Immobilization of type-I collagen and basic fibroblast growth factor (bFGF) onto poly (HEMA-co-MMA) hydrogel surface and its cytotoxicity study.

Type-I collagen and bFGF were immobilized onto the surface of poly (HEMA-co-MMA) hydrogel by grafting and coating methods to improve its cytotoxicity. The multi-layered structure of the biocompatible layer was confirmed by FTIR, AFM and static water contact angles. The layers were stable in body-like environment (pH 7.4). Human skin fibroblast cells (HSFC) were seeded onto Col/bFGF-poly (HEMA-co-MMA), Col-poly (HEMA-co-MMA) and poly (HEMA-co-MMA) films for 1, 3 and 5 day. MTT assay was performed to evaluate the extraction toxicity of the materials. Results showed that the cell attachment, proliferation and differentiation on Col/bFGF-poly (HEMA-co-MMA) film were higher than those of the control group, which indicated the improvement of cell-material interaction. The extraction toxicity of the modified materials was also lower than that of the unmodified group. The protein and bFGF immobilized poly (HEMA-co-MMA) hydrogel might hold great promise to be a biocompatible material.

The morphological and biomechanical changes of keratocytes cultured on modified p (HEMA-MMA) hydrogel studied by AFM.

The poor integration with host cornea tissue and the low mechanical properties of pHEMA hydrogel for artificial cornea remains a difficult problem to solve. A modified pHEMA hydrogel, MMA copolymerized and type-I collagen and bFGF immobilized, was previously prepared in an attempt to solve the problems. In this study, the cytotoxicity of Col/bFGF-p (HEMA-MMA) and p (HEMA-MMA) was studied by cell adhesion assay and atomic force microscopy (AFM). The results of cell adhesion assay show that the attachment of keratocytes on the modified membrane is much higher than that of the unmodified membrane. This indicates that the material after modification have better cell-material interaction. The AFM images reveal that the morphology of keratocytes cultured on different substrate is obviously different. The cell cultured on modified membrane presented a completely elongated and spindle-shape morphology. The force-distance indicates that the biomechanical of keratocytes changes significantly after culturing on different substrates. The adhesion force (2328+/-523 pN) and Young's modulus (0.51+/-0.125 kPa) of the cell cultured on modified membrane are much higher, and the stiffness (0.08+/-0.022 mN/m) is lower than those of the cell cultured on unmodified membrane. These results show that the cytotoxicity of Col/bFGF-p (HEMA-MMA) for keratocytes is much improved.

[Implantation of modified polyhydroxyethyl methacrylate-polymethyl methacrylate keratoprostheses in rabbit and monkey corneas].

OBJECTIVE: To investigate the biocolonization of polyhydroxyethyl methacrylate (PHEMA) sponge with cornea tissue and evaluate the therapeutic effects of modified porous PHEMA-PMMA (polymethyl methacrylate) Keratoprostheses (KPro) on rabbit and monkey corneas. METHODS: The KPro were made using two-stage polymerization combined with mechanical cutting. The experiment was divided into two groups. In the control group (A group), ten normal rabbit eyes received lamellar implantation of PHEMA sponges. The sponges were obtained 2 weeks, 1, 2, 3 and 4 months after operation. The cell proliferation and neovascularization inside the sponges were observed using light and transmission electron microscopy (TEM) and immunohistochemistry. In the experimental group (B group), the porous PHEMA-PMMA KPros were inserted into the lamellar pockets of ten rabbit corneas and two monkey corneas (stage I operation). The healing process was investigated by slit-lamp microscopy. The anterior lamellar cornea tissues were removed 3 months after surgery, exposing the underneath transparent core (stage II operation). The operated eyes were then followed up for 3 - 6 months. RESULTS: No complication was observed in A group. Under the light microscope, fibroblasts started to grow into the cornea 2 weeks after operation; lots of cells, accompanied with new blood vessels, invaded into the cornea 2 - 3 months after surgery. Invading cells of sponge, as well as keratocyte, were positive for vimentin. Under the electron microscope, the invading cells looked healthy and were surrounded by extracellular matrix and collagen. In B group, eight rabbit eyes which have received KPro implantation, anterior lamellar cornea melting happened in two eyes after the stage I operation. The remaining six corneas retained their central cores during observation after the stage II operation. Two monkey operated eyes were found no complication throughout the whole follow-up. CONCLUSIONS: The PHEMA sponge can obtain a tight fusion with the host cornea. The modified PHEMA-PMMA KPros have obtained a relatively stable therapeutic results after implantation into animal corneas.