The MT1 receptor as the target of ramelteon neuroprotection in ischemic stroke.

Stroke is the leading cause of death and disability worldwide. Novel and effective therapies for ischemic stroke are urgently needed. Here, we report that melatonin receptor 1A (MT1) agonist ramelteon is a neuroprotective drug candidate as demonstrated by comprehensive experimental models of ischemic stroke, including a middle cerebral artery occlusion (MCAO) mouse model of cerebral ischemia in vivo, organotypic hippocampal slice cultures ex vivo, and cultured neurons in vitro; the neuroprotective effects of ramelteon are diminished in MT1-knockout (KO) mice and MT1-KO cultured neurons. For the first time, we report that the MT1 receptor is significantly depleted in the brain of MCAO mice, and ramelteon treatment significantly recovers the brain MT1 losses in MCAO mice, which is further explained by the Connectivity Map L1000 bioinformatic analysis that shows gene-expression signatures of MCAO mice are negatively connected to melatonin receptor agonist like Ramelteon. We demonstrate that ramelteon improves the cerebral blood flow signals in ischemic stroke that is potentially mediated, at least, partly by mechanisms of activating endothelial nitric oxide synthase. Our results also show that the neuroprotection of ramelteon counteracts reactive oxygen species-induced oxidative stress and activates the nuclear factor erythroid 2-related factor 2/heme oxygenase-1 pathway. Ramelteon inhibits the mitochondrial and autophagic death pathways in MCAO mice and cultured neurons, consistent with gene set enrichment analysis from a bioinformatics perspective angle. Our data suggest that Ramelteon is a potential neuroprotective drug candidate, and MT1 is the neuroprotective target for ischemic stroke, which provides new insights into stroke therapy. MT1-KO mice and cultured neurons may provide animal and cellular models of accelerated ischemic damage and neuronal cell death.

Extended characterization of IL-33/ST2 as a predictor for wound age determination in skin wound tissue samples of humans and mice.

Interleukin (IL)-33, an important inflammatory cytokine, is highly expressed in skin wound tissue and serum of humans and mice, and plays an essential role in the process of skin wound healing (SWH) dependent on the IL-33/suppression of tumorigenicity 2 (ST2) pathway. However, whether IL-33 and ST2 themselves, as well as their interaction, can be applied for skin wound age determination in forensic practice remains incompletely characterized. Human skin samples with injured intervals of a few minutes to 24 hours (hs) and mouse skin samples with injured intervals of 1 h to 14 days (ds) were collected. Herein, the results demonstrated that IL-33 and ST2 are increased in the human skin wounds, and that in mice skin wounds, there is an increase over time, with IL-33 expression peaking at 24 hs and 10 ds, and ST2 expression peaking at 12 hs and 7 ds. Notably, the relative quantity of IL-33 and ST2 proteins < 0.35 suggested a wound age of 3 hs; their relative quantity > 1.0 suggested a wound age of 24 hs post-mouse skin wounds. In addition, immunofluorescent staining results showed that IL-33 and ST2 were consistently expressed in the cytoplasm of F4/80-positive macrophages and CD31-positive vascular endothelial cells with or without skin wounds, whereas nuclear localization of IL-33 was absent in α-SMA-positive myofibroblasts with skin wounds. Interestingly, IL-33 administration facilitated the wound area closure by increasing the proliferation of cytokeratin (K) 14 -positive keratinocytes and vimentin-positive fibroblasts. In contrast, treating with its antagonist (i.e., anti-IL-33) or receptor antagonist (e.g., anti-ST2) exacerbated the aforementioned pathological changes. Moreover, treatment with IL-33 combined with anti-IL-33 or anti-ST2 reversed the effect of IL-33 on facilitating skin wound closure, suggesting that IL-33 administration facilitated skin wound closure through the IL-33/ST2 signaling pathway. Collectively, these findings indicate that the detection of IL-33/ST2 might be a reliable biomarker for the determination of skin wound age in forensic practice.

ACSL4-Mediated Ferroptosis and Its Potential Role in Central Nervous System Diseases and Injuries.

As an iron-dependent regulated form of cell death, ferroptosis is characterized by iron-dependent lipid peroxidation and has been implicated in the occurrence and development of various diseases, including nervous system diseases and injuries. Ferroptosis has become a potential target for intervention in these diseases or injuries in relevant preclinical models. As a member of the Acyl-CoA synthetase long-chain family (ACSLs) that can convert saturated and unsaturated fatty acids, Acyl-CoA synthetase long-chain familymember4 (ACSL4) is involved in the regulation of arachidonic acid and eicosapentaenoic acid, thus leading to ferroptosis. The underlying molecular mechanisms of ACSL4-mediated ferroptosis will promote additional treatment strategies for these diseases or injury conditions. Our review article provides a current view of ACSL4-mediated ferroptosis, mainly including the structure and function of ACSL4, as well as the role of ACSL4 in ferroptosis. We also summarize the latest research progress of ACSL4-mediated ferroptosis in central nervous system injuries and diseases, further proving that ACSL4-medicated ferroptosis is an important target for intervention in these diseases or injuries.

Molecular Toxicology and Pathophysiology of Comorbid Alcohol Use Disorder and Post-Traumatic Stress Disorder Associated with Traumatic Brain Injury.

The increasing comorbidity of alcohol use disorder (AUD) and post-traumatic stress disorder (PTSD) associated with traumatic brain injury (TBI) is a serious medical, economic, and social issue. However, the molecular toxicology and pathophysiological mechanisms of comorbid AUD and PTSD are not well understood and the identification of the comorbidity state markers is significantly challenging. This review summarizes the main characteristics of comorbidity between AUD and PTSD (AUD/PTSD) and highlights the significance of a comprehensive understanding of the molecular toxicology and pathophysiological mechanisms of AUD/PTSD, particularly following TBI, with a focus on the role of metabolomics, inflammation, neuroendocrine, signal transduction pathways, and genetic regulation. Instead of a separate disease state, a comprehensive examination of comorbid AUD and PTSD is emphasized by considering additive and synergistic interactions between the two diseases. Finally, we propose several hypotheses of molecular mechanisms for AUD/PTSD and discuss potential future research directions that may provide new insights and translational application opportunities.

Autophagy inhibition facilitates wound closure partially dependent on the YAP/IL-33 signaling in a mouse model of skin wound healing.

Autophagy is a self-phagocytic and highly evolutionarily conserved intracellular lysosomal catabolic system, which plays a vital role in a variety of trauma models, including skin wound healing (SWH). However, the roles and potential mechanisms of autophagy in SWH are still controversial. We firstly investigated the role of autophagy in SWH-induced wound closure rate, inflammatory response, and histopathology, utilizing an inhibitor of autophagy 3-methyladenine (3-MA) and its agonist rapamycin (RAP). As expected, we found 3-MA treatment remarkably increased the wound closure rate, combated inflammation response, and mitigated histopathological changes, while RAP delivery aggravated SWH-induced pathological damage. To further exploit the underlying mechanism of autophagy regulating inflammation, the specific inhibitors of yes-associated protein (YAP), Verteporfin, and Anti-IL-33 were applied. Herein, treating with 3-MA markedly suppressed the expression of tumor necrosis factor-α (TNF-α), IL-1ß, and IL-6, promoted that of IL-10, IL-33, and ST2, while RAP administration reverted SWH-induced the up-regulation of these inflammatory cytokines mentioned above. Importantly, Verteporfin administration not only down-regulated the expression levels of YAP, TNF-α, and IL-6 but also up-regulated that of IL-33 and IL-10. Unexpectedly, 3-MA or RAP retreatment did not have any impact on the changes in IL-33 among these inflammatory indicators. Furthermore, elevated expression of IL-33 promoted wound closure and alleviated the pathological damage, whereas, its antagonist Anti-IL-33 treatment overtly reversed the above-mentioned effects of IL-33. Moreover, 3-MA in combination with anti-IL-33 treatment reversed the role of 3-MA alone in mitigated pathological changes, but they failed to revert the effect of anti-IL-33 alone on worsening pathological damage. In sum, emerging data support the novel contribution of the YAP/IL-33 pathway in autophagy inhibition against SWH-induced pathological damage, and highlight that the autophagy/YAP/IL-33 signal axis is expected to become a new therapeutic target for SWH.

Emerging Effects of IL-33 on COVID-19.

Since the start of COVID-19 pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), more than 6 million people have lost their lives worldwide directly or indirectly. Despite intensified efforts to clarify the immunopathology of COVID-19, the key factors and processes that trigger an inflammatory storm and lead to severe clinical outcomes in patients remain unclear. As an inflammatory storm factor, IL-33 is an alarmin cytokine, which plays an important role in cell damage or infection. Recent studies have shown that serum IL-33 is upregulated in COVID-19 patients and is strongly associated with poor outcomes. Increased IL-33 levels in severe infections may result from an inflammatory storm caused by strong interactions between activated immune cells. However, the effects of IL-33 in COVID-19 and the underlying mechanisms remain to be fully elucidated. In this review, we systematically discuss the biological properties of IL-33 under pathophysiological conditions and its regulation of immune cells, including neutrophils, innate lymphocytes (ILCs), dendritic cells, macrophages, CD4+ T cells, Th17/Treg cells, and CD8+ T cells, in COVID-19 phagocytosis. The aim of this review is to explore the potential value of the IL-33/immune cell pathway as a new target for early diagnosis, monitoring of severe cases, and clinical treatment of COVID-19.

Deletion of ferritin H in neurons counteracts the protective effect of melatonin against traumatic brain injury-induced ferroptosis.

Accumulating evidence demonstrates that ferroptosis may be important in the pathophysiological process of traumatic brain injury (TBI). As a major hormone of the pineal gland, melatonin exerts many beneficial effects on TBI, but there is no information regarding the effects of melatonin on ferroptosis after TBI. As expected, TBI resulted in the time-course changes of ferroptosis-related molecules expression and iron accumulation in the ipsilateral cortex. Importantly, we found that treating with melatonin potently rescued TBI induced the changes mentioned above and improved functional deficits versus vehicle. Similar results were obtained with a ferroptosis inhibitor, liproxstatin-1. Moreover, the protective effect of melatonin is likely dependent on melatonin receptor 1B (MT2). Although ferritin plays a vital role in iron metabolism by storing excess cellular iron, its precise function in the brain, and whether it involves melatonin's neuroprotection remain unexplored. Considering ferritin H (Fth) is expressed predominantly in the neurons and global loss of Fth in mice induces early embryonic lethality, we then generated neuron-specific Fth conditional knockout (Fth-KO) mice, which are viable and fertile but have altered iron metabolism. In addition, Fth-KO mice were more susceptible to ferroptosis after TBI, and the neuroprotection by melatonin was largely abolished in Fth-KO mice. In vitro siFth experiments further confirmed the results mentioned above. Taken together, these data indicate that melatonin produces cerebroprotection, at least partly by inhibiting neuronal Fth-mediated ferroptosis following TBI, supporting the notion that melatonin is an excellent ferroptosis inhibitor and its anti-ferroptosis provides a potential therapeutic target for treating TBI.

Mdivi-1 alleviates brain damage and synaptic dysfunction after intracerebral hemorrhage in mice.

As a selective inhibitor of mitochondrial fission protein dynamin-related protein-1 (Drp1), mitochondrial division inhibitor 1 (mdivi-1) can cross the blood-brain barrier (BBB) and exert neuroprotection. However, it remains unclear whether mdivi-1 can attenuate intracerebral hemorrhage (ICH)-induced secondary brain injury. This study was undertaken to characterize the roles of mdivi-1 in short-term and long-term behavioral outcomes, along with synaptic plasticity changes in mice after ICH. The results indicated mdivi-1 reversed Drp1 translocation and the morphologic changes of mitochondria, as well as ameliorated short-term neurobehavioral deficits, the BBB disruption and brain edema remarkably. In addition, mdivi-1 could rescue ICH-induced motor and memory dysfunctions. Mdivi-1 could also prevent ICH-induced reductions in synaptic proteins (synapsin I, PSD95) and phosphorylated cAMP-response element binding (p-CREB). In vitro, mdivi-1 inhibited hemin-induced hippocampal neuron death and improved neurite outgrowth. In conclusion, we found that mdivi-1 can alleviate short-term and long-term neurological deficits, synaptic dysfunction. These findings demonstrate that mdivi-1 may be beneficial in the treatment of secondary brain injury, synaptic dysfunction and neurological outcomes caused by ICH.

Platelet regulates neuroinflammation and restores blood-brain barrier integrity in a mouse model of traumatic brain injury.

Neuroinflammation accompanied by microglial activation triggers multiple cell death after traumatic brain injury (TBI). The secondary injury caused by inflammation may persist for a long time. Recently, platelet C-type lectin-like 2 receptor (CLEC-2) has been shown to regulate inflammation in certain diseases. However, its possible effects on TBI remain poorly understood. Here, we aimed to investigate the role of platelet CLEC-2 in the pathological process of neuroinflammation after TBI. In this study, mice were subjected to sham or controlled cortical impact injury, and arbitrarily received recombinant platelet CLEC-2. In parallel, BV2 cells were treated with lipopolysaccharide (LPS) to mimic microglial activation after TBI. Primary endothelial cells were also subjected to LPS in order to replicate the inflammatory damage caused by TBI. We used western blot analysis, reverse transcription polymerase chain reaction (RT-PCR), and immunostaining to evaluate the role of platelet CLEC-2 in TBI. In conditional knock out platelet CLEC-2 mice, trauma worsened the integrity of the blood-brain barrier and amplified the release of inflammatory cytokines. In wild type mice subjected to controlled cortical impact injury, recombinant platelet CLEC-2 administration altered the secretion of inflammatory cytokines, reduced brain edema, and improved neurological function. In vitro, the polarization phenotype of microglia induced by LPS was transformed by recombinant platelet CLEC-2, and this conversion depended on the mammalian target of rapamycin (mTOR) pathway. Endothelial cell injury by LPS was ameliorated when microglia expressed mostly M2 phenotype markers. In conclusion, platelet CLEC-2 regulates trauma-induced neuroinflammation and restores blood-brain barrier integrity.

Chd8 Rescued TBI-Induced Neurological Deficits by Suppressing Apoptosis and Autophagy Via Wnt Signaling Pathway.

Traumatic brain injury (TBI) and autism spectrum disorder (ASDs) share several same biochemical mechanisms and symptoms, such as learning memory impairments and communication deficits. Chromodomain helicase DNA binding protein 8 (CHD8), a member of the CHD family of ATP-dependent chromatin remodeling factors, is one of the top risk genetic factors in ASDs and is highly associated with Wnt/ß-catenin signaling. Yet, the possible effect of CHD8 on TBI remains poorly understood. In vivo, we found that Chd8 co-localized in neurons, astrocytes, and microglia, but predominantly presented in neurons in the prefrontal cortex, hippocampus, and cortex. Both Chd8 and ß-catenin expression peaked at 12 h and shared the similar change tendency after TBI. Chd8 knockdown inhibited wnt pathway, promoted the activation of apoptosis and autophagy, and caused learning and memory impairments both at normal and TBI condition. In addition, overexpression of Chd8 via 17ß-estrogen (E2) treatment enhanced wnt signaling pathway and suppressed TBI-induced apoptosis and autophagic activation. In vitro, a significant increase of Chd8 and ß-catenin expression was observed in HT22 cells after lipopolysaccharide (lps) treatment or mechanical injury, respectively. Chd8 knockdown inhibited wnt signaling pathway and increased apoptosis and autophagy activation in lps-stimulated HT22 cells. But activation of wnt signaling inverted the effects of Chd8-siRNA. Our results demonstrated that Chd8 exerted neuroprotection and promoted cognitive recovery through inhibiting apoptosis and autophagy activation following TBI, at least partially by wnt signaling pathway.

Chronic restraint stress exacerbates neurological deficits and disrupts the remodeling of the neurovascular unit in a mouse intracerebral hemorrhage model.

Growing evidences have shown that patients recovering from stroke experience high and unremitting stress. Chronic restraint stress (CRS) has been found to exacerbate neurological impairments in an experimental focal cortical ischemia model. However, there have been no studies reporting the effect and mechanism of CRS on intracerebral hemorrhage (ICH). This study aimed to evaluate the effect of CRS on a mouse ICH model. Adult male C57BL mice were subjected to infusion of collagenase IV (to induce ICH) or saline (for sham) into the left striatum. After ICH, animals were stressed with application of CRS protocol for 21 days. Our results showed that CRS significantly exacerbated neurological deficits (Garcia test, corner turn test, and wire grip test) and the ipsilateral brain atrophy and reduced body weight gain after ICH. Immunofluorescence staining indicated that CRS exerted significant suppressive effects on neuron, astrocyte, vascular endothelial cell and pericyte and excessively activated microglia post ICH. All of the key cellular components mentioned above are involved in the neurovascular unit (NVU) remodeling in the peri-hemorrhagic region after ICH. Western blot results showed that matrix metalloproteinase (MMP)-9 and tight junction (TJ) proteins including zonula occludens-1, occludin and claudin-5 were increased after ICH, but MMP-9 protein was further up-regulated and TJ-related proteins were down-regulated by CRS. In addition, ICH-induced activation of endoplasmic reticulum stress and apoptosis were further strengthened by CRS. Collectively, CRS exacerbates neurological deficits and disrupts the remodeling of the peri-hemorrhagic NVU after ICH, which may be associated with TJ proteins degradation and excessive activation of MMP-9 and endoplasmic reticulum stress-apoptosis.LAY SUMMARYCRS exacerbates neurological deficits and disrupts the remodeling of the NVU in the recovery stage after ICH, which suggest that monitoring chronic stress levels in patients recovering from ICH may merit consideration in the future.

The Function and Mechanisms of Autophagy in Traumatic Brain Injury.

Traumatic brain injury (TBI) is one of the most common causes of long-term disability and death worldwide. Autophagy is activated and autophagic flux is impaired following TBI. But the controversial roles and underlying mechanisms of autophagy after TBI are not clear. This chapter will update the current state of knowledge in the process of autophagy, the roles of autophagy in TBI as well as some upstream moleculars and pharmacological regulators of autophagy involved in TBI. We also discuss autophagy mechanism-based preclinical pharmacological intervention. These observations make autophagy an attractive therapeutic target for developing new therapeutic strategies to achieve better outcomes for patients suffering from TBI.

The Function and Mechanisms of Autophagy in Spinal Cord Injury.

Spinal cord injury (SCI) is one of the major causes of death and long-term disability in the world. Numerous studies have reported that autophagy plays an important role in SCI. However, the understanding of underlying mechanisms of autophagy after SCI and autophagy mechanism-based preclinical pharmacological intervention needs to be updated. This part provides an overview of current knowledge about the mechanisms of autophagy and autophagy flux as well as its potential molecular mechanisms based on the pharmacological regulation of autophagy. Although autophagic activation and the disruption of autophagy flux exist in SCI, whether autophagy is beneficial and detrimental is still under debate. We also focus on the existing and developing therapeutic options based on the potential molecular mechanisms of autophagy.

The Function and Mechanisms of Autophagy in Trauma of Other Parts of the Body.

Trauma is common in modern society. Besides TBI and SCI, trauma can lead to severe cardiopulmonary injury and even to death. Fracture and skin injury are also very likely to occur in our daily life. Limited studies have reported the levels of autophagy after heart and trauma, fracture, and skin injury. In this chapter, we update the current state of knowledge and recent advances in the study of autophagy after trauma including heart and lung trauma, fracture, and skin injury which we try to clarify how autophagy levels are affected by injury or trauma and how their manipulation may represent potential novel protective targets for treatments.

Salubrinal offers neuroprotection through suppressing endoplasmic reticulum stress, autophagy and apoptosis in a mouse traumatic brain injury model.

Traumatic brain injury (TBI) is a complex injury that can cause severe disabilities and even death. TBI can induce secondary injury cascades, including but not limited to endoplasmic reticulum (ER) stress, apoptosis and autophagy. Although the investigators has previously shown that salubrinal, the selective phosphatase inhibitor of p-eIF2α, ameliorated neurologic deficits in murine TBI model, the neuroprotective mechanisms of salubrinal need further research to warrant the preclinical value. This study was undertaken to characterize the effects of salubrinal on cell death and neurological outcomes following TBI in mice and the potential mechanisms. In the current study, ER stress-related proteins including p-eIF2α, GRP78 and CHOP showed peak expressions both in the cortex and hippocampus from day 2 to day 3 after TBI, indicating ER stress was activated in our TBI model. Immunofluorescence staining showed that CHOP co-located NeuN-positive neuron, GFAP-positive astrocyte, Iba-1-positive microglia, CD31-positive vascular endothelial cell and PDGFR-ß-positive pericyte in the cortex on day 2 after TBI, and these cells mentioned above constitute the neurovascular unit (NVU). We also found TBI-induced plasmalemma permeability, motor dysfunction, spatial learning and memory deficits and brain lesion volume were alleviated by continuous intraperitoneal administration of salubrinal post TBI. To investigate the underlying mechanisms further, we determined that salubrinal suppressed the expression of ER stress, autophagy and apoptosis related proteins on day 2 after TBI. In addition, salubrinal administration decreased the number of CHOP+/TUNEL+ and CHOP+/LC3+ cells on day 2 after TBI, detected by immunofluorescence. In conclusion, these data imply that salubrinal treatment improves morphological and functional outcomes caused by TBI in mice and these neuroprotective effects may be associated with inhibiting apoptosis, at least in part by suppressing ER stress-autophagy pathway.

Upregulation of 3-MST Relates to Neuronal Autophagy After Traumatic Brain Injury in Mice.

3-mercaptopyruvate sulfurtransferase (3-MST) was a novel hydrogen sulfide (H2S)-synthesizing enzyme that may be involved in cyanide degradation and in thiosulfate biosynthesis. Over recent years, considerable attention has been focused on the biochemistry and molecular biology of H2S-synthesizing enzyme. In contrast, there have been few concerted attempts to investigate the changes in the expression of the H2S-synthesizing enzymes with disease states. To investigate the changes of 3-MST after traumatic brain injury (TBI) and its possible role, mice TBI model was established by controlled cortical impact system, and the expression and cellular localization of 3-MST after TBI was investigated in the present study. Western blot analysis revealed that 3-MST was present in normal mice brain cortex. It gradually increased, reached a peak on the first day after TBI, and then reached a valley on the third day. Importantly, 3-MST was colocalized with neuron. In addition, Western blot detection showed that the first day post injury was also the autophagic peak indicated by the elevated expression of LC3. Importantly, immunohistochemistry analysis revealed that injury-induced expression of 3-MST was partly colabeled by LC3. However, there was no colocalization of 3-MST with propidium iodide (cell death marker) and LC3 positive cells were partly colocalized with propidium iodide. These data suggested that 3-MST was mainly located in living neurons and may be implicated in the autophagy of neuron and involved in the pathophysiology of brain after TBI.

Blocking B7-1/CD28 Pathway Diminished Long-Range Brain Damage by Regulating the Immune and Inflammatory Responses in a Mouse Model of Intracerebral Hemorrhage.

Acute brain injuries can activate bidirectional crosstalk between the injured brain and the immune system. The immune system, particularly T lymphocytes and cytokines, has been implicated in the progression of brain injury after intracerebral hemorrhage (ICH). Co-stimulatory molecules B7-1 (CD80)/B7-2 (CD86) binding cognate receptor provides a secondary signaling to T cell activation. The aim of our study was to explore the effects of anti-B7-1 antibody on the development and prognosis of cerebral hemorrhage and to investigate the possible underlying mechanism. Mice were inner canthus veniplex administered with anti-B7-1 antibody at 10 min and 24 h after ICH and sacrificed on the third day after ICH. Immune function was assessed via splenocyte proliferation assay and organism index, respectively. IFN-γ and IL-4 were detected by enzyme-linked immuno sorbent assay. The cerebral edema was evaluated via brain water content. The levels of autophagy and apoptosis related proteins were measured by western blotting analysis. In addition, functional outcome was studied with pole-climbing test and morris water maze. The mice were weighed on 0, 1, 3, 14 and 21 days after ICH. The treatment with anti-B7-1 antibody significantly lowered immune function, and reduced the latency of water maze on 18 and 20 days, the ratio of IFN-γ/IL-4 as well as body weight on day 3 after cerebral hemorrhage. Our study suggests that in the cerebral hemorrhage mice brain anti-B7-1 antibody may reduce long-range brain damage by reversing immune imbalance.

N-acetyl-serotonin offers neuroprotection through inhibiting mitochondrial death pathways and autophagic activation in experimental models of ischemic injury.

N-acetylserotonin (NAS) is an immediate precursor of melatonin, which we have reported is neuroprotective against ischemic injury. Here we test whether NAS is a potential neuroprotective agent in experimental models of ischemic injury. We demonstrate that NAS inhibits cell death induced by oxygen-glucose deprivation or H2O2 in primary cerebrocortical neurons and primary hippocampal neurons in vitro, and organotypic hippocampal slice cultures ex vivo and reduces hypoxia/ischemia injury in the middle cerebral artery occlusion mouse model of cerebral ischemia in vivo. We find that NAS is neuroprotective by inhibiting the mitochondrial cell death pathway and the autophagic cell death pathway. The neuroprotective effects of NAS may result from the influence of mitochondrial permeability transition pore opening, mitochondrial fragmentation, and inhibition of the subsequent release of apoptogenic factors cytochrome c, Smac, and apoptosis-inducing factor from mitochondria to cytoplasm, and activation of caspase-3, -9, as well as the suppression of the activation of autophagy under stress conditions by increasing LC3-II and Beclin-1 levels and decreasing p62 level. However, NAS, unlike melatonin, does not provide neuroprotection through the activation of melatonin receptor 1A. We demonstrate that NAS reaches the brain subsequent to intraperitoneal injection using liquid chromatography/mass spectrometry analysis. Given that it occurs naturally and has low toxicity, NAS, like melatonin, has potential as a novel therapy for ischemic injury.

JMJD2A contributes to breast cancer progression through transcriptional repression of the tumor suppressor ARHI.

INTRODUCTION: Breast cancer is a worldwide health problem and the leading cause of cancer death among females. We previously identified Jumonji domain containing 2A (JMJD2A) as a critical mediator of breast cancer proliferation, migration and invasion. We now report that JMJD2A could promote breast cancer progression through transcriptional repression of the tumor suppressor aplasia Ras homolog member I (ARHI). METHODS: Immunohistochemistry was performed to examine protein expressions in 155 cases of breast cancer and 30 non-neoplastic tissues. Spearman correlation analysis was used to analyze the correlation between JMJD2A expression and clinical parameters as well as several tumor regulators in 155 cases of breast cancer. Gene and protein expressions were monitored by quantitative polymerase chain reaction (qPCR) and Western blot. Results from knockdown of JMJD2A, overexpression of JMJD2A, Co-immunoprecipitation (Co-IP) assay, dual luciferase reporter gene assay and chromatin immunoprecipitation (ChIP) elucidated molecular mechanisms of JMJD2A action in breast cancer progression. Furthermore, the effects of ARHI overexpression on JMJD2A-mediated tumor progression were investigated in vitro and in vivo. For in vitro experiments, cell proliferation, wound-healing, migration and invasion were monitored by cell counting, scratch and Boyden Chamber assays. For in vivo experiments, control cells and cells stably expressing JMJD2A alone or together with ARHI were inoculated into mammary fat pads of mice. Tumor volume, tumor weight and metastatic nodules were measured by caliper, electronic balance and nodule counting, respectively. RESULTS: JMJD2A was highly expressed in human breast cancers and positively correlated with tumor progression. Knockdown of JMJD2A increased ARHI expression whereas overexpression of JMJD2A decreased ARHI expression at both protein and mRNA levels. Furthermore, E2Fs and histone deacetylases were involved in the transcriptional repression of ARHI expression by JMJD2A. And the aggressive behavior of JMJD2A in breast cancers could be reversed by re-expression of ARHI in vitro and in vivo. CONCLUSION: We demonstrated a cancer-promoting effect of JMJD2A and defined a novel molecular pathway contributing to JMJD2A-mediated breast cancer progression.

The Interplay between Ferroptosis and Neuroinflammation in Central Neurological Disorders.

Central neurological disorders are significant contributors to morbidity, mortality, and long-term disability globally in modern society. These encompass neurodegenerative diseases, ischemic brain diseases, traumatic brain injury, epilepsy, depression, and more. The involved pathogenesis is notably intricate and diverse. Ferroptosis and neuroinflammation play pivotal roles in elucidating the causes of cognitive impairment stemming from these diseases. Given the concurrent occurrence of ferroptosis and neuroinflammation due to metabolic shifts such as iron and ROS, as well as their critical roles in central nervous disorders, the investigation into the co-regulatory mechanism of ferroptosis and neuroinflammation has emerged as a prominent area of research. This paper delves into the mechanisms of ferroptosis and neuroinflammation in central nervous disorders, along with their interrelationship. It specifically emphasizes the core molecules within the shared pathways governing ferroptosis and neuroinflammation, including SIRT1, Nrf2, NF-κB, Cox-2, iNOS/NO·, and how different immune cells and structures contribute to cognitive dysfunction through these mechanisms. Researchers' findings suggest that ferroptosis and neuroinflammation mutually promote each other and may represent key factors in the progression of central neurological disorders. A deeper comprehension of the common pathway between cellular ferroptosis and neuroinflammation holds promise for improving symptoms and prognosis related to central neurological disorders.