Change in neuron aggregation and neurite fasciculation on EVAL membranes modified with different diamines.

In this study, we modified poly(ethylene-co-vinyl alcohol) (EVAL) membranes by the covalent bonding of diamines via epoxidation of surface hydroxyl groups of EVAL to analyze the effect of immobilized diamines with different carbon chain length on the cultured cerebellar granule neurons. Morphological studies showed that neurons seeded on the diamine-immobilized EVAL membrane were able to survive and regenerate with formation of an extensive neuritic network. Furthermore, cultured neurons showed that the presence of diamine with different carbon chain length was able to effectively regulate the neuron adhesion, migration, aggregation, and neurite growth pattern, but mediated neuronal activity with equal efficacy. The short-chain amine stimulated neuron migration, aggregation, and neurite fasciculation, whereas the long carbon chain diamine maintained single neuron distribution with the defasciculated feature of the neurite. Although it is known that positively charged amine molecules can interact directly with cell surface proteoglycans to mediate cell attachment, this study further demonstrated that the terminal primary amine with different carbon chain length is involved in mediating cell-substrate interaction to further regulate neuron aggregation and neurite fasciculation. This indicates a delicate interaction of neuron with the immobilized diamine molecules on the EVAL membrane surface. This work is encouraging because the diamine- immobilized EVAL membranes can be applied for the establishment of different neural culture systems useful for future investigations of neuron biology under in vitro conditions.

Immobilization of L-lysine on microporous PVDF membranes for neuron culture.

Microporous poly(vinylidene fluoride) (PVDF) membranes with dense or porous surface were prepared by immersion precipitation of PVDF/TEP solutions in coagulation baths containing different amounts of water. Onto the membrane surface, poly(glycidyl methacrylate) (PGMA) was grafted by plasma-induced free radical polymerization. Then, L-lysine was covalently bonded to the as-grafted PGMA through ring-opening reactions between epoxide and amine to form amino alcohol. The highest attainable graft density of PGMA on a PVDF membrane was 0.293 mg/cm2. This was obtained when the reaction was carried out on a porous surface under an optimized reaction condition. For immobilization of L-lysine, the yield was found to depend on the reaction temperature and L-lysine concentration. The maximal yield was 0.226 mg/cm2, a value considerably higher than reported in the literature using other immobilization methods. Furthermore, neurons were cultured on L-lysine-immobilized PVDF membranes. The results indicated that these membrane surfaces were suited to the growth of neurons, with a MTT value higher than that of the standard culture dish.