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DNA methylation is an important way to regulate gene expression in eukaryotes. In order to reveal the role of DNA methylation in the regulation of germ cell-specific piwi gene expression during spermatogenesis of Japanese flounder (Paralichthys olivaceus), the expression profiles of piwil1 (piwi-like 1) and piwil2 (piwi-like 2) genes in the gonads of female, male, and sex-reversed pseudo-male P. olivaceus were analyzed, and the dynamic of DNA methylation was investigated. As a result, piwil1 and piwil2 genes were highly expressed in the testis of both male and pseudo-male P. olivaceus, with significant variation among male individuals. The DNA methylation levels in the promoter regions of both piwil1 and piwil2 were negatively correlated with their expression levels, which may contribute to the transcriptional regulation of piwi genes during spermatogenesis. There was also sperm quality variation among male P. olivaceus, and the sperm curvilinear velocity was positively correlated with the expression of both piwil1 and piwil2 genes. These results indicated that the DNA methylation in piwil1 and piwil2 promoter regions may affect the initiation of piwi gene transcription, thereby regulating gene expression and further affecting the spermatogenesis process and gamete quality in P. olivaceus.

Abstract Background The repair of critical-sized bone defects is always a challenging problem. Electromagnetic fields (EMFs), used as a physiotherapy for bone defects, have been suspected to cause potential hazards to human health due to the long-term exposure. To optimize the application of EMF while avoiding its adverse effects, a combination of EMF and tissue engineering techniques is critical. Furthermore, a deeper understanding of the mechanism of action of EMF will lead to better applications in the future. Methods In this research, bone marrow mesenchymal stem cells (BMSCs) seeded on 3D-printed scaffolds were treated with sinusoidal EMFs in vitro. Then, 5.5 mm critical-sized calvarial defects were created in rats, and the cell scaffolds were implanted into the defects. In addition, the molecular and cellular mechanisms by which EMFs regulate BMSCs were explored with various approaches to gain deeper insight into the effects of EMFs. Results The cell scaffolds treated with EMF successfully accelerated the repair of critical-sized calvarial defects. Further studies revealed that EMF could not directly induce the differentiation of BMSCs but improved the sensitivity of BMSCs to BMP signals by upregulating the quantity of specific BMP (bone morphogenetic protein) receptors. Once these receptors receive BMP signals from the surrounding milieu, a cascade of reactions is initiated to promote osteogenic differentiation via the BMP/Smad signalling pathway. Moreover, the cytokines secreted by BMSCs treated with EMF can better facilitate angiogenesis and osteoimmunomodulation which play fundamental roles in bone regeneration. Conclusion In summary, EMF can promote the osteogenic potential of BMSCs and enhance the paracrine function of BMSCs to facilitate bone regeneration. These findings highlight the profound impact of EMF on tissue engineering and provide a new strategy for the clinical treatment of bone defects.

Abstract Background Intervertebral fusion is the most common surgery to treat lumbar degenerative disease (LDD). And the graft material used in the operation is derived from the iliac crest to promote fusion. However, autografts possess the fatal disadvantage of lack of source. Therefore, economical and practical bone substitutes are urgently needed to be developed. Sinusoidal electromagnetic fields (EMF) combined with tissue engineering techniques may be an appropriate way to promote intervertebral fusion. Methods In this research, porous scaffolds made of polycaprolactone (PCL) and nano-hydroxyapatite (nHA) were used as cell carriers. Then, the scaffolds loaded with bone marrow mesenchymal stem cells (BMSCs) were treated with sinusoidal electromagnetic field and the osteogenic capability of BMSCs was tested later. In addition, an intervertebral disc of the tail vertebra of the rat was removed to construct a spinal intervertebral fusion model with a cell-scaffold implanted. The intervertebral fusion was observed and analyzed by X-ray, micro-CT, and histological methods. Results BMSCs stimulated by EMF possess splendid osteogenic capability under an osteogenic medium (OM) in vitro. And the conditioned medium of BMSCs treated with EMF can further promote osteogenic differentiation of the primitive BMSCs. Mechanistically, EMF regulates BMSCs via BMP/Smad and mitogen-activated protein kinase (MAPK)-associated p38 signaling pathways. In vivo experiments revealed that the scaffold loaded with BMSCs stimulated by EMF accelerated intervertebral fusion successfully. Conclusion In summary, EMF accelerated intervertebral fusion by improving the osteogenic capacity of BMSCs seeded on scaffolds and might boost the paracrine function of BMSCs to promote osteogenic differentiation of the homing BMSCs at the injured site. EMF combined with tissue engineering techniques may become a new clinical treatment for LDD.