Co-modified 3D printed ß-tricalcium phosphate with magnesium and selenium promotes bone defect regeneration in ovariectomized rat.

Magnesium (Mg) and Selenium (Se) are essential elements for bone health and have been studied extensively for its powerful osteogenesis and promoting bone regeneration. The purpose was to observe whether Co-modified 3D-printed ß-tricalcium phosphate with Mg and Se could promote bone defect regeneration in an ovariectomized(OVX) rat model. The MC3T3-E1 cells were co-cultured with the leachate of ß-TCP, Mg-TCP, and Mg/Se-TCP and induced to osteogenesis, and the cell viability, ROS, and osteogenic activity were observed by Cell Count Kit-8(CCK-8), fluorescent probe 2', 7'-dichlorofluorescin diacetate, Alkaline phosphatase (ALP) staining, Alizarin Red(RES) staining, western blotting(WB), and immunofluorescence. Then the ß-TCP, Mg-TCP, and Mg/Se-TCP were implanted into the femoral epiphysis bone defect model of OVX rats for 12 weeks. Micro-CT and histology analysis were used to observe the therapeutic effect. In vitro results show that the cell mineralization and osteogenic activity of the Mg/Se-TCP group is significantly higher than the ß-TCP group and Mg-TCP group. Protein expressions such as FOxO1, SIRT1, SOD2, Runx-2, Cola1a, and OC of the Mg/Se-TCP group are significantly higher than the Con group and the ß-TCP group. The results of intracellular ROS and SIRT1 and SOD2 immunofluorescence showed that Mg/Se-TCP can restore the oxidative stress balance of osteoblasts. Micro-CT and histology analysis showed that treatment with Mg/Se-TCP showed the largest amount of bone tissue in the defect area (p < 0.05), and exhibited lower values of residual biological material (p < 0.05), compared to that of the ß-TCP group and Mg-TCP group. Our research results confirm that Mg/Se-TCP can improve the activity and function of osteoblasts and enhance bone regeneration mediated by reducing intracellular ROS in OVX rat models. The release of Mg and Se during the degradation of Mg/Se-TCP can improve the local bone repair ability. At the same time, it can also inhibit cell ROS, and ultimately greatly promote local bone repair.

Probucol promotes osteoblasts differentiation and prevents osteoporosis development through reducing oxidative stress.

Probucol (PBC) is a potent cholesterol-lowering drug and has been studied extensively for its powerful antioxidative stress. The purpose of this study is to investigate the role of PBC in ovariectomized rat model and to explore the mechanism of osteogenic differentiation of MC3TE-E1 Cells. RT-qPCR and Immunofluorescence were used to measure the expression level of SOD2, SIRT1, intracellular oxidative stress levels and osteogenic markers proteins. Moreover, CCK-8 assay was conducted to detect cell viability. Alizarin red staining and alkaline phosphatase staining were applied to examine osteogenic function and calcium deposits. The ovariectomized rat model was set up successfully and HE staining were employed to examine femoral trabeculae tissue. Our results showed that PBC suppressed MC3TE-E1 resist oxidative stress to promote osteogenic differentiation. Additionally, it was confirmed that PBC promoted osteogenic differentiation of MC3TE-E1 through inhibiting oxidative stress. Further study indicated that PBC exerted its beneficial function by suppressing oxidative stress-mediated alter bone metabolism to alleviate osteoporosis in vivo. Our research suggested that the PBC-modulated oxidative stress inhibition is responsible for activation of the process of osteogenic differentiation, providing a novel insight into the treatment of osteoporosis.

Changes in pedometer-measured physical activity are associated with weight loss and changes in body composition and fat distribution in response to reduced-energy diet interventions: The POUNDS Lost trial.

AIMS: To examine whether changes in objectively measured physical activity (PA) are associated with weight loss and changes in body composition and fat distribution in response to weight-loss diet interventions. METHODS: This study included 535 participants with overweight/ obesity, who were randomly assigned to four weight-loss diets varying in macronutrients. PA was measured objectively with pedometers, and body composition and fat distribution were measured using dual-energy X-ray absorptiometry and computed tomography scans at baseline, 6 months and 24 months. RESULTS: From baseline to 6 months, when the maximum weight loss was achieved, each 1000-steps/d increment in PA was associated with a greater reduction in body weight (ß[SE] = -0.48[0.11]) and waist circumference (ß[SE] = -0.49[0.12]). Similar inverse associations were found in changes in body composition and fat distribution (P < 0.05 and false discovery rate qvalue < 0.1 for all). The trajectory of the above adiposity measures across the 24-month intervention period differed between the patterns of PA change. Participants with the largest increase in PA maintained their weight loss from 6 months to 24 months, while those with a smaller increase in PA regained their weight. In addition, dietary fat or protein intake significantly modified the associations between changes in PA and changes in body weight and waist circumference over 24 months (P∆PA*diet < 0.05). CONCLUSIONS: Changes in objectively measured PA were inversely related to changes in body weight, body composition and fat distribution in response to weight-loss diets, and such associations were more evident in people on a high-fat or average-protein diet compared with a low-fat or high-protein diet.

Silibinin-modified Hydroxyapatite coating promotes the osseointegration of titanium rods by activation SIRT1/SOD2 signaling pathway in diabetic rats.

The purpose of this study is to investigate the role of Silibinin (SIL)-modified Hydroxyapatite coating on osseointegration in diabetes in vivo and in vitro and explore the mechanism of osteogenic differentiation of MC3T3-E1. RT-qPCR, Immunofluorescence, and Western blot were used to measure the expression level of oxidative Stress Indicators and osteogenic markers proteins. Moreover, CCK-8 assay was conducted to detect cell viability in hyperglycemia. Alizarin red staining and alkaline phosphatase staining were used to examine osteogenic function and calcium deposits. The diabetic rat model receive titanium rod implantation was set up successfully and Von-Gieson staining was used to examine femoral bone tissue around titanium rod. Our results showed that intracellular oxidative stress in hyperglycemia was overexpressed, while FoxO1, SIRT1, GPX1, and SOD2 were downregulated. SIL suppressed oxidative stress to promote osteogenic differentiation. Additionally, it was confirmed that SIL promoted osteogenic differentiation of MC3T3-E1 and obviously restored the osseointegration ability of diabetic rats. Further study indicated that SIL exerted its beneficial function through activation SIRT1/SOD2 signaling pathway to restore osteoblast function, and improved the osseointegration and stability of titanium rods in vivo. Our research suggested that the SIL-modulated oxidative Stress inhibition is responsible for the activation of the process of osteogenic differentiation through activation SIRT1/SOD2 signaling pathway in hyperglycemia, providing a novel insight into improving prosthetic osseointegration in diabetic patients. Hyperglycemia impaired the activity and function of MC3T3-E1 and inhibits bone formation by up-regulating intracellular ROS levels through inhibition of SIRT1/SOD2 signaling pathway. Local administrator SIL can improve the activity and function of osteoblasts and enhance osseointegration by reducing intracellular ROS through activation of SIRT1/SOD2 signaling pathway in DM rat models.

Selenium-modified calcium phosphate cement can accelerate bone regeneration of osteoporotic bone defect.

OBJECTIVE: The purpose is to observe whether local administration with selenium (Se) can enhance the efficacy of calcium phosphate cement (CPC) in the treatment of osteoporotic bone defects. METHODS: Thirty ovariectomized (OVX) rats with two defects were generated and randomly allocated into the following graft study groups: (1) OVX group (n = 10), (2) CPC group (n = 10); and (3) Se-CPC group (n = 10). Then, these selenium-modified calcium phosphate cement (Se-CPC) scaffolds were implanted into the femoral epiphysis bone defect model of OVX rats for 12 weeks. Micro-CT, history, western blot and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis were used to observe the therapeutic effect and to explore the possible mechanism. RESULT: Micro-CT and histological analysis evaluation showed that the Se-CPC group presented the strongest effect on bone regeneration and bone mineralization when compared with the CPC group and the OVX group. Protein expressions showed that the oxidative stress protein expressions, such as SOD2 and GPX1 of the Se-CPC group, are significantly higher than those of the OVX group and the CPC group, while Se-CPC remarkably reduced the expression of CAT. RT-qPCR analysis showed that the Se-CPC group displayed more OPG than the OVX and CPC groups (p < 0.05), while Se-CPC exhibited less RANKL than the OVX and CPC groups (p < 0.05). CONCLUSION: Our current study demonstrated that Se-CPC is a scheme for rapid repair of femoral condylar defects, and these effects may be achieved by inhibiting local oxidative stress and through OPG/RANKL signaling pathway.

Simvastatin reverses the harmful effects of high fat diet on titanium rod osseointegration in ovariectomized rats.

INTRODUCTION: The objectives of the present study were to determine whether simvastatin (SIM) could reverse the harmful effects on titanium rod osseointegration in ovariectomized rats fed high-fat diet (HFD). MATERIALS AND METHODS: Ovariectomized female Sprague-Dawley rats were randomly allocated to three groups and received SIM treatment plus HFD for 12 weeks. We then evaluated the microstructure parameters, histological parameters, biomechanical parameters, bone turnover, and blood lipid level. RESULTS: After 12 weeks of treatment, SIM can significantly improve bone formation around the titanium rod and osseointegration including higher values of maximum push-out force, bone area ratio (BAR), bone-to-implant contact (BIC), bone mineral density (BMD), bone volume (BV/TV), trabecular number (Tb.N), trabecular thickness (Tb.Th), mean connective density (Conn.D) when compared with the HFD group. In addition, system administration of SIM showed positive effects on collagen type 1 cross-linked C-telopeptide (CTX-1), procollagen I N-terminal propeptide (PINP), total cholesterol (TC), triglycerides (TGL), low-density lipoprotein (LDL) cholesterol and high-density lipoprotein (HDL) cholesterol. Compared with the HFD group, lower values of CTX-1, P1NP, TC, TGL and LDL were observed in the SIM+HFD group (P < 0.05). CONCLUSION: Our findings revealed that HFD may have an adverse effect on osseointegration in osteoporotic conditions, and the harmful effect of HFD on osseointegration could be reversed by SIM.

Hydrogel contained valproic acid accelerates bone-defect repair via activating Notch signaling pathway in ovariectomized rats.

The purpose was to observe whether valproic acid (VPA) has a positive effect on bone-defect repair via activating the Notch signaling pathway in an OVX rat model. The MC3T3-E1 cells were cocultured with VPA and induced to osteogenesis, and the osteogenic activity was observed by alkaline phosphatase (ALP) staining, Alizarin Red (RES) staining and Western blotting (WB). Then the hydrogel-containing VPA was implanted into the femoral epiphysis bone-defect model of ovariectomized (OVX) rats for 12 weeks. Micro-CT, biomechanical testing, histology, immunofluorescence, RT-qPCR, and WB analysis were used to observe the therapeutic effect and explore the possible mechanism. ALP and ARS staining and WB results show that the cell mineralization, osteogenic activity, and protein expression of ALP, OPN, RUNX-2, OC, Notch 1, HES1, HEY1, and JAG1 of VPA group is significantly higher than the control group. Micro-CT, biomechanical testing, histology, immunofluorescence, and RT-qPCR evaluation show that group VPA presented the stronger effect on bone strength, bone regeneration, bone mineralization, higher expression of VEGFA, BMP-2, ALP, OPN, RUNX-2, OC, Notch 1, HES1, HEY1, and JAG1 of VPA when compared with OVX group. Our current study demonstrated that local treatment with VPA could stimulate repair of femoral condyle defects, and these effects may be achieved by activating Notch signaling pathway and acceleration of blood vessel and bone formation.

Relationship Between COVID-19 Infection and Risk Perception, Knowledge, Attitude, and Four Nonpharmaceutical Interventions During the Late Period of the COVID-19 Epidemic in China: Online Cross-Sectional Survey of 8158 Adults.

BACKGROUND: So far, there have been no published population studies on the relationship between a COVID-19 infection and public risk perception, information source, knowledge, attitude, and behaviors during the COVID-19 outbreak in China. OBJECTIVE: This study aims to understand the relationships between COVID-19 infection; four personal nonpharmaceutical interventions (NPIs; handwashing, proper coughing habits, social distancing, and mask wearing); and public risk perception, knowledge, attitude, and other social demographic variables. METHODS: An online survey of 8158 Chinese adults between February 22 and March 5, 2020, was conducted. Bivariate associations between categorical variables were examined using Fisher exact test. We also explored the determinants of four NPIs as well as their association with COVID-19 infection using logistic regression. RESULTS: Of 8158 adults included, 57 (0.73%) were infected with COVID-19. The overwhelming majority of respondents showed a positive attitude (n=8094, 99.2%), positive risk perception (n=8146, 99.9%), and high knowledge levels that were among the strongest predictors of the four adopted NPIs (handwashing: n=7895, 96.8%; proper coughing: 5997/6444, 93.1%; social distancing: n=7104/8158, 87.1%; and mask wearing: 5011/5120, 97.9%). There was an increased risk of COVID-19 infection for those who did not wash their hands (2.28% vs 0.65%; risk ratio [RR] 3.53, 95% CI 1.53-8.15; P=.009), did not practice proper coughing (1.79% vs 0.73%; RR 2.44, 95% CI 1.15-5.15; P=.03), did not practice social distancing (1.52% vs 0.58%; RR 2.63, 95% CI 1.48-4.67; P=.002), and did not wear a mask (7.41% vs 0.6%; RR 12.38, 95% CI 5.81-26.36; P<.001). For those who did practice all other three NPIs, wearing a mask was associated with a significantly reduced risk of infection compared to those who did not wear a mask (0.6% vs 16.7%; P=.04). Similarly, for those who did not practice all or part of the other three NPIs, wearing a mask was also associated with a significantly reduced risk of infection. In a penalized logistic regression model including all four NPIs, wearing a mask was the only significant predictor of COVID-19 infection among the four NPIs (odds ratio 7.20, 95% CI 2.24-23.11; P<.001). CONCLUSIONS: We found high levels of risk perception, positive attitude, desirable knowledge, as well as a high level of adopting the four NPIs. The relevant knowledge, risk perception, and attitude were strong predictors of adapting the four NPIs. Mask wearing, among the four personal NPIs, was the most effective protective measure against COVID-19 infection, with added preventive effect among those who practiced all or part of the other three NPIs.

Resveratrol reverses the negative effect of alcohol on hydroxyapatite-coated implant osseointegration in senile female rats.

Previous studies have demonstrated the damaging effect of alcohol (ALH) consumption on bone tissue and bone metabolism. Resveratrol (RES) promotes osteoblast proliferation and inhibits osteoclast proliferation and positively affects bone regeneration; however, reports about effects of RES on osseointegration in aged female rats with ALH consumption are limited. This study was designed to investigate the impact of treatment with RES on osseointegration for aged female rats with ALH consumption. This study included 30 female Sprague-Dawley rats (22 months old), weighing approximately 520 g. All animals were randomly divided into 3 groups of 10: a control group (CON) receiving saline, a group receiving ALH and a group receiving ALH + RES until death after 12 weeks. The results of enhanced osseointegration in senile female rats with RES consumption were evaluated by histology, microcomputerized tomography (micro-CT), gene expression analysis and a biomechanical test. The results of this study indicated that ALH + RES showed stronger effects on the improvement of osseointegration in senile female rats with ALH consumption. The ALH + RES produced stronger effects on bone volume per total volume (BV/TV), mean trabecular thickness (Tb.Th), mean trabecular number (Tb.N) and mean trabecular separation (Tb.Sp), connective tissue density (Conn.D) and maximum push-out force for implants, and regulation of osteogenesis and bone resorption-related gene expression. These results seem to indicate that RES intervention reverses the negative effect of alcohol on hydroxyapatite-coated implant osseointegration in senile female rats with ALH consumption.

Melatonin improves the osseointegration of hydroxyapatite-coated titanium implants in senile female rats.

The aim of this study was to confirm the effect of the systemic administration of melatonin on hydroxyapatite-coated titanium (HA-Ti) implants in senile osteopenic rats. For this study 24-month-old female Sprague-Dawley rats were used. The animals were randomly divided into two groups: a control group and a melatonin group and the bilateral femurs of all the rats received HA-Ti implants. Animals in the melatonin group received treatment with melatonin (30 mg/kg day). After a 12-week healing period, rats in the melatonin group revealed improved osseointegration compared to the control group, with the bone area ratio (BAR) and bone to implant contact (BIC) increased by 1.87-fold and 1.65-fold in histomorphometry, the quantitative results of implant osseointegration and peri-implant trabeculae, such as a higher bone volume per total volume (BV/TV), trabecular number (Tb.N), the mean connective density (Conn.D), trabecular thickness (Tb.Th), and a lower trabecular spacing (Tb.Sp) in micro-computed tomography (CT) evaluation and the maximum push-out force by 1.75-fold in push out tests. Additionally, compared with the control group, melatonin could significantly up-regulate the expression of the runt-related transcription factor 2 (Runx2), osteocalcin (OC) and osteoprotegerin (OPG) genes and down-regulate the expression of the RANKL gene. These findings suggest that systemic administration with melatonin is useful to improve the fixation of HA-coated implants even in osteopenic rats through promoting Runx2, OC and OPG gene expression and inhibiting RANKL gene expression.

Rapamycin could increase the effects of melatonin against age-dependent bone loss.

Previous studies have demonstrated the beneficial effect of melatonin (MEL) on bone tissue and bone metabolism. Rapamycin (RAP) promotes osteoblast proliferation and inhibits osteoclast proliferation, and positively affects bone regeneration; however, reports about effects of RAP on bone loss for aged female rats with MEL administration are limited. This study investigated the impact of treatment with RAP on bone loss for aged female rats with MEL administration. Female Sprague-Dawley rats weighing approximately 520 g were randomly divided into 3 groups of 10: group CON, group MEL and group MEL + RAP and received saline, MEL, RAP plus MEL treatment until death at 12 weeks, respectively. The results of maintaining bone mass and bone strength with RAP plus MEL administration were evaluated by histology, microcomputerized tomography (Micro-CT), gene expression analysis and biomechanical testing. Results from this study indicated that MEL + RAP had stronger effects on the prevention and treatment of osteoporosis than MEL administration. Administration of MEL + RAP produced the strongest effects on bone parameters and strength for distal femurs and regulation of OPG/RANKL signalling pathway-related gene expression. These results seemed to indicate that RAP could increase the effects of MEL on age-dependent bone loss.

[Bletilla striata ecological planting patterns and technical principle analysis].

Bletilla striata is a shallow-rooted herbaceous plant, which is highly dependent on the planting environment.In order to improve the quality of B.striata effectively and solve the contradiction between the production of traditional Chinese medicine and the ecological environment, a variety of simulated cultivation patterns have been explored in various producing areas. Through field investigation and literature collation, this paper sorted out 24 sets of simulated cultivation modes of B.striata, including 4 kinds of sub-forest imitative wild planting techniques, 10 kinds of B.striata-economic forest cooperative development mode, 14 kinds of intercropping techniques of B.striata and 1 kind of ecological rotation technique of B.striata-edible fungi under economic forest. Combining economic benefit, social benefit and ecological benefit, the advantages and disadvantages of these planting patterns were compared. The ecological principles of B.striata simulative cultivation are analyzed in detail, as follows:(1) the wild cultivation of B.striata simulative cultivation under forest is to utilize the complementary principle of land resources under forest and spatial light and heat resources, and carry out the compound cultivation of cultivating forest with medicine and growing with short growth;(2) the intercropping of B.striata makes full use of the difference in height and height of the two plants to meet their needs for light, humidity and ventilation, reduce the cost of shading and weeding, improve the risk-resistant ability of Chinese medicinal materials cultivation, and increase the utilization rate of land, and give consideration to the benefits of both length and length;(3) the ecological rotation technology of "B.striata-edible fungi" in economic forests is to realize the effective utilization of waste bacteria and residues according to the mutually beneficial relationship between B.striata and edible fungi. The promotion and optimization of B.striata imitated wild cultivation technology will further promote the sustainable development of B.striata industry.

Local administration of aspirin with ß-tricalcium phosphate/poly-lactic-co-glycolic acid (ß-TCP/PLGA) could enhance osteoporotic bone regeneration.

Composite materials ß-tricalcium phosphate (ß-TCP) and poly-lactic-co-glycolic acid (PLGA) have achieved stable bone regeneration without cell transplantation in previous studies. Recent research shows that aspirin (ASP) has great potential in promoting bone regeneration. The objective of the present study was to incorporate PLGA into ß-TCP combined with a lower single-dose local administration of ASP to enhance its in vivo biodegradation and bone tissue growth. After the creation of a rodent critical-sized femoral metaphyseal bone defect, PLGA -modified ß-TCP (TP) was prepared by mixing sieved granules of ß-TCP and PLGA (50:50, v/v) for medical use, then TP with dripped 50 µg/0.1 ml and 100 µg/0.1 ml aspirin solution was implanted into the defect of OVX rats until death at 8 weeks. The defected area in distal femurs of rats was harvested for evaluation by histology, micro-CT, biomechanics and real time RT-PCR. The results of our study show that a single-dose local administration of ASP combined with the local usage of TP can increase the healing of defects in OVX rats. Single-dose local administration of aspirin can improve the transcription of genes involved in the regulation of bone formation and vascularization in the defect area, and inhibits osteoclast activity. Furthermore, treatments with a higher single-dose local administration of ASP and TP showed a stronger effect on accelerating the local bone formation than while using a lower dose of ASP. The results from our study demonstrate that the combination of a single-dose local administration of ASP and ß-TCP/PLGA had an additive effect on local bone formation in osteoporosis rats, and bone regeneration by PLGA/ß-TCP/ASP occured in a dose-dependent manner.

Single-dose local administration of parathyroid hormone (1-34, PTH) with ß-tricalcium phosphate/collagen (ß-TCP/COL) enhances bone defect healing in ovariectomized rats.

Parathyroid hormone (1-34, PTH) combined ß-tricalcium phosphate (ß-TCP) achieves stable bone regeneration without cell transplantation in previous studies. Recently, with the development of tissue engineering slow release technology, PTH used locally to promote bone defect healing become possible. This study by virtue of collagen with a combination of drugs and has a slow release properties, and investigated bone regeneration by ß-TCP/collagen (ß-TCP/COL) with the single local administration of PTH. After the creation of a rodent critical-sized femoral metaphyseal bone defect, ß-TCP/COL was prepared by mixing sieved granules of ß-TCP and atelocollagen for medical use, then ß-TCP/COL with dripped PTH solution (1.0 µg) was implanted into the defect of OVX rats until death at 4 and 8 weeks. The defected area in distal femurs of rats was harvested for evaluation by histology, micro-CT, and biomechanics. The results of our study show that single-dose local administration of PTH combined local usage of ß-TCP/COL can increase the healing of defects in OVX rats. Furthermore, treatments with single-dose local administration of PTH and ß-TCP/COL showed a stronger effect on accelerating the local bone formation than ß-TCP/COL used alone. The results from our study demonstrate that combination of single-dose local administration of PTH and ß-TCP/COL had an additive effect on local bone formation in osteoporosis rats.

Prevention of ovariectomy-induced osteoporosis in rats : Comparative study of zoledronic acid, parathyroid hormone (1-34) and strontium ranelate.

Recently, the use of the pharmacological agents strontium ranelate (SR), parathyroid hormone (1-34, PTH) and zoledronic acid (ZA) has come to prominence for the treatment of osteoporosis due to their ability to prevent bone loss in osteoporotic patients. Although much emphasis has been placed on using pharmacological agents for the prevention of disease, much less attention has been placed on which one is more effective. There is still no direct comparative study on these three drugs. The aim of the present study was to investigate the effect of SR, PTH, ZA on preventing ovariectomy-induced osteoporosis in rats. After bilateral ovariectomy the rats randomly received vehicle, SR (500 mg/kg body weight/day, orally), PTH (20 µg/kg/day, subcutaneously) or a single injection of ZA (0.1 mg/kg, i.v.) until death at 12 weeks. The distal femurs were harvested for evaluation of bone metabolism. The rats treated with ZA demonstrated the highest levels of new bone formation as assessed by microcomputed tomography (CT), biomechanical strength, histological analysis and bone metabolism. Furthermore, PTH and SR showed a stronger effect on improving trabecular bone mass at 12 weeks. The results from the present study demonstrate that systemic administration of PTH, SR and ZA could prevent bone loss, while a single dose of ZA has a better effect on preventing ovariectomy-induced osteoporosis than either PTH or SR.

Combined treatment with alendronate and Drynaria rhizome extracts : Effect on fracture healing in osteoporotic rats.

The effects of both alendronate (ALN) and Drynaria rhizome extracts (DRE) alone could promote bone healing in osteoporotic fractures but there are no reports about the combined use of ALN and DRE for promotion of bone healing of fractures in osteoporotic settings. This study investigated the effects of ALN plus DRE on fractures in osteopenic rats. Osteopenic rats underwent unilateral transverse osteotomy on the femur fixed by a sterilized Kirschner wire 2 weeks after intragastric administration of retinoic acid (80 mg/kg body weight/day). Subsequently, the animals were randomly divided into four groups: control, ALN, DRE and ALN + DRE. All rats from groups ALN, DRE and ALN + DRE received ALN (40 mg/kg, weekly), DRE (90 mg/kg/day), or both for 2, 4 and 6 weeks. The results of our study indicated that all treatment promoted fracture healing and callus formation compared to controls but ALN + DRE treatment showed significantly stronger effects than ALN or DRE alone in histological, X­ray and biomechanical tests. These results seem to indicate that combined treatment with ALN and DRE has an additive effect on fracture healing and callus formation in osteoporotic rats.

A novel micro-CT-based method to monitor the morphology of blood vessels in the rabbit endplate.

PURPOSE: The aim of this study was to develop a novel method for observing the morphology of the blood vessels in the rabbit endplate. METHODS: Twenty 6-month-old rabbits were used in this study. The blood vessels in the L5 endplate in Group A were injected with iohexol and Group B with barium sulfate. Group C was the control group with saline. To optimize the study, Group B was divided into two subgroups: Group B-1 was injected with 100% (w/v) barium sulfate and Group B-2 with 50% (w/v). After injection, the L4-L5 vertebral body was excised and the cranial endplate of L5 was scanned using a micro-CT scanner. Models of the vertebral endplate and vessels were reconstructed using the 3D reconstruction software (Mimics 16.0) by calculating a bone threshold value, and then merged these two models to create a superimposed model. RESULTS: The 3D vessel models could not be created in Groups A and C, but they were clearly visualized in Group B. In the 3D model, the blood vessels extended from the subchondral bone to the endplate, and the density of the blood vessels in the area of the nucleus pulposus (NP) was higher than in the annulus fibrosus. CONCLUSIONS: The results of this study suggest that the blood vessels in the rabbit endplate can be clearly observed by the method described using barium sulfate [the 50% (w/v) gave better results compared with the 100% (w/v)]. The information from the 3D vessel structure could provide essential data to help us understand the nutrient pathways within the vertebral endplate.

The effects of combined human parathyroid hormone (1-34) and simvastatin treatment on the interface of hydroxyapatite-coated titanium rods implanted into osteopenic rats femurs.

The effect of human parathyroid hormone 1-34 (PTH) and simvastatin (SIM) alone could promote bone healing in osteoporotic implant fixation, but there are no reports about the combined use of PTH and SIM for promotion of bone healing around implant in osteoporotic settings. This study aims to investigate effects of PTH + SIM on implant stabilization in osteopenic rats. Fourteen weeks after chronically fed a low protein diet, osteopenic rats randomly received implants. Subsequently, the animals were randomly divided into four groups: Control, SIM, PTH and PTH + SIM. Then all rats from groups PTH, SIM and PTH + SIM received PTH (40 µg/kg, three times a week), SIM (25 mg/kg, daily), or both for 12 weeks. The results of our study indicated that all treatments promoted bone healing around implant compared to Control, but PTH + SIM treatment showed significantly stronger effects than PTH or SIM alone in histological, micro-CT, and biomechanical tests. The results indicated additive effects of PTH and SIM on implant fixation in osteoporotic rats.

Effect of teriparatide on repair of femoral metaphyseal defect in ovariectomized rats.

OBJECTIVE: This study aimed to investigate the effect exerted by teriparatide on the repair of femoral metaphyseal defect in ovariectomized rats. METHOD: Female Sprague-Dawley rats were ovariectomized and after 3 months a critically sized defect of 3 mm in diameter-a through-hole bone defect-was drilled into each distal femur of the ovariectomized rats. The rats were injected with teriparatide (30 µg/kg) parathyroid hormone (PTH) in the peritoneum three times per week. After 4 and 8 weeks the animals were killed and the blood and bilateral femora were harvested for biochemical analysis, histopathological observation, and micro-computed tomography (CT) examination. RESULTS: The PTH group and control group were compared 4  and 8 weeks after surgery. PTH increased bone formation in the defect area. Moreover, PTH showed the strongest effects on bone volume per total volume, trabecular number, trabecular thickness, trabecular separation, and total fluorescence-marked new bone area. Additionally, the PTH treatment group showed inhibited serum concentrations of C-terminal telopeptide of type I collagen and enhanced expression of calcium, phosphorus, and bone alkaline phosphatase. CONCLUSION: Our findings suggest a positive effect of PTH on defect healing in ovariectomized rats.

Protease Omi cleaving Hax-1 protein contributes to OGD/R-induced mitochondrial damage in neuroblastoma N2a cells and cerebral injury in MCAO mice.

AIM: In the penumbra after focal cerebral ischemia, an increase of protease Omi is linked to a decrease of Hs1-associated protein X-1 (Hax-1), a protein belonging to the Bcl-2 family. In this study we investigated the mechanisms underlying the regulation of Hax-1 by protease Omi in cerebral ischemia/reperfusion (I/R) injury. METHODS: Mouse neuroblastoma N2a cells were subjected to oxygen-glucose deprivation and reoxygenation (OGD/R); cell viability was assessed with MTT assay. Mice underwent 2-h middle cerebral artery occlusion (MCAO) and reperfusion, and the infarct volume was determined with TTC staining. The expression of Omi and Hax-1 was detected using immunoblot and immunofluorescence assays. The mitochondrial membrane potential was measured using TMRM staining. RESULTS: In the brains of MCAO mice, the protein level of Omi was significantly increased, while the protein level of Hax-1 was decreased. Similar changes were observed in OGD/R-treated N2a cells, but the mRNA level of Hax-1 was not changed. Furthermore, in OGD/R-treated N2a cells, knockdown of Omi significantly increased Hax-1 protein level. Immunofluorescence assay showed that Omi and Hax-1 were co-localized in mitochondria of N2a cells. OGD/R caused marked mitochondrial damage and apoptosis in N2a cells, while inhibition of Omi protease activity with UCF-101 (10 µmol/L) or overexpression of Hax-1 could restore the mitochondrial membrane potential and attenuate cell apoptosis. Moreover, pretreatment of MCAO mice with UCF-101 (7.15 mg/kg, ip) could restore Hax-1 expression, inhibit caspase activation, and significantly reduce the infarct volume. CONCLUSION: Protease Omi impairs mitochondrial function by cleaving Hax-1, which induces apoptosis in OGD/R-treated N2a cells and causes I/R injury in MCAO mice.