RESUMO
Collagen XVII (COL17) is a transmembrane protein that mediates skin homeostasis. Due to expression of full length collagen was hard to achieve in microorganisms, arising the needs for selection of collagen fragments with desired functions for microbial biosynthesis. Here, COL17 fragments (27-33 amino acids) were extracted and replicated 16 times for recombinant expression in Escherichia coli. Five variants were soluble expressed, with the highest yield of 223 mg/L. The fusion tag was removed for biochemical and biophysical characterization. Circular dichroism results suggested one variant (sample-1707) with a triple-helix structure at >37 °C. Sample-1707 can assemble into nanofiber (width, 5.6 nm) and form hydrogel at 3 mg/mL. Sample-1707 was shown to induce blood clotting and promote osteoblast differentiation. Furthermore, sample-1707 exhibited high capacity to induce mouse hair follicle stem cells differentiation and osteoblast migration, demonstrating a high capacity to induce skin cell regeneration and promote wound healing. A strong hydrogel was prepared from a chitosan and sample-1707 complex with a swelling rate of >30 % higher than simply using chitosan. Fed-batch fermentation of sample-1707 with a 5-L bioreactor obtained a yield of 600 mg/L. These results support the large-scale production of sample-1707 as a biomaterial for use in the skin care industry.
RESUMO
Hair follicle stem cells (HFSCs) contribute to the regeneration of hair follicles (HFs), thus accelerating hair growth. microRNAs (miRs) are potential regulators in various cellular processes, including HFSC proliferation and differentiation. This study proposed a potential target, enhancer of zeste homolog 2 (EZH2) for facilitating hair growth, due to its function over HFSC activities by mediating the miR-22/serine/threonine kinase 40 (STK40)/myocyte enhancer factor 2 (MEF2)/alkaline phosphatase (ALP) axis. Gain- and loss-of-function approaches were adopted to explore the roles of EZH2, miR-22, and STK40 in the proliferation and apoptosis of HFSCs, along with the functional relevance of MEF2-ALP activity. STK40 was elevated during HFSC differentiation, which was found to facilitate HFSC proliferation, but impede their apoptosis by activating MEF2-ALP. Mechanically, miR-22 targeted and inversely regulated STK40, which inhibited MEF2-ALP activity to impede HFSC proliferation and differentiation. Moreover, EZH2 elevated the STK40 expression by repressing miR-22 to promote the proliferation and differentiation of HFSCs. Furthermore, in vivo experiments further validated the roles of EZH2 and STK40 on hair follicle neogenesis and hair growth. Collectively, EZH2 elevated the STK40 expression by downregulating miR-22, consequently accelerating differentiation of HFSCs and hair growth, which sheds light on the underlying molecular mechanism responsible for hair growth.