Bone Mass In Rats Research Articles

The Effect of Zonisamide on Rat Bone Mass, Structure, and Metabolism

Introduction: Our study aimed to investigate the effect of zonisamide (ZNS) on bone metabolism in the rat model. Methods: Eight-week-old rats were divided into four groups. The sham-operated control group (SHAM) and the control group after orchidectomy (ORX) received the standard laboratory diet (SLD). The experimental group after orchidectomy (ORX+ZNS) and the sham-operated control group (SHAM+ZNS) received SLD enriched with ZNS for 12 weeks. Bone marker concentrations in serum of receptor activator of nuclear factor kappa B ligand, PINP, and osteoprotegerin, and the levels of sclerostin and bone alkaline phosphatase in bone homogenate, were measured using an enzyme-linked immunosorbent assay. Bone mineral density (BMD) was measured by dual-energy X-ray absorptiometry. The femurs were used for biomechanical testing. Results: We found a statistically significant reduction in BMD and biomechanical strength 12 weeks after orchidectomy of the rats (ORX). After ZNS administration to orchidectomized rats (ORX+ZNS) and the sham-operated control rats (SHAM+ZNS), there were no statistically significant changes in BMD, bone turnover markers, or biomechanical properties as compared with the ORX group and SHAM group. Conclusions: The results suggest that administration of ZNS to rats exerts no negative effect on BMD, bone metabolism markers, or biomechanical properties.

A Preliminary Study on the Effects of Black Cohosh Preparations on Bone Metabolism of Rat Models With GnRH-a-Induced Peri-Menopausal Symptoms.

A Preliminary Study on the Effects of Black Cohosh Preparations on Bone Metabolism of Rat Models With GnRH-a-Induced Peri-Menopausal Symptoms.

Resveratrol promotes bone mass in ovariectomized rats and the SIRT1 rs7896005 SNP is associated with bone mass in women during perimenopause and early postmenopause

Objective This study aimed to examine the effects of SIRT1 agonist resveratrol on bone mass in ovariectomized (OVX) rats and the SIRT1 single-nucleotide polymorphism (SNP) rs7896005 on bone mass in women during menopause and early postmenopause. Methods An animal experiment was conducted on rats that were sham-operated (SHAM), OVX or OVX and different administered doses of resveratrol. Serum markers and femur microstructure and staining were assessed. A cross-sectional study was conducted in women undergoing menopause. SIRT1 protein and SIRT1 SNP rs7896005 were evaluated. Results OVX rats administered resveratrol, especially high doses, showed lower bone loss than OVX rats. Serum osteoprotegerin (OPG) and femur SIRT1, β-catenin and bone mineral density (BMD) were significantly increased, whereas receptor activator of NF-κB ligand (RANKL) was significantly decreased. Serum SIRT1 levels were significantly lower in women with low bone mass (p < 0.01). Women with the CA genotype of rs7896005 had lower bone mass than those with the CC genotype. The A allele showed a significant negative effect on bone loss risk (odds ratio = 3.48; p = 0.025). Conclusions Resveratrol stimulated SIRT1 expression and Wnt/β-catenin signaling to promote bone mass in rat femurs. Among women in perimenopause and early postmenopause, SIRT1 protected bone mass, and the A allele of SIRT1 rs7896005 was a risk factor for reduced bone mass.

Bisphosphonate-enoxacin inhibit osteoclast formation and function by abrogating RANKL-induced JNK signalling pathways during osteoporosis treatment.

Osteoporosis is an age‐related disease characterized by low mineral density, compromised bone strength and increased risk of fragility fracture. Most agents for treating osteoporosis focus primarily on anti‐resorption by inhibiting osteoclast activity. Bisphosphonate (BP) is a potent anti‐resorptive agent that has been used clinically for decades and is proven to be effective. However, BP has a variety of side effects and is far from being an ideal anti‐osteoporosis agent. BP selectively binds to calcium crystals, which are subsequently taken up or released by osteoclasts. Based on the action of BP, we previously demonstrated the inhibitory effect of a novel bone‐targeting BP derivative, bisphosphonate‐enoxacin (BE). In the current study, we used bone marrow‐derived osteoclast cultures to further assess the inhibitory effect of BE on osteoclastogenesis and employed reverse transcription PCR and real‐time PCR to examine expression of osteoclast‐specific genes. Additionally, we used bone resorption and F‐actin immunofluorescence assays to evaluate the effect of BE on osteoclast function and investigated the potential mechanisms affecting osteoclast differentiation and function in vitro. Furthermore, an ovariectomized (OVX) rat model was established to evaluate the therapeutic effects of BE on preventing bone loss. Results showed that BE exerted potent inhibitory effects on osteoclast formation and bone resorption by specifically abrogating RANKL‐induced JNK signalling, and that it preserved OVX rat bone mass in vivo without any notable side effects. Collectively, these results indicated that the BP derivative BE may have significant potential as a treatment for osteoporosis and other osteolytic diseases.

Puerarin improves graft bone defect through microRNA‑155‑3p‑mediated p53/TNF‑α/STAT1 signaling pathway.

Bone graft defects may lead to dysfunction of bone regeneration and metabolic disorders of bone mesenchymal stem cells (BMSCs). Puerarin has demonstrated pharmacological activities in the treatment of human metabolic diseases. The purpose of the present study was to investigate the role of puerarin and to explore its possible protective mechanism of action in rats with bone grafts. A bone graft rat model was established using bone grafting surgery and the rats received puerarin or PBS. Reverse transcription-quantitative PCR, western blot, TUNEL, immunofluorescence and immunohistochemistry assays were used to analyze the beneficial effects of puerarin on bone repair. The results demonstrated that puer-arin effectively ameliorated pathological graft bone defects, decreased bone loss and apoptosis of BMSCs, promoted BMSC proliferation and differentiation, and increased bone mass and the parameters of bone formation in rats with bone grafts. Puerarin decreased the levels of pro-inflammatory cytokines [tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-17A, IL-6 and transforming growth factor (TGF)-β1] and increased the levels of anti-inflammatory cytokines (IL-2 and IL-10) in the serum compared with the PBS group. Puerarin treatment was associated with lower serum alanine transaminase, glutamic oxaloacetic transaminase, γ-glutamyl transferase, alkaline phosphatase, direct bilirubin and total bilirubin levels compared with those in the PBS group in experimental rats. The expression of microRNA-155-3p (miR-155-3p) was upregulated, whereas that of p53, TNF-α and signal transducer and activator of transcription (STAT)1 was downregulated in BMSC cultures of puerarin-treated rats. In vitro assay demonstrated that knockdown of miR-155-3p increased p53, TNF-α and STAT1 expression in BMSCs, and blocked puerarin-regulated p53/TNF-α/STAT1 signaling. Most importantly, miR-155-3p knockdown inhibited puer-arin-regulated apoptosis, proliferation and differentiation of BMSCs. Moreover, the results demonstrated that puerarin regulated vascular endothelial growth factor expression via the miR-155-3p signaling pathway. In conclusion, the results of the present study demonstrated that the upregulation of miR-155-3p induced by puerarin promoted BMSC differentiation and bone formation and increased bone mass in rats with bone grafts, thereby supporting the potential application of puerarin in the prevention of bone graft defects.

Applying vibration in early postmenopausal osteoporosis promotes osteogenic differentiation of bone marrow-derived mesenchymal stem cells and suppresses postmenopausal osteoporosis progression.

We aimed to evaluate whether applying low magnitude vibration (LMV) in early postmenopausal osteoporosis (PMO) suppresses its progression, and to investigate underlying mechanisms. Rats were randomly divided into Sham (Sham-operated), Sham+V, OVX (ovariectomized), OVX+E2 (estradiol benzoate), OVX+V (LMV at 12–20 weeks postoperatively), and OVX+Vi (LMV at 1–20 weeks postoperatively) groups. LMV was applied for 20 min once daily for 5 days weekly. V rats were loaded with LMV at 12–20 weeks postoperatively. Vi rats were loaded with LMV at 1–20 weeks postoperatively. Estradiol (E2) rats were intramuscularly injected at 12–20 weeks postoperatively once daily for 3 days. The bone mineral densities (BMDs), biomechanical properties, and histomorphological parameters of tibiae were analyzed. In vitro, rat bone marrow-derived mesenchymal stem cells (rBMSCs) were subjected to LMV for 30 min daily for 5 days, or 17β-E2 with or without 1-day pretreatment of estrogen receptor (ER) inhibitor ICI 182,780 (ICI). The mRNA and protein expresion were performed. Data showed that LMV increased BMD, bone strength, and bone mass of rats, and the effects of Vi were stronger than those of E2. In vitro, LMV up-regulated the mRNA and protein expressions of Runx2, Osx, Col I, and OCN and down-regulated PPARγ, compared with E2. The effects of both LMV and E2 on rBMSCs were inhibited by ICI. Altogether, LMV in early PMO suppresses its progression, which is associated with osteogenic differentiation of rBMSCs via up-regulation of ERα and activation of the canonical Wnt pathway. LMV may therefore be superior to E2 for the suppression of PMO progression.

Wnt Signaling Mediated Inhibition of Osteoclastogenesis by Dietary Phenolic Hippuric Acid and 3-(3-hydroxyphenyl) Propionic Acid (3–3-PPA) (P01-037-19)

ObjectivesNutritional factors influence bone development. We have previously reported that Blueberry (BB) and BB-associated phenolic acids had significant effects on increasing bone mass in rats. We also found BB-associated phenolic acids [hippuric acid (HA) and 3-(3-hydroxyphenyl) propionic acid (3-3-PPA)] inhibited osteoclastogenesis in vitro and in ex vivo. Here, we investigated the mechanisms by which HA or 3-3-PPA inhibit osteoclast formation and if HA or 3-3-PPA inhibits bone resorption in vivo. MethodsMurine osteoclast (macrophage) cell line, RAW 264.7 cells, and hematopoietic osteoclast progenitor cells (isolated from 4 weeks old C57BL6/J mice) were cultured. Cells were treated with HA or 3-3-PPA (0.6, 6, 60, 600 μg/dL) for 4 days in the presence of 50 ng/ml of RANKL (Receptor activator of nuclear factor kappa-Β ligand). In animal studies, dietary HA or 3-3-PPA was provided at 1, 5 or 10 mg/kg/d to female mice for 4 weeks, starting at postnatal day 28. ResultsIn RAW 264.7 cell and primary cell cultures, TRAPase staining showed that RANKL-stimulated osteoclast number/well dose-dependently decreased with treatments of HA or 3-3-PPA. HA and 3-3-PPA significantly inhibited RANKL-induced β-catenin and GPR109A (G protein coupled receptor 109A) protein expression. Moreover, RANKL-induced β-catenin and GPR109A mRNA expression were also suppressed by HA and 3-3-PPA. Furthermore, after pre-treatment with HA or 3-3-PPA, RANKL-stimulated increases of Wnt/β-catenin down-stream genes such as c-myc, and osteoclastogenic genes such as NFkB and NFATc1 gene expression were blunted. In mice, we found significantly increased bone mass in dietary HA and 3-3-PPA supplemented groups compared to control group (P < 0.05). Increased bone mass in HA and 3-3-PPA treated mice was accompanied by decreased bone resorption (bone resorption markers and osteoclastogenic gene expression), but increased β-catenin expression in total protein isolated from bone. ConclusionsThese results indicate significant inhibition of osteoclastogenesis and bone resorption by HA and 3-3-PPA, and suggest that HA and 3-3-PPA inhibit bone marrow the hematopoietic cell differentiation program through non-canonical Wnt signaling to protect against increased bone resorption. Funding SourcesUSDA-ARS Project.

Effects of compound medicine of icariin and puerarin on peak bone mass in growing rats

Objective To study the effects of the compound medicine of icariin and puerarin on peak bone mass in rats during growth period, and to explore its possible mechanism. Methods Forty female Sprague-Dawley rats aged 1 month were randomly divided into normal control group(C), icariin group(I), puerarin group(P), icariin and puerarin compound groupc(I+ P), 10 in each group. The body weights were recorded once every two weeks, and the bone mineral density was measured by dual-energy X-ray absorptiometry every month. After the bone mineral density of the whole body was significantly different between the control group and drug groups, the animals were sacrificed. The right femur and vertebrae were separated to measure the bone mineral density. The biomechanical properties of the femur and vertebra were detected by AG-IS series desktop electronic universal testing machine. The bone formation index osteocalcin, PINP and bone resorption index were determined by ELISA. Changes in the contents of tartrate-resistant acid phosphatase 5b(TRACP 5b) and CTX-1; and changes in trabecular bone related parameters were recorded after magenta-picric acid staining. Results There was no significant difference in body weight between the two groups (P>0.05). There was no significant difference in whole body bone density after 1 month of treatment (P>0.05). After 2 months of treatment, the body bone density of the drug-administered group was higher than that of the control group. Whole body bone density, femur and vertebral bone density, femur maximum load value, maximum vertebrae load value and trabecular bone number and area, serum OC and PINP levels increased, while TRACP 5b and CTX-1 levels decreased(P<0.01) in drug group. The difference from the control group was statistically significant (P<0.05). There were significant differences in biochemical parameters and bone histomorphology between the compound drug group and the two-flavor monomer group (P<0.01). There was no significant difference in bone mineral density and biomechanics, but the average value was higher than that of the monomer group. Conclusion The combination of icariin and puerarin can effectively increase the peak bone mass in rats. Key words: Icariin; Puerarin; Compound; Peak bone mass; Rats

Sinusoidal Electromagnetic Fields Increase Peak Bone Mass in Rats by Activating Wnt10b/β-Catenin in Primary Cilia of Osteoblasts

Extremely low-frequency electromagnetic fields have been considered a potential candidate for the prevention and treatment of osteoporosis; however, their action mechanism and optimal magnetic flux density (intensity) parameter are still elusive. The present study found that 50-Hz sinusoidal electromagnetic fields (SEMFs) at 1.8 mT increased the peak bone mass of young rats by increasing bone formation. Gene array expression studies with femoral bone samples showed that SEMFs increased the expression levels of collagen-1α1 and Wnt10b, a critical ligand of the osteogenic Wnt/β-catenin pathway. Consistently, SEMFs promoted osteogenic differentiation and maturation of rat calvarial osteoblasts (ROBs) in vitro through activating the Wnt10b/β-catenin pathway. This osteogenesis-promoting effect of SEMFs via Wnt10b/β-catenin signaling was found to depend on the functional integrity of primary cilia in osteoblasts. When the primary cilia were abrogated by small interfering RNA (siRNA) targeting IFT88, the ability of SEMFs to promote the osteogenic differentiation of ROBs through activating Wnt10b/β-catenin signaling was blocked. Although the knockdown of Wnt10b expression with RNA interference had no effect on primary cilia, it significantly suppressed the promoting effect of SEMFs on osteoblastic differentiation/maturation. Wnt10b was normally localized at the bases of primary cilia, but it disappeared (or was released) from the cilia upon SEMF treatment. Interestingly, primary cilia were elongated to different degrees by different intensities of 50-Hz SEMFs, with the window effect observed at 1.8 mT, and the expression level of Wnt10b increased in accord with the lengths of primary cilia. These results indicate that 50-Hz 1.8-mT SEMFs increase the peak bone mass of growing rats by promoting osteogenic differentiation/maturation of osteoblasts, which is mediated, at least in part, by Wnt10b at the primary cilia and the subsequent activation of Wnt/β-catenin signaling. © 2019 American Society for Bone and Mineral Research.

Siglec-15-targeting therapy increases bone mass in rats without impairing skeletal growth.

The treatment of juvenile osteoporosis has not been established due to a lack of data regarding the efficacy and adverse effects of therapeutic agents. The possible adverse effects of the long-term use of antiresorptive therapies on skeletal growth in children is of particular concern. Sialic acid-binding immunoglobulin-like lectin 15 (Siglec-15) is an immunoreceptor that regulates osteoclast development and bone resorption, and its deficiency suppresses bone remodeling in the secondary spongiosa, but not in the primary spongiosa, due to a compensatory mechanism of osteoclastogenesis. This prompted us to develop an anti-Siglec-15 therapy for juvenile osteoporosis because most anti-resorptive drugs have potential adverse effects on skeletal growth. Using growing rats, we investigated the effects of an anti-Siglec-15 neutralizing antibody (Ab) on systemic bone metabolism and skeletal growth, comparing this drug to bisphosphonate, a first-line treatment for osteoporosis. Male 6-week-old F344/Jcl rats were randomized into six groups: control (PBS twice per week), anti-Siglec-15 Ab (0.25, 1, or 4 mg/kg every 3 weeks), and alendronate (ALN) (0.028 or 0.14 mg/kg twice per week). Treatment commenced at 6 weeks of age and continued for the next 6 weeks. Changes in bone mass, bone metabolism, bone strength, and skeletal growth during treatment were analyzed. Both anti-Siglec-15 therapy and ALN increased bone mass and the mechanical strength of both the femora and lumbar spines in a dose-dependent manner. Anti-Siglec-15 therapy did not have a significant effect on skeletal growth as evidenced by micro-CT-based measurements of femoral length and histology, whereas high-dose ALN resulted in growth retardation with histological abnormalities in the growth plates of femurs. This unique property of the anti-Siglec-15 Ab can probably be attributed to compensatory signaling for Siglec-15 inhibition in the primary spongiosa, but not in the secondary spongiosa. Thus, anti-Siglec-15 therapy could be a safe and effective for juvenile osteoporosis.

Resveratrol and puerarin on peak bone mass in young rats

To compare effects of resveratrol, puerarin and the compounds on peak bone mass in rats. Forty SPF Wistar rats weighed 109.45 g to 119.44 g with an average of 115.87 g were selected. After 3 days' adaption, rats were divided into control group (the same volume of distilled water per day), puerarin group(15.4 mg/kg puerarin daily), resveratrol group (8.4 mg/kg resveratrol daily), compound drug group (daily dose of 8.4 resveratrol added 15.4 mg/kg of puerarin) and 10 in each group. The body weight of the rats was monitored at every 7 days and body bone density was measured at every month. All rats were sacrificed after 3 months. The bone mineral density of femur and vertebrae was detected by dual energy X-ray absorptiometry; bone biomechanics, VG staining was used to analyze bone histomorphometry;ELISA was used to detect serum bone metabolic index and microstructure of femur were scanned with Micro-CT scanner. There were no significant differences in body weight among groups during exoeriment. Bone mineral density results showed BMD of femur and vertebrae in the other three groups were significantly increased, and R+P group was significantly higher than PR group and RES group(P<0.05) by compared with CON group;three-point bending and compression test results showed compared with CON group, other three groups of femoral and vertebral maximum load values were significantly increased, and P+R group was higher than PR group and RES group, but elastic modulus was not statistically significant. Bone histomorphometry showed that number of trabecular bone in other three groups were significantly increased compared with CON group, separation of trabecular bone were significantly reduced, continuity was improved, and R+P group was significantly better than RES and PR group. The results of Micro-CT scan showed that separation of trabecular bone were significantly reduced, continuity were improved in other three groups, and R+P group was significantly better than RES and PR group. The numbers of trabecular bone (Tb.N), trabecular bone thickness (Tb.Th), volume of trabecular bone (BV/TV) in PR group, RES group and R+P group were significantly higher than CON group, but trabecular bone separation (Tb.Sp) was significantly reduced. Serum levels results showed, level of OC in the other three groups were higher than control group(P<0.05), content of TRACP 5b decreased, and level of OC in P+R group was significantly higher than PR group and RES group, content of TRACP 5b was no significant change. Compound of puerarin and resveratrol assigned in a 1:1 ratio could improve bone mineral density and bone mass in young rats, enhance biomechanical properties of bone, promote mineralization and maturation of osteoblasts, inhibit osteoblastic bone resorption, and is better than the role of their respective monomers. The paper showed that traditional Chinese medicine compound medicine will be used as a new way to prevent and treat osteoporosis.

Intensity‑dependent effect of treadmill running on differentiation of rat bone marrow stromal cells.

The effect of running on bone mass depends on its intensity. However, the underlying molecular mechanism that associates running intensity with bone mass is unclear. The current study examined the effects of treadmill running at different intensities on bone mass and osteogenic differentiation of bone marrow stromal cells (BMSCs) in a rat model. A total of 24 male Wistar rats were randomly divided into groups and subjected to no running (Con group), low-intensity running (LIR group), moderate-intensity running (MIR group), and high-intensity running (HIR group). Histological, immunohistochemistry and micro-CT examinations were performed on the femora harvested after 8 weeks of treadmill running. The study demonstrated that treadmill running affected trabecular bone mass in an intensity-dependent manner. In addition, such an intensity-dependent effect was also demonstrated on the osteogenic and adipogenic differentiation and proliferation of BMSCs. Furthermore, the Wnt/β-catenin signaling pathway may be involved in the running-induced increase in bone mass in rats in the MIR group. There appears to be a biomechanical ‘window’, in which running-induced strain signals can increase the number of BMSCs and progenitor cells (specific to the osteoblast lineage) causing upregulation of osteogenesis and downregulation of adipogenesis of BMSCs. This finding may provide insight into the molecular and cellular mechanisms responsible for bone homeostasis.

Impacts of Ultra-early Hyperbaric Oxygen Therapy on Bone Mass of Rats With Complete Spinal Cord Transection.

Seventy-five SD rats were randomly assigned to four groups: sham-operated group (Sham, n = 15), complete spinal cord transection (CSCT, n = 20) group, hyperbaric oxygenation group 1 (HBO1, n = 20), and hyperbaric oxygenation group 2 (HBO2, n = 20). The aim of this study was to analyze the impacts of ultra-early hyperbaric oxygen therapy on bone mass of rats with CSCT. Treatment of patients with complete SCI is still an unresolved medical issue and needs to be further investigated. Studies on changes in bone mass as well as osteoporosis prevention after SCI have important clinical significance. Rats in the sham group only underwent T10 laminectomy, without damaging the spinal cord. Rats in CSCT, HBO1, and HBO2 groups underwent T10 laminectomy and spinal cord transection at T10 level. Rats in HBO1 and HBO2 groups received three courses of hyperbaric oxygen therapy with 10 days per course starting at 3 and 12 hours after spinal cord injury, respectively. The femoral biomechanical characteristics, the bone calcium, and the bone hydroxyproline (B-HYP) contents were determined. Morphology of the femur bone trabecula and the bone collagen were observed by HE staining and by masson triad color staining, respectively. After 6 weeks of treatment, rats in the CSCT group showed significant decreases in femur structural and material mechanics parameters, calcium and B-HYP contents, (P < 0.01), as well as sparse, fractured, malaligned trabecular bone and collagen compared with rats in the sham group. After treatments, compared with rats in the CSCT and HBO2 groups, rats in HBO1 group showed enhancement in femur structural and material mechanics parameters, calcium and B-HYP contents, (P < 0.05), as well as trabecular bone and collagen with better continuity and neater arrangement. Ultra-early HBO therapy can significantly improve bone mass in CSCT rats. N/A.

The Optimal Duration of PTH(1-34) Infusion Is One Hour per Day to Increase Bone Mass in Rats.

Parathyroid hormone (PTH) is a potential medicine for osteoporosis, and subcutaneous (s.c.) PTH treatment enhances bone mass; however, continuous infusion of PTH elicits bone resorption and induces bone loss. To clarify this contradictory phenomenon, we examined bone markers and bone mass in rats to assess the optimal duration of PTH(1-34) infusion. Continuous infusion of PTH at 1 µg/kg/h (Css, steady-state concentration ca. 300 pg/mL) for 1-4 h clearly stimulated the expression both of bone formation-related genes (c-fos, Wnt4, EphrinB2) and of bone resorption-related genes (tnfsf11, tnfsf11b, encoding receptor activator of nuclear factor-kappaB ligand (RANKL), osteoprotegerin (OPG)), but s.c. treatment stimulated these genes only 1-h after the injection. Rats were treated with 1-, 2-, or 4-h infusions of PTH daily using a totally implanted catheter system, and the femoral bone mineral density (BMD) was measured at 4 weeks. The 1-h infusion of PTH significantly stimulated serum bone formation markers (procollagen I N-terminal propeptide (PINP) and osteocalcin) on day 14 and femoral BMD at 2 and 4 weeks, but the 4-h infusion of PTH did not enhance BMD. Since the 4-h infusion increased the levels of both the bone formation markers and a bone resorption marker (urinary C-terminal telopeptide of type 1 collagen (CTx)), the increased bone resorption may predominate over bone formation. The intermittent elevation of plasma PTH to 300 pg/mL for 1-h each day is optimal for increasing bone mass in rats. In osteoporosis therapy in human, using the optimal duration for the clinical dose of PTH may selectively stimulate bone formation.

Altered material properties are responsible for bone fragility in rats with chronic kidney injury

Chronic kidney disease (CKD) is associated with an increased risk of fragility fractures, but the underlying pathophysiological mechanism remains obscure. We performed an in vivo experimental study to examine the roles of uremia and abnormal mineral/parathyroid metabolism in the development of bone metabolic abnormalities in uremic rats. Male Sprague-Dawley rats were divided into four groups, comprising sham operation (high turnover bone control=HTB-Cont), 5/6-nephrectomy (high turnover bone nephrectomized=HTB-Nx), thyroparathyroidectomy (low turnover bone control=LTB-Cont), and thyroparathyroidectomy plus 5/6 nephrectomy (low turnover bone nephrectomized=LTB-Nx), and maintained for 16weeks. Uremia was successfully created in the LTB-Nx and HTB-Nx groups, while hyperparathyroidism was only found in the HTB-Nx group. Cancellous bone histomorphometry revealed significantly higher bone turnover in the HTB-Nx group than in the LTB-Nx group. Storage modulus at 1Hz and tan delta in cortical bone of the femur, which represent the viscoelastic mechanical properties, were significantly lower in both Nx groups than in the Cont groups regardless of bone metabolism. Pentosidine-to-matrix ratio was increased and crystallinity was decreased in both Nx groups regardless of bone turnover. Mineral-to-matrix ratio was significantly decreased in the HTB-Nx group, but increased in the LTB-Nx group. Enzymatic collagen crosslinks were decreased in the HTB-Nx group. The degree of orientation of the c-axis in carbonated hydroxyapatite (biological apatite=BAp) crystallites was decreased in both Nx groups regardless of bone metabolism. Stepwise multivariate regression revealed that pentosodine-to-matrix ratio and BAp preferential c-axis orientation were significantly associated with storage modulus and tan delta. In conclusion, bone elastic mechanical properties deteriorated regardless of bone metabolism or bone mass in rats with chronic kidney injury. Various changes in bone mineral properties were associated with CKD, including abnormal parathyroid function, impaired bone turnover, and uremia associated with the accumulation of uremic toxins, were responsible for these changes. Pentosidine-to-matrix ratio and BAp orientation at position 5 were the two meaningful determinants of elastic bone mechanical strength, and both factors were associated with the severity of uremia, but not parathyroid function or bone metabolism. These two factors may account for the increased bone fragility among CKD patients.

The effect of topiramate and lamotrigine on rat bone mass, structure and metabolism

There is only limited data concerning the effect of the newer antiepileptic drugs on bone. The objective of this study was to determine the effect of topiramate (TPM) and lamotrigine (LTG) monotherapy on bone mineral density (BMD), mineral content (BMC), bone markers, body composition and bone mechanical strength in the orchidectomized (ORX) rat model.24 orchidectomized Wistar rats were divided into control and test groups, 8 rats in each group. The control rats received standard laboratory diet (SLD) while rats in the test group were fed with SLD enriched with LTG or TPM for 12weeks. Dual energy X-ray absorptiometry was used to measure bone mineral density. The concentrations of bone metabolism markers were assayed in bone homogenate. In addition, both femurs were measured and used for biomechanical testing.Compared to the control group, both test groups had significantly lower weight, fat mass, whole body and femur BMD, BMC and reduced mechanical strength of bone. All of these changes were more pronounced in rats exposed to LTG.In conclusion, both LTG and TPM significantly reduce BMD and body weight and impair mechanical strength of bone. A question arises as to the degree of dependence of the effect on the dose. Further studies are warranted to establish whether LTG and TPM may have a clinically significant effect on BMD exclusively in the model of gonadectomized rats, or whether the effect applies also in the model of gonadally intact animals, and in the respective human models.

Effects of icariin and genistein on peak bone mass in rats

To compare the effects of icariin (ICA) and genistein (GEN) on rats bone peak mass and thus screen for a drug that can more effectively prevent osteoporosis. Totally 36 one-month SD rats were randomly divided into three groups: ICA group [25 mg/(kg·d), intragastric administration], GEN group [10 mg/(kg·d), intragastric administration], and control group (fed with equal volume of distilled water). The body weight was monitored weekly and the bone mineral density of total body was measured monthly. All rats were sacrificed three months later. The femoral bone mineral density and the serum levels of osteocalcin and anti-tartaric acid phosphatase 5b, N-terminal propeptide of type 1 procollagen, and C-terminal propeptide of type 1collagen were measured. The bone microarchitectures were analyzed with micro-CT and the bone biomechanics properties were tested with universal material machine. The body weight and organ index showed no significant difference among these three groups(P>0.05). No obvious pathological change was found. The bone mineral density was also not significantly different in the first and second months; however, in the third months, the ICA group had significant higher bone mineral density for both total body and femur than those in the control and GEN group (P<0.05). The same trends were found for both femur bone mineral density and whole-body bone mineral density (P<0.05). The ICA group also had significantly higher serum levels of osteocalcin (P<0.05) and lower level of anti-tartaric acid phosphatase 5b(P<0.05). Besides, rats in the ICA group had significantly larger bone volume/tissue volume, trabecular thickness, and trabecular number than the control group, whereas the trabecular spacing and model coefficients were signicantly lower(all P<0.05), which, however, were not significantly different between ICA group and GEN group (P>0.05). Femoral maximum load, Youg's modulus, and yield load were significantly higher in these two groups than in the control group (P<0.05), which, again, were not significantly different between ICA group and GEN group (P>0.05). Orally administered ICA is more efficient than GEN in inhibiting resorption and promoting bone formation, and thus can dramatically improve the peak bone mineral density and bone quality.

The effect of levetiracetam on rat bone mass, structure and metabolism

To determine the effect of levetiracetam (LEV) Lon bone mineral density (BMD), mineral content (BMC), bone markers, body composition and bone mechanical strength in the orchidectomised (ORX) rat model. 16 orchidectomised Wistar rats were divided into control and test groups, 8 rats in each group. The control rats received standard laboratory diet (SLD) while rats in the test group were fed with SLD enriched with LEV for 12 weeks. BMD was measured by dual energy X-ray absorptiometry at the whole body, lumbar spine and femur. Bone marker concentrations were examined of osteoprotegerin (OPG) and insulin-like growth factor 1 (IGF-1) in serum, and amino-terminal propeptide of procollagen type I (PINP), carboxy-terminal cross-linking telopeptide of type I collagen (CTX-I), bone alkaline phosphatase (ALPL), and bone morphogenetic protein 2 (BMP-2) in bone homogenate. The femurs were used for biomechanical testing. Compared to the control group we found lower fat mass, lower BMD in the area of the left femur, lower BMC in both femurs, a reduced concentration of OPG, and an increased concentration of CTX-I of borderline statistical significance (p=0.0661). Biomechanical parameters did not differ between groups. Significant loss of BMD or BMC was seen at the left and right femur area in the LEV group. Administration of LEV in the ORX-rat model significantly decreased levels of OPG (marker of bone formation) in serum and increased levels of CTX-I (marker of bone resorption) in bone homogenate, but results in this study did not reveal any change in biomechanical bone strength. Administration of LEV in the ORX-rat model may reduce adipose tissue. Further studies in animals and humans will be needed to confirm these findings.

Effect of Sodium Fluoride Administration to Rats on Bone Phosphorous Content and Phosphatemia

Fluoride (F) has a known mitogenic effect on bone cells. The daily administration of 40 micromol NaF per 100 g of body weight (bw) increases bone mass in rats. Nevertheless, the quality and composition of bone formed under F stimulus is still matter of study. The objective of this work was to investigate the effect of sodium fluoride (NaF, CAS 7681-49-4) administration on phosphate metabolism and its impact on bone. Experiments were carried out in female fasted 50-day-old rats. Unless otherwise stated, NaF dose was 40 micromol NaF/day . 100 g bw, administered by gastric tube. Four groups of 4 rats each were given the following daily NaF doses: Group A: 0, B: 20, C: 40 and D: 60 micromol NaF/day x 100 g bw. After 30 days rats were killed saving their femora and plasma. Bone phosphorous contents (BPC) and phosphatemia (mg/dl) were measured. BPC decreased significantly as a function of NaF dose. A: 93.3+/-14.1; B: 78.7+/-15.5; C: 61.1+/-14.7**; D: 59.6+/-8.6 **mg P/g dry bone (** significant difference to group A, p < 0.01). Phosphatemia (mg/dl) increased significantly with a peak at 90 min after NaF dose (basal: 5.34+/-0.06; 90 min: 8.15 +/-0.43, p < 0.001). Phosphaturia (microg/min) increased though differences were not significant (basal: 46.7+/-42.8; 4 h: 1275 +/-757, p > 0.05). In thyroparathyroidectomized rats, plasma phosphate increased continuously for at least 240 min. Renal plasmatic flow, glomerular filtration rate and renal blood flow were not affected by NaF treatment. In isolated perfused rat kidneys, urinary phosphate excretion remained unaffected after NaF administration. Phosphate concentration was measured in the plasma and erythrocytes of rats after one dose of NaF (n = 8). Phosphate content of erythrocyte was not affected by NaF, in spite of the concurrent increase in phosphatemia. It is concluded that the treatment with NaF causes a transitory increase in plasma phosphate levels. Neither renal hemodynamic factors nor the inhibitory effect on parathormone actions appear to be the causes of hyperphosphatemia. Efflux of phosphate from cells might not be the cause of the increase in phosphatemia. The loss of phosphorous from bone appears as the most probable determinant of hyperphosphatemia after fluoride administration.

GSK-3 inhibition by an orally active small molecule increases bone mass in rats

Glycogen synthase kinase 3β (GSK-3β) actions are central in the canonical Wnt pathway, important in many biological processes and a potential drug target for treating several diseases. It is appreciated that a balanced Wnt canonical signaling is crucial for the maintenance of normal bone mass. In this study we investigated the effects of a potent orally active GSK-3 inhibitor, AZD2858, on bone mass in rats.Treatment (1μM) of human osteoblast cells with AZD2858 in vitro increased β-catenin levels after a short period of time. In rats, oral AZD2858 treatment caused a dose-dependent increase in trabecular bone mass compared to control after a two-week treatment with a maximum effect at a dose of 20mg/kg once daily (total BMC: 172% of control; p<0.001). A small but significant effect was also seen at cortical sites (total BMC: 111% of control; p<0.001). Biomechanical testing demonstrated an increase in both vertebral compression strength at a dose of 20mg/kg once daily (Load at failure: 370% of control, p<0.001) and diaphyseal strength of femora subjected to a three point bending test (Load at failure: 115% of control; p<0.01). Furthermore, histomorphometry showed a dramatic increase in bone formation indices, and serum markers of both bone formation (Osteocalcin, 146% of control; p<0.001) and resorption (CTX, 189% of control; p<0.001) were elevated.Our conclusion is that a GSK-3 inhibitor drug may prove effective as an anabolic strategy in the treatment of diseases characterized by low bone mass, since AZD2858 has extensive bone building effects at predominantly trabecular sites.