Local Delivery Of Flavopiridol Repairs Spinal Cord Injury By Regulation Of Astrocytes And Inflammation

Spinal cord injury(SCI)is one of the most devastating traumas.Current treatments receive very limited neurological recovery.SCI brings a heavy burden for both the family and society of each patient.It is very important to discover a solution for SCI therapy.SCI results in severe and extensive local inflammation.The repair of SCI is closely related to inflammatory cytokines,among which quite a few have been demonstrated detrimental or beneficial to SCI repair.The sequential changes of local inflammatory cytokine protein levels after rat SCI are still not clear.Therefore in chapter one,we studied the sequential changes of inflammatory cytokines in the local microenvironment after rat SCI.We found that 7 inflammatory cytokines that might be beneficial to SCI repair had differential changes after SCI,while 11 inflammatory cytokines that might be detrimental to repair also had differential changes after SCI.Importantly,4 cytokines that might be beneficial to repair decreased after injury,and 9 cytokines that might be detrimental to repair increased.This study provided basic biological data for drug screening in the pre-clinical studies of SCI therapy,as well as the clinical therapy and prognosis.In many experimental models of diseases including SCI,it has been shown that pathological activation of the cell-cycle not only causes post-mitotic cell death(neurons and mature oligodendrocytes),but also initiates mitotic cell proliferation and activation(microglia/macrophages,astrocytes,and precursor cells).Flavopiridol,a nonselective CDK inhibitor,has been reported to significantly improve motor recovery,reduce lesion volume,decrease astrocytic reactivity,and attenuate neuronal and oligodendroglial apoptosis after SCI.Astrocytes are the key cell type in neuroinflammation.It is also the important source of inflammatory cytokines.So in chapter two,we asked whether flavopiridol could regulate the synthesis of inflammatory cytokines in astrocytes.We found that flavopiridol inhibited proliferation,scratch-wound healing,and inflammatory factor synthesis in astrocytes,while permitting the survival of neurons.Hence,flavopiridol has differential effects on astrocytes and neurons,indicating its potential promotion on SCI repair.Flavopiridol has been reported to significantly improve motor recovery after SCI,when administered systemically by daily intraperitoneal injection or centrally by continuous intrathecal delivery.However,since flavopiridol is a broad-spectrum inhibitor,its high systemic dose may cause strong side-effects.The mini-osmotic pump used for intrathecal delivery is costly and may cause problems with histocompatibility,as the pump is foreign matter in corpore.Therefore,it is necessary to develop a more efficient and economical method for the sustained local delivery of flavopiridol without side-effects.Biodegradable and injectable Poly(lactic-co-glycolic acid)(PLGA)-based methylprednisolone nanoparticles have been used in the treatment of rat SCI,and the nanoparticle-enabled local delivery is significantly more effective than systemic delivery.So chapter three aimed to develop a strategy for the local delivery of flavopiridol and to evaluate this strategy.The results showed that flavopiridol Nanoparticles had a sustained release of up to 3 days.Flavopiridol Nanoparticles significantly decreased the pro-inflammatory factor synthesis by astrocytes,while the IL-10 expression was elevated.In vivo study demonstrated that flavopiridol nanoparticles decreased the cell-cycle related protein expressions of astrocytes,neurons and macrophages.They also increased the integrity of spinal cord gross tissue structure,inhibited the glial scarring and cavitation,and facilitated neuronal survival and regeneration.In the epicenter,flavopiridol nanoparticles increased the protein levels of GM-CSF,while decreased that of IP-10.Administration of flavopiridol nanoparticles eventually improved the functional recovery of injured rats.These findings demonstrated that local delivery of flavopiridol in PLGA nanoparticles improves recovery from SCI by regulation of astrocytes and inflammation.