Neuroprotective Response Research Articles

Interspecies Organoids Reveal Human-Specific Molecular Features of Dopaminergic Neuron Development and Vulnerability.

The disproportionate expansion of telencephalic structures during human evolution involved tradeoffs that imposed greater connectivity and metabolic demands on midbrain dopaminergic neurons. Despite the central role of dopaminergic neurons in human-enriched disorders, molecular specializations associated with human-specific features and vulnerabilities of the dopaminergic system remain unexplored. Here, we establish a phylogeny-in-a-dish approach to examine gene regulatory evolution by differentiating pools of human, chimpanzee, orangutan, and macaque pluripotent stem cells into ventral midbrain organoids capable of forming long-range projections, spontaneous activity, and dopamine release. We identify human-specific gene expression changes related to axonal transport of mitochondria and reactive oxygen species buffering and candidate cis- and trans -regulatory mechanisms underlying gene expression divergence. Our findings are consistent with a model of evolved neuroprotection in response to tradeoffs related to brain expansion and could contribute to the discovery of therapeutic targets and strategies for treating disorders involving the dopaminergic system.

Docosahexaenoic acid protects against ischemic stroke in diabetic mice by inhibiting inflammatory responses and apoptosis

This study was to explore whether docosahexaenoic acid (DHA) protects against ischemic stroke in diabetic mice and its mechanisms. DHA was administered to mice and its effects on stroke outcomes in type 1 diabetes mellitus were assessed 24 h and 3 days post-reperfusion using RNA sequencing, flow cytometry, multiplex immunoassays, and western-blotting analysis. In diabetic mice, DHA administration post-ischemic stroke significantly reduced cerebral infarct size, brain edema, and neurological impairments. Flow cytometric analysis demonstrated a notable decrease in the percentage of neutrophils in the ischemic brain, suggesting a mitigated inflammatory response. Western blotting assay revealed that pro-apoptotic protein Bax was reduced whereas anti-apoptotic protein Bcl-2 was increased, indicating the attenuation of apoptosis. Additionally, RNA sequencing of brain tissue highlighted significant transcriptomic changes, with downregulation of genes for several inflammatory pathways such as NF-kappa B signaling and upregulation of genes for neuroprotective pathways such as neuroactive ligand-receptor interaction. Similar transcriptomic changes in peripheral blood mononuclear cells indicated that DHA treatment resulted the systemic anti-inflammatory and neuroprotective response. DHA treatment mitigated cerebral ischemic injuries by dampening inflammatory responses and apoptosis in diabetic mice after ischemic stroke, highlighting its therapeutic potential for clinical management of stroke in diabetic patients.

CB1 Receptor Activation Provides Neuroprotection in an Animal Model of Glutamate-Induced Excitotoxicity Through a Reduction of NOX-2 Activity and Oxidative Stress.

Excitotoxicity is a process in which NADPH oxidase-2 (NOX-2) plays a pivotal role in the generation of reactive oxygen species (ROS). Oxidative stress influences the expression of Aquaporin 4 (AQP4), a water channel implicated in blood-brain barrier (BBB) permeability and edema formation. The endocannabinoid system is widely distributed in the brain, particularly through the cannabinoid receptor type 1 (CB1) and type 2 (CB2), which have been shown to have a neuroprotective function in brain injury. Given the significant involvement of NOX-2 in ROS production during excitotoxicity, our research aims to assess the participation of NOX-2 in the neuroprotective effect of the cannabinoid receptor agonist WIN55,212-2 against glutamate-induced excitotoxicity damage in the striatum using invivo model. Wild-type mice (C57BL/6) and NOX-2 KO (gp91Cybbtm1Din/J) were stereotactically injected in the striatum with monosodium glutamate or vehicle. Subsequently, a group of mice was administered an intraperitoneal dose of WIN55,212-2, AM251, or AM251/WIN55,212-2 following the intracerebral injection. Motor activity was assessed, and the lesion was examined through histological sections stained with cresyl violet. Additionally, brain water content and Evans blue assay were conducted. The activity of NOX was quantified, and the protein expression of CB1, gp91phox, AQP4, Iba-1, TNF-α, and NF-κB was analyzed using Western blot. Furthermore, ROS formation was measured through the DHE assay. The activation of the endocannabinoid receptors demonstrated a neuroprotective response during excitotoxicity, meditated by NOX-2. The reduction in ROS production led to a decrease in neuroinflammation, and AQP4 expression, resulting in reduced edema formation, and BBB permeability. During excitotoxic damage, WIN55,212-2 inhibits NOX-2-induced ROS production, reducing brain injury.

Selective blockade of cannabinoid receptors influence motoneuron survival and glial reaction after neonatal axotomy

Sciatic nerve crush in neonatal rats leads to an extensive death of motor and sensory neurons, serving as a platform to develop new neuroprotective approaches. The endocannabinoid system plays important neuromodulatory roles and has been involved in neurodevelopment and neuroprotection. The present work investigated the role of the cannabinoid receptors CB1 and CB2 in the neuroprotective response after neonatal axotomy. CB1 and CB2 antagonists (AM251 and AM630, respectively) were used after sciatic nerve crush in 2-day-old Wistar rats. Five days after lesion and treatment, the rats were perfused, and the spinal cords and dorsal root ganglia (DRG) were obtained and processed to investigate neuronal survival and immunohistochemistry changes, or RT-qPCR analysis. Motoneuron survival analysis showed that blocking CB2 alone or in combination with CB1 was neuroprotective. This effect was associated with a decrease in astrogliosis and microglial reaction. Interestingly, Cnr1 (CB1) and Bdnf gene transcripts were downregulated in the spinal cords of the antagonist-treated groups. Despite no intergroup difference regarding neuronal survival in the DRG, the simultaneous blockade of CB1 and CB2 receptors led to an increased expression of both Cnr1 and Cnr2, combined with Gdnf upregulation. The results indicate that the selective antagonism of cannabinoid receptors facilitates neuroprotection and decreases glial reactivity, suggesting new potential treatment approaches.

Dimethyl Fumarate Strongly Ameliorates Gray and White Matter Brain Injury and Modulates Glial Activation after Severe Hypoxia-Ischemia in Neonatal Rats.

Neonatal hypoxia-ischemia is a major cause of infant death and disability. The only clinically accepted treatment is therapeutic hypothermia; however, cooling is less effective in the most severely encephalopathic infants. Here, we wanted to test the neuroprotective effect of the antioxidant dimethyl fumarate after severe hypoxia-ischemia in neonatal rats. We used a modified Rice-Vannucci model to generate severe hypoxic-ischemic brain damage in day 7 postnatal rats, which were randomized into four experimental groups: Sham, Sham + DMF, non-treated HI, and HI + DMF. We analyzed brain tissue loss, global and regional (cortex and hippocampus) neuropathological scores, white matter injury, and microglial and astroglial reactivity. Compared to non-treated HI animals, HI + DMF pups showed a reduced brain area loss (p = 0.0031), an improved neuropathological score (p = 0.0016), reduced white matter injuries by preserving myelin tracts (p < 0.001), and diminished astroglial (p < 0.001) and microglial (p < 0.01) activation. After severe hypoxia-ischemia in neonatal rats, DMF induced a strong neuroprotective response, reducing cerebral infarction, gray and white matter damage, and astroglial and microglial activation. Although further molecular studies are needed and its translation to human babies would need to evaluate the molecule in piglets or lambs, DMF may be a potential treatment against neonatal encephalopathy.

1H-NMR Analysis of Amino Acid Metabolism in Cerebrospinal Fluid of Dogs with Neurological Distemper

Abstract Canine Distemper Virus (CDV) infection causes a multifocal demyelinating progressive disease within the central nervous system (CNS) that results in wide range of neurological symptoms. Pathological changes in the brain or CNS could be observed by analyzing cerebrospinal fluid (CSF). Therefore, this study aimed to investigate the concentrations of amino acids in the CSF of dogs with neurological form of CDV infection, and to identify amino acids as markers that can be used in the diagnosis, pathogenesis, and treatment of the disease. Heathy dogs (n=6), confirmed by clinical and laboratory examinations (Healthy group), and CDV-infected dogs (n=10) with neurological symptoms, confirmed by clinical, laboratory, and rapid diagnostic test kits (Distemper group), were used. CSF samples were obtained with an appropriate method and were subjected to 1H-NMR analysis. Identification was made on 10, whereas quantification on 8 amino acids. L-tyrosine, L-phenylalanine, L-threonine, and L-alanine concentrations were significantly lower, while L-histidine and L-tryptophan were significantly higher than the Healthy group (p&lt;0.05). It was concluded that L-tyrosine could be used for assessing mental status changes, L-phenylalanine for evaluating neuroprotective responses, L-threonine and L-histidine for gauging the extent of neurodegeneration and ventricular degeneration, L-alanine for exploring cellular stress and energy metabolism, and L-tryptophan for understanding the process of sympathetic nervous system activation.

Exploring the protective potential of NRF2 overexpressed neural extracellular vesicles against cisplatin-induced neurotoxicity via NRF2/ARE pathway

Neurotoxicity is characterized by the accumulation of harmful chemicals such as heavy metals and drugs in neural tissue, resulting in subsequent neuronal death. Among chemicals platinum-based cancer drugs are frequently used due to their antineoplastic effects, but this drug is also known to cause a wide range of toxicities, such as neurotoxicity. The nuclear-factor-erythroid 2-related factor-2 (NRF2) is crucial in combating oxidative stress and maintaining cellular homeostasis. This study thoroughly explores the protective effects of extracellular vesicles derived from NRF2 gene overexpressed neural progenitor cells (NEVs) on cisplatin-induced neurotoxicity. Therefore, extracellular vesicles derived from neural progenitor cells were isolated and characterized. The Cisplatin neurotoxicity dose was 75 µM in mature, post-mitotic neurons. 1.25 µM of tert-butyl hydroquinone that induces NRF2/ARE pathway was used as the positive control. The effects of extracellular vesicles (EVs) were investigated using functional and molecular assays such as PCR and protein-based assays. Here, we observed that NEVs dose-dependently protected post-mitotic neuron cells in response to cisplatin. The study also examined whether the effect was EV-induced by limiting EV biogenesis. The molecular basis of preventive treatment was established. When pre-administered, 1×108 particles/ml of NEVs maintained antioxidant and detoxifying gene and protein expression levels similar to control cell levels. Furthermore, NEVs reduced both cellular and mitochondrial ROS levels and preserved mitochondrial membrane potential. In addition, Catalase and SOD levels were found higher in NEV-treated cells compared to cisplatin control. The findings in NRF2-based protection of cisplatin-induced neurotoxicity may provide further evidence for the relationship between EVs and inhibition of neuronal stress through the NRF2/ARE pathway, increasing the understanding of neuroprotective responses and the development of gene-engineered EV therapy options for peripheral neuropathy or other neurodegenerative diseases. This is the first study in the literature to investigate the neutralizing potency of NRF2 overexpressed neural EVs against cisplatin-induced neurotoxicity.

Age-dependent Powassan virus lethality is linked to glial cell activation and divergent neuroinflammatory cytokine responses in a murine model.

Powassan virus is an emerging tick-borne flavivirus causing lethal encephalitis in aged individuals. We reveal an age-dependent POWV murine model that mirrors human POWV encephalitis and long-term CNS damage in the elderly. We found that POWV is neuroinvasive and directs reactive gliosis in all age mice, but at acute stages selectively induces pro-inflammatory Th1 cytokine responses in 50-week-old mice and neuroprotective Th2 cytokine responses in 10-week-old mice. Our findings associate CNS viral loads and divergent cytokine responses with age-dependent POWV lethality and survival outcomes. Responses of young mice suggest potential therapeutic targets and approaches for preventing severe POWV encephalitis that may be broadly applicable to other neurodegenerative diseases. Our age-dependent murine POWV model permits analysis of vaccines that prevent POWV lethality, and therapeutics that resolve severe POWV encephalitis.

C5aR1 antagonism suppresses inflammatory glial responses and alters cellular signaling in an Alzheimer’s disease mouse model

Alzheimer’s disease (AD) is the leading cause of dementia in older adults, and the need for effective, sustainable therapeutic targets is imperative. The complement pathway has been proposed as a therapeutic target. C5aR1 inhibition reduces plaque load, gliosis, and memory deficits in animal models, however, the cellular bases underlying this neuroprotection were unclear. Here, we show that the C5aR1 antagonist PMX205 improves outcomes in the Arctic48 mouse model of AD. A combination of single cell and single nucleus RNA-seq analysis of hippocampi derived from males and females identified neurotoxic disease-associated microglia clusters in Arctic mice that are C5aR1-dependent, while microglial genes associated with synapse organization and transmission and learning were overrepresented in PMX205-treated mice. PMX205 also reduced neurotoxic astrocyte gene expression, but clusters associated with protective responses to injury were unchanged. C5aR1 inhibition promoted mRNA-predicted signaling pathways between brain cell types associated with cell growth and repair, while suppressing inflammatory pathways. Finally, although hippocampal plaque load was unaffected, PMX205 prevented deficits in short-term memory in female Arctic mice. In conclusion, C5aR1 inhibition prevents cognitive loss, limits detrimental glial polarization while permitting neuroprotective responses, as well as leaving most protective functions of complement intact, making C5aR1 antagonism an attractive therapeutic strategy for AD.

Effects of metal oxide inhalation on the transcription of some hormone receptors in the brain, examined in an in vivo mouse model

Respirable metal oxide nanoparticles in welding fumes pose significant health risks upon inhalation, potentially leading to neurodegenerative diseases. While the exact mechanisms remain unclear, it is evident that metal oxide nanoparticles can disrupt cellular functions, including metabolism and inflammatory responses after crossing the blood–brain barrier (BBB). Our study investigates the impact of manual metal arc welding fumes on hormone receptor transcription in an in vivo mouse model. After collecting samples from six different brain regions at 24 and 96 h upon exposure, we focused on expression levels of estrogen receptors (ERs), thyroid hormone receptors (TRs), and peroxisome proliferator-activated receptors (PPARs) due to their roles in modulating neuroprotective responses and neuroinflammatory processes. Analysis revealed differential susceptibility of brain regions to hormonal disruption induced by welding fumes, with the hypothalamus (HT) and olfactory bulb (OB) showing prominent changes in receptor expression. Considering ERs, 24 h sampling showed an elevation in OB, with later increases in both ERα and ERβ. HT showed significant ERβ change only by 96 h. TRs mirrored ER patterns, with notable changes in OB and less in HT. PPARγ followed TR trends, with early upregulation in HT and downregulation elsewhere. These findings suggest a compensatory response within the CNS aimed at mitigating neuroinflammatory effects, as evidenced by the upregulation of ERβ, TRα, and PPARγ. The coordinated increase in ERs, TRs, and PPARs in the hypothalamus and olfactory bulb also highlights their potential neuroprotective roles in response to welding fume exposure. Our results also support the theory of metal oxide penetration to the CNS via the lungs-blood-BBB pathway, making HT and OB more vulnerable to welding fume exposure.

Does the brain make prolactin?

The prolactin receptor (Prlr) is widely expressed in the brain, particularly in the hypothalamus. Prolactin also has an increasing range of well-characterised effects on central nervous system function. Because of this, over many years, there has been interest in whether the hormone itself is also expressed within the brain, perhaps acting as a neuropeptide to regulate brain function via its receptor in neurons. The aim of this invited review is to critically evaluate the evidence for brain production of prolactin. Unlike the evidence for the Prlr, evidence for brain prolactin is inconsistent and variable. A range of different antibodies have been used, each characterising a different distribution of prolactin-like immunoreactivity. Prolactin mRNA has been detected in the brain, but only at levels markedly lower than seen in the pituitary gland. Importantly, it has largely only been detected by highly sensitive amplification-based techniques, and the extreme sensitivity means there is a risk of false-positive data. Modern in situ hybridisation methods and single-cell RNA sequencing have not provided supporting evidence, but it is hard to prove a negative! Finally, I acknowledge and discuss the possibility that prolactin might be produced in the brain under specific circumstances, such as to promote a neuroprotective response to cell damage. Collectively, however, based on this analysis, I have formed the opinion that brain production of prolactin is unlikely, and even if occurs, it is of little physiological consequence. Most, if not all of the brain actions of prolactin can be explained by pituitary prolactin gaining access to the brain.

The impact of astrocytic NF-κB on healthy and Alzheimer’s disease brains

Astrocytes play a role in healthy cognitive function and Alzheimer’s disease (AD). The transcriptional factor nuclear factor-κB (NF-κB) drives astrocyte diversity, but the mechanisms are not fully understood. By combining studies in human brains and animal models and selectively manipulating NF-κB function in astrocytes, we deepened the understanding of the role of astrocytic NF-κB in brain health and AD. In silico analysis of bulk and cell-specific transcriptomic data revealed the association of NF-κB and astrocytes in AD. Confocal studies validated the higher level of p50 NF-κB and phosphorylated-p65 NF-κB in glial fibrillary acidic protein (GFAP)+-astrocytes in AD versus non-AD subjects. In the healthy mouse brain, chronic activation of astrocytic NF-κB disturbed the proteomic milieu, causing a loss of mitochondrial-associated proteins and the rise of inflammatory-related proteins. Sustained NF-κB signaling also led to microglial reactivity, production of pro-inflammatory mediators, and buildup of senescence-related protein p16INK4A in neurons. However, in an AD mouse model, NF-κB inhibition accelerated β-amyloid and tau accumulation. Molecular biology studies revealed that astrocytic NF-κB activation drives the increase in GFAP and inflammatory proteins and aquaporin-4, a glymphatic system protein that assists in mitigating AD. Our investigation uncovered fundamental mechanisms by which NF-κB enables astrocytes' neuroprotective and neurotoxic responses in the brain.

Stem cell grafts enhance endogenous extracellular vesicle expression in the stroke brain

Endogenous brain repair occurs following an ischemic stroke but is transient, thus unable to fully mount a neuroprotective response against the evolving secondary cell death. Finding a treatment strategy that may render robust and long-lasting therapeutic effects stands as a clinically relevant therapy for stroke. Extracellular vesicles appear to be upregulated after stroke, which may represent a candidate target for neuroprotection. In this study, we probed whether transplanted stem cells could enhance the expression of extracellular vesicles to afford stable tissue remodeling in the ischemic stroke brain. Aged rats were initially exposed to the established ischemic stroke model of middle cerebral artery occlusion then received intravenous delivery of either bone marrow-derived mesenchymal stem cell transplantation or vehicle. A year later, the animals were assayed for brain damage, inflammation, and extracellular vesicle expression. Our findings revealed that while core infarction was not reduced, the stroke animals transplanted with stem cells displayed a significant reduction in peri-infarct cell loss that coincided with downregulated Iba1-labeled inflammatory cells and upregulated CD63-positive extracellular vesicles that appeared to be co-localized with GFAP-positive astrocytes. Interestingly, grafted stem cells were not detected at one year post-transplantation period, suggesting that the extracellular vesicles likely originated within the host brain. That long-lasting functional benefits persisted in the absence of surviving transplanted stem cells, but with upregulation of endogenous extracellular vesicles, advances the concept that transplantation of stem cells acutely after stroke propels host extracellular vesicles to the ischemic brain, altogether promoting chronic brain remodeling.

Impacts of PI3K/protein kinase B pathway activation in reactive astrocytes: from detrimental effects to protective functions.

Astrocytes are the most abundant type of glial cell in the central nervous system. Upon injury and inflammation, astrocytes become reactive and undergo morphological and functional changes. Depending on their phenotypic classification as A1 or A2, reactive astrocytes contribute to both neurotoxic and neuroprotective responses, respectively. However, this binary classification does not fully capture the diversity of astrocyte responses observed across different diseases and injuries. Transcriptomic analysis has revealed that reactive astrocytes have a complex landscape of gene expression profiles, which emphasizes the heterogeneous nature of their reactivity. Astrocytes actively participate in regulating central nervous system inflammation by interacting with microglia and other cell types, releasing cytokines, and influencing the immune response. The phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) signaling pathway is a central player in astrocyte reactivity and impacts various aspects of astrocyte behavior, as evidenced by in silico , in vitro , and in vivo results. In astrocytes, inflammatory cues trigger a cascade of molecular events, where nuclear factor-κB serves as a central mediator of the pro-inflammatory responses. Here, we review the heterogeneity of reactive astrocytes and the molecular mechanisms underlying their activation. We highlight the involvement of various signaling pathways that regulate astrocyte reactivity, including the PI3K/AKT/mammalian target of rapamycin (mTOR), α v β 3 integrin/PI3K/AKT/connexin 43, and Notch/PI3K/AKT pathways. While targeting the inactivation of the PI3K/AKT cellular signaling pathway to control reactive astrocytes and prevent central nervous system damage, evidence suggests that activating this pathway could also yield beneficial outcomes. This dual function of the PI3K/AKT pathway underscores its complexity in astrocyte reactivity and brain function modulation. The review emphasizes the importance of employing astrocyte-exclusive models to understand their functions accurately and these models are essential for clarifying astrocyte behavior. The findings should then be validated using in vivo models to ensure real-life relevance. The review also highlights the significance of PI3K/AKT pathway modulation in preventing central nervous system damage, although further studies are required to fully comprehend its role due to varying factors such as different cell types, astrocyte responses to inflammation, and disease contexts. Specific strategies are clearly necessary to address these variables effectively.

Neuroprotective Strategies and Cell-Based Biomarkers for Manganese-Induced Toxicity in Human Neuroblastoma (SH-SY5Y) Cells.

Manganese (Mn) is an essential heavy metal in the human body, while excess Mn leads to neurotoxicity, as observed in this study, where 100 µM of Mn was administered to the human neuroblastoma (SH-SY5Y) cell model of dopaminergic neurons in neurodegenerative diseases. We quantitated pathway and gene changes in homeostatic cell-based adaptations to Mn exposure. Utilizing the Gene Expression Omnibus, we accessed the GSE70845 dataset as a microarray of SH-SY5Y cells published by Gandhi et al. (2018) and applied statistical significance cutoffs at p < 0.05. We report 74 pathway and 10 gene changes with statistical significance. ReactomeGSA analyses demonstrated upregulation of histones (5 out of 10 induced genes) and histone deacetylases as a neuroprotective response to remodel/mitigate Mn-induced DNA/chromatin damage. Neurodegenerative-associated pathway changes occurred. NF-κB signaled protective responses via Sirtuin-1 to reduce neuroinflammation. Critically, Mn activated three pathways implicating deficits in purine metabolism. Therefore, we validated that urate, a purine and antioxidant, mitigated Mn-losses of viability in SH-SY5Y cells. We discuss Mn as a hypoxia mimetic and trans-activator of HIF-1α, the central trans-activator of vascular hypoxic mitochondrial dysfunction. Mn induced a 3-fold increase in mRNA levels for antioxidant metallothionein-III, which was induced 100-fold by hypoxia mimetics deferoxamine and zinc.

The Microglial Transcriptome of Age-Associated Deep Subcortical White Matter Lesions Suggests a Neuroprotective Response to Blood-Brain Barrier Dysfunction.

Age-associated deep-subcortical white matter lesions (DSCLs) are an independent risk factor for dementia, displaying high levels of CD68+ microglia. This study aimed to characterize the transcriptomic profile of microglia in DSCLs and surrounding radiologically normal-appearing white matter (NAWM) compared to non-lesional control white matter. CD68+ microglia were isolated from white matter groups (n = 4 cases per group) from the Cognitive Function and Ageing Study neuropathology cohort using immuno-laser capture microdissection. Microarray gene expression profiling, but not RNA-sequencing, was found to be compatible with immuno-LCM-ed post-mortem material in the CFAS cohort and identified significantly differentially expressed genes (DEGs). Functional grouping and pathway analysis were assessed using the Database for Annotation Visualization and Integrated Discovery (DAVID) software, and immunohistochemistry was performed to validate gene expression changes at the protein level. Transcriptomic profiling of microglia in DSCLs compared to non-lesional control white matter identified 181 significant DEGs (93 upregulated and 88 downregulated). Functional clustering analysis in DAVID revealed dysregulation of haptoglobin-haemoglobin binding (Enrichment score 2.5, p = 0.017), confirmed using CD163 immunostaining, suggesting a neuroprotective microglial response to blood-brain barrier dysfunction in DSCLs. In NAWM versus control white matter, microglia exhibited 347 DEGs (209 upregulated, 138 downregulated), with significant dysregulation of protein de-ubiquitination (Enrichment score 5.14, p < 0.001), implying an inability to maintain protein homeostasis in NAWM that may contribute to lesion spread. These findings enhance understanding of microglial transcriptomic changes in ageing white matter pathology, highlighting a neuroprotective adaptation in DSCLs microglia and a potentially lesion-promoting phenotype in NAWM microglia.

Abstract P116: Exploring The Cardioprotective Potential Of Roflumilast (pde-4 Inhibitor) Against Experimental Epirubicin-induced Cardiotoxicity.

Introduction: Cardiotoxicity is an important side effect of the treatment of a malignant tumour with Anthracyclines. The anthracycline family drug epirubicin (EPI) is one of the most potent chemotherapeutic agents for treating a range of solid and hematologic malignant tumours. However, its anti-tumor efficacy is dose-dependent, and the development of cardiac-toxicities limits its clinical use. In various Pharmacological Studies Roflumilast (ROF), a PDE-4 Inhibitor has been shown a Neuroprotective and Cardioprotective response but its role in EPI-induced cardiotoxicity and the underlying mechanisms has never been Evaluated. Hypothesis: The Study hypothesized that Roflumilast exerts cardioprotection against EPI-induced cardiotoxicity through attenuating cardiac mitochondrial dysfunction and inflammatory markers. Methods: Wistar rats of either sex were employed into 4 groups 3 groups comprised 7 animals while the control group had 5 animals, EPI-treated rats were intraperitoneally injected with 10 mg/kg for 1 day, while control rats were on normal diet. EPI+ROF (Low Dose) treated rats were intraperitoneally injected with 10 mg/kg of EPI for 1 day and 1mg/kg of ROF via oral gavage for 7 days. While in the higher dose of EPI+ROF treated rats were intraperitoneally injected with 10 mg/kg of EPI for 1 day and 2mg/kg of ROF via oral gavage for 7 days. The rat hearts were rapidly removed to determine cardiac mitochondrial functions and various cardiac biomarkers. Results: EPI induced significantly excessive mitochondrial Oxidative Stress, increased tissue TBARS and decreased levels of SOD and GSH. Conversely, a 7-day treatment with ROF (1mg/kg/day , p.o .) and ROF (2mg/kg/day, p.o .) significantly reduced TBARS levels while increasing SOD and GSH concentrations. The levels of TNF-α, CK-MB AST, ALT, and LDH were noted to be markedly increased in EPI-induced rats. While treatment with ROF (1mg/kg/day, p.o., 7 days ) and ROF (2mg/kg/day, p.o., 7 days) showed significantly decreased levels of TNF-α, CK-MB AST, ALT, LDH. Conclusion: In the current investigation, it can be concluded that administration of EPI leads to Oxidative Stress, Inflammation and Cardiotoxicity. Treatment with the ROF 2mg/kg showed cardioprotective potential by significantly decreasing inflammatory markers, oxidative stress parameters and cardiotoxic parameters. Our findings suggest that ROF protects the heart from EPI-induced cardiotoxicity, and it may serve as a guide for future cardioprotective research strategies.

“Immunohistochemical analysis of Sigma-1 receptor (σ-1R) expression in human pineal gland in relation to different causes of death”

Sigma-1 receptor (σ-1R) modulates cellular signaling pathways, probably acting as a ligand operated chaperone. When activated, the receptor translocates from the interface mitochondrion associated membrane of the endoplasmic reticulum to the cell membrane. σ-1R was demonstrated in some brain regions, including the pineal gland, and was proposed to be involved in several cerebral processes, including neuroprotective responses against homeostasis alterations. On this basis, the immunohistochemical expression of σ-1R in human pineal glands was evaluated, with particular regard to the different causes of death. Thirty-eight pineal glands obtained from forensic autopsies were divided into five groups according to the cause of death: sudden death, drowning, fire fatality, hanging, and hemorrhagic shock, and examined with hematoxylin-eosin stain and immunohistochemistry for σ-1R. Both pinealocytes and perivascular spaces were evaluated. The pineal glands from sudden death were only mildly positive for σ-1R, while a more evident immunopositivity was observed in hanging, fire fatality, hemorrhagic shock, and drowning. These results were confirmed in a two-by-two comparison between the sudden death group and other groups. Our data demonstrate for the first time with immunohistochemical techniques the presence of σ-1R expression in the human pineal gland and propose a direct correlation between σ-1R expression and duration of the death process, in particular when hypoxic conditions and/or excessive psychological stress are present.

P17/C18-ceramide-mediated mitophagy is an endogenous neuroprotective response in preclinical and clinical brain injury.

Repeat concussions (or repetitive mild traumatic brain injury [rmTBI]) are complex pathological processes consisting of a primary insult and long-term secondary complications and are also a prerequisite for chronic traumatic encephalopathy (CTE). Recent evidence implies a significant role of autophagy-mediated dysfunctional mitochondrial clearance, mitophagy, in the cascade of secondary deleterious events resulting from TBI. C18-ceramide, a bioactive sphingolipid produced in response to cell stress and damage, and its synthesizing enzyme (CerS1) are precursors to selective stress-mediated mitophagy. A transporter, p17, mediates the trafficking of CerS1, induces C18-ceramide synthesis in the mitochondrial membrane, and acts as an elimination signal in cell survival. Whether p17-mediated mitophagy occurs in the brain and plays a causal role in mitochondrial quality control in secondary disease development after rmTBI are unknown. Using a novel repetitive less-than-mild TBI (rlmTBI) injury paradigm, ablation of mitochondrial p17/C18-ceramide trafficking in p17 knockout (KO) mice results in a loss of C18-ceramide-induced mitophagy, which contributes to susceptibility and recovery from long-term secondary complications associated with rlmTBI. Using a ceramide analog with lipid-selenium conjugate drug, LCL768 restored mitophagy and reduced long-term secondary complications, improving cognitive deficits in rlmTBI-induced p17KO mice. We obtained a significant reduction of p17 expression and a considerable decrease of CerS1 and C18-ceramide levels in cortical mitochondria of CTE human brains compared with age-matched control brains. These data demonstrated that p17/C18-ceramide trafficking is an endogenous neuroprotective mitochondrial stress response following rlmTBI, thus suggesting a novel prospective strategy to interrupt the CTE consequences of concussive TBI.

Abstract WP9: Ischemic Stroke Progression After Witnessed Onset in Daytime vs Night-Time

Introduction: Early neurologic deterioration (END) and poor functional outcomes are more frequent with night than daytime onset of ischemic stroke. This variation may be related to: 1) circadian variation endogenous bodily rhythms and/or 2) extrinsic environmental time cues. Methods: To assess the contribution of extrinsic environmental time to ischemic stroke course, we evaluated patients with acute cerebral ischemia (ACI) onset within last 2h, enrolled in the NIH FAST-MAG trial of prehospital neuroprotection, consisting of patients with witnessed/wake onset. Results: Among 1235 ACI patients (ischemic stroke 1033, TIA 202), age was 70.8 [±13.2], 45.3% female, initial NIHSS 7 (IQR 2-15), LKW to randomization 59 mins (IQR 46-80), and 473 (38%) treated with IV lytics. Onset occurred during clock daytime (07:00-22:59) in 1147 (92.9%) and night-time (23:00-06:59) in 88 (7.1%). Patients were similar in 18 baseline characteristics, but night-onset had higher initial SBP (162 vs 155, p=0.01) and longer onset to paramedic times (33m vs 23m, p = 0.0002). Among non-lytic patients, both unadjusted and adjusted clinical outcomes were similar in night vs daytime patients, including END (13.6% vs 16.4%), 90d mRS 0-2 (63.6% vs 58.5%), and 90d mortality (10.2% vs 12.8%). Among lytic patients, clinical outcomes were also similar in night vs daytime onset patients, including unadjusted END (36% vs 21.7%), 90d mRS 0-2 (52% vs 46.9%), and 90d mortality (12% vs 17.4%). Night vs daytime onset did not modify the effect of magnesium vs placebo on outcome among either non-thrombolytic or thrombolytic-treated patients (Figure). Conclusion: Among acute cerebral ischemia patients, efficacy, safety, and neuroprotective agent response outcomes were not modified by witnessed onset during active versus inactive phase clock times. These findings suggest biological wake-sleep state rather than chronologic clock time is the driver of known circadian rhythmicity in stroke course.