A Reliable Nonhuman Primate Model of Ischemic Stroke with Reproducible Infarct Size and Long-term Sensorimotor Deficits.

A nonhuman primate model of ischemic stroke is considered as an ideal preclinical model to replicate various aspects of human stroke because of their similarity to humans in genetics, neuroanatomy, physiology, and immunology. However, it remains challenging to produce a reliable and reproducible stroke model in nonhuman primates due to high mortality and variable outcomes. Here, we developed a focal cerebral ischemic model induced by topical application of 50% ferric chloride (FeCl3) onto the MCA-M1 segment through a cranial window in the cynomolgus monkeys. We found that FeCl3 rapidly produced a stable intraarterial thrombus that caused complete occlusion of the MCA, leading to the quick decrease of the regional cerebral blood flow in 10 min. A typical cortical infarct was detected 24 hours by magnetic resonance imaging (MRI) and was stable at least for 1 month after surgery. The sensorimotor deficit assessed by nonhuman primate stroke scale was observed at 1 day and up to 3 months after ischemic stroke. No spontaneous revascularization or autolysis of thrombus was observed, and vital signs were not affected. All operated cynomolgus monkeys survived. Our data suggested that FeCl3-induced stroke in nonhuman primates was a replicable and reliable model that is necessary for the correct prediction of the relevance of experimental therapeutic approaches in human beings.

LDC7559 Exerts Neuroprotective Effects by Inhibiting GSDMD-Dependent Pyroptosis of Microglia in Mice with Traumatic Brain Injury.

Abstract Pyroptosis is considered one of a critical factor in the recovery of neurological function following traumatic brain injury. Brain injury activates a molecular signaling cascade associated with pyroptosis and inflammation, including NLRP3, inflammatory cytokines, caspase-1, gasdermin D (GSDMD), and other pyroptosis-related proteins. In this study, we explored the neuroprotective effects of LDC7559, a GSDMD inhibitor. Briefly, LDC7559, siRNA-GSDMD (si-GSDMD), or equal solvent was administrated to mice with a lipopolysaccharide + nigericin (LPS + Nig) model in vitro or with controlled cortical impact brain injury. The findings revealed that inflammation and pyroptosis levels were decreased by LDC7559 or si-GSDMD treatment both in vitro and in vivo. Immunofluorescence staining, brain water content, hematoxylin and eosin staining, and behavioral investigations suggested that LDC7559 or si-GSDMD inhibited microglial proliferation, ameliorated cerebral edema, reduced brain tissue loss, and promoted brain function recovery. Taken together, LDC7559 may inhibit pyroptosis and reduce inflammation by inhibiting GSDMD, thereby promoting the recovery of neurological function.

Conventional craniotomy versus conservative treatment in patients with minor spontaneous intracerebral hemorrhage in the basal ganglia.

BACKGROUND: The treatment for spontaneous intracerebral hemorrhage (ICH) is still controversial, especially for hematomas in the basal ganglia. A retrospective case-control study with propensity score matching was performed to compare the outcomes of conventional craniotomy and conservative treatment for patients with minor ICH in the basal ganglia. METHODS: We retrospectively collected the data of consecutive patients with minor basal ganglia hemorrhage from January 2018 to August 2019. We compared clinical outcomes of two groups using propensity score matching. The extended Glasgow outcome scale obtained by phone interviews based on questionnaires at a 12-month follow-up was used as the primary outcome measure. According to a previous prognosis algorithm, patients were divided into good and poor prognosis groups to obtain a dichotomized (favorable or unfavorable) outcome as the primary outcome. Secondary outcomes included hospitalized complications, mortality, and modified Rankin score at 12 months. RESULTS: A total of 54 patients were analyzed, and the baseline characteristics of patients in the surgery and conservative treatment groups were well matched. The primary favorable outcome at 12 months was significantly higher in the conservative treatment group than in the surgery group (81% vs 44%; OR 1.833, 95% CI 1.159-2.900; P=0.005). The incidence of pneumonia in the surgery group was significantly higher than that in the conservative treatment group (P=0.005). CONCLUSIONS: It is not recommended to undertake conventional craniotomy for patients with a minor hematoma (25-40 ml) in the basal ganglia. An open craniotomy might induce worse long-term functional outcomes than the conservative treatment.

Non-invasive Liver Fibrosis Scores Are Associated With Recurrence of Postoperative Chronic Subdural Hematoma.

Objective: Although liver diseases have already been identified as a risk factor for increased recurrence and mortality in patients with chronic subdural hematoma (CSDH), the association between subclinical liver disease, specifically liver fibrosis (LF), and CSDH remains unknown. In the present study, we aimed to investigate the association between the LF scores and CSDH recurrence. Methods: We retrospectively analyzed consecutive patients with CSDH who underwent burr-hole irrigation in the First Affiliated Hospital of Wenzhou Medical University between January 2015 and December 2018. The clinical data were collected, and the LF scores were calculated including aspartate aminotransferase-platelet ratio index (APRI), fibrosis-4 (FIB-4), and Forns index. Multivariable logistic regression analysis was applied to identify the association between the LF scores and CSDH recurrence, and Cox regression model and Fine-Gray competing risks model were performed to calculate hazard ratios (HRs) for CSDH recurrence based on time-to-event outcomes. The C-statistic, the integrated discrimination improvement (IDI), and the net reclassification improvement (NRI) evaluated the additive value of the LF scores to predict the recurrence of CSDH. Results: A total of 419 patients with CSDH were included, hematoma recurrence was observed in 62 patients (14.80%) within 1 year after surgery. The LF scores were significantly higher in those who recurred, whereas the standard hepatic assays were mostly normal. The patients were assigned to groups of high and low LF scores based on the validated cut-offs; compared with the subjects with low scores, those with high score levels had significantly higher recurrence rates. After adjusting for potential confounders, the LF scores were independently associated with CSDH recurrence, multivariable-adjusted HRs (95% CI) for those with higher levels of APRI, FIB-4, and Forns score were 4.32 (1.37-13.60), 2.56 (1.20-5.43), and 2.02 (1.07-3.79) for the recurrence of CSDH, respectively. Moreover, adding the APRI to the conventional model improved the C-statistic from 0.731 to 0.763, with an NRI and IDI of 7.50 and 1.35%, respectively. Two further commonly-used LF score indices (FIB-4 score and Forns index) yielded comparable results. Conclusions: The data from this study first indicated that the high LF scores were significantly associated with the recurrence of CSDH and that careful follow-up in these patients may be needed.

Shikonin regulates autophagy via the AMPK/mTOR pathway and reduces apoptosis of human umbilical cord mesenchymal stem cells to improve survival in tissues surrounding brain contusion.

Shikonin has been reported to regulate autophagy via the AMP-activated protein kinase (AMPK)/mTOR signalling pathway and decrease apoptosis in transplanted human umbilical cord mesenchymal stem cells (HUMSCs). In the present study, HUMSCs were exposed to oxygen glucose deprivation (OGD) in vitro for 12 h, and TUNEL fluorescence staining was used to detect apoptosis. Differences in autophagy and AMPK/mTOR pathway-related protein expression following treatment with shikonin were quantitatively analyzed by western blotting. Green fluorescent protein-labelled stem cells were implanted into traumatic brain injury-model mice and the survival of HUMSCs was observed after 7 days. Shikonin increased the number of cells in brain tissue surrounding the contusion 7 days after transplantation. Furthermore, shikonin treatment decreased apoptosis, increased the expression of autophagy-related proteins, increased phosphorylated AMPK expression and downregulated phosphorylated mTOR expression. In addition, the autophagy inhibitor 3-methyladenine attenuated these effects and aggravated apoptosis. Subsequently, shikonin upregulated autophagy and protected HUMSCs in the area surrounding contused brain tissue. Shikonin may regulate autophagy via the AMPK/mTOR signalling pathway and protect transplanted HUMSCs from apoptosis induced by hypoxia/ischemia.

Ablation of GSDMD Improves Outcome of Ischemic Stroke Through Blocking Canonical and Non-canonical Inflammasomes Dependent Pyroptosis in Microglia.

Ischemia/reperfusion (I/R) injury is a significant cause of mortality and long-term disability worldwide. Recent evidence has proved that pyroptosis, a novel cell death form, contributes to inflammation-induced neuron death and neurological function impairment following ischemic stroke. Gasdermin D (GSDMD) is a newly discovered key molecule of cell pyroptosis, but its biological function and precise role in ischemic stroke are still unclear. The present study investigates the cleavage activity of GSDMD, localization of pyroptotic cells, and global neuroinflammation in gsdmd -/- mice after I/R. The level of cell pyroptosis around the infarcted area was significantly increased in the acute phase of cerebral I/R injury. The ablation of GSDMD reduced the infraction volume and improved neurological function against cerebral I/R injury. Furthermore, we confirmed I/R injury induced cell pyroptosis mainly in microglia. Knockdown of GSDMD effectively inhibited the secretion of mature IL-1ß and IL-18 from microglia cells but did not affect the expression of caspase-1/11 in vitro and in vivo. In summary, blocking GSDMD expression might serve as a potential therapeutic strategy for ischemic stroke.

Elevated blood urea nitrogen is associated with recurrence of post-operative chronic subdural hematoma.

BACKGROUND: Chronic subdural hematoma (CSDH) is fundamentally treatable with about a 2-31% recurrence rate. Recently, there has been renewed interest in the association between Blood Urea Nitrogen (BUN) and intracranial lesion. Therefore, this paper attempts to show the relationship between BUN and CSDH recurrence. METHODS: A total of 653 CSDH cases with Burr-hole Irrigation (BHI) were enrolled from December 2014 to April 2019. The analyzed parameters included age, gender, comorbidities, laboratory investigations, medication use and hematoma location. The cases were divided into recurrence and non-recurrence groups while postoperative BUN concentration was further separated into quartiles (Q1 ≤ 4.0 mmol/L, 4.0 < Q2 ≤ 4.9 mmol/L, 4.9 < Q3 ≤ 6.4 mmol/L, Q4 > 6.4 mmol/L). Restricted cubic spline regressions and logistic regression models were performed to estimate the effect of BUN on CSDH recurrence. RESULTS: CSDH recurrence was observed in 96 (14.7%) cases. Significant distinctions were found between recurrence and non-recurrence groups in postoperative BUN quartiles of cases (P = 0.003). After adjusting for the potential confounders, the odds ratio of recurrence was 3.069 (95%CI =1.488-6.330, p = 0.002) for the highest quartile of BUN compared with the lowest quartile. In multiple-adjusted spline regression, a high BUN level visually showed a significantly high OR value of recurrence risk. CONCLUSIONS: Elevated BUN at post-operation is significantly associated with the recurrence of CSDH, and it is indicated that high levels of serum BUN after evacuation may serve as a risk factor for CSDH recurrence.

Distribution pattern of invasion-related bio-markers in head Marjolin's ulcer.

Marjolin's ulcer (MU) is a rare and aggressive cutaneous malignancy that typically presented in an area of traumatized or chronically inflamed skin and particularly in burn scars. Among them, the MU in the scalp with extensive invasion of the skull is exceptional and severe. The principle of management for MU is to obtain an early diagnosis and perform prompt surgical interventions. The invasive capacity of MU may vary among different sites of the scalp, which may require different therapeutic strategies for surgical excision. However, no clear evidence has been provided to determine the invasion ability of MU at different regions of the lesion as a surgical guidance. In present study, a 41-year-old female with a 40-year history of scalp ulceration has been examined. After resection of the MU lesion, hematoxylin and eosin (H&E) staining was performed to confirm the pathology of the cutaneous malignancy after surgical excision. Furthermore, reverse transcription-quantitative PCR experiment was performed out to determine the expression levels of invasion-associated biomarkers at different sites of the scalp affected by MU. Pathological analysis with H&E staining indicated a differentiated squamous cell carcinoma with invasion of the skull. The invasion-associated biomarkers were highly expressed in the core region compared to the middle region as well as the edge of MU tissue. Taken together, the present study suggests that the expression pattern of invasion-associated biomarkers varies between different regions of the MU lesion. High expression levels in the core region of MU indicates that the resection of the center area may be critical for the successful surgical treatment of MU.

Immunohistochemical Profiles of Matrix Metalloproteinases and Vascular Endothelial Growth Factor Overexpression in the Antoni B Area of Vestibular Schwannomas.

OBJECTIVE: To evaluate the clinical manifestations of cystic vestibular schwannomas (VSs), investigate the immunohistochemical profiles of matrix metalloproteinases (MMPs) and vascular endothelial growth factor (VEGF) expression in Antoni A and B areas, and speculate the pathogenesis of cystic formation and intratumoral hemorrhage. METHODS: Clinical features and outcomes of 24 cases of cystic VSs and 38 cases of solid VSs were retrospectively compared. Immunohistochemical studies were conducted to evaluate the characteristics of MMPs and VEGF in cystic and solid VSs. RESULTS: The tumor size was 38.92 ± 1.86 mm and 31.95 ± 1.74 mm in the cystic and solid VSs group, respectively (P = 0.011). Cystic VSs were rich in the Antoni B area. MMP-9 expression was low in the Antoni A and B areas. MMP-2 was moderately expressed. No significant difference in MMP-2 expression existed between the Antoni A and B areas (P > 0.05). VEGF and MMP-14 expression were moderate in the Antoni A area and intense in the Antoni B area, and the expression of both was significantly greater in the Antoni B area than in the Antoni A area (P < 0.001). CONCLUSIONS: MMP-14 and VEGF expression were significantly greater in the Antoni B area than in the Antoni A area. Upregulated MMP-14 may degrade loose collagen in the Antoni B area and contribute to cystic formation. MMP-14 can enhance VEGF activity, which may induce extravasation of a plasma ultrafiltrate, cystic expansion, and intratumoral hemorrhage. Therefore, MMP-14 inhibition may be a therapeutic strategy for treating cystic VSs.

New Measures for the Coronavirus Disease 2019 Response: A Lesson From the Wenzhou Experience.

As the outbreak of coronavirus disease 2019 (COVID-19) has spread globally, determining how to prevent the spread is of paramount importance. We reported the effectiveness of different responses of 4 affected cities in preventing the COVID-19 spread. We expect the Wenzhou anti-COVID-19 measures may provide information for cities around the world that are experiencing this epidemic.

Extracellular vesicles extracted from young donor serum attenuate inflammaging via partially rejuvenating aged T-cell immunotolerance.

Biologic aging results in a chronic inflammatory condition, termed inflammaging, which establishes a risk for such age-related diseases as neurocardiovascular diseases; therefore, it is of great importance to develop rejuvenation strategies that are able to attenuate inflammaging as a means of intervention for age-related diseases. A promising rejuvenation factor that is present in young blood has been found that can make aged neurons younger; however, the component in the young blood and its mechanism of action are poorly elucidated. We assessed rejuvenation in naturally aged mice with extracellular vesicles (EVs) or exosomes extracted from young murine serum on the basis of different spectrums of microRNAs in these vesicles from young and old sera. We found that EVs extracted from young donor mouse serum, rather than EVs extracted from old donor mouse serum or non-EV supernatant extracted from young donor mouse serum, were able to attenuate inflammaging in old mice. Inflammaging is attributed to multiple factors, one of which is thymic aging-released self-reactive T cell-induced pathology. We found that the attenuation of inflammaging after treatment with EVs from young serum partially contributed to the rejuvenation of thymic aging, which is characterized by partially reversed thymic involution, enhancement of negative selection signals, and reduced autoreactions in the periphery. Our results provide evidence for understanding of the potential rejuvenation factor in the young donor serum, which holds great promise for the development of novel therapeutics to reduce morbidity and mortality caused by age-related inflammatory diseases.-Wang, W., Wang, L., Ruan, L., Oh, J., Dong, X., Zhuge, Q., Su, D.-M. Extracellular vesicles extracted from young donor serum attenuate inflammaging via partially rejuvenating aged T-cell immunotolerance.

Impact of aging immune system on neurodegeneration and potential immunotherapies.

The interaction between the nervous and immune systems during aging is an area of avid interest, but many aspects remain unclear. This is due, not only to the complexity of the aging process, but also to a mutual dependency and reciprocal causation of alterations and diseases between both the nervous and immune systems. Aging of the brain drives whole body systemic aging, including aging-related changes of the immune system. In turn, the immune system aging, particularly immunosenescence and T cell aging initiated by thymic involution that are sources of chronic inflammation in the elderly (termed inflammaging), potentially induces brain aging and memory loss in a reciprocal manner. Therefore, immunotherapeutics including modulation of inflammation, vaccination, cellular immune therapies and "protective autoimmunity" provide promising approaches to rejuvenate neuroinflammatory disorders and repair brain injury. In this review, we summarize recent discoveries linking the aging immune system with the development of neurodegeneration. Additionally, we discuss potential rejuvenation strategies, focusing aimed at targeting the aging immune system in an effort to prevent acute brain injury and chronic neurodegeneration during aging.

Preclinical Studies and Translational Applications of Intracerebral Hemorrhage.

Intracerebral hemorrhage (ICH) which refers to bleeding in the brain is a very deleterious condition with high mortality and disability rate. Surgery or conservative therapy remains the treatment option. Various studies have divided the disease process of ICH into primary and secondary injury, for which knowledge into these processes has yielded many preclinical and clinical treatment options. The aim of this review is to highlight some of the new experimental drugs as well as other treatment options like stem cell therapy, rehabilitation, and nanomedicine and mention some translational clinical applications that have been done with these treatment options.

T2 relaxation time for intervertebral disc degeneration in patients with upper back pain: initial results on the clinical use of 3.0 Tesla MRI.

BACKGROUND: Magnetic resonance imaging (MRI) is a useful non-invasive tool for evaluating abnormalities of intervertebral discs. However, there are few studies which applied functional MRI techniques to investigate degenerative changes in cervical and cervicothoracic junction (CTJ) spine among adults. The aim of this study was to compare T2 relaxation time measurement evaluation with morphological grading for assessing cervical and CTJ intervertebral discs (IVD) in the patients suffering neck, shoulder, and upper back pain. METHODS: Sixty-three patients (378 IVDs) and 60 asymptomatic volunteers (360 IVDs) of the cervical and CTJ discs were assessed using a 3.0 T magnetic resonance imaging (MRI) protocol, including an sagittal T2 relaxation time protocol. The relaxation time values of the nucleus pulposus (NP) were recorded and all discs were visually graded according to Pfirrman's grading system. The correlation between T2 relaxation time values and qualitative clinical grading of degeneration, patient age, sex and anatomic level were analyzed RESULTS: There is a clear trend of decreasing mean T2 values of the NP associate with increasing Pfirrmann grades (C2-T1) for both patients and asymptotic volunteers. Significant T2 differences were seen among grades I-V (P < 0.05). However, grade V was not observed in the CTJ. Linear correlation analysis revealed a strong negative association between T2 values of the NP and Pfirrmann grade (r = -0.588, r = -0.808) of C2-7 and C7T1. Age were also significantly correlated NP T2 values (r = -0.525, r = -0.723) for patients and volunteers. Moreover, the receiver operating characteristic analysis for average measures in a range from 0.70-0.79 (C2-7) to 0.84-0.89 (C7T1) for patients. CONCLUSIONS: T2 quantitation provides a more sensitive and robust approach for detecting and characterizing the early stage of IVD degeneration and age-associated disc changes.

Thymic involution perturbs negative selection leading to autoreactive T cells that induce chronic inflammation.

Thymic involution and the subsequent amplified release of autoreactive T cells increase the susceptibility toward developing autoimmunity, but whether they induce chronic inflammation with advanced age remains unclear. The presence of chronic low-level proinflammatory factors in elderly individuals (termed inflammaging) is a significant risk factor for morbidity and mortality in virtually every chronic age-related disease. To determine how thymic involution leads to the persistent release and activation of autoreactive T cells capable of inducing inflammaging, we used a Foxn1 conditional knockout mouse model that induces accelerated thymic involution while maintaining a young periphery. We found that thymic involution leads to T cell activation shortly after thymic egress, which is accompanied by a chronic inflammatory phenotype consisting of cellular infiltration into non-lymphoid tissues, increased TNF-α production, and elevated serum IL-6. Autoreactive T cell clones were detected in the periphery of Foxn1 conditional knockout mice. A failure of negative selection, facilitated by decreased expression of Aire rather than impaired regulatory T cell generation, led to autoreactive T cell generation. Furthermore, the young environment can reverse age-related regulatory T cell accumulation in naturally aged mice, but not inflammatory infiltration. Taken together, these findings identify thymic involution and the persistent activation of autoreactive T cells as a contributing source of chronic inflammation (inflammaging).

Coupling of neurogenesis and angiogenesis after ischemic stroke.

Stroke is a leading cause of mortality and severe long-term disability worldwide. Development of effective treatment or new therapeutic strategies for ischemic stroke patients is therefore crucial. Ischemic stroke promotes neurogenesis by several growth factors including FGF-2, IGF-1, BDNF, VEGF and chemokines including SDF-1, MCP-1. Stroke-induced angiogenesis is similarly regulated by many factors most notably, eNOS and CSE, VEGF/VEGFR2, and Ang-1/Tie2. Important findings in the last decade have revealed that neurogenesis is not the stand-alone consideration in the fight for full functional recovery from stroke. Angiogenesis has been also shown to be critical in improving post-stroke neurological functional recovery. More than that, recent evidence has shown a highly possible interplay or dependence between stroke-induced neurogenesis and angiogenesis. Moving forward, elucidating the underlying mechanisms of this coupling between stroke-induced neurogenesis and angiogenesis will be of great importance, which will provide the basis for neurorestorative therapy. This article is part of a Special Issue entitled SI: Cell Interactions In Stroke.

Transplanted neural stem cells modulate regulatory T, γδ T cells and corresponding cytokines after intracerebral hemorrhage in rats.

The immune system, particularly T lymphocytes and cytokines, has been implicated in the progression of brain injury after intracerebral hemorrhage (ICH). Although studies have shown that transplanted neural stem cells (NSCs) protect the central nervous system (CNS) from inflammatory damage, their effects on subpopulations of T lymphocytes and their corresponding cytokines are largely unexplored. Here, rats were subjected to ICH and NSCs were intracerebrally injected at 3 h after ICH. The profiles of subpopulations of T cells in the brain and peripheral blood were analyzed by flow cytometry. We found that regulatory T (Treg) cells in the brain and peripheral blood were increased, but γδT cells (gamma delta T cells) were decreased, along with increased anti-inflammatory cytokines (IL-4, IL-10 and TGF-ß) and decreased pro-inflammatory cytokines (IL-6, and IFN-γ), compared to the vehicle-treated control. Our data suggest that transplanted NSCs protect brain injury after ICH via modulation of Treg and γδT cell infiltration and anti- and pro-inflammatory cytokine release.

Inhibition of mammalian target of rapamycin improves neurobehavioral deficit and modulates immune response after intracerebral hemorrhage in rat.

BACKGROUND: Mammalian target of rapamycin (mTOR), a serine/threonine kinase, regulates many processes, including cell growth and the immune response. mTOR is also dysregulated in several neurological diseases, such as traumatic brain injury (TBI), stroke, and neurodegenerative disease. However, the role of mTOR in intracerebral hemorrhage (ICH) remains unexplored. The aims of our study were to determine whether inhibiting mTOR signaling could affect the outcome after ICH and to investigate the possible underlying mechanism. METHODS: A rat ICH model was induced by intracerebral injection of collagenase IV into the striatum, and mTOR activation was inhibited by administration of rapamycin. mTOR signaling activation was determined by western blotting. Neurobehavioral deficit after ICH was determined by a set of modified Neurological Severity Scores (mNSS). The levels of CD4+CD25+Foxp3+ regulatory T cells (Tregs) and cytokines were examined using flow cytometry and ELISA, respectively. RESULTS: Our results demonstrated thatmTOR signaling was activated 30 minutes and returned to its basal level 1 day after ICH. Increased p-mTOR, which mean that mTOR signaling was activated, was predominantly located around the hematoma. Rapamycin treatment significantly improved the neurobehavioral deficit after ICH, increased the number of Tregs, increased levels of interleukin-10 and transforming growth factor-ß and reduced interferon-γ both in peripheral blood and brain. CONCLUSIONS: Our study suggests that mTOR improves ICH outcome and modulates immune response after ICH.