8/19 activity, osteoblast differentiation and femoral bone using micro-computed tomography in female rats. KY-273 potently inhibited CDK8/19 activity, promoted osteoblast differentiation with an increase in alkaline phosphatase (ALP) activity, and gene expression of type I collagen, ALP and BMP-4 in mesenchymal stem cells (ST2 cells). In female rat femur, ovariectomy decreased metaphyseal trabecular bone volume (Tb.BV), mineral content (Tb.BMC), yet had no effect on metaphyseal and diaphyseal cortical bone volume (Ct.BV), mineral content (Ct.BMC) and strength parameters (BSPs). In ovaries-intact and ovariectomized rats, oral administration of KY-273 (10 mg/kg/d) for 6 weeks increased metaphyseal and diaphyseal Ct.BV, Ct.BMC, and BSPs without affecting medullary volume (Med.V), but did not affect Tb.BV and Tb.BMC. In ovariectomized rats, alendronate (3 mg/kg/d) caused marked restoration of Tb.BV, Tb.BMC and structural parameters after ovariectomy, and increased metaphyseal but not diaphyseal Ct.BV, Ct.BMC, and BSPs. In ovaries-intact and ovariectomized rats, by the last week, KY-273 increased bone formation rate/bone surface at the periosteal but not the endocortical side. These findings indicate that KY-273 causes osteogenesis in cortical bone at the periosteal side without reducing Med.V. In conclusion, KY-273 has cortical-bone-selective osteogenic effects by osteoblastogenesis via CDK8/19 inhibition in ovaries-intact and ovariectomized rats, and is an orally active drug candidate for bone diseases such as osteoporosis in monotherapy and combination therapy.

Mechanistic study on the alleviation of postmenopausal osteoporosis by Lactobacillus acidophilus through butyrate-mediated inhibition of osteoclast activity.

In China, traditional medications for osteoporosis have significant side effects, low compliance, and high costs, making it urgent to explore new treatment options. Probiotics have demonstrated superiority in the treatment of various chronic diseases, and the reduction of bone mass in postmenopausal osteoporosis (PMOP) is closely related to the degradation and metabolism of intestinal probiotics. It is crucial to explore the role and molecular mechanisms of probiotics in alleviating PMOP through their metabolites, as well as their therapeutic effects. We aim to identify key probiotics and their metabolites that affect bone loss in PMOP through 16srDNA sequencing combined with non-targeted metabolomics sequencing, and explore the impact and possible mechanisms of key probiotics and their metabolites on the progression of PMOP in the context of osteoporosis caused by estrogen deficiency. The sequencing results showed a significant decrease in Lactobacillus acidophilus and butyrate in PMOP patients. In vivo experiments confirmed that the intervention of L. acidophilus and butyrate significantly inhibited osteoclast formation and bone resorption activity, improved intestinal barrier permeability, suppressed B cells, and the production of RANKL on B cells, effectively reduced systemic bone loss induced by oophorectomy, with butyric acid levels regulated by L. acidophilus. Consistently, in vitro experiments have confirmed that butyrate can directly inhibit the formation of osteoclasts and bone resorption activity. The above research results indicate that there are various pathways through which L. acidophilus inhibits osteoclast formation and bone resorption activity through butyrate. Intervention with L. acidophilus may be a safe and promising treatment strategy for osteoclast related bone diseases, such as PMOP.

The bone-protective benefits of kaempferol combined with metformin by regulation of osteogenesis-angiogenesis coupling in OVX rats.

This study was to investigate the potential mechanisms of treatment with metformin (Met) combined with kaempferol (Kae) against postmenopausal osteoporosis. Experiments were conducted in both ovariectomy (OVX)-induced osteoporosis rats and in vitro using RAW264.7 cells, MC3T3-E1 cells, and HUVECs. Results demonstrated the therapeutic effect of Met combined with Kae on osteoporosis. In vivo, Kae alone and in combination with Met treatments enhanced tibial trabecular microstructure, bone mineral density (BMD), and mechanical properties in OVX rats without causing hepatotoxicity and nephrotoxicity. It also reduced bone resorption markers (CTX-1 and TRAP) and increased the bone formation marker (PINP) level in the serum of OVX rats. The expression of bone resorption marker TRAP was reduced, while bone formation markers Runx2 and ALP were enhanced in the bone tissue of OVX rats. Furthermore, Met combined with Kae also promoted the expression of angiogenesis-related markers CD31 and VEGF in OVX rats. In vitro, MC3T3-E1s cells treated with Met combined with Kae showed higher expression of ALP, Runx2, and VEGF. Interestingly, the treatment did not directly promote HUVECs migration and angiogenesis, but enhanced osteoblast-mediated angiogenesis by upregulating VEGF levels. Additionally, Met combined with Kae treatment promoted VEGF secretion in MC3T3-E1, and activated the Notch intracelluar pathway by upregulating HES1 and HEY1 in HUVECs. Meantime, their stimulation on CD31 expression were inhibited by DAPT, a Notch signaling inhibitor. Overall, this study demonstrates the positive effects of Met combined with Kae on osteoporotic rats by promoting osteogenesis-angiogenesis coupling, suggesting their potential application in postmenopausal osteoporosis.

Linking alterations in estrogen receptor expression to memory deficits and depressive behavior in an ovariectomy mouse model.

The high risk of neurological disorders in postmenopausal women is an emerging medical issue. Based on the hypothesis of altered estrogen receptors (ERα and ß) after the decline of estrogen production, we investigated the changes in ERs expressions across brain regions and depressive/amnesic behaviors. C57BL/6J female mice were ovariectomized (OVX) to establish a menopausal condition. Along with behavior tests (anxiety, depression, and memory), the expression of ERs, microglial activity, and neuronal activity was measured in six brain regions (hippocampus, prefrontal cortex, striatum, raphe nucleus, amygdala, and hypothalamus) from 4 to 12 weeks after OVX. Mice exhibited anxiety- and depressive-like behaviors, as well as memory impairment. These behavioral alterations have been linked to a suppression in the expression of ERß. The decreased ERß expression coincided with microglial-derived neuroinflammation, as indicated by notable activations of Ionized calcium-binding adapter molecule 1 and Interleukin-1beta. Additionally, the activity of brain-derived neurotrophic factor (BDNF), particularly in the hippocampus, decreased in a time-dependent manner from 4 to 12 weeks post-OVX. Our study provides evidence shedding light on the susceptibility to memory impairment and depression in women after menopause. This susceptibility is associated with the suppression of ERß and alteration of ERα in six brain regions.

Evaluation of the analgesic effects of diclofenac as a premedication drug in dogs undergoing ovariohysterectomy.

Pre-emptive analgesia consists of administering drugs such as opioids and nonsteroid anti-inflammatory drugs. This study aims to evaluate the intraoperative antinociceptive effects of diclofenac administered alone in premedication or combined with morphine along with its potential influence on recovery of dogs undergoing ovariohysterectomy. A total of 34 dogs (ASA I or II) admitted for ovariohysterectomy were randomly allocated into three groups according to the drugs given in premedication: Diclofenac (D) (n = 11), Morphine (M) (n = 13) and Diclofenac-Morphine (DM) (n = 10) groups. Induction and maintenance of anesthesia were standardized in all dogs. To assess intraoperative nociception, the heart rate (HR) and mean arterial pressure (MAP) were recorded during the surgery and at predefined time points: St (steady-state), Cut (cutaneous incision), P1 (first ovarian manipulation), P2 (second ovarian manipulation) and Cerv (cervical manipulation). The dynamic variation of HR (ΔHR) and MAP (ΔMAP) over 2 min was calculated at each time point. After extubation, early quality of recovery was assessed. Compared to St, a significant increase in HR and MAP at P1, P2 and Cerv was shown in all groups. MAP in the M group was lower at St than in the other groups. The dynamic variation of HR (ΔHR) and MAP (ΔMAP) was significantly less important at P2 and Cerv compared to P1 only in the DM group. Also, a better quality of recovery was shown in the D group compared to the M and DM groups. Diclofenac may be considered a suitable premedication drug and a part of a multimodal anesthetic approach in dogs.

Ovariectomy-Induced Arterial Stiffening Differs From Vascular Aging and Is Reversed by GPER Activation.

BACKGROUND: Arterial stiffness is a cardiovascular risk factor and dramatically increases as women transition through menopause. The current study assessed whether a mouse model of menopause increases arterial stiffness in a similar manner to aging and whether activation of the G-protein-coupled estrogen receptor could reverse stiffness. METHODS: Female C57Bl/6J mice were ovariectomized at 10 weeks of age or aged to 52 weeks, and some mice were treated with G-protein-coupled estrogen receptor agonists. RESULTS: Ovariectomy and aging increased pulse wave velocity to a similar extent independent of changes in blood pressure. Aging increased carotid wall thickness, while ovariectomy increased material stiffness without altering vascular geometry. RNA-sequencing analysis revealed that ovariectomy downregulated smooth muscle contractile genes. The enantiomerically pure G-protein-coupled estrogen receptor agonist, LNS8801, reversed stiffness in ovariectomy mice to a greater degree than the racemic agonist G-1. In summary, ovariectomy and aging induced arterial stiffening via potentially different mechanisms. Aging was associated with inward remodeling, while ovariectomy-induced material stiffness independent of geometry and a loss of the contractile phenotype. CONCLUSIONS: This study enhances our understanding of the impact of estrogen loss on vascular health in a murine model and warrants further studies to examine the ability of LNS8801 to improve vascular health in menopausal women.

Konjac Oligosaccharides Alleviated Ovariectomy-Induced Bone Loss through Gut Microbiota Modulation and Treg/Th17 Regulation.

Oligosaccharides from the plant Amorphophallus konjac were potentially effective in menopausal osteoporosis due to their prebiotic attributes. The present work mainly studied the regulation of konjac oligosaccharides (KOS) on menopausal bone loss. Experiments were carried out in ovariectomized (OVX) rats, and various contents of KOS were correlated with diet. After 3 months of treatment, the degree of osteoporosis was determined by bone mineral density and femoral microarchitecture. The research data showed that the 8% dietary KOS significantly alleviated bone loss in OVX rats, as it promoted the bone trabecular number by 134.2% and enhanced the bone bending stiffness by 103.1%. From the perspective of the gut-bone axis, KOS promoted gut barrier repair and decreased pro-inflammatory cytokines. Besides, KOS promoted the growth of Bifidobacterium longum and restored Treg/Th17 balance in bone marrow. The two aspects contributed to decreased osteoclastogenic activity and thus inhibited inflammation-related bone loss. This work extended current knowledge of prebiotic inhibition on bone loss and provide an alternative strategy for osteoporosis prevention.

Litsea glutinosa extract promotes fracture healing and prevents bone loss via BMP2/SMAD1 signaling.

Estrogen deficiency is one of the main causes for postmenopausal osteoporosis. Current osteoporotic therapies are of high cost and associated with serious side effects. So there is an urgent need for cost-effective anti-osteoporotic agents. Anti-osteoporotic activity of Litsea glutinosa extract (LGE) is less explored. Moreover, its role in fracture healing and mechanism of action is still unknown. In the present study we explore the osteoprotective potential of LGE in osteoblast cells and fractured and ovariectomized (Ovx) mice models. Alkaline phosphatase (ALP), MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and mineralization assays revealed that LGE treatment increased osteoblast cell differentiation, viability and mineralization. LGE treatment at 0.01 µg increased the expression of BMP2, PSMAD, RUNX2 and type 1 col. LGE also mitigated RANKL-induced osteoclastogenesis. Next, drill hole injury Balb/C mice model was treated with LGE for 12 days. Micro-CT analysis and Calcein labeling at the fracture site showed that LGE (20 mg/kg) enhanced new bone formation and bone regeneration, also increased expression of BMP2/SMAD1 signaling genes at fracture site. Ovx mice were treated with LGE for 1 month. µCT analysis indicated that the treatment of LGE at 20 mg/kg dose prevented the alteration in bone microarchitecture and maintained bone mineral density and bone mineral content. Treatment also increased bone strength and restored the bone turnover markers. Furthermore, in bone samples, LGE increased osteogenesis by enhancing the expression of BMP2/SMAD1 signaling components and decreased osteoclast number and surface. We conclude that LGE promotes osteogenesis via modulating the BMP2/SMAD1 signaling pathway. The study advocates the therapeutic potential of LGE in osteoporosis treatment.

Cichoric acid targets RANKL to inhibit osteoclastogenesis and prevent ovariectomy-induced bone loss.

BACKGROUND AND AIM: Osteoporosis, a systemic metabolic bone disease, is characterized by the decline of bone mass and quality due to excessive osteoclast activity. Currently, drug-targeting osteoclasts show promising therapy for osteoporosis. In this study, we investigated the effect of cichoric acid (CA) on receptor activator of nuclear kappa-B ligand (RANKL)-induced osteoclastogenesis and the bone loss induced by ovariectomy in mice. EXPERIMENTAL PROCEDURE: Molecular docking technologies were employed to examine the interaction between CA and RANKL. CCK8 assay was used to evaluate the cell viability under CA treatment. TRAcP staining, podosome belt staining, and bone resorption assays were used to test the effect of CA on osteoclastogenesis and osteoclast function. Further, an OVX-induced osteoporosis mice model was employed to identify the effect of CA on bone loss using micro-CT scanning and histological examination. To investigate underlying mechanisms, network pharmacology was applied to predict the downstream signaling pathways, which were verified by Western blot and immunofluorescence staining. KEY RESULTS: The molecular docking analysis revealed that CA exhibited a specific binding affinity to RANKL, engaging multiple binding sites. CA inhibited RANKL-induced osteoclastogenesis and bone resorption without cytotoxic effects. Mechanistically, CA suppressed RANKL-induced intracellular reactive oxygen species, nuclear factor-kappa B, and mitogen-activated protein kinase pathways, followed by abrogated nuclear factor activated T-cells 1 activity. Consistent with this finding, CA attenuated post-ovariectomy-induced osteoporosis by ameliorating osteoclastogenesis. CONCLUSIONS AND IMPLICATIONS: CA inhibited osteoclast activity and bone loss by targeting RANKL. CA might represent a promising candidate for treating osteoclast-related diseases, such as osteoporosis.

Prolame produces anxiolytic- and antidepressant-like effects in middle-aged female rats with less uterotrophic effects than 17ß-estradiol.

Estrogen hormone replacement therapy (EHRT), improving women's life quality at menopause, reduces anxiety and depression symptoms associated with ovarian hormonal decline. However, its potential adverse effects, like thromboembolism and cancer risk, limit its use. Prolame is a synthetic 17ß-amino estrogen with antithrombotic actions that exerts anxiolytic- and antidepressant-like effects on young adult ovariectomized female rats. It is unknown if prolame's effects may be observed in age and endocrine conditions emulating menopause. This study aimed to identify the antidepressant- and anxiolytic-like effects of prolame and E2 (used as a reference estrogen treatment) in middle-aged female rats coursing with irregular cycles, in two different conditions: ovariectomized or gonadally intact. Results were compared with those from young adult ovariectomized rats. Prolame (60 or 120 µg/kg), 17ß-estradiol (E2, 40 or 80 µg/kg), or vehicle were chronically administered, and their effects were evaluated in the elevated plus-maze, defensive burying behavior test, open field test, and forced swimming test. Uterotrophic actions were estimated by uterine weight related to body weight. Prolame and E2 produced robust anxiolytic- and antidepressant-like effects in young adult ovariectomized rats, but these effects were absent in gonadally intact middle-aged rats. Interestingly, only prolame induced anxiolytic- and antidepressant-like effects in middle-aged ovariectomized rats. Uterotrophic effects of prolame were weaker than E2 effects, notably in middle-aged females. Altogether, present data support the notion that prolame has the potential to be considered an EHRT with relevant psychoactive actions and with apparently lower adverse-side effects, especially in middle-aged populations.

Protocol for ovariectomy and estradiol replacement in mice.

Ovariectomy, involving the surgical removal of ovaries, and estradiol replacement facilitate the understanding of sexual dimorphism-related physiological changes, encompassing reproductive biology, metabolism, and hormone-related diseases. In this study, we present a protocol for conducting ovariectomy and estradiol replacement in mice. We describe steps for performing sham and ovariectomy operations, outline preoperative preparations, and provide details on postoperative care, including analgesia administration and the removal of surgical clips. Additionally, we elaborate on the procedures for performing vehicle and estradiol injections. For complete details on the use and execution of this protocol, please refer to Luengo-Mateos et al.1.