A protein-permeable scaffold of a collagen vitrigel membrane useful for reconstructing crosstalk models between two different cell types.
Soft and turbid collagen gel disks were previously converted into strong and transparent gel membranes utilizing a concept for the vitrification of heat-denatured of proteins. This novel stable and transparent gel has been termed 'vitrigel'. By encompassing the collagen vitrigel membrane in a nylon frame, it can be easily handled with tweezers, and functions as an excellent scaffold for three-dimensional cell culture models, as cells can be cultured on both sides. Here, we investigated the molecular permeability of the collagen vitrigel membrane in a time course-dependent manner using glucose and serum proteins. Glucose penetrated through the collagen vitrigel membrane to the opposite side, and concentrations on each side were found to be equilibrated within 24 h. Serum proteins up to a molecular weight >100 kDa also gradually passed through the collagen vitrigel membrane. In addition, human microvascular endothelial cells (HMVECs) were cultured on one surface of the collagen vitrigel membrane with a nylon frame, and human dermal fibroblasts (HDFs) or HT-29 (a human colon carcinoma cell line) cells were cocultured on the opposite surface. Histomorphological observations revealed the formation of three-dimensional crosstalk models composed of HMVECs and HDFs or HMVECs and HT-29 cells. Resulting data suggest that the protein-permeable scaffold composed of the collagen vitrigel membrane is useful for the reconstruction and/or modeling of 'crosstalk' between two different cells types. Hereafter, such crosstalk models in vitro could be applied to research not only of paracrine factors, but also to epithelial- or endothelial-mesenchymal transitions.
Takezawa
Toshiaki
T
Laboratory of Animal Cell Biology, National Institute of Agrobiological Sciences, Tsukuba, Ibaraki, Japan. t.takezawa@affrc.go.jp
Nitani
Aya
A
Shimo-Oka
Tadashi
T
Takayama
Yoshiharu
Y
D001798:Blood Proteins / Q000737:chemistry / Q000378:metabolism*; D002452:Cell Count; D018929:Cell Culture Techniques; D002463:Cell Membrane Permeability; D002478:Cells, Cultured; D018920:Coculture Techniques; D024042:Collagen Type I / Q000737:chemistry*; D003470:Culture Media; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology; D004847:Epithelial Cells / Q000166:cytology; D005240:Feasibility Studies; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D005452:Fluoresceins; D005456:Fluorescent Dyes; D005782:Gels; D019073:HT29 Cells; D006801:Humans; D008567:Membranes, Artificial; D008648:Mesoderm / Q000166:cytology; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D008970:Molecular Weight; D012867:Skin / Q000166:cytology; D013997:Time Factors; D014471:Umbilical Veins / Q000166:cytology
D001798:Blood Proteins; D024042:Collagen Type I; D003470:Culture Media; D005452:Fluoresceins; D005456:Fluorescent Dyes; D005782:Gels; D008567:Membranes, Artificial; C007740:fluorexon