Peripheral cytokine interleukin-10 alleviates perihematomal edema after intracerebral hemorrhage via interleukin-10 receptor/JAK1/STAT3 signaling.

AIMS: The extent of perihematomal edema following intracerebral hemorrhage (ICH) significantly impacts patient prognosis, and disruption of the blood-brain barrier (BBB) exacerbates perihematomal edema. However, the role of peripheral IL-10 in mitigating BBB disruption through pathways that link peripheral and central nervous system signals remains poorly understood. METHODS: Recombinant IL-10 was administered to ICH model mice via caudal vein injection, an IL-10-inhibiting adeno-associated virus and an IL-10 receptor knockout plasmid were delivered intraventricularly, and neurobehavioral deficits, perihematomal edema, BBB disruption, and the expression of JAK1 and STAT3 were evaluated. RESULTS: Our study demonstrated that the peripheral cytokine IL-10 mitigated BBB breakdown, perihematomal edema, and neurobehavioral deficits after ICH and that IL-10 deficiency reversed these effects, likely through the IL-10R/JAK1/STAT3 signaling pathway. CONCLUSIONS: Peripheral IL-10 has the potential to reduce BBB damage and perihematomal edema following ICH and improve patient prognosis.

Endothelial EGLN3-PKM2 signaling induces the formation of acute astrocytic barrier to alleviate immune cell infiltration after subarachnoid hemorrhage.

BACKGROUND: Most subarachnoid hemorrhage (SAH) patients have no obvious hematoma lesions but exhibit blood-brain barrier dysfunction and vasogenic brain edema. However, there is a few days between blood‒brain barrier dysfunction and vasogenic brain edema. The present study sought to investigate whether this phenomenon is caused by endothelial injury induced by the acute astrocytic barrier, also known as the glial limitans. METHODS: Bioinformatics analyses of human endothelial cells and astrocytes under hypoxia were performed based on the GEO database. Wild-type, EGLN3 and PKM2 conditional knock-in mice were used to confirm glial limitan formation after SAH. Then, the effect of endothelial EGLN3-PKM2 signaling on temporal and spatial changes in glial limitans was evaluated in both in vivo and in vitro models of SAH. RESULTS: The data indicate that in the acute phase after SAH, astrocytes can form a temporary protective barrier, the glia limitans, around blood vessels that helps maintain barrier function and improve neurological prognosis. Molecular docking studies have shown that endothelial cells and astrocytes can promote glial limitans-based protection against early brain injury through EGLN3/PKM2 signaling and further activation of the PKC/ERK/MAPK signaling pathway in astrocytes after SAH. CONCLUSION: Improving the ability to maintain glial limitans may be a new therapeutic strategy for improving the prognosis of SAH patients.

TIMP-3 Alleviates White Matter Injury After Subarachnoid Hemorrhage in Mice by Promoting Oligodendrocyte Precursor Cell Maturation.

Subarachnoid hemorrhage (SAH) is associated with high mortality and disability rates, and secondary white matter injury is an important cause of poor prognosis. However, whether brain capillary pericytes can directly affect the differentiation and maturation of oligodendrocyte precursor cells (OPCs) and subsequently affect white matter injury repair has still been revealed. This study was designed to investigate the effect of tissue inhibitor of metalloproteinase-3 (TIMP-3) for OPC differentiation and maturation. PDGFRßret/ret and wild-type C57B6J male mice were used to construct a mouse model of SAH via endovascular perforation in this study. Mice were also treated with vehicle, TIMP-3 RNAi or TIMP-3 RNAi + TIMP-3 after SAH. The effect of TIMP-3 on the differentiation and maturation of OPCs was determined using behavioral score, ELISA, transmission electron microscopy, immunofluorescence staining and cell culture. We found that TIMP-3 was secreted mainly by pericytes and that SAH and TIMP-3 RNAi caused a significant decrease in the TIMP-3 content, reaching a nadir at 24 h, followed by gradual recovery. In vitro, the myelin basic protein content of oligodendrocytes after oxyhemoglobin treatment was increased by TIMP-3 overexpression. The data indicates TIMP-3 could promote the differentiation and maturation of OPCs and subsequently improve neurological outcomes after SAH. Therefore, TIMP-3 could be beneficial for repair after white matter injury and could be a potential therapeutic target in SAH.

Hydrophobic Polystyrene-Modified Gelatin Enhances Fast Hemostasis and Tissue Regeneration in Traumatic Brain Injury.

Hemostatic sealant is required to deal with blood loss, especially in the scenario of traumatic brain injury (TBI), which presents high rates of morbidity and disability. Hemostasis in surgery with traditional gelatin-based sealants often leads to blood loss and other issues in brain because of the hydrophilic gelatin swelling. Herein, hydrophobic effects on the hemostasis in TBI surgery are studied by tuning the chain length of polystyrene (PS) onto methylacrylated gelatin (Gel-MA). The hydrophobicity and hemostatic efficiency can be tuned by controlling the length of PS groups. The platelet activation of modified sealants Gel-MA-2P, Gel-MA-P, and Gel-MA-0.5P is as much as 17.5, 9.1, and 2.1 times higher than Gel-MA in vitro. The hemostatic time of Gel-MA-2P, Gel-MA-P, and Gel-MA-0.5P groups is 2.0-, 1.6-, and 1.1-folds faster than that in Gel-MA group in TBI mice. Increased formation of fibrins and platelet aggregation can also be observed in vitro by scanning electron microscopy. Animal's mortality is lowered by 46%, neurologic deficiency is reduced by 1.5 times, and brain edema is attenuated by 10%. Protein expression is further investigated to exhibit toxic iron-related processes caused by delayed hemostasis and activation of platelets via PI3K/PKC-α signaling. The hydrophobic Gel-MA has the potential in hemostatic TBI and promotes nervous system recovery in brain with the potentials in clinics.

Myelin sheath injury and repairment after subarachnoid hemorrhage.

Subarachnoid hemorrhage (SAH) can lead to damage to the myelin sheath in white matter. Through classification and analysis of relevant research results, the discussion in this paper provides a deeper understanding of the spatiotemporal change characteristics, pathophysiological mechanisms and treatment strategies of myelin sheath injury after SAH. The research progress for this condition was also systematically reviewed and compared related to myelin sheath in other fields. Serious deficiencies were identified in the research on myelin sheath injury and treatment after SAH. It is necessary to focus on the overall situation and actively explore different treatment methods based on the spatiotemporal changes in the characteristics of the myelin sheath, as well as the initiation, intersection and common action point of the pathophysiological mechanism, to finally achieve accurate treatment. We hope that this article can help researchers in this field to further clarify the challenges and opportunities in the current research on myelin sheath injury and treatment after SAH.

A redox-reactive delivery system via neural stem cell nanoencapsulation enhances white matter regeneration in intracerebral hemorrhage mice.

Intracerebral hemorrhage (ICH) poses a great threat to human health because of its high mortality and morbidity. Neural stem cell (NSC) transplantation is promising for treating white matter injury following ICH to promote functional recovery. However, reactive oxygen species (ROS)-induced NSC apoptosis and uncontrolled differentiation hindered the effectiveness of the therapy. Herein, we developed a single-cell nanogel system by layer-by-layer (LbL) hydrogen bonding of gelatin and tannic acid (TA), which was modified with a boronic ester-based compound linking triiodothyronine (T3). In vitro, NSCs in nanogel were protected from ROS-induced apoptosis, with apoptotic signaling pathways downregulated. This process of ROS elimination by material shell synergistically triggered T3 release to induce NSC differentiation into oligodendrocytes. Furthermore, in animal studies, ICH mice receiving nanogels performed better in behavioral evaluation, neurological scaling, and open field tests. These animals exhibited enhanced differentiation of NSCs into oligodendrocytes and promoted white matter tract regeneration on Day 21 through activation of the αvß3/PI3K/THRA pathway. Consequently, transplantation of LbL(T3) nanogels largely resolved two obstacles in NSC therapy synergistically: low survival and uncontrolled differentiation, enhancing white matter regeneration and behavioral performance of ICH mice. As expected, nanoencapsulation with synergistic effects would efficiently provide hosts with various biological benefits and minimize the difficulty in material fabrication, inspiring next-generation material design for tackling complicated pathological conditions.

NLRP3-dependent lipid droplet formation contributes to posthemorrhagic hydrocephalus by increasing the permeability of the blood-cerebrospinal fluid barrier in the choroid plexus.

Hydrocephalus is a severe complication that can result from intracerebral hemorrhage, especially if this hemorrhage extends into the ventricles. Our previous study indicated that the NLRP3 inflammasome mediates cerebrospinal fluid hypersecretion in the choroid plexus epithelium. However, the pathogenesis of posthemorrhagic hydrocephalus remains unclear, and therapeutic strategies for prevention and treatment are lacking. In this study, an Nlrp3-/- rat model of intracerebral hemorrhage with ventricular extension and primary choroid plexus epithelial cell culture were used to investigate the potential effects of NLRP3-dependent lipid droplet formation and its role in the pathogenesis of posthemorrhagic hydrocephalus. The data indicated that NLRP3-mediated dysfunction of the blood-cerebrospinal fluid barrier (B-CSFB) accelerated neurological deficits and hydrocephalus, at least in part, through the formation of lipid droplets in the choroid plexus; these lipid droplets interacted with mitochondria and increased the release of mitochondrial reactive oxygen species that destroyed tight junctions in the choroid plexus after intracerebral hemorrhage with ventricular extension. This study broadens the current understanding of the relationship among NLRP3, lipid droplets and the B-CSFB and provides a new therapeutic target for the treatment of posthemorrhagic hydrocephalus. Strategies to protect the B-CSFB may be effective therapeutic approaches for posthemorrhagic hydrocephalus.

Discovering hematoma-stimulated circuits for secondary brain injury after intraventricular hemorrhage by spatial transcriptome analysis.

Introduction: Central nervous system (CNS) diseases, such as neurodegenerative disorders and brain diseases caused by acute injuries, are important, yet challenging to study due to disease lesion locations and other complexities. Methods: Utilizing the powerful method of spatial transcriptome analysis together with novel algorithms we developed for the study, we report here for the first time a 3D trajectory map of gene expression changes in the brain following acute neural injury using a mouse model of intraventricular hemorrhage (IVH). IVH is a common and representative complication after various acute brain injuries with severe mortality and mobility implications. Results: Our data identified three main 3D global pseudospace-time trajectory bundles that represent the main neural circuits from the lateral ventricle to the hippocampus and primary cortex affected by experimental IVH stimulation. Further analysis indicated a rapid response in the primary cortex, as well as a direct and integrated effect on the hippocampus after IVH stimulation. Discussion: These results are informative for understanding the pathophysiological changes, including the spatial and temporal patterns of gene expression changes, in IVH patients after acute brain injury, strategizing more effective clinical management regimens, and developing novel bioinformatics strategies for the study of other CNS diseases. The algorithm strategies used in this study are searchable via a web service (www.combio-lezhang.online/3dstivh/home).

Knockout of transient receptor potential ankyrin 1 (TRPA1) modulates the glial phenotype and alleviates perihematomal neuroinflammation after intracerebral hemorrhage in mice via MAPK/NF-κB signaling.

The objective is to explore the role of astrocytic transient receptor potential ankyrin 1 (TRPA1) in glial phenotype transformation in neuroinflammation after intracerebral hemorrhage (ICH). Wild-type astrocytes and TRPA1-/- astrocytes were subjected to 6-h hemin treatment, and the calcium ions and transcriptome sequencing were assessed. A mouse autologous blood injection ICH model was established to evaluate the proliferation and phenotypes of astrocytes and microglia around the hematoma. The neuroinflammation and behavioral performance of wild-type ICH mice and TRPA1-/- ICH mice were assessed. Knockout of astrocytic TRPA1 decreased calcium ions of astrocytes after hemin treatment in-vitro, and microglial and astrocytes around the hematoma proliferated after the ICH model. Furthermore, RNA-sequencing (RNA-seq), immunofluorescence, and Western blotting results showed that the activated astrocytes transformed into the A2 phenotype in TRPA1-/- ICH mice. The 'ameboid' microglia were observed around the hematoma in TRPA1-/- ICH mice. The proliferation of A2 astrocytes and 'ameboid' microglia ameliorated the neuroinflammation after ICH. The inflammatory response was reduced by inhibiting the mitogen-activated protein kinase/nuclear factor kappa-B signaling pathway, and neurologic deficits were improved in TRPA1-/- ICH mice compared with wild-type ICH mice. This research suggests that astrocytic TRPA1 is a new therapeutic target to rescue neuroinflammation by modulating the glial phenotype after ICH.

EphA4/EphrinB2 signaling mediates pericyte-induced transient glia limitans formation as a secondary protective barrier after subarachnoid hemorrhage in mice.

BACKGROUND: Most patients with subarachnoid hemorrhage (SAH) do not exhibit brain parenchymal injury upon imaging but present significant blood-brain barrier (BBB) disruption and secondary neurological deficits. The aim of this study was to investigate whether stressed astrocytes act as a secondary barrier to exert a protective effect after SAH and to investigate the mechanism of glial limitan formation. METHODS: A total of 204 adult male C57BL/6 mice and an endovascular perforation SAH model were employed. The spatiotemporal characteristics of glial limitan formation after SAH were determined by immunofluorescence staining and transmission electron microscopy. The molecular mechanisms by which pericytes regulate glia limitans formation were analyzed using polymerase chain reaction, Western blotting, immunofluorescence staining and ELISA in a pericyte-astrocyte contact coculture system. The findings were validated ex vivo and in vivo using lentiviruses and inhibitors. Finally, pericytes were targeted to regulate glial limitan formation, and the effect of the glia limitans on secondary brain injury after SAH was evaluated by flow cytometry and analysis of neurological function. RESULTS: Stress-induced glial limitan formation occurred 1 day after SAH and markedly subsided 3 days after ictus. Pericytes regulated astrocyte glia limitan formation via EphA4/EphrinB2 signaling, inhibited inflammatory cell infiltration and altered neurological function. CONCLUSIONS: Astrocyte-derived glia limitans serve as a secondary protective barrier following BBB disruption after SAH in mice, and pericytes can regulate glial limitan formation and alter neurological function via EphA4/EphrinB2 signaling. Strategies for maintaining this secondary protective barrier may be novel treatment approaches for alleviating early brain injury after SAH.

Lipocalin-2-Mediated Insufficient Oligodendrocyte Progenitor Cell Remyelination for White Matter Injury After Subarachnoid Hemorrhage via SCL22A17 Receptor/Early Growth Response Protein 1 Signaling.

Insufficient remyelination due to impaired oligodendrocyte precursor cell (OPC) differentiation and maturation is strongly associated with irreversible white matter injury (WMI) and neurological deficits. We analyzed whole transcriptome expression to elucidate the potential role and underlying mechanism of action of lipocalin-2 (LCN2) in OPC differentiation and WMI and identified the receptor SCL22A17 and downstream transcription factor early growth response protein 1 (EGR1) as the key signals contributing to LCN2-mediated insufficient OPC remyelination. In LCN-knockdown and OPC EGR1 conditional-knockout mice, we discovered enhanced OPC differentiation in developing and injured white matter (WM); consistent with this, the specific inactivation of LCN2/SCl22A17/EGR1 signaling promoted remyelination and neurological recovery in both atypical, acute WMI due to subarachnoid hemorrhage and typical, chronic WMI due to multiple sclerosis. This potentially represents a novel strategy to enhance differentiation and remyelination in patients with white matter injury.

Secondary White Matter Injury and Therapeutic Targets After Subarachnoid Hemorrhage.

Aneurysmal subarachnoid hemorrhage (SAH) is one of the special stroke subtypes with high mortality and mobility. Although the mortality of SAH has decreased by 50% over the past two decades due to advances in neurosurgery and management of neurocritical care, more than 70% of survivors suffer from varying degrees of neurological deficits and cognitive impairments, leaving a heavy burden on individuals, families, and the society. Recent studies have shown that white matter is vulnerable to SAH, and white matter injuries may be one of the causes of long-term neurological deficits caused by SAH. Attention has recently focused on the pivotal role of white matter injury in the pathophysiological processes after SAH, mainly related to mechanical damage caused by increased intracerebral pressure and the metabolic damage induced by blood degradation and hypoxia. In the present review, we sought to summarize the pathophysiology processes and mechanisms of white matter injury after SAH, with a view to providing new strategies for the prevention and treatment of long-term cognitive dysfunction after SAH.

MiR-706 alleviates white matter injury via downregulating PKCα/MST1/NF-κB pathway after subarachnoid hemorrhage in mice.

Increasing numbers of patients with spontaneous subarachnoid hemorrhage(SAH) who recover from surgery and intensive care management still live with cognitive impairment after discharge, indicating the importance of white matter injury at the acute stage of SAH. In the present study, standard endovascular perforation was employed to establish an SAH mouse model, and a microRNA (miRNA) chip was used to analyze the changes in gene expression in white matter tissue after SAH. The data indicate that 17 miRNAs were downregulated, including miR-706, miR-669a-5p, miR-669p-5p, miR-7116-5p and miR-195a-3p, while 13 miRNAs were upregulated, including miR-6907-5p, miR-5135, miR-6982-5p, miR-668-5p, miR-8119. Strikingly, miR-706 was significantly downregulated with the highest fold change. Further experiments confirmed that miR-706 could alleviate white matter injury and improve neurological behavior, at least partially by inhibiting the PKCα/MST1/NF-κB pathway and the release of inflammatory cytokines. These results might provide a deeper understanding of the pathophysiological processes in white matter after SAH, as well as potential therapeutic strategies for the translational research.

Developing the novel bioinformatics algorithms to systematically investigate the connections among survival time, key genes and proteins for Glioblastoma multiforme.

BACKGROUND: Glioblastoma multiforme (GBM) is one of the most common malignant brain tumors and its average survival time is less than 1 year after diagnosis. RESULTS: Firstly, this study aims to develop the novel survival analysis algorithms to explore the key genes and proteins related to GBM. Then, we explore the significant correlation between AEBP1 upregulation and increased EGFR expression in primary glioma, and employ a glioma cell line LN229 to identify relevant proteins and molecular pathways through protein network analysis. Finally, we identify that AEBP1 exerts its tumor-promoting effects by mainly activating mTOR pathway in Glioma. CONCLUSIONS: We summarize the whole process of the experiment and discuss how to expand our experiment in the future.

Development of an Early Prediction Model for Subarachnoid Hemorrhage With Genetic and Signaling Pathway Analysis.

Subarachnoid hemorrhage (SAH) is devastating disease with high mortality, high disability rate, and poor clinical prognosis. It has drawn great attentions in both basic and clinical medicine. Therefore, it is necessary to explore the therapeutic drugs and effective targets for early prediction of SAH. Firstly, we demonstrate that LCN2 can effectively intervene or treat SAH from the perspective of cell signaling pathway. Next, three potential genes that we explored have been validated by manually reviewed experimental evidences. Finally, we turn out that the SAH early ensemble learning predictive model performs better than the classical LR, SVM, and Naïve-Bayes models.

Nrf1 Is Endowed with a Dominant Tumor-Repressing Effect onto the Wnt/ß-Catenin-Dependent and Wnt/ß-Catenin-Independent Signaling Networks in the Human Liver Cancer.

Our previous work revealed that Nrf1α exerts a tumor-repressing effect because its genomic loss (to yield Nrf1α-/- ) results in oncogenic activation of Nrf2 and target genes. Interestingly, ß-catenin is concurrently activated by loss of Nrf1α in a way similar to ß-catenin-driven liver tumor. However, a presumable relationship between Nrf1 and ß-catenin is not yet established. Here, we demonstrate that Nrf1 enhanced ubiquitination of ß-catenin for targeting proteasomal degradation. Conversely, knockdown of Nrf1 by its short hairpin RNA (shNrf1) caused accumulation of ß-catenin so as to translocate the nucleus, allowing activation of a subset of Wnt/ß-catenin signaling responsive genes, which leads to the epithelial-mesenchymal transition (EMT) and related cellular processes. Such silencing of Nrf1 resulted in malgrowth of human hepatocellular carcinoma, along with malignant invasion and metastasis to the lung and liver in xenograft model mice. Further transcriptomic sequencing unraveled significant differences in the expression of both Wnt/ß-catenin-dependent and Wnt/ß-catenin-independent responsive genes implicated in the cell process, shape, and behavior of the shNrf1-expressing tumor. Notably, we identified that ß-catenin is not a target gene of Nrf1, but this CNC-bZIP factor contributes to differential or opposing expression of other critical genes, such as CDH1, Wnt5A, Wnt11A, FZD10, LEF1, TCF4, SMAD4, MMP9, PTEN, PI3K, JUN, and p53, each of which depends on the positioning of distinct cis-regulatory sequences (e.g., ARE and/or AP-1 binding sites) in the gene promoter contexts. In addition, altered expression profiles of some Wnt/ß-catenin signaling proteins were context dependent, as accompanied by decreased abundances of Nrf1 in the clinic human hepatomas with distinct differentiation. Together, these results corroborate the rationale that Nrf1 acts as a bona fide dominant tumor repressor, by its intrinsic inhibition of Wnt/ß-catenin signaling and relevant independent networks in cancer development and malignant progression.

Unification of Opposites between Two Antioxidant Transcription Factors Nrf1 and Nrf2 in Mediating Distinct Cellular Responses to the Endoplasmic Reticulum Stressor Tunicamycin.

The water-soluble Nrf2 (nuclear factor, erythroid 2-like 2, also called Nfe2l2) is accepted as a master regulator of antioxidant responses to cellular stress, and it was also identified as a direct target of the endoplasmic reticulum (ER)-anchored PERK (protein kinase RNA-like endoplasmic reticulum kinase). However, the membrane-bound Nrf1 (nuclear factor, erythroid 2-like 1, also called Nfe2l1) response to ER stress remains elusive. Herein, we report a unity of opposites between these two antioxidant transcription factors, Nrf1 and Nrf2, in coordinating distinct cellular responses to the ER stressor tunicamycin (TU). The TU-inducible transcription of Nrf1 and Nrf2, as well as GCLM (glutamate cysteine ligase modifier subunit) and HO-1 (heme oxygenase 1), was accompanied by activation of ER stress signaling networks. Notably, the unfolded protein response (UPR) mediated by ATF6 (activating transcription factor 6), IRE1 (inositol requiring enzyme 1) and PERK was significantly suppressed by Nrf1-specific knockout, but hyper-expression of Nrf2 and its target genes GCLM and HO-1 has retained in Nrf1a-/- cells. By contrast, Nrf2-/-ΔTA cells with genomic deletion of its transactivation (TA) domain resulted in significant decreases of GCLM, HO-1 and Nrf1; this was accompanied by partial decreases of IRE1 and ATF6, rather than PERK, but with an increase of ATF4 (activating transcription factor 4). Interestingly, Nrf1 glycosylation and its trans-activity to mediate the transcriptional expression of the 26S proteasomal subunits, were repressed by TU. This inhibitory effect was enhanced by Nrf1a-/- and Nrf2-/-ΔTA, but not by a constitutive activator caNrf2ΔN (that increased abundances of the non-glycosylated and processed Nrf1). Furthermore, caNrf2ΔN also enhanced induction of PERK and IRE1 by TU, but reduced expression of ATF4 and HO-1. Thus, it is inferred that such distinct roles of Nrf1 and Nrf2 are unified to maintain cell homeostasis by a series of coordinated ER-to-nuclear signaling responses to TU. Nrf1 (i.e., a full-length form) acts in a cell-autonomous manner to determine the transcription of most of UPR-target genes, albeit Nrf2 is also partially involved in this process. Consistently, transactivation of ARE (antioxidant response element)-driven BIP (binding immunoglobulin protein)-, PERK- and XBP1 (X-box binding protein 1)-Luc reporter genes was mediated directly by Nrf1 and/or Nrf2. Interestingly, Nrf1 is more potent than Nrf2 at mediating the cytoprotective responses against the cytotoxicity of TU alone plus tBHQ (tert-butylhydroquinone). This is also further supported by the evidence that the intracellular reactive oxygen species (ROS) levels are increased in Nrf1a-/- cells, but rather are, to our surprise, decreased in Nrf2-/-ΔTA cells.

Distinct isoforms of Nrf1 diversely regulate different subsets of its cognate target genes.

The single Nrf1 gene has capability to be differentially transcripted alongside with alternative mRNA-splicing and subsequent translation through different initiation signals so as to yield distinct lengths of polypeptide isoforms. Amongst them, three of the most representatives are Nrf1α, Nrf1ß and Nrf1γ, but the putative specific contribution of each isoform to regulating ARE-driven target genes remains unknown. To address this, we have herein established three cell lines on the base of the Flp-In T-REx system, which are allowed for the tetracycline-inducibly stable expression of Nrf1α, Nrf1ß and Nrf1γ. Consequently, the RNA-Sequencing results have demonstrated that a vast majority of differentially expressed genes (i.e. >90% DEGs detected) were dominantly up-regulated by Nrf1α and/or Nrf1ß following induction by tetracycline. By contrast, the other DEGs regulated by Nrf1γ were far less than those regulated by Nrf1α/ß (i.e. ~11% of Nrf1α and ~7% of Nrf1ß). However, further transcriptomic analysis revealed that the tetracycline-induced expression of Nrf1γ significantly increased the percentage of down-regulated genes in total DEGs. These statistical data were further validated by quantitative real-time PCR. The experimental results indicate that distinct Nrf1 isoforms make diverse and even opposing contributions to regulating different subsets of target genes, such as those encoding 26S proteasomal subunits and others involved in various biological processes and functions. Collectively, Nrf1γ acts as a major dominant-negative inhibitor competitively against Nrf1α/ß activity, such that a number of DEGs regulated by Nrf1α/ß are counteracted by Nrf1γ.

Oncogenic Activation of Nrf2, Though as a Master Antioxidant Transcription Factor, Liberated by Specific Knockout of the Full-Length Nrf1α that Acts as a Dominant Tumor Repressor.

Liver-specific knockout of Nrf1 in the mouse leads to spontaneous development of non- alcoholic steatohepatitis with dyslipidemia, and then its deterioration results in hepatoma, but the underlying mechanism remains elusive to date. A similar pathological model is reconstructed here by using human Nrf1α-specific knockout cell lines. Our evidence has demonstrated that a marked increase of the inflammation marker COX2 definitely occurs in Nrf1α-/- cells. Loss of Nrf1α leads to hyperactivation of Nrf2, which results from substantial decreases in Keap1, PTEN and most of 26S proteasomal subunits in Nrf1α-/- cells. Further investigation of xenograft model mice showed that malignant growth of Nrf1α-/--derived tumors is almost abolished by silencing of Nrf2, while Nrf1α+/⁺-tumor is markedly repressed by an inactive mutant (i.e., Nrf2-/-ΔTA), but largely unaffected by a priori constitutive activator (i.e., caNrf2ΔN). Mechanistic studies, combined with transcriptomic sequencing, unraveled a panoramic view of opposing and unifying inter-regulatory cross-talks between Nrf1α and Nrf2 at different layers of the endogenous regulatory networks from multiple signaling towards differential expression profiling of target genes. Collectively, Nrf1α manifests a dominant tumor-suppressive effect by confining Nrf2 oncogenicity. Though as a tumor promoter, Nrf2 can also, in turn, directly activate the transcriptional expression of Nrf1 to form a negative feedback loop. In view of such mutual inter-regulation by between Nrf1α and Nrf2, it should thus be taken severe cautions to interpret the experimental results from loss of Nrf1α, Nrf2 or both.

Nexilin Regulates Oligodendrocyte Progenitor Cell Migration and Remyelination and Is Negatively Regulated by Protease-Activated Receptor 1/Ras-Proximate-1 Signaling Following Subarachnoid Hemorrhage.

Progressive white matter (WM) impairments caused by subarachnoid hemorrhage (SAH) contribute to cognitive deficits and poor clinical prognoses; however, their pathogenetic mechanisms are poorly understood. We investigated the role of nexilin and oligodendrocyte progenitor cell (OPC)-mediated repair in a mouse model of experimental SAH generated via left endovascular perforation. Nexilin expression was enhanced by the elevated migration of OPCs after SAH. Knocking down nexilin by siRNA reduced OPC migration both in vitro and in vivo and abridged WM repair. In contrast, the protease-activated receptor 1 (PAR1), Ras-proximate-1 (RAP1) and phosphorylated RAP1 (pRAP1) levels in WM were elevated after SAH. The genetic inhibition of PAR1 reduced RAP1 and pRAP1 expression, further enhancing nexilin expression. When delivered at an early stage at a concentration of 25 µg/kg, thrombin receptor antagonist peptide along with PAR1 knockdown rescued the down-regulation of myelin basic protein and improved remyelination at the later stage of SAH. Our results suggest that nexilin is required for OPC migration and remyelination following SAH, as it negatively regulates PAR1/RAP1 signaling, thus providing a promising therapeutic target in WM repair and functional recovery.