Supramolecular Hybrid Hydrogel Delivers Exosomes Derived From IL-1β Stimulated BMSCs To Attenuate Neuroinflammation In Cerebral Ischemia-Reperfusion Injury

Background:Long-term neuroinflammation is a major obstacle to neurological recovery after cerebral ischemia/reperfusion(I/R)injury.Preliminary studies of our group have demonstrated that exosomes derived from bone marrow mesenchymal stem cells(BMSC-Exos)can effectively inhibit the dysfunction after cerebral I/R injury.This study intends to use IL-1β to simulate the inflammatory environment,which improve the production and anti-inflammatory ability of exosomes secreted by bone marrow mesenchymal stem cells(IL-1β-stimulated-BMSC-Exos,βExos).A thermosensitive supramolecular injectable hybrid hydrogel(HDU/SF hydrogel)has been devloped to sustainably release exosomes,which can solve the problems of poor retention and short half-life of exosomes at the injury site.Object:To study IL-1β-induced BMSC to increase exosome production and anti-inflammatory ability;to verify that HDU/SF hydrogel by sustained release ofβExos inhibits the inflammatory response and improves neurological dysfunction after cerebral I/R injury.Methods:1.In this study,exosomes were extracted by ultracentrifugation;the physical characterization of HDU/SF hydrogels was tested by rheometer and scanning electron microscope;2.Cell live/dead staining,CCK-8,HE staining and immunofluorescence were used to verify the cytocompatibility and histocompatibility of HDU/SF hydrogels;the sustained release effect of DIR-labeled exosomes in HDU/SF hydrogels was observed by in IVIS;3.Nanoflow and RT-qPCR were used to verify the production and anti-inflammatory effect of IL-1 β-induced Exos;4.Immunofluorescence and Nissl staining were used to detect the therapeutic effect of HDU/SF hydrogel sustained-release βExos on the reactive proliferation of astrocytes and microglia and neuron loss after brain I/R injury.Results:1.BMSCs and βExos were successfully extracted;HDU/SF has injectability,stability and self-healing properties,and can quickly gel at 37℃;2.HDU/SF hydrogel has good cytocompatibility and histocompatibility,and enhance the sustained release effect of BMSC-Exos in vivo;3.IL-1β can increase the production of BMSC-Exos;βExos can inhibit LPS-induced BV2 cells to secrete pro-inflammatory factors(IL-1β and TNF-α)and M1 microglia(CD32 and CD68)mRNA expression;4.HDU/SF hydrogel inhibits the reactive proliferation of astrocytes and microglia and the loss of neurons in the ischemic penumbra after cerebral I/R injury by sustained release of βExos.Conclusion:IL-1β-induced BMSCs can increase the production of exosomes and anti-inflammatory ability;HDU/SF hydrogel can inhibit the neuroinflammation and neuron loss after cerebral I/R injury by sustained release of βExos.