Regulatory T cells ameliorate intracerebral hemorrhage-induced inflammatory injury by modulating microglia/macrophage polarization through the IL-10/GSK3ß/PTEN axis.

Inflammation mediated by the peripheral infiltration of inflammatory cells plays an important role in intracerebral hemorrhage (ICH) induced secondary injury. Previous studies have indicated that regulatory T lymphocytes (Tregs) might reduce ICH-induced inflammation, but the precise mechanisms that contribute to ICH-induced inflammatory injury remain unclear. Our results show that the number of Tregs in the brain increases after ICH. Inducing Tregs deletion using a CD25 antibody or Foxp3DTR-mice increased neurological deficient scores (NDS), the level of inflammatory factors, hematoma volumes, and neuronal degeneration. Meanwhile, boosting Tregs using a CD28 super-agonist antibody reduced the inflammatory injury. Furthermore, Tregs depletion shifted microglia/macrophage polarization toward the M1 phenotype while boosting Tregs shifted this transition toward the M2 phenotype. In vitro, a transwell co-culture model of microglia and Tregs indicated that Tregs changed the polarization of microglia, decreased the expression of MHC-II, IL-6, and TNF-α and increased CD206 expression. IL-10 originating from Tregs mediated the microglia polarization by increasing the expression of Glycogen Synthase Kinase 3 beta (GSK3ß), which phosphorylates and inactivates Phosphatase and Tensin homologue (PTEN) in microglia, TGF-ß did not participate in this conversion. Thus, Tregs ameliorated ICH-induced inflammatory injury by modulating microglia/macrophage polarization toward the M2 phenotype through the IL-10/GSK3ß/PTEN axis.

Interleukin-23 Secreted by Activated Macrophages Drives γδT Cell Production of Interleukin-17 to Aggravate Secondary Injury After Intracerebral Hemorrhage.

BACKGROUND: Neuroinflammation plays a key role in intracerebral hemorrhage (ICH)-induced secondary brain injury, but the specific roles of peripheral inflammatory cells such as macrophages and lymphocytes remain unknown. The purpose of this study was to explore the roles of macrophages, T lymphocytes, and the cytokines they secrete as potential targets for treating secondary brain injury after ICH. METHODS AND RESULTS: Our results showed that peripheral macrophages and T lymphocytes successively infiltrated the brain, with macrophage counts peaking 1 day after ICH and T-lymphocyte counts peaking after 4 days. These peaks in cellular infiltration corresponded to increases in interleukin (IL)-23 and IL-17 expression, respectively. We found that hemoglobin from the hematoma activated IL-23 secretion by infiltrating macrophages by inducing the formation of toll-like receptor (TLR) 2/4 heterodimer. This increased IL-23 expression stimulated γδT-cell production of IL-17, which increased brain edema and neurologic deficits in the model mice as a proinflammatory factor. Finally, we found that sparstolonin B (SsnB) could ameliorate brain edema and neurologic deficits in ICH model mice via inhibition of TLR2/TLR4 heterodimer formation, and notably, SsnB interacted with myeloid differentiation factor 88 Arg196. CONCLUSIONS: Together, our results reveal the importance of the IL-23/IL-17 inflammatory axis in secondary brain injury after ICH and thus provide a new therapeutic target for ICH treatment.

Toll-Like Receptor 4/MyD88-Mediated Signaling of Hepcidin Expression Causing Brain Iron Accumulation, Oxidative Injury, and Cognitive Impairment After Intracerebral Hemorrhage.

BACKGROUND: Disturbance of brain iron metabolism after intracerebral hemorrhage (ICH) results in oxidative brain injury and cognition impairment. Hepcidin plays an important role in regulating iron metabolism, and we have reported that serum hepcidin is positively correlated with poor outcomes in patients with ICH. However, the roles of hepcidin in brain iron metabolism after ICH remain largely unknown. METHODS: Parabiosis and ICH models combined with in vivo and in vitro experiments were used to investigate the roles of hepcidin in brain iron metabolism after ICH. RESULTS: Increased hepcidin-25 was found in serum and primarily in astrocytes after ICH. The brain iron efflux, oxidative brain injury, and cognition impairment were improved in Hepc-/- ICH mice but aggravated by the human hepcidin-25 peptide in C57BL/6 ICH mice. Data obtained in in vitro studies showed that increased hepcidin inhibited the intracellular iron efflux of brain microvascular endothelial cells but was rescued by a hepcidin antagonist, fursultiamine. Using parabiosis ICH models also shows that increased serum hepcidin prevents brain iron efflux. In addition, Toll-like receptor 4 (TLR4)/MyD88 signaling pathway increased hepcidin expression by promoting interleukin-6 expression and signal transducer and activator of transcription 3 phosphorylation. TLR4-/- and MyD88-/- mice exhibited improvement in brain iron efflux at 7, 14, and 28 days after ICH, and the TLR4 antagonist (6R)-6-[N-(2-chloro-4-fluorophenyl) sulfamoyl] cyclohex-1-ene-1-carboxylate significantly decreased brain iron levels at days 14 and 28 after ICH and improved cognition impairment at day 28. CONCLUSIONS: The results presented here show that increased hepcidin expression caused by inflammation prevents brain iron efflux via inhibition of the intracellular iron efflux of brain microvascular endothelial cells entering into circulation and aggravating oxidative brain injury and cognition impairment, which identifies a mechanistic target for muting inflammation to promote brain iron efflux and to attenuate oxidative brain injury after ICH.

Safety and Efficacy of Wingspan Stenting for Severe Symptomatic Atherosclerotic Stenosis of the Middle Cerebral Artery: Analysis of 278 Continuous Cases.

OBJECTIVE: Our objective is to investigate the safety and long-term efficacy of the Wingspan stent (Boston Scientific, Natick, MA, USA) for treating severe atherosclerotic stenosis of the middle cerebral artery (MCA). METHODS: A total of 278 consecutive patients from our stroke database with clinical symptoms within the prior 90 days and intracranial atherosclerotic stenosis of 70% or above of the MCA were enrolled in this study between September 2012 and November 2014, and these patients were followed until the end of June 2015. The endpoint events included any stroke or death within 30 days after stenting and any subsequent ipsilateral ischemic stroke. RESULTS: Among the 278 enrolled patients, 277 patients (99.6%) successfully underwent stenting. The mean rate of stenosis decreased from 82.5 ± 7.9% to 9.0 ± 3.2% following treatment. Within 30 days after stenting, 12 patients (4.3%) experienced endpoint events, including 8 cases (2.9%) of hemorrhagic stroke and 4 cases (1.4%) of ischemic stroke; 2 perioperative deaths occurred. During 8-33 months of follow-up, 19 patients developed endpoint events. The 1- and 2-year endpoint event rates were 5.8% (95% confidence interval [CI], 5.0%-15.7%) and 7.2% (95% CI, 4.3%-10.1%), respectively. CONCLUSIONS: From this study, we can conclude that the treatment of severe symptomatic atherosclerotic stenosis of the MCA using the Wingspan stent was safe and effective and that the long-term stroke recurrence rate after stenting was low.

Association of CYP2C19 Polymorphisms with the Clinical Efficacy of Clopidogrel Therapy in Patients Undergoing Carotid Artery Stenting in Asia.

The CYP2C19 gene plays a detrimental role in the metabolism of clopidogrel. This study aimed to investigate the association between CYP2C19 polymorphisms and the clinical efficacy of clopidogrel therapy in patients who have undergone carotid artery stenting (CAS). CYP2C19 genotype screening was performed on 959 ischemic stroke patients. Of these patients, 241 who had undergone CAS were enrolled in the study. They were all followed up for 1 year after stent surgery, and the primary clinical end-points were ischemic events. The frequencies of the CYP2C19*2 and *3 alleles among the 959 patients were 31.80% and 5.06%, respectively. Regarding the 241 participants who had undergone CAS, multivariate Cox regression analysis showed that the CYP2C19 loss-of-function (LOF) alleles (*2 and *3) were risk factors for post-CAS prognosis. Within 1 year of follow-up, the patients carrying the CYP2C19 LOF alleles were more likely to experience ischemic events than those carrying none. The occurrence of ischemic events did not significantly differ between the *2 and *3 allele carriers. Our results suggest that CYP2C19 LOF alleles (*2 and *3) significantly impact the prognosis of patients on clopidogrel therapy after CAS and that the CYP2C19*2 and CYP2C19*3 alleles have the same effects on prognosis.

Serum hepcidin concentrations correlate with serum iron level and outcome in patients with intracerebral hemorrhage.

Iron plays a detrimental role in the intracerebral hemorrhage (ICH)-induced brain damage, while hepcidin is the most important iron-regulated hormone. Here, we investigate the association between serum hepcidin and serum iron, outcome in patients with ICH. Serum samples of 81 cases with ICH were obtained on consecutive days to detect the levels of hepcidin, iron, interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α). The National Institutes of Health Stroke Scale score (NIHSS) was measured at admission and on days 7 and 30, and the modified Rankin Scale (mRS) score was evaluated at 3 months after ICH. Additionally, the correlations of serum hepcidin with serum iron and the mRS score were analyzed by a generalized linear model. Higher serum hepcidin levels were detected in patients with poor outcomes (P < 0.001), and the mRS score increased by a mean of 1.135 points (95% CI 1.021-1.247, P < 0.001) for every serum hepcidin quartile after adjusting for other prognostic variables. Pearson correlation analysis showed that serum hepcidin was negatively correlated with serum iron (r = -0.5301, P < 0.001), and a significantly lower concentration of serum iron was found in patients with poor outcomes (P = 0.007). Additionally, serum hepcidin was independently correlated with mRS scores of ICH patients (OR 1.115, 95% CI 0.995-1.249, P = 0.021). Our results suggest that serum hepcidin is closely related to the outcome of patients with ICH and may be a biological marker for outcome prediction.