Perilipin-2 mediates ferroptosis in oligodendrocyte progenitor cells and myelin injury after ischemic stroke.

JOURNAL/nrgr/04.03/01300535-202507000-00024/figure1/v/2024-09-09T124005Z/r/image-tiff Differentiation of oligodendrocyte progenitor cells into mature myelin-forming oligodendrocytes contributes to remyelination. Failure of remyelination due to oligodendrocyte progenitor cell death can result in severe nerve damage. Ferroptosis is an iron-dependent form of regulated cell death caused by membrane rupture induced by lipid peroxidation, and plays an important role in the pathological process of ischemic stroke. However, there are few studies on oligodendrocyte progenitor cell ferroptosis. We analyzed transcriptome sequencing data from GEO databases and identified a role of ferroptosis in oligodendrocyte progenitor cell death and myelin injury after cerebral ischemia. Bioinformatics analysis suggested that perilipin-2 (PLIN2) was involved in oligodendrocyte progenitor cell ferroptosis. PLIN2 is a lipid storage protein and a marker of hypoxia-sensitive lipid droplet accumulation. For further investigation, we established a mouse model of cerebral ischemia/reperfusion. We found significant myelin damage after cerebral ischemia, as well as oligodendrocyte progenitor cell death and increased lipid peroxidation levels around the infarct area. The ferroptosis inhibitor, ferrostatin-1, rescued oligodendrocyte progenitor cell death and subsequent myelin injury. We also found increased PLIN2 levels in the peri-infarct area that co-localized with oligodendrocyte progenitor cells. Plin2 knockdown rescued demyelination and improved neurological deficits. Our findings suggest that targeting PLIN2 to regulate oligodendrocyte progenitor cell ferroptosis may be a potential therapeutic strategy for rescuing myelin damage after cerebral ischemia.

Unveiling the impact of SUMOylation at K298 site of heat shock factor 1 on glioblastoma malignant progression.

BACKGROUND: Glioblastoma (GBM) poses a significant medical challenge due to its aggressive nature and poor prognosis. Mitochondrial unfolded protein response (UPRmt) and the heat shock factor 1 (HSF1) pathway play crucial roles in GBM pathogenesis. Post-translational modifications, such as SUMOylation, regulate the mechanism of action of HSF1 and may influence the progression of GBM. Understanding the interplay between SUMOylation-modified HSF1 and GBM pathophysiology is essential for developing targeted therapies. METHODS: We conducted a comprehensive investigation using cellular, molecular, and in vivo techniques. Cell culture experiments involved establishing stable cell lines, protein extraction, Western blotting, co-immunoprecipitation, and immunofluorescence analysis. Mass spectrometry was utilized for protein interaction studies. Computational modeling techniques were employed for protein structure analysis. Plasmid construction and lentiviral transfection facilitated the manipulation of HSF1 SUMOylation. In vivo studies employed xenograft models for tumor growth assessment. RESULTS: Our research findings indicate that HSF1 primarily undergoes SUMOylation at the lysine residue K298, enhancing its nuclear translocation, stability, and downstream heat shock protein expression, while having no effect on its trimer conformation. SUMOylated HSF1 promoted the UPRmt pathway, leading to increased GBM cell proliferation, migration, invasion, and reduced apoptosis. In vivo studies have confirmed that SUMOylation of HSF1 enhances its oncogenic effect in promoting tumor growth in GBM xenograft models. CONCLUSION: This study elucidates the significance of SUMOylation modification of HSF1 in driving GBM progression. Targeting SUMOylated HSF1 may offer a novel therapeutic approach for GBM treatment. Further investigation into the specific molecular mechanisms influenced by SUMOylated HSF1 is warranted for the development of effective targeted therapies to improve outcomes for GBM patients.

Prognostic prediction and immune checkpoint profiling in glioma patients through neddylation-associated features.

BACKGROUND: Gliomas are the most common primary malignant tumours of the central nervous system, and neddylation may be a potential target for the treatment of gliomas. Our study analysed neddylation's potential role in gliomas of different pathological types and its correlation with immunotherapy. METHODS: Genes required for model construction were sourced from existing literature, and their expression data were extracted from the TCGA and CGGA databases. LASSO regression was employed to identify genes associated with the prognosis of glioma patients in TCGA and to establish a clinical prognostic model. Biological changes in glioma cell lines following intervention with hub genes were evaluated using the CCK-8 assay and transwell assay. The genes implicated in the model construction were validated across various cell lines using Western blot. We conducted analyses to examine correlations between model scores and clinical data, tumor microenvironments, and immune checkpoints. Furthermore, we investigated potential differences in molecular functions and mechanisms among different groups. RESULTS: We identified 249 genes from the Reactome database and analysed their expression profiles in the TCGA and CGGA databases. After using LASSO-Cox, four genes (BRCA1, BIRC5, FBXL16 and KLHL25, p < 0.05) with significant correlations were identified. We selected FBXL16 for validation in in vitro experiments. Following FBXL16 overexpression, the proliferation, migration, and invasion abilities of glioma cell lines all showed a decrease. Then, we constructed the NEDD Index for gliomas. The nomogram indicated that this model could serve as an independent prognostic marker. Analysis of the tumour microenvironment and immune checkpoints revealed that the NEDD index was also correlated with immune cell infiltration and the expression levels of various immune checkpoints. CONCLUSION: The NEDD index can serve as a practical tool for predicting the prognosis of glioma patients, and it is correlated with immune cell infiltration and the expression levels of immune checkpoints.

The role of nitric oxide and hydrogen sulfide in spinal cord injury: an updated review.

Medical gases play an important role in the pathophysiology of human diseases and have received extensive attention for their role in neuroprotection. Common pathological mechanisms of spinal cord injury include excitotoxicity, inflammation, cell death, glial scarring, blood-spinal cord barrier disruption, and ischemia/reperfusion injury. Nitric oxide and hydrogen sulfide are important gaseous signaling molecules in living organisms; their pathological role in spinal cord injury models has received more attention in recent years. This study reviews the possible mechanisms of spinal cord injury and the role of nitric oxide and hydrogen sulfide in spinal cord injury.

Dystrophin 71 deficiency causes impaired aquaporin-4 polarization contributing to glymphatic dysfunction and brain edema in cerebral ischemia.

OBJECTIVE: The glymphatic system serves as a perivascular pathway that aids in clearing liquid and solute waste from the brain, thereby enhancing neurological function. Disorders in glymphatic drainage contribute to the development of vasogenic edema following cerebral ischemia, although the molecular mechanisms involved remain poorly understood. This study aims to determine whether a deficiency in dystrophin 71 (DP71) leads to aquaporin-4 (AQP4) depolarization, contributing to glymphatic dysfunction in cerebral ischemia and resulting in brain edema. METHODS: A mice model of middle cerebral artery occlusion and reperfusion was used. A fluorescence tracer was injected into the cortex and evaluated glymphatic clearance. To investigate the role of DP71 in maintaining AQP4 polarization, an adeno-associated virus with the astrocyte promoter was used to overexpress Dp71. The expression and distribution of DP71 and AQP4 were analyzed using immunoblotting, immunofluorescence, and co-immunoprecipitation techniques. The behavior ability of mice was evaluated by open field test. Open-access transcriptome sequencing data were used to analyze the functional changes of astrocytes after cerebral ischemia. MG132 was used to inhibit the ubiquitin-proteasome system. The ubiquitination of DP71 was detected by immunoblotting and co-immunoprecipitation. RESULTS: During the vasogenic edema stage following cerebral ischemia, a decline in the efflux of interstitial fluid tracer was observed. DP71 and AQP4 were co-localized and interacted with each other in the perivascular astrocyte endfeet. After cerebral ischemia, there was a notable reduction in DP71 protein expression, accompanied by AQP4 depolarization and proliferation of reactive astrocytes. Increased DP71 expression restored glymphatic drainage and reduced brain edema. AQP4 depolarization, reactive astrocyte proliferation, and the behavior of mice were improved. After cerebral ischemia, DP71 was degraded by ubiquitination, and MG132 inhibited the decrease of DP71 protein level. CONCLUSION: AQP4 depolarization after cerebral ischemia leads to glymphatic clearance disorder and aggravates cerebral edema. DP71 plays a pivotal role in regulating AQP4 polarization and consequently influences glymphatic function. Changes in DP71 expression are associated with the ubiquitin-proteasome system. This study offers a novel perspective on the pathogenesis of brain edema following cerebral ischemia.

Coxsackievirus A10 impairs nail regeneration and induces onychomadesis by mimicking DKK1 to attenuate Wnt signaling.

Coxsackievirus A10 (CV-A10) infection, a prominent cause of childhood hand-foot-and-mouth disease (HFMD), frequently manifests with the intriguing phenomenon of onychomadesis, characterized by nail shedding. However, the underlying mechanism is elusive. Here, we found that CV-A10 infection in mice could suppress Wnt/ß-catenin signaling by restraining LDL receptor-related protein 6 (LRP6) phosphorylation and ß-catenin accumulation and lead to onychomadesis. Mechanistically, CV-A10 mimics Dickkopf-related protein 1 (DKK1) to interact with Kringle-containing transmembrane protein 1 (KRM1), the CV-A10 cellular receptor. We further found that Wnt agonist (GSK3ß inhibitor) CHIR99021 can restore nail stem cell differentiation and protect against nail shedding. These findings provide novel insights into the pathogenesis of CV-A10 and related viruses in onychomadesis and guide prognosis assessment and clinical treatment of the disease.

Targeting miR-31 represses tumourigenesis and dedifferentiation of BRAFV600E-associated thyroid carcinoma.

BACKGROUND: BRAFV600E is the most common genetic mutation in differentiated thyroid cancer (DTC) occurring in 60% of patients and drives malignant tumour cell phenotypes including proliferation, metastasis and immune-escape. BRAFV600E-mutated papillary thyroid cancer (PTC) also displays greatly reduced expression of thyroid differentiation markers, thus tendency to radioactive iodine (RAI) refractory and poor prognosis. Therefore, understanding the molecular mechanisms and main oncogenic events underlying BRAFV600E will guide future therapy development. METHODS: Bioinformatics and clinical specimen analyses, genetic manipulation of BRAFV600E-induced PTC model, functional and mechanism exploration guided with transcriptomic screening, as well as systematic rescue experiments were applied to investigate miR-31 function within BRAFV600E-induced thyroid cancer development. Besides, nanoparticles carrying miR-31 antagomirs were testified to alleviate 131I iodide therapy on PTC models. RESULTS: We identify miR-31 as a significantly increased onco-miR in BRAFV600E-associated PTC that promotes tumour progression, metastasis and RAI refractoriness via sustained Wnt/ß-catenin signalling. Mechanistically, highly activated BRAF/MAPK pathway induces miR-31 expression via c-Jun-mediated transcriptional regulation across in vitro and transgenic mouse models. MiR-31 in turn facilitates ß-catenin stabilisation via directly repressing tumour suppressors CEBPA and DACH1, which direct the expression of multiple essential Wnt/ß-catenin pathway inhibitors. Genetic functional assays showed that thyroid-specific knockout of miR-31 inhibited BRAFV600E-induced PTC progression, and strikingly, enhanced expression of sodium-iodide symporter and other thyroid differentiation markers, thus promoted 131I uptake. Nanoparticle-mediated application of anti-miR-31 antagomirs markedly elevated radio-sensitivity of BRAFV600E-induced PTC tumours to 131I therapy, and efficiently suppressed tumour progression in the pre-clinical mouse model. CONCLUSIONS: Our findings elucidate a novel BRAF/MAPK-miR-31-Wnt/ß-catenin regulatory mechanism underlying clinically BRAFV600E-associated DTC tumourigenesis and dedifferentiation, also highlight a potential adjuvant therapeutic strategy for advanced DTC.

SELENOI Functions as a Key Modulator of Ferroptosis Pathway in Colitis and Colorectal Cancer.

Ferroptosis plays important roles both in normal physiology and multiple human diseases. It is well known that selenoprotein named glutathione peroxidase 4 (GPX4) is a crucial regulator for ferroptosis. However, it remains unknown whether other selenoproteins responsible for the regulation of ferroptosis, particularly in gut diseases. In this study, it is observed that Selenoprotein I (Selenoi) prevents ferroptosis by maintaining ether lipids homeostasis. Specific deletion of Selenoi in intestinal epithelial cells induced the occurrence of ferroptosis, leading to impaired intestinal regeneration and compromised colonic tumor growth. Mechanistically, Selenoi deficiency causes a remarkable decrease in ether-linked phosphatidylethanolamine (ePE) and a marked increase in ether-linked phosphatidylcholine (ePC). The imbalance of ePE and ePC results in the upregulation of phospholipase A2, group IIA (Pla2g2a) and group V (Pla2g5), as well as arachidonate-15-lipoxygenase (Alox15), which give rise to excessive lipid peroxidation. Knockdown of PLA2G2A, PLA2G5, or ALOX15 can reverse the ferroptosis phenotypes, suggesting that they are downstream effectors of SELENOI. Strikingly, GPX4 overexpression cannot rescue the ferroptosis phenotypes of SELENOI-knockdown cells, while SELENOI overexpression can partially rescue GPX4-knockdown-induced ferroptosis. It suggests that SELENOI prevents ferroptosis independent of GPX4. Taken together, these findings strongly support the notion that SELENOI functions as a novel suppressor of ferroptosis during colitis and colon tumorigenesis.

ANKZF1 knockdown inhibits glioblastoma progression by promoting intramitochondrial protein aggregation through mitoRQC.

Protein homeostasis is fundamental to the development of tumors. Ribosome-associated quality-control (RQC) is able to add alanine and threonine to the stagnant polypeptide chain C-terminal (CAT-tail) when protein translation is hindered, while Ankyrin repeat and zinc-finger domain-containing-protein 1 (ANKZF1) can counteract the formation of the CAT-tail, preventing the aggregation of polypeptide chains. In particular, ANKZF1 plays an important role in maintaining mitochondrial protein homeostasis by mitochondrial RQC (mitoRQC) after translation stagnation of precursor proteins targeting mitochondria. However, the role of ANKZF1 in glioblastoma is unclear. Therefore, the current study was aimed to investigate the effects of ANKZF1 in glioblastoma cells and a nude mouse glioblastoma xenograft model. Here, we reported that knockdown of ANKZF1 in glioblastoma cells resulted in the accumulation of CAT-tail in mitochondria, leading to the activated mitochondrial unfolded protein response (UPRmt) and inhibits glioblastoma malignant progression. Excessive CAT-tail sequestered mitochondrial chaperones HSP60, mtHSP70 and proteases LONP1 as well as mitochondrial respiratory chain subunits ND1, Cytb, mtCO2 and ATP6, leading to mitochondrial oxidative phosphorylation dysfunction, membrane potential impairment, and mitochondrial apoptotic pathway activation. Our study highlights ANKZF1 as a valuable target for glioblastoma intervention and provides an innovative insight for the treatment of glioblastoma through the regulating of mitochondrial protein homeostasis.

mTORC2-AKT signaling to PFKFB2 activates glycolysis that enhances stemness and tumorigenicity of intestinal epithelial cells.

Although elevated glycolysis has been widely recognized as a hallmark for highly proliferating cells like stem cells and cancer, its regulatory mechanisms are still being updated. Here, we found a previously unappreciated mechanism of mammalian target of rapamycin complex 2 (mTORC2) in regulating glycolysis in intestinal stem cell maintenance and cancer progression. mTORC2 key subunits expression levels and its kinase activity were specifically upregulated in intestinal stem cells, mouse intestinal tumors, and human colorectal cancer (CRC) tissues. Genetic ablation of its key scaffolding protein Rictor in both mouse models and cell lines revealed that mTORC2 played an important role in promoting intestinal stem cell proliferation and self-renewal. Moreover, utilizing mouse models and organoid culture, mTORC2 loss of function was shown to impair growth of gut adenoma and tumor organoids. Based on these findings, we performed RNA-seq and noticed significant metabolic reprogramming in Rictor conditional knockout mice. Among all the pathways, carbohydrate metabolism was most profoundly altered, and further studies demonstrated that mTORC2 promoted glycolysis in intestinal epithelial cells. Most importantly, we showed that a rate-limiting enzyme in regulating glycolysis, 6-phosphofructo-2-kinase (PFKFB2), was a direct target for the mTORC2-AKT signaling. PFKFB2 was phosphorylated upon mTORC2 activation, but not mTORC1, and this process was AKT-dependent. Together, this study has identified a novel mechanism underlying mTORC2 activated glycolysis, offering potential therapeutic targets for treating CRC.

Dysregulation of LINC00324 promotes poor prognosis in patients with glioma.

BACKGROUND: LINC00324 is a long-stranded non-coding RNA, which is aberrantly expressed in various cancers and is associated with poor prognosis and clinical features. It involves multiple oncogenic molecular pathways affecting cell proliferation, migration, invasion, and apoptosis. However, the expression, function, and mechanism of LINC00324 in glioma have not been reported. MATERIAL AND METHODS: We assessed the expression of LINC00324 of LINC00324 in glioma patients based on data from The Cancer Genome Atlas (TCGA) and Genotype-Tissue Expression (GTEx) to identify pathways involved in LINC00324-related glioma pathogenesis. RESULTS: Based on our findings, we observed differential expression of LINC00324 between tumor and normal tissues in glioma patients. Our analysis of overall survival (OS) and disease-specific survival (DSS) indicated that glioma patients with high LINC00324 expression had a poorer prognosis compared to those with low LINC00324 expression. By integrating clinical data and genetic signatures from TCGA patients, we developed a nomogram to predict OS and DSS in glioma patients. Gene set enrichment analysis (GSEA) revealed that several pathways, including JAK/STAT3 signaling, epithelial-mesenchymal transition, STAT5 signaling, NF-κB activation, and apoptosis, were differentially enriched in glioma samples with high LINC00324 expression. Furthermore, we observed significant correlations between LINC00324 expression, immune infiltration levels, and expression of immune checkpoint-related genes (HAVCR2: r = 0.627, P = 1.54e-77; CD40: r = 0.604, P = 1.36e-70; ITGB2: r = 0.612, P = 6.33e-7; CX3CL1: r = -0.307, P = 9.24e-17). These findings highlight the potential significance of LINC00324 in glioma progression and suggest avenues for further research and potential therapeutic targets. CONCLUSION: Indeed, our results confirm that the LINC00324 signature holds promise as a prognostic predictor in glioma patients. This finding opens up new possibilities for understanding the disease and may offer valuable insights for the development of targeted therapies.

Cathepsin B as a key regulator of ferroptosis in microglia following intracerebral hemorrhage.

Intracerebral hemorrhage (ICH) is a subtype of stroke marked by elevated mortality and disability rates. Recently, mounting evidence suggests a significant role of ferroptosis in the pathogenesis of ICH. Through a combination of bioinformatics analysis and basic experiments, our goal is to identify the primary cell types and key molecules implicated in ferroptosis post-ICH. This aims to propel the advancement of ferroptosis research, offering potential therapeutic targets for ICH treatment. Our study reveals pronounced ferroptosis in microglia and identifies the target gene, cathepsin B (Ctsb), by analyzing differentially expressed genes following ICH. Ctsb, a cysteine protease primarily located in lysosomes, becomes a focal point in our investigation. Utilizing in vitro and in vivo models, we explore the correlation between Ctsb and ferroptosis in microglia post-ICH. Results demonstrate that ICH and hemin-induced ferroptosis in microglia coincide with elevated levels and activity of Ctsb protein. Effective alleviation of ferroptosis in microglia after ICH is achieved through the inhibition of Ctsb protease activity and protein levels using inhibitors and shRNA. Additionally, a notable increase in m6A methylation levels of Ctsb mRNA post-ICH is observed, suggesting a pivotal role of m6A methylation in regulating Ctsb translation. These research insights deepen our comprehension of the molecular pathways involved in ferroptosis after ICH, underscoring the potential of Ctsb as a promising target for mitigating brain damage resulting from ICH.

ERBB1 alleviates secondary brain injury induced by experimental intracerebral hemorrhage in rats by modulating neuronal death via PLC-γ/PKC pathway.

AIMS: Intracerebral hemorrhage (ICH) is a disease with high rates of disability and mortality. The role of epidermal growth factor receptor 1 (ERBB1) in ICH was elucidated in this study. METHODS: ICH model was constructed by injecting autologous arterial blood into the right basal ganglia. The protein level of ERBB1 was detected by western blot analysis. To up- and downregulation of ERBB1 in rats, intraventricular injection of a lentivirus overexpression vector of ERBB1 and AG1478 (a specific inhibitor of ERBB1) was used. The cell apoptosis, neuronal loss, and pro-inflammatory cytokines were assessed by TUNEL, Nissl staining, and ELISA. Meanwhile, behavioral cognitive impairment of ICH rats was evaluated after ERBB1-targeted interventions. RESULTS: ERBB1 increased significantly in brain tissue of ICH rats. Overexpression of ERBB1 remarkably reduced cell apoptosis and neuronal loss induced by ICH, as well as pro-inflammatory cytokines and oxidative stress. Meanwhile, the behavioral and cognitive impairment of ICH rats were alleviated after upregulation of ERBB1; however, the secondary brain injury (SBI) was aggravated by AG1478 treatment. Furthermore, the upregulation of PLC-γ and PKC in ICH rats was reversed by AG1478 treatment. CONCLUSIONS: ERBB1 can improve SBI and has a neuroprotective effect in experimental ICH rats via PLC-γ/PKC pathway.

RACK1 promotes autophagy via the PERK signaling pathway to protect against traumatic brain injury in rats.

AIMS: Neuronal cell death is a primary factor that determines the outcome after traumatic brain injury (TBI). We previously revealed the importance of receptor for activated C kinase (RACK1), a multifunctional scaffold protein, in maintaining neuronal survival after TBI, but the specific mechanism remains unclear. The aim of this study was to explore the mechanism underlying RACK1-mediated neuroprotection in TBI. METHODS: TBI model was established using controlled cortical impact injury in Sprague-Dawley rats. Genetic intervention and pharmacological inhibition of RACK1 and PERK-autophagy signaling were administrated by intracerebroventricular injection. Western blotting, coimmunoprecipitation, transmission electron microscopy, real-time PCR, immunofluorescence, TUNEL staining, Nissl staining, neurobehavioral tests, and contusion volume assessment were performed. RESULTS: Endogenous RACK1 was upregulated and correlated with autophagy induction after TBI. RACK1 knockdown markedly inhibited TBI-induced autophagy, whereas RACK1 overexpression exerted the opposite effects. Moreover, RACK1 overexpression ameliorated neuronal apoptosis, neurological deficits, and cortical tissue loss after TBI, and these effects were abrogated by the autophagy inhibitor 3-methyladenine or siRNAs targeting Beclin1 and Atg5. Mechanistically, RACK1 interacted with PERK and activated PERK signaling. Pharmacological and genetic inhibition of the PERK pathway abolished RACK1-induced autophagy after TBI. CONCLUSION: Our findings indicate that RACK1 protected against TBI-induced neuronal damage partly through autophagy induction by regulating the PERK signaling pathway.

Acetylation of mtHSP70 at Lys595/653 affecting its interaction between GrpEL1 regulates glioblastoma progression via UPRmt.

BACKGROUND: The mitochondrial unfolded protein response (UPRmt) is a vital biological process that regulates mitochondrial protein homeostasis and enables glioblastoma cells to cope with mitochondrial oxidative stress in the tumor microenvironment. We previously reported that the binding of mitochondrial stress-70 protein (mtHSP70) to GrpE protein homolog 1 (GrpEL1) is involved in the regulation of the UPRmt. However, the mechanisms regulating their binding remain unclear. Herein, we examined the UPRmt in glioblastoma and explored whether modulating the interaction between mtHSP70 and GrpEL1 affects the UPRmt. METHODS: Western blot analysis, aggresome staining, and transmission electron microscopy were used to detect the activation of the UPRmt and protein aggregates within mitochondria. Molecular dynamics simulations were performed to investigate the impact of different mutations in mtHSP70 on its binding to GrpEL1. Endogenous site-specific mutations were introduced into mtHSP70 in glioblastoma cells using CRISPR/Cas9. In vitro and in vivo experiments were conducted to assess mitochondrial function and glioblastoma progression. RESULTS: The UPRmt was activated in glioblastoma cells in response to oxidative stress. mtHSP70 regulated mitochondrial protein homeostasis by facilitating UPRmt-progress protein import into the mitochondria. Acetylation of mtHSP70 at Lys595/653 enhanced its binding to GrpEL1. Missense mutations at Lys595/653 increased mitochondrial protein aggregates and inhibited glioblastoma progression in vitro and in vivo. CONCLUSIONS: We identified an innovative mechanism in glioblastoma progression by which acetylation of mtHSP70 at Lys595/653 influences its interaction with GrpEL1 to regulate the UPRmt. Mutations at Lys595/653 in mtHSP70 could potentially serve as therapeutic targets and prognostic indicators of glioblastoma.

Dehydroevodiamine ameliorates neurological dysfunction after traumatic brain injury in mice via regulating the SIRT1/FOXO3a/Bim pathway.

BACKGROUND: Traumatic Brain Injury (TBI) poses a considerable public health challenge, resulting in mortality, disability, and economic strain. Dehydroevodiamine (DEDM) is a natural compound derived from a traditional Chinese herbal medicine. Prior studies have substantiated the neuroprotective attributes of this compound in the context of TBI. Nevertheless, a comprehensive comprehension of the exact mechanisms responsible for its neuroprotective effects remains elusive. It is imperative to elucidate the precise intrinsic mechanisms underlying the neuroprotective actions of DEDM. PURPOSE: The aim of this investigation was to elucidate the mechanism underlying DEDM treatment in TBI utilizing both in vivo and in vitro models. Specifically, our focus was on comprehending the impact of DEDM on the Sirtuin1 (SIRT1) / Forkhead box O3 (FOXO3a) / Bcl-2-like protein 11 (Bim) pathway, a pivotal player in TBI-induced cell death attributed to oxidative stress. STUDY DESIGN AND METHODS: We established a TBI mouse model via the weight drop method. Following continuous intraperitoneal administration, we assessed the neurological dysfunction using the Modified Neurological Severity Score (mNSS) and behavioral assay, followed by sample collection. Secondary brain damage in mice was evaluated through Nissl staining, brain water content measurement, Evans blue detection, and Western blot assays. We scrutinized the expression levels of oxidative stress-related indicators and key proteins for apoptosis. The intricate mechanism of DEDM in TBI was further explored through immunofluorescence, Co-immunoprecipitation (Co-IP) assays, real-time quantitative PCR (RT-qPCR), dual-luciferase assays and western blotting. Additionally, we further investigated the specific therapeutic mechanism of DEDM in an oxidative stress cell model. RESULTS: The results indicated that DEDM effectively ameliorated oxidative stress and apoptosis post-TBI, mitigating neurological dysfunction and brain injury in mice. DEDM facilitated the deacetylation of FOXO3a by up-regulating the expression of the deacetylase SIRT1, consequently suppressing Bim expression. This mechanism contributed to the alleviation of neurological injury and symptom improvement in TBI-afflicted mice. Remarkably, SIRT1 emerged as a central mediator in the overall treatment mechanism. CONCLUSIONS: DEDM exerted significant neuroprotective effects on TBI mice by modulating the SIRT1/FOXO3a/Bim pathway. Our innovative research provides a basis for further exploration of the clinical therapeutic potential of DEDM in the context of TBI.

Excessive nucleic acid R-loops induce mitochondria-dependent epithelial cell necroptosis and drive spontaneous intestinal inflammation.

Oxidative stress, which can be activated by a variety of environmental risk factors, has been implicated as an important pathogenic factor for inflammatory bowel disease (IBD). However, how oxidative stress drives IBD onset remains elusive. Here, we found that oxidative stress was strongly activated in inflamed tissues from both ulcerative colitis patients and Crohn's disease patients, and it caused nuclear-to-cytosolic TDP-43 transport and a reduction in the TDP-43 protein level. To investigate the function of TDP-43 in IBD, we inducibly deleted exons 2 to 3 of Tardbp (encoding Tdp-43) in mouse intestinal epithelium, which disrupted its nuclear localization and RNA-processing function. The deletion gave rise to spontaneous intestinal inflammation by inducing epithelial cell necroptosis. Suppression of the necroptotic pathway with deletion of Mlkl or the RIP1 inhibitor Nec-1 rescued colitis phenotypes. Mechanistically, disruption of nuclear TDP-43 caused excessive R-loop accumulation, which triggered DNA damage and genome instability and thereby induced PARP1 hyperactivation, leading to subsequent NAD+ depletion and ATP loss, consequently activating mitochondrion-dependent necroptosis in intestinal epithelial cells. Importantly, restoration of cellular NAD+ levels with NAD+ or NMN supplementation, as well as suppression of ALKBH7, an α-ketoglutarate dioxygenase in mitochondria, rescued TDP-43 deficiency-induced cell death and intestinal inflammation. Furthermore, TDP-43 protein levels were significantly inversely correlated with γ-H2A.X and p-MLKL levels in clinical IBD samples, suggesting the clinical relevance of TDP-43 deficiency-induced mitochondrion-dependent necroptosis. Taken together, these findings identify a unique pathogenic mechanism that links oxidative stress to intestinal inflammation and provide a potent and valid strategy for IBD intervention.