A Protein-Permeable Scaffold of a Collagen Vitrigel Membrane Useful for Reconstructing Crosstalk Models between Two Different Cell Types

Abstract

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.

Predictive performance of the Vitrigel‐eye irritancy test method using 118 chemicals

“…In addition, it possesses excellent transparency and permeability of protein with high molecular weight and consequently the various studies utilizing it as a cell culture scaffold advances so well (2004, 2007a, 2007b, 2007c). We established a preparation method of a corneal epithelium model utilizing an air–liquid interface culture system that facilitates induction of layering rabbit corneal epithelial cells cultured on the CVM scaffold (Takezawa et al ., 2008).…”

Collagen vitrigel scaffold for regenerative medicine of the trachea: Experimental study and quantitative evaluation

“…However, from the results of clinical applications, as at least 2 months are required for sufficient epithelialization on the luminal side of the scaffold, further promotion of epithelialization is important for the prevention of infection in the early stages after implantation. Takezawa et al [13,14] improved the properties of collagen gel by utilizing vitrification technology. A collagen vitrigel membrane showing excellent mechanical strength, transparency, and protein permeability was successfully prepared by vitrifying a disk of conventional collagen gel and subsequently rehydrating the vitrified material.…”

“…Fabrication and surface evaluation of the collagen vitrigel scaffold Vitrigel was initially developed by Takezawa [13,14] through the application of vitrification technology. The advantages are that the collagen vitrigel membrane is smooth in appearance and is about 20 times stronger than conventional collagen gel.…”

Effects of a cell‐free method using collagen vitrigel incorporating TGF‐β1 on articular cartilage repair in a rabbit osteochondral defect model

“…Takezawa et al applied this vitrification process to conventional collagen gel and extended this work by rehydrating the vitrified collagen gel to produce a high‐density gel comprising collagen fibrils—a novel biomaterial named “collagen vitrigel.” Using this technology, the disk‐shaped conventional gel of 0.25% (w/v) collagen fibrils can be concentrated up to 100–133 times and converted into a round collagen vitrigel membrane of 25–33% (w/v) collagen fibrils, which is equivalent to the composition of connective tissue found in the body . Relative to conventional collagen gels, this collagen vitrigel membrane offers superior handling, strength, transparency, and permeability to proteins with a molecular weight of 100 kDa or higher . The collagen vitrigel membrane comprises mainly collagen type I and has been reported to be an effective carrier for paracrine assays in three‐dimensional in vitro culture and in vivo drug‐delivery systems (DDSs) …”