Nitrogen-doped carbon nanocages and human umbilical cord mesenchymal stem cells cooperatively inhibit neuroinflammation and protect against ischemic stroke

Aims: This study aimed to explore the synergistic effects of nitrogen-doped carbon nanocages (NCNCs) and human umbilical cord mesenchymal stem cells (HUC-MSCs) on ischemic stroke and investigate the potential underlying mechanisms. Main methods: The properties of NCNCs were analyzed by transmission electron microscopy, and the markers of HUC-MSCs were detected by flow cytometry. The cell toxicity of NCNCs was evaluated by MTT. Mice were induced cerebral infarction by transient middle cerebral artery occlusion (MCAO). NCNCs or HUC-MSCs or HUC-MSCs-NCNCs were intravenously injected thirty minutes after reperfusion. The infarct volume was examined by 2,3,5-triphenyltetrazolium chloride (TTC) staining, and behavior tests were evaluated by the modified Neurological Severity Score (mNSS) and rotarod test. The mRNA levels of TNF-alpha and IL-10 were detected by real-time PCR. The protein levels of TNF-alpha stimulated gene/protein 6 (TSG-6) and prostaglandin 2 (PGE2) were detected by ELISA. The microglia markers (CD86 and CD206) and the protein levels of TNF-alpha and IL-10 were examined by flow cytometry. The protein levels of Iba1 and CD16 were determined by immunostaining. Key findings: NCNCs enhanced the therapeutic effects of HUC-MSCs on MCAO mice, including reducing infarct volume, improving behavior scores and inhibiting inflammation response. In addition, NCNCs and HUC-MSCs cooperatively inhibit the mRNA and protein levels of TNF-alpha, and increased the mRNA and protein levels of IL-10 and protein levels of PGE2 and TSG-6 in LPS-treated microglia. Furthermore, NCNCs exerted synergistic effects with HUC-MSCs on remodeling microglia polarization. Significance: NCNCs enhance the therapeutic effects of HUC-MSCs on cerebral infarction in a mouse MCAO model, and inhibit the microglia reactivation and neuroinflammation, which indicates it as a potential treatment for ischemic stroke.