1H, 13C, and 15N resonance assignments of human glutathione peroxidase 4.

Glutathione peroxidase 4 (GPx4) behaves as an antioxidant enzyme capable of directly reducing peroxidized phospholipids within cell membranes. Recently, GPx4 has attracted attention as a target molecule for cancer therapy because it induces the immortalization of cancer cells suppressing ferroptosis. In this study, to analyze the function and structure of GPx4 by solution NMR, we performed resonance assignments of GPx4 and assigned almost all backbone 1H, 13C, and 15N resonances and most of the side chain 1H and 13C resonances. Using these assignments, the secondary structure of GPx4 was analyzed by the TALOS + program. GPx4 has six helices and seven strands. Then, the backbone dynamics were examined by the {1H}-15N heteronuclear NOE experiment. GPx4 was found to be rigid except for a short loop region. These results will provide basis for functional analysis and the first solution structure determination of GPx4.

Co-Culture of Osteoblasts and Endothelial Cells on a Microfiber Scaffold to Construct Bone-Like Tissue with Vascular Networks.

Bone is based on an elaborate system of mineralization and vascularization. In hard tissue engineering, diverse biomaterials compatible with osteogenesis and angiogenesis have been developed. In the present study, to examine the processes of osteogenesis and angiogenesis, osteoblast-like MG-63 cells were co-cultured with human umbilical vein endothelial cells (HUVECs) on a microfiber scaffold. The percentage of adherent cells on the scaffold was more than 60% compared to the culture plate, regardless of the cell type and culture conditions. Cell viability under both monoculture and co-culture conditions was constantly sustained. During the culture periods, the cells were spread along the fibers and extended pseudopodium-like structures on the microfibers three-dimensionally. Compared to the monoculture results, the alkaline phosphatase activity of the co-culture increased 3-6 fold, whereas the vascular endothelial cell growth factor secretion significantly decreased. Immunofluorescent staining of CD31 showed that HUVECs were well spread along the fibers and formed microcapillary-structures. These results suggest that the activation of HUVECs by co-culture with MG-63 could enhance osteoblastic differentiation in the microfiber scaffold, which mimics the microenvironment of the extracellular matrix. This approach can be effective for the construction of tissue-engineered bone with vascular networks.