| Nerve regeneration is still remain many questions to uncover. It is a focus. However, neurons are in highly differentiated state, its regeneration is more difficult than the other cells, so ideal repair treatments of nervous system are needed to be found. Microenvironment is considered to play an important role in nervous system repair. Moreover, interaction between cells play an important part in nervous system repair, but the mechanism remain unclear. Co-culture of neurons and astrocytes in vitro is an indispensable method to explore . It can provide an excellent model for nervous system injury research. With the development of biotechnology, biological tissue engineering have begun to use in the treatment of nerve injury, including use of biocompatible materials to support the regeneration of CNS.Recent evidences suggest that astrocytes have important neuroregulatory functions in addition to their classic functions of support and segregation of neurons. These newly revealed functions including regulation of neuron communication, neurosecretion, and synaptic plasticity. In this study, we first successfully cultured astrocytes, neurons in vitro and then co-cultured the astrocytes and neurons in vitro to study glials function of support the growth of neurons. The results showed that the total number of branches and number of primary dendrites of co-culture group was more than the group cultured alone. It indicated that astrocytes could stimulate neurons in neurite outgrowth. In addition, neurons in co-culture group, exhibited mature morphlogical features, could form the neural network.Silk fibrion possesses good biocompatibility. It could be used to form a variety of biomaterials and could be obtained easily, in addition, they possess structures modified with immobilized RGD prptide to increase cells attachment, so they provide promising new therapeutic approaches for many medical researches.So far, studies have revealed that the astrocytes on the silk showed very high compatibility and they also had high cell adhesion, proliferation and migration capacities.Moreover, neurons cultured with glial cells could improve the nutrition outgrowth,Therefore, following experiment aimed to find the effect of the silk fibroin and astrocytes on the growth and development of neurons. The results showed that the combination of non-woven, SF/TSF Silk Fibroin and astrocytes supported neuron growth, and they had good biocompatibility. It was observed that the neurons co-cultured with the astrocytes in the SF fibers had a higher complexity than those in the TSF fibers. It was reported previously that the astrocytes in different phases had different effects on the development of neurons,astrocytes in the embryonic stage promoted the development of neurons,while, astrocytes in the adult stage inhibited the development of neurons. Considering glial cells in different stages have different roles on the development of neurons, we co-cultured the glial cells from a model of brain injury and the neurons coming from SVZ to study the effect of astrocytes in different stages on the development of neurons.The result suggested that in the early period of co-culture, the glial cells promoted the development of the neurons, this trend of promotion, however, would decreased with the time of co-culturing prolong.On one hand, regeneration of axon is prevented by glial scar formed after nerve injury, on the other hand, astrocytes have important neuroregulatory functions. This is one of the main factors causing the failure of nerve regeneration. So studies are needed to understand how to convert the function of glial scar to protect neural development. Our results show that glial cells inoculated on non-woven silk fibroin material can promote the development of neurons. Fibroin nanofiber material including tussah silk nanofibers can be used as a biological scaffold material for neuron growth, having a optimistic future in their practical application to nerve injury.
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