Effects of pre-osteogenic differentiation on the bone regeneration potentiality of marrow mesenchymal stem cells/poly(ethylene glycol)-diacrylate hydrogel using a rat cranial defect model.

Transplanting cell/hydrogel constructs into a bone defect site is an effective strategy to repair the damaged tissues. However, before transplantation, there are various methods to culture cell/hydrogel constructs. Especially, the preferred pre-osteogenic differentiation period to achieve satisfied bone regeneration should be determined. To this end, Bone marrow mesenchymal stem cells (BMSCs) were firstly photo-encapsulated into poly(ethylene glycol)-diacrylate (PEGDA) hydrogel. Then the constructs were implanted in rat calvarial defects after being osteogenically induced for 0, 7, 14, and 21 days. In vitro experiments demonstrated that the proliferation of BMSCs in the hydrogels deceased significantly from 0 day to 7 days. The activity and the gene expression of alkaline phosphatase, besides the gene expression of bone morphogenetic protein-2 peaked at day 14, whereas the gene expression of osteocalcin and the formation of calcium nodules increased with the prolongation of differentiation time. In vivo results showed that limited areas of newly formed bone were found in the day0 and day21 groups. In the day7 group, obvious new bone with bone marrow space was found, while the day14 group nearly achieved complete bone healing. Our data suggested that the period of in vitro pre-osteogenic differentiation played a crucial role for the osteogenesis of BMSCs/PEGDA hydrogels. Furthermore, we found that a pre-differentiation for 14 days is preferable for bone regeneration in the rat cranial defects.

Curcumin regulates EZH2/Wnt/ß-Catenin pathway in the mandible and femur of ovariectomized osteoporosis rats.

Osteoporosis (OP) behaves in different manners in different parts of the skeleton. This study aims to investigate the effects of curcumin on bone mass of the mandibular and femur from ovariectomized OP rats and to validate whether enhancer of zeste homolog 2 (EZH2)/Wnt/ß-Catenin pathway is involved in this process. Curcumin was administered intragastrically into ovariectomized rats for 12 weeks. The bone parameters and the morphology of the trabecular bone of the left mandible and left femur were assessed by micro-computed tomography assay. Morphological changes of the left mandible and left femur were evaluated by hematoxylin and eosin staining. The mRNA levels of EZH2, ß-Catenin, and Runx2 in the right mandible and right femur were examined by quantitative real-time polymerase chain reaction. Immunohistochemistry was performed to assess EZH2 expression. Both the mandible and femur exhibited OP-like changes in ovariectomized rats, while the mandible bone resorption was less than the femur bone resorption. Curcumin intragastric administration improved bone microstructure and promoted bone formation in the mandible and femur. Curcumin inhibited EZH2 mRNA level and induced that of ß-Catenin and Runx2 in the mandible and femur. Collectively, curcumin exerts protective effects against OP, possibly by regulating the EZH2/Wnt/ß-Catenin pathway.

Influence of low modulus Co-Zr alloys surface modification on protein adsorption and MC3T3-E1, NIH3T3 and RAW264.7 cell behaviour.

Three types of Co-xZr (x = 5, 7.5, and 10 wt.%) were treated with hydroxyapatite (HA) and used as an object to investigate the effect of HA coating on the surface and biocompatibility of Co-xZr alloys. And the protein adsorption and the subsequent biological behaviour of osteoblast, fibroblast and macrophages were also investigated. The surface microstructure and wettability were assessed by scanning electron microscopy (SEM) and static angle profilometer. To evaluate the biocompatibility of Co-xZr and Co-xZr-HA, we quantified plasma proteins adsorption by bicinchoninic acid assay (BCA), cytotoxicity and cell proliferation by cell counting kit-8 (CCK-8) and scanning electron microscopy (SEM). The results indicated that Co-xZr-HA alloy surfaces were more hydrophilic and had higher affinity to plasma proteins. Higher protein concentrations were found adsorbed onto Co-7.5Zr-HA and Co-10Zr-HA alloys. Cytotoxicity analysis indicated that HA coating improved the biocompatibility of Co-xZr alloys. Furthermore, the comparable results of co-incubation of Co-xZr-HA alloys with cells reveal cellular attachments to HA surfaces. HA was successfully formed on Co-xZr alloys and modified the surface structure and biocompatibility of the alloys. Co-10Zr-HA and Co-7.5Zr-HA had the most favourable properties and cytocompatibility, and therefore can be potentially used for dental implants.

[Effects of curcumin on EZH2 mRNA expression in the mandible and femur of ovariectomized osteoporosis rats].

PURPOSE: To study the effects of curcumin on EZH2 mRNA expression in the mandible and femur of ovariectomized osteoporosis rats,and to investigate its protective effect and mechanism. METHODS: Thirty female 6-month old SD rats were randomly divided into sham group,OVX group and experimental group. The rats in the experimental groups were given curcumin (110 mg/kg) by intragastric administration after ovariectomy, while rats in the sham group and OVX group were given the same dosage of carboxymethylcellulose sodium solution, once a day for 12 weeks. All rats were sacrificed after the last intragastric administration. The serum samples were collected for detemination of biochemistrical parameters. Micro-CT was used for bone parameters and the morphology of the trabecular bone of the left mandibles and femurs. The expression level of EZH2mRNA in right mandible and femurs tissue was detected by reverse transcription polymerase chain reaction (RT-PCR). SPSS22.0 software package was used for statistical analysis. RESULTS: The expression of EZH2mRNA in the OVX group was significantly higher than the sham group (P<0.05). Compared with the OVX group,curcumin increased BMD and improved bone microstructure, decreased serum contents of alkaline phosphatase,and down-regulated the expression levels of EZH2mRNA in bone tissues of rats with osteoporosis (P<0.05). CONCLUSIONS: Curcumin can effectively prevent the lose of bone volume of ovariectomized rats, and repaire bone microstructure. Its mechanism is related with down -regulation of EZH2mRNA.

[Effect of different surface zirconium oxide treatments on binding strength between zirconia and veneering ceramics].

OBJECTIVE: To compare the effect of three different surface zirconium oxide treatments on binding strength and fracture patterns between zirconia and veneering ceramics.
 Methods: A total of 40 zirconia specimens and 10 nickel-chromium alloy were divided into 5 groups (n=10 in each group): a treatment group by zirconium oxide sand-blaste (Group A), a zirconia bonded porcelain group (Group B), a hot-etching solution group (Group C), a non-treatment zirconia (Group D) and a non-treatment nickel-chromium alloy group (Group E). After all treatments, a veneering porcelain (4 mm×4 mm×2 mm) was formed onto the center of all substrate specimens by plastic coating method. Shear bond strength (SBS) test with a universal testing machine was used in each group. Scanning electron microscopy was used to observe the surface morphology of the damaged specimen interface, which was randomly selected from each group.
 Results: The SBS test showed that there was no significant difference in SBS results between the Group A, the Group B and the Group D (both P>0.05), and both of them were significant less than that in the group E (both P<0.05). The SBS results in the Group C were significantly higher than that in the Group D, the Group A, and the Group B (all P<0.05), but there were no significant difference compared with that in the Group E (P<0.05). 
 Conclusion: Sand-blaste and liner application on zirconia ceramic contribute no effect to binding strength between zirconia and veneering ceramics, but hot-etching solution can increase the binding strength between zirconia and veneering ceramics.

In vitro cell response and in vivo primary osteointegration of highly porous Ta-Nb alloys as implant materials.

By combining the biocompatibility of Ta and excellent performance of Nb, the ß-type Ta-xNb (x = 5, 10, and 15 wt %) alloys with three-dimensional network structure was successfully fabricated. This work was designed to carry out the systematic study of in vitro/vivo biocompatibility of highly porous Ta-Nb alloys. In the present study, the in vitro biological response was characterized by cell attachment, cytotoxicity test, and gene expression. Cells were well distributed on porous alloy surface and both bulk/porous Ta-15Nb alloys showd a 0-1 RGR level. Moreover, the upregulated gene expression may reveal an increase in bone regeneration. An assessment of primary osteogenic response in vivo was achieved by histological evaluation and pull-out test. In vivo results demonstrated that the porous Ta-15Nb alloy showed a higher pull-out force than bulk Ta-15Nb alloy and exhibited a more extensive contact between implants and bones after 12 weeks. Based on the present experiments, porous Ta-Nb alloy scaffolds had a good biocompatibility and were promising for biomedical applications. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2018. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 573-581, 2019.

[Evaluation of osseointegration of domestic porous tantalum-niobium alloy].

PURPOSE: To evaluate the osseointegration of domestic porous tantalum-niobium(PTa-Nb) alloy. METHODS: A total of 36 adult New Zealand rabbits were selected and divided into 3 groups. Pta-Nb rods(3.5 mm×10 mm) were implanted into the femoral condyle of each rabbit.The rabbits were sacrificed successively at 4,8,12 weeks after operation. X-ray, hard tissue slices stained with toluidine blue, scanning electron microscope(SEM), X-ray energy disperse spectroscopy(XEDS) and push-out test were used to test the osseointegration of PTa-Nb. SPSS19.0 software package was used for statistical analysis. RESULTS: X-ray films showed no obvious inflammation,as well as implants loosing and bone resorption. Density of the bone around implants increased. Hard tissue slices displayed chimeric shape in the implant-bone interface. New bone contacted directly to the surface of PTa-Nb and got more and more closer with the increase of healing time. Osteoid formed in the inner pores at 4 weeks after operation,and more mature bone tissue grew into inner part of PTa-Nb at 12 weeks. SEM showed bone tissue was more denser and closer to the Pta-Nb,which was similar to the results of hard tissue slices. XEDS analysis showed the percent content of the calcium and phosphate of the tissue within PTa-Nb increased gradually in the three groups.The ratio of Ca/P at 8 and 12 weeks was significantly higher than at 4 weeks (P<0.05); Push-out test showed the shear-strength of PTa-Nb implants increased from (8.26±0.75) MPa at 4 weeks to (21.04±1.46) MPa at 12 weeks (P<0.05). CONCLUSIONS: The domestic PTa-Nb alloy has good osseointegration with both bone tissue ongrowth and ingrowth,which may be a high potential biomimetic bone material.

[Expressions of RANK, RANKL, and osteoprotegerin in male rats at different ages].

OBJECTIVE: To investigate the expression of receptor activator of nuclear factor-κB (RANK), its ligand RANKL, and osteoprotegerin, and observe the effects of αD3 on their expressions in male rats at different ages. METHODS: Wistar rats at 6 weeks, 6 months, and 24 months (n=15) were examined for mRNA expressions of RANK/RANKL and osteoprotegerin in the left proximal femur using RT-PCR and for their protein expressions in the right femur using immunohistochemistry. RANK/RANKL and osteoprotegerin expressions were also detected in another 15 rats aged 24 months following intragastric administration of 0.05 µg/kg αD3 (3 times a week for 10 weeks). RESULTS: Compared with 6-week-old rats, 6-month- and 24-month-old rats showed a 6.2-fold and 7.3-fold increase of RANKL mRNA expression, respectively (P<0.05), and osteoprotegerin mRNA levels increased slightly with age. αD3 treatment resulted in significantly increased expression of RANK in 24-month-old rats with a lowered RANKL/osteoprotegerin ratio. RANKL and osteoprotegerin were co-localized in the osteoblasts and chondrocytes. αD3 treatment also caused an increased expression of osteoprotegerin mRNA in 24-month-old rats. CONCLUSION: The age-related increase of the ratio of RANKL/osteoprotegerin mRNA promotes osteoclast activity and bone turnover. αD3 has favorable effect on osteogenesis and suppress bone absorption in the femur possibly by reducing RANK expression and lowering RANKL/osteoprotegerin ratio.

[Expression of RANKL/OPG in male mandible of different ages].

PURPOSE: To investigate the relationship between male osteoporosis and the expression of receptor activator of NF-κB ligand(RANKL) and osteoprotegerin(OPG)-mRNA in male mandible at different ages. METHODS: Between May 2008 and January 2009, bone tissues of the mandible were collected as the experimental material from 46 patients suffering from jaw facial deformity and extraction. The patients with periodontist, systemic disorder and smoking as well as drinking were excluded.They were divided into three groups: young group, whose age was 10-29 years old ;middle age group, whose age was 30-59 years old; aged group, whose age was 60-89 years old. The expression of RANKL mRNA and OPG mRNA was examined by real-time PCR. The data was analyzed using ANOVA followed by least significant difference (LSD) test with SPSS16.0 software package. RESULTS: (1)Compared with the young group, RANKL mRNA level in mandible was 2.1-fold and 5.3-fold higher in the middle age and aged groups, respectively, whereas OPG mRNA level was 3.3-fold and 4.8-fold higher in middle age and aged groups, respectively. RANKL and OPG were positively correlated with age.(2)The ratio of RANKL/OPG in middle age group was lower than that of young group and old group,respectively. CONCLUSIONS: (1)The expression of RANKL and OPG increases with age remarkably.(2)Bone formation of the mandible is activated in middle aged group. The formation is over absorption.(3)Bone formation of the mandible in aged group is at low level. Bone absorption exceeds bone formation.

The relationship between bone turnover markers and BMD decreasing rates in Chinese middle-aged women.

BACKGROUND: The relationship between bone turnover markers (BTMs) and BMD decreasing rate (BDR) in Chinese women is unclear. Wu investigated the relationship between (BTMs) and BDR at various skeletal sites in Chinese middle-aged women. METHODS: A cross-section study of 555 healthy Chinese women over 35-60years of age. BMD at posteroanterior spine, the left hip, and the left forearm were measured with a DXA. Levels of serum osteocalcin (OC), bone-specific alkaline phosphatase (BAP), cross-linked N-terminal telopeptides of type I collagen (sNTX) and total urinary deoxypyridinoline (uDPD) were determined. RESULTS: BDR at various skeletal sites had significant negative correlation with serum OC(r=-0.395 to -0.530), BAP(r=-0.297 to -0.486), and sNTX(r=-0.207 to -0.272). After adjustment of age and weight, serum OC, BAP, and sNTX rather than total uDPD still exhibited significant correlations with BDR. Stepwise regression analyses showed that, serum OC and BAP were the significantly negative determinants of BDR. Between 4.7-27.7% and 1.2-16.1% of the changes in BDR were determined by serum OC and BAP, respectively. However, sNTX and total uDPD had no significant effect on BDR at various skeletal sites. CONCLUSIONS: This study indicated the correlation between BTMs and early-stage BDR in Chinese middle-aged women and suggested that serum OC and BAP, rather than sNTX and total uDPD, are the key determining factors of early BMD decreases.

[Effects of monofluorphosphate and alphaD3 on the bone mineral density of mandibular of ovariectomy rats].

OBJECTIVE: To investigate the effects of monofluorphosphate and alphaD3 on the bone mineral density of mandibular of ovariectomy rats. METHODS: 39 female Sprague-Dawley rats about 90 days of age were randomly divided into the following treatment groups and treated for 12 weeks: SHAM group, OVX group, MFP group, MFP+alphaD3 group. Rats of MFP group received monofluorphosphate by gastric feeding. Rats of MFP+alphaD3 group received monofluorphosphate and alphatD3 by gastric feeding. The femur and the mandibles were collected for histomorphometry and bone mineral density measurement. RESULTS: The bone area, bone thick, and bone mineral density of femur in the OVX group were significantly lower than those of other groups, while the trabecular separation of the OVX group was significantly higher than that of other groups. The bone mineral density of mandible was increased in OVX group and MFP+alphaD3 group compared with SHAM group. CONCLUSION: Monofluorphosphate and alphaD3 can obviously increase the mass of mandible. The bone mineral density of femur decreased after ovariectomy while it increased in mandibular.