Stable system in vitro culture of cerebellar granule cell precursors
In vitro culture system of cerebellar granule precursor cells (granule cell progenitors, GCP) simple, reliable, easily reproducible and stable both time and with respect to the gene / biochemical expression profile of the obtained GCP cell cultures, similar to that of cultures of fresh and pure GCPs cells with respect to the possibility of freezing and thawing of the same.
Valid in vitro model for the study of the pathophysiology of cerebellar granules, for the study of cerebellar diseases resulting from damage or neurodegeneration, and potentially for their treatment by means of gene therapy approaches. After surgical resection from mice on the 7th day of postnatal life, the cerebellum is mechanically triturated in a buffer containing HBSS, glucose and antibiotics, and the single cells thus obtained plated in a medium composed of DMEM-F12, glucose, B27 W / O VitA, insulin, NALC, heparin, antibiotics and growth factors. In this medium, the cells can be expanded and maintained for an unlimited time, after periodic dissociation of the spheres and propagation by dilution (to prevent excessive growth and consequent death from lack of nutrients and hypoxia). According to known culture models, cultured GCPs either stop proliferating rapidly, exposing the need to generate new cultures each time, or are of tumor origin and therefore cannot correctly represent the physiology of GCPs.
- Generation of in vitro models for the study of the pathophysiology of cerebellar granules;
- Evaluation of the toxic effects of new drugs on ‘normal’ GCPs compared to cancer in vitro before starting in vivo studies;
- Treatment of cerebellar diseases due to damage or neurodegeneration, using gene therapy approaches.
- GCP cultures in a state of unlimited proliferation and with a gene / biochemical expression profile comparable to that of fresh GCP cultures, even after thawing;
- Preservation of the physiological characteristics of GCPs;
- Stability over time;