p38 Signaling Mediates Naringin-Induced Osteogenic Differentiation of Porcine Metanephric Mesenchymal Cells

Objective:To explore the potential of metanephric mesenchymal cells(MMCs)for osteogenesis and naringin’s ability to enhance this process and its molecular mechanism.Methods:Porcine MMCs at 70 days of gestation were used as tool cells,cultured in osteogenic induction medium,identified by immunocytochemistry staining.Osteogenic potential of porcine MMCs and naringin’s ability to enhance this process was tested by detecting changes in cell viability,alkaline phosphatase(ALP)activity,the expression of runt-related transcription factor 2(Runx2),osteopontin(OPN)and osteocalcin(OCN),and the formation of mineralized nodules,and the application of the p38 signaling pathway inhibitor SB203580 vitiated the osteogenesis-promoting effect of naringin.Results:Immunocytochemical staining showed that the cells were Vimentin and Six2(+),E-cadherin and CK-18(−).Naringin can activate the p38 signaling pathway to enhance the osteogenesis of porcine MMCs by increasing cell viability,ALP activity,the expressions of Runx2,OPN and OCN,and the formation of mineralized nodules(P<0.05).The application of p38 signaling pathway inhibitor SB203580 vitiated the osteogenesis-promoting effect of naringin,manifested by decreased ALP activity,the expressions of Runx2,OPN and OCN,and the formation of mineralized nodules(P<0.05).Conclusion:Naringin,the active ingredient of Chinese herbal medicine Rhizoma Drynariae for nourishing Shen(Kidney)and strengthening bone,enhances the osteogenic differentiation of renal MMCs through the p38 signaling pathway.

Three-dimensional printing of β-tricalcium phosphate/calcium silicate composite scaffolds for bone tissue engineering

Bioactive scaffolds with interconnected porous structures are essential for guiding cell growth and new bone formation. In this work, we successfully fabricated three-dimensional （3D） porous β-tricalcium phosphate （β-TCP）/calcium silicate （CS） composite scaffolds with different ratios by 3D printing technique and further investigated the physiochemical properties, in vitro apatite mineralization properties and degradability of porous β-TCP/CS scaffolds. Moreover, a series of in vitro cell experiments including the attachment, proliferation and osteogenic differentiation of mouse bone marrow stromal cells were conducted to testify their biological performances. The results showed that 3D printed β-TCP/CS scaffolds possessed of controllable internal porous structures and external shape. Furthermore, the introduction of CS decreased the shrinkage of scaffolds and improved the in vitro apatite formation activity and degradation rate. Meanwhile, compared with pure β- TCP scaffold, the β-TCP/CS composite scaffolds were more conducive to promote cell adhesion, spread and osteogenesis differentiation. However, when the content of CS was increased to 45%, the ions dissolution rate of the composite scaffolds was so high that leaded to the increase in pH value, which inhibited the proliferation of cells. Our results suggested that the introduction of appropriate CS into β-TCP bioceramic is an effective strategy to prepare bioactive 3D printed bioceramic scaffolds for hard tissue regeneration.

Magnetic hydroxyapatite nanobelt-stem cell hybrid spheroids for remotely patterning bone tissues

The low survival rate and poor differentiation efficiency of stem cells,as well as the insufficient integration of implanted stem cells,limit the regeneration of bone defects.Here,we have developed magnetic ferroferric oxidehydroxyapatitepolydopamine(Fe3O4-HAp-PDA)nanobelts to assemble mesenchymal stem cells(MSCs)into a three-dimensional hybrid spheroid for patterning bone tissue.These nanobelts,which are featured by their highaspect ratio and contain Fe3O4 nanospheres with a PDA coating,can be manipulated by a magnetic field and foster enhanced cell-nanobelt interactions.This strategy has been demonstrated to be effective for both bone marrow mesenchymal stem cells and adipose-derived mesenchymal stem cells,enabling remote manipulation of stem cell spheroids and efficient spheroid fusion,which in turn accelerates osteogenic differentiation.Consequently,this methodology serves as an efficient and general tool for bone tissue printing and can potentially overcome the low survival rate and poor differentiation efficiency of stem cells,as well as mismatched interface fusion issues.

Influence of the intensity and loading time of direct current electric field on the directional migration of rat bone marrow mesenchymal stem cells

Exogenic electric fields can effectively accelerate bone healing and remodeling through the enhanced migration of bone marrow mesenchymal stem cells （BMSCs） toward the injured area. This study aimed to determine the following： （1） the direction of rat BMSC （rBMSC） migration upon exposure to a direct current electric field （DCEF）, （2） the optimal DCEF intensity and duration, and （3） the possible regulatory role of SDF-1/ CXCR4 axis in rBMSC migration as induced by DCEF. Results showed that rBMSCs migrated to the positive electrode of the DCEF, and that the DCEF of 200 mV/mm for 4 h was found to be optimal in enhancing rBMSC migration. This DCEF strength and duration also upregulated the expression of osteoblastic genes, including ALP and OCN, and upregulated the expression of ALP and Runx2 proteins. Moreover, when CXCR4 was inhibited, rBMSC migration due to DCEF was partially blocked. These findings indicated that DCEF can effectively induce rBMSC migration. A DCEF of 200 mV/mm for 4 h was recommended because of its ability to promote rBMSC migration, proliferation, and osteogenic differentiation. The SDF-1/CXCR4 signaling pathway may play an important role in regulating the DCEF-induced migration of rBMSCs.

Biomimetic cell-adhesive ligand-functionalized peptide composite hydrogels maintain stemness of human amniotic mesenchymal stem cells

In vivo,stem cells reside in a three-dimensional(3D)extracellular microenvironment in which complicated biophysical and biochemical factors regulate their behaviors.Biomimicking of the stem cellmatrix interactions is an ideal approach for controlling the stem cell fate.This study investigates the effects of the incorporation of cell-adhesive ligands in 3D self-assembling peptide hydrogels to modulate stem cell survival,proliferation,maintenance of stemness,and osteogenic differentiation.The results show that the composite hydrogels were non-cytotoxic and effective for maintaining human amniotic mesenchymal stem cell(hAMSC)survival,proliferation and phenotypic characterization.The expression levels of pluripotent markers were also upregulated in the composite hydrogels.Under inductive media conditions,mineral deposition and mRNA expression levels of osteogenic genes of hAMSCs were enhanced.The increasing expression of integrin aand b-subunits for hAMSCs indicates that the ligandintegrin interactions may modulate the cell fate for hAMSCs in composite hydrogels.