Neuronal regeneration material screening method by ex vivo model
View Patent ↗The present invention relates to a neuronal regeneration material screening method using ex vivo models, more precisely a neuronal regeneration material screening method using an organotypic spinal cord slice culture. The neuronal regeneration material screening method using spinal cord slices of living bodies of the present invention can be effectively applied for the development of a novel therapeutics for neurodegenerative diseases, such as spinal cord injury.
1. A method for screening for a neuronal regeneration material comprising the following steps:
1) culturing, on a filter, a single tissue slice obtained from the lumbar spinal cord of a mammal in a well, wherein the thickness of the tissue slice is 400 μm;
2) damaging the tissue slice in the course of culturing the single tissue slice of step 1) by treating with a toxic compound, which induces demyelination of axons in the spinal cord;
3) contacting neuronal regeneration material candidates to the damaged tissue slice; and
4) selecting a neuronal regeneration material candidate that induces significant directional neuronal regeneration, from the tissue slice to the neuronal regeneration material candidate, compared with the negative control untreated with the neuronal regeneration material candidates; wherein a spinal cord slice treated with mesenchymal stem cells is used as a positive control and the negative control group is a spinal cord slice untreated with the mesenchymal stem cells.
2. The method according to claim 1 , wherein the neuronal regeneration material candidate includes a compound, a growth factor, a cytokine, a mesenchymal stem cell-originated factor, an extracellular matrix protein, a protein involved in the axonal growth and a factor involved in intracellular signal transduction.
3. The method according to claim 1 , wherein the spinal cord tissue slice is live tissue cut by a tissue chopper.
4. The method according to claim 1 , wherein the toxic compound of step 2 is lysolecithin.
5. The method according to claim 1 , wherein the mammal is a rat.
6. The method according to claim 5 , wherein the spinal cord is isolated from a rat at 16 days old.