Synergistic effects of repeated transcranial magnetic stimulation and mesenchymal stem cells transplantation on alleviating neuroinflammation and PANoptosis in cerebral ischemia.

Abstract

Background: Neuronal death is the primary cause of poor outcomes in cerebral ischemia. The inflammatory infiltration in the early phase of ischemic stroke plays a vital role in triggering neuronal death. Either transplantation of mesenchymal stem cells (MSCs) derived from humans or repetitive transcranial magnetic stimulation (rTMS) have respectively proved to be neuroprotective and anti-inflammatory in cerebral ischemia. However, either treatment above has its limitations. Whether these two therapies have synergistic effects on improving neurological function and the underlying mechanisms remains unclear. This investigation aims to elucidate the synergistic effects and underlying mechanisms of MSCs combined with rTMS treatment on the neurological function recovery post-ischemia.

Methods: A Sprague-Dawley rat model of cerebral infarction was induced via transient middle cerebral artery occlusion (tMCAO). The rats were divided into five groups (n = 50): sham, tMCAO, rTMS, MSCs, and MSCs + rTMS groups. Transplantation of human umbilical cord MSCs and rTMS intervention were performed 24 h post-stroke. Neurological function was further assessed via several behavioral tests and the 2,3,5-triphenyltetrazolium chloride (TTC) staining companied with Nissl staining were used to assess neuronal survival. TUNEL staining, western blotting, immunofluorescence, immunohistochemistry, ELISA, and flow cytometry were employed to measure the levels of neuroinflammation and PANoptosis. The molecular mechanisms underlying the special role of rTMS in the combined therapy were distinguished with transcriptome sequencing via PC12 cells in oxygen-glucose deprivation/reoxygenation (OGD/R) conditions.

Results: The combined therapy efficiently reduced lesion volume and improved neuronal survival (P < 0.05), subsequently improving functional recovery after ischemic stroke. MSCs + rTMS treatment ameliorated the PANoptosis in neurons (P < 0.05), accompanied by decreased levels of inflammatory factors in the cerebral tissue and serum during the subacute phase of cerebral infarction. To further explore the roles of either therapy on synergistic effect, we found that the transplanted MSCs primarily localized in the spleen and reduced cerebral inflammatory infiltration after ischemia via suppressed splenic inflammation. Meanwhile, rTMS significantly protects neurons from PANoptosis in MSCs-inhibited inflammatory conditions by downregulating REST unveiled by transcriptome sequencing.

Conclusions: Our study elucidates an unidentified mechanism by which the combination of MSCs and rTMS could synergistically promote neuronal survival and suppress neuroinflammation during the subacute phase of cerebral infarction, thus improving neurological outcomes. The downregulating REST induced by rTMS may potentially contribute to the neuroprotective effect against PANoptosis in MSCs-inhibited inflammatory conditions. These results are expected to provide novel insights into the mechanisms of MSCs and rTMS combination therapy in synergistically protecting against cerebral ischemia injury and potential targets underlying neuronal PANoptosis in the early phase of stroke.