Selective autophagy as a therapeutic target for neurological diseases.

The neurological diseases primarily include acute injuries, chronic neurodegeneration, and others (e.g., infectious diseases of the central nervous system). Autophagy is a housekeeping process responsible for the bulk degradation of misfolded protein aggregates and damaged organelles through the lysosomal machinery. Recent studies have suggested that autophagy, particularly selective autophagy, such as mitophagy, pexophagy, ER-phagy, ribophagy, lipophagy, etc., is closely implicated in neurological diseases. These forms of selective autophagy are controlled by a group of important proteins, including PTEN-induced kinase 1 (PINK1), Parkin, p62, optineurin (OPTN), neighbor of BRCA1 gene 1 (NBR1), and nuclear fragile X mental retardation-interacting protein 1 (NUFIP1). This review highlights the characteristics and underlying mechanisms of different types of selective autophagy, and their implications in various forms of neurological diseases.

Rh-relaxin-2 attenuates degranulation of mast cells by inhibiting NF-κB through PI3K-AKT/TNFAIP3 pathway in an experimental germinal matrix hemorrhage rat model.

BACKGROUND: Mast cells play an important role in early immune reactions in the brain by degranulation and the consequent inflammatory response. Our aim of the study is to investigate the effects of rh-relaxin-2 on mast cells and the underlying mechanisms in a germinal matrix hemorrhage (GMH) rat model. METHODS: One hundred seventy-three P7 rat pups were subjected to GMH by an intraparenchymal injection of bacterial collagenase. Clodronate liposome was administered through intracerebroventricular (i.c.v.) injections 24 h prior to GMH to inhibit microglia. Rh-relaxin-2 was administered intraperitoneally at 1 h and 13 h after GMH. Small interfering RNA of RXFP1 and PI3K inhibitor LY294002 were given by i.c.v. injection. Post-GMH evaluation included neurobehavioral function, Western blot analysis, immunofluorescence, Nissl staining, and toluidine blue staining. RESULTS: Our results demonstrated that endogenous relaxin-2 was downregulated and that RXFP1 level peaked on the first day after GMH. Administration of rh-relaxin-2 improved neurological functions, attenuated degranulation of mast cells and neuroinflammation, and ameliorated post-hemorrhagic hydrocephalus (PHH) after GMH. These effects were associated with RXFP1 activation, increased expression of PI3K, phosphorylated AKT and TNFAIP3, and decreased levels of phosphorylated NF-κB, tryptase, chymase, IL-6, and TNF-α. However, knockdown of RXFP1 and PI3K inhibition abolished the protective effects of rh-relaxin-2. CONCLUSIONS: Our findings showed that rh-relaxin-2 attenuated degranulation of mast cells and neuroinflammation, improved neurological outcomes, and ameliorated hydrocephalus after GMH through RXFP1/PI3K-AKT/TNFAIP3/NF-κB signaling pathway.

Inhibition of mast cell tryptase attenuates neuroinflammation via PAR-2/p38/NFκB pathway following asphyxial cardiac arrest in rats.

BACKGROUND: Cardiac arrest survivors suffer from neurological dysfunction including cognitive impairment. Cerebral mast cells, the key regulators of neuroinflammation contribute to neuroinflammation-associated cognitive dysfunction. Mast cell tryptase was demonstrated to have a proinflammatory effect on microglia via the activation of microglial protease-activated receptor-2 (PAR-2). This study investigated the potential anti-neuroinflammatory effect of mast cell tryptase inhibition and the underlying mechanism of PAR-2/p-p38/NFκB signaling following asphyxia-induced cardiac arrest in rats. METHODS: Adult male Sprague-Dawley rats resuscitated from 10 min of asphyxia-induced cardiac arrest were randomized to four separate experiments including time-course, short-term outcomes, long-term outcomes and mechanism studies. The effect of mast cell tryptase inhibition on asphyxial cardiac arrest outcomes was examined after intranasal administration of selective mast cell tryptase inhibitor (APC366; 50 µg/rat or 150 µg/rat). AC55541 (selective PAR-2 activator; 30 µg/rat) and SB203580 (selective p38 inhibitor; 300 µg/rat) were used for intervention. Short-term neurocognitive functions were evaluated using the neurological deficit score, number of seizures, adhesive tape removal test, and T-maze test, while long-term cognitive functions were evaluated using the Morris water maze test. Hippocampal neuronal degeneration was evaluated by Fluoro-Jade C staining. RESULTS: Mast cell tryptase and PAR-2 were dramatically increased in the brain following asphyxia-induced cardiac arrest. The inhibition of mast cell tryptase by APC366 improved both short- and long-term neurological outcomes in resuscitated rats. Such behavioral benefits were associated with reduced expressions of PAR-2, p-p38, NFκB, TNF-α, and IL-6 in the brain as well as less hippocampal neuronal degeneration. The anti-neuroinflammatory effect of APC366 was abolished by AC55541, which when used alone, indeed further exacerbated neuroinflammation, hippocampal neuronal degeneration, and neurologic deficits following cardiac arrest. The deleterious effects aggregated by AC55541 were minimized by p38 inhibitor. CONCLUSIONS: The inhibition of mast cell tryptase attenuated neuroinflammation, led to less hippocampal neuronal death and improved neurological deficits following cardiac arrest. This effect was at least partly mediated via inhibiting the PAR-2/p-p38/NFκB signaling pathway. Thus, mast cell tryptase might be a novel therapeutic target in the management of neurological impairment following cardiac arrest.

Insights into major facilitator superfamily domain-containing protein-2a (Mfsd2a) in physiology and pathophysiology. What do we know so far?

Major facilitator superfamily domain-containing protein-2a (Mfsd2a) which was considered as an orphan transporter has recently gained attention for its regulatory role in the maintenance of proper functioning of the blood-brain barrier. Besides the major role of Mfsd2a in maintaining the barrier function, increasing evidence has emerged with regard to the contributions of Mfsd2a to various biological processes such as transport, cell fusion, cell cycle, inflammation and regeneration, managing tumor growth, functioning of other organs with barrier functions or responses to injury. The purpose of this article is to review the different roles of Mfsd2a and its involvement in the physiological and pathophysiological processes primarily in the central nervous system and throughout the mammalian body under the lights of the current literature.

Rh-IFN-α attenuates neuroinflammation and improves neurological function by inhibiting NF-κB through JAK1-STAT1/TRAF3 pathway in an experimental GMH rat model.

Neuroinflammation occurs after germinal matrix hemorrhage (GMH) and induces secondary brain injury. Interferon-α (IFN-α) has been shown to exert anti-inflammatory effects in infectious diseases via activating IFNAR and its downstream signaling. We aimed to investigate the anti-inflammatory effects of Recombinant human IFN-α (rh-IFN-α) and the underlying mechanisms in a rat GMH model. Two hundred and eighteen P7 rat pups of both sexes were subjected to GMH by an intraparenchymal injection of bacterial collagenase. Rh-IFN-α was administered intraperitoneally. Small interfering RNA (siRNA) of IFNAR, and siRNA of tumor necrosis factor receptor associated factor 3 (TRAF3) were administered through intracerebroventricular (i.c.v.) injections. JAK1 inhibitor ruxolitinib was given by oral lavage. Post-GMH evaluation included neurobehavioral function, Nissl staining, Western blot analysis, and immunofluorescence. Our results showed that endogenous IFN-α and phosphorylated IFNAR levels were increased after GMH. Administration of rh-IFN-α improved neurological functions, attenuated neuroinflammation, inhibited microglial activation, and ameliorated post-hemorrhagic hydrocephalus after GMH. These observations were concomitant with IFNAR activation, increased expression of phosphorylated JAK1, phosphorylated STAT1 and TRAF3, and decreased levels of phosphorylated NF-κB, IL-6 and TNF-α. Specifically, knockdown of IFNAR, JAK1 and TRAF3 abolished the protective effects of rh-IFN-α. In conclusion, our findings demonstrated that rh-IFN-α treatment attenuated neuroinflammation, neurological deficits and hydrocephalus formation through inhibiting microglial activation after GMH, which might be mediated by IFNAR/JAK1-STAT1/TRAF3/NF-κB signaling pathway. Rh-IFN-α may be a promising therapeutic agent to attenuate brain injury via its anti-inflammatory effect.

Targeting mast cell as a neuroprotective strategy.

Background: Mast cells (MCs) are perivascularly located immune cells of haematopoietic origin. Emerging evidences suggest that the activation of MCs play important roles in the pathogenesis of blood brain barrier disruption, neuroinflammation, and neurodegeneration. Objectives: In this review, we aimed to discuss the detrimental effects of MCs in response to various types of brain injury, as well as the therapeutic potential and neuroprotective effects of targeting the activation and degranulation of MCs, particularly in the management of the acute phase. Methods: An extensive online literature search was conducted through Pubmed/Central on March 2018. Then, we comprehensively summarized the effects of the activation of brain MCs in acute brain injury along with current pharmacological strategies targeting at the activation of MCs. Results: The review of the current literature indicated that the activation and degranulation of brain MCs significantly contribute to the acute pathological process following different types of brain injury including focal and global cerebral ischaemia, intracerebral haemorrhage, subarachnoid haemorrhage, and traumatic brain injury. Conclusions: Brain MCs significantly contribute to the acute pathological processes following brain injury. In that regard, targeting brain MCs may provide a novel strategy for neuroprotection.

Endovascular Treatment of Ruptured Tiny Intracranial Aneurysms with Low-Profile Visualized Intraluminal Support Device.

BACKGROUND: Ruptured tiny intracranial aneurysms (TIAs) have been challenging both for endovascular and neurosurgical interventions. Thus, we aimed to evaluate the safety and efficacy of low-profile visualized intraluminal support (LVIS) device in the treatment of ruptured TIAs (rTIAs). MATERIAL AND METHODS: Among 761 intracranial aneurysms which were treated either surgically or endovascularly, 32 rTIAs underwent stent-assisted coiling with LVIS device between 2014 and 2017. Patient data were reviewed retrospectively. Clinical and radiological outcomes were recorded at discharge and mid-term follow-up. RESULTS: Mean patient ages were 53 ± 14.5 years. Mean aneurysm size was 2.28 ± .53 mm (range, 1-2.9 mm) with a mean dome:neck ratio of 1.08 (range, .75-2.14). The LVIS stents were successfully implanted in all patients. Mean follow-up period was 9.3 ± 1.9 months (range, 6-15 months). Immediate angiographic evaluation demonstrated complete occlusion in 13 (40.6%) patients, while neck remnant and residual sac were observed in 12 (37.5%) and 7 (21.9%), respectively. All patients had moderate disability (mRS 2-3) at discharge. Number of aneurysms with complete occlusion significantly increased and 82.1% of the patients (23 of 28) demonstrated complete occlusion at follow-up (P = .0015). Among these, 27 had good outcome (mRS 0-1; 96.9%) with significant improvement compared to discharge (P = .0001). There was no recurrence or enlargement of the residual aneurysms. Additionally, there were no procedure-related complications except the one (3.6%) showing asymptomatic stenosis of the posterior cerebral artery in follow-up imagings. CONCLUSIONS: Stent-assisted coiling of rTIAs with LVIS device provides high rates of technical success and complete occlusion at mid-term follow-up with an excellent safety profile.

Facial nerve outcome and extent of resection in cystic versus solid vestibular schwannomas in radiosurgery era.

OBJECTIVE Cystic vestibular schwannomas (CVSs) are a subgroup of vestibular schwannomas (VSs) that are reported to be associated with unpredictable clinical behavior and unfavorable postoperative outcomes. The authors aimed to review their experience with microsurgical treatment of CVSs in terms of extent of resection and postoperative facial nerve (FN) function and compare these outcomes with those of their solid counterparts. METHODS Two hundred-eleven VS patients were treated surgically between 2006 and 2017. Tumors were defined as cystic when preoperative neuroimaging demonstrated cyst formation that was confirmed by intraoperative findings. Solid VS (SVSs) with similar classes were used for comparison. Clinical data of the patients were reviewed retrospectively, including clinical notes and images, as well as operative, pathology, and neuroradiology reports. RESULTS Thirty-two patients (20 males and 12 females) with a mean age of 52.2 years (range 17-77 years) underwent microsurgical resection of 33 CVSs (mean size 3.6 cm, range 1.5-5 cm). Forty-nine patients (26 males and 23 females) with a mean age of 49.9 years (range 21-75 years) underwent microsurgical resection of 49 SVSs (mean size 3 cm, range 2-4.5 cm). All operations were performed via either a retrosigmoid or a translabyrinthine approach. Gross-total resection was achieved in 30 cases in the CVS group (90.9%) and 37 in the SVS group (75.5%). The main reason for subtotal and near-total resection was adherence of the tumor to the brainstem and/or FN in both groups. None of the patients with subtotal or near-total resection in the CVS group demonstrated symptomatic regrowth of the tumor during the mean follow-up period of 41.6 months (range 18-82 months). The FN was anatomically preserved in all patients in both groups. Good FN outcomes were achieved in 15 of CVS (grade I-II; 45.5%) and 35 of SVS (71.4%) surgeries at discharge. Good and fair FN functions were noted in 22 (grade I-II; 81.5%) and 5 (grade III only; 18.5%) of the CVS patients, respectively, at the 1-year follow-up; none of the patients showed poor FN function. CONCLUSIONS Surgery of CVSs does not necessarily result in poor outcomes in terms of the extent of resection and FN function. Special care should be exercised to preserve anatomical continuity of the FN during surgery, since long-term FN function outcomes are much more satisfactory than short-term results. High rates of gross-total resection and good FN outcomes in our study may also suggest that microsurgery stands as the treatment of choice in select cases of large CVSs and SVSs in the era of radiosurgery.

Evaluation of the content, quality, reliability and accuracy of YouTube videos regarding endotracheal intubation techniques.

BACKGROUND: Today, the internet is widely used to obtain any type of information. The use of internet may facilitate healthcare professionals' education as well. OBJECTIVES: We aimed to evaluate the content, accuracy, reliability and quality of YouTube videos regarding intubation, one of the most important skills for healthcare professionals. MATERIALS AND METHODS: Of the 54,000 videos found using the keyword 'intubation', the first 50 were included in the study. The sources were divided into three categories: academicians, healthcare professionals and medical. The view ratio, like ratio, and video power index were used to determine the popularity while Journal of American Medical Association (JAMA) benchmark criteria were used for reliability. The extent of the information was evaluated based on the scoring system provided by us. RESULTS: The majority of the videos were uploaded by healthcare professionals (92%) including academicians. Twenty-seven (54%) videos had training purposes. The information point was highest in the academic group (4.6 ± 2.7); however, there was no significance between groups (P = 0.2). The mean JAMA score was highest in the academic group (1.9 ± 0.8), with a statistical significance (P = 0.00055). The JAMA score and information points were significantly higher in training videos compared to non-training videos (p=<0.001 and P = 0.003, respectively). Popularity ratios were similar between groups. CONCLUSION: Videos regarding medical skills should be accurate. Information on YouTube regarding intubation is limited and is of low quality. The establishment of an organization authorized to evaluate the content, quality, accuracy and reliability of the information on the internet regarding medical skills is warranted.

Acute coronary syndrome presenting with earache and sore throat.

Chest pain and discomfort are regarded as the hallmark symptoms of acute coronary syndrome, and the absence of these symptoms are described as "atypical" presentation. In this case, we aimed to report an acute coronary syndrome­diagnosed patient who was admitted to our emergency service with unilateral earache.

Met-RANTES preserves the blood-brain barrier through inhibiting CCR1/SRC/Rac1 pathway after intracerebral hemorrhage in mice.

BACKGROUND: C-C chemokine receptor type 1 (CCR1) and its endogenous ligand, CCL5, participate in the pathogenesis of neuroinflammatory diseases. However, much remains unknown regarding CCL5/CCR1 signaling in blood-brain barrier (BBB) permeability after intracerebral hemorrhage (ICH). METHODS: A total of 250 CD1 male mice were used and ICH was induced via autologous whole blood injection. Either Met-RANTES, a selective CCR1 antagonist, or Met-RANTES combined with a Rac1 CRISPR activator was administered to the mice 1 h after ICH. Post-ICH assessments included neurobehavioral tests, brain water content, BBB integrity, hematoma volume, Western blot, and immunofluorescence staining. The CCR1 ligand, rCCL5, and SRC CRISPR knockout in naïve mice were used to further elucidate detrimental CCL5/CCR1/SRC signaling. RESULTS: Brain endogenous CCR1 and CCL5 were upregulated after ICH in mice with a peak at 24 h, and CCR1 was expressed in endothelial cells, astrocytes, and neurons. Met-R treatment reduced brain edema and neurobehavioral impairment, as well as preserved BBB integrity and tight junction protein expression in ICH mice. Met-R treatment decreased expression of p-SRC, Rac1, albumin, and MMP9, but increased claudin-5, occludin, and ZO-1 tight junction proteins after ICH. These effects were regressed using the Rac1 CRISPR activator. Administration of rCCL5 in naïve mice increased expression of p-SRC, Rac1, albumin, and MMP9, but decreased levels of claudin-5, occludin, and ZO-1 tight junction proteins. These effects in naïve mice were reversed with SRC CRISPR (KO). CONCLUSIONS: Our findings demonstrate that CCR5 inhibition by Met-R improves neurological deficits after ICH by preserving BBB integrity through inhibiting CCR1/SRC/Rac1 signaling pathway in mice. Thus, Met-R has therapeutic potential in the management of ICH patients.

Melanocortin 1 receptor attenuates early brain injury following subarachnoid hemorrhage by controlling mitochondrial metabolism via AMPK/SIRT1/PGC-1α pathway in rats.

Mitochondria-mediated oxidative stress and apoptosis contribute greatly to early brain injury (EBI) following subarachnoid hemorrhage (SAH). This study hypothesized that activation of melanocortin 1 receptor (MC1R), using BMS-470539, attenuates EBI by controlling mitochondrial metabolism after SAH. Methods: We utilized BMS-470539, MSG-606, selisistat, and PGC-1α to verify the neuroprotective effects of MC1R. We evaluated short- and long-term neurobehavior after SAH. Western blotting, immunofluorescence, and Golgi staining techniques were performed to assess changes in protein levels. Results: The results of western blotting suggested that the expression of SIRT1 and PGC-1α were increased, reaching their peaks at 24 h following SAH. Moreover, BMS-470539 treatment notably attenuated neurological deficits, and also reduced long-term spatial learning and memory impairments caused by SAH. The underlying neuroprotective mechanisms of the BMS-470539/MC1R system were mediated through the suppression of oxidative stress, apoptosis, and mitochondrial fission by increasing the levels of SIRT1, PGC-1α, UCP2, SOD, GPx, Bcl-2, cyto-Drp1, and ATP, while decreasing the levels of cleaved caspase-3, Bax, mito-Drp1, ROS, GSH/GSSG, and NADPH/NADP+ ratios. The neuroprotective effects of the BMS-470539/MC1R system were significantly abolished by MSG-606, selisistat, and PGC-1α siRNA. Conclusions: The activation of MC1R with BMS-470539 significantly attenuated EBI after SAH by suppressing the oxidative stress, apoptosis, and mitochondrial fission through the AMPK/SIRT1/PGC-1α signaling pathway.

FSLLRY-NH2 Improves Neurological Outcome After Cardiac Arrest in Rats.

AIM: To evaluate the effect of FSLLRY-NH2, a protease-activated receptor 2 (PAR2) inhibitor, on neurocognitive impairment and hippocampal neuronal degeneration in the setting of asphyxial cardiac arrest (ACA)-induced global cerebral ischemia (GCI) in rats. MATERIAL AND METHODS: A total of 43 Sprague-Dawley male rats were used. Shams and rats resuscitated from 9 minutes of ACA were randomized to two separate experiments including time course and short-term neurological outcomes. FSLLRY-NH2 (50 microgram [µg] per rat) was administered intranasally at 1 hour postresuscitation. Neurological function and hippocampal neuronal degeneration were evaluated after ACA. RESULTS: Significant neurological function decline and hippocampal neuron degeneration were observed in ACA animals as compared with the shams. Treatment with FSLLRY-NH2 significantly improved neurological outcome and reduced the number of degenerating hippocampal neurons after ACA. CONCLUSION: Targeting PAR2 may be a novel therapeutic approach in the management of neurological dysfunction after cardiac arrest-associated ischemic injury.

Peroxisomal Dysfunction Contributes to White Matter Injury Following Subarachnoid Hemorrhage in Rats via Thioredoxin-Interacting Protein-Dependent Manner.

BACKGROUND AND PURPOSE: White matter injury (WMI) exists in the early stage of subarachnoid hemorrhage (SAH) and has not been well addressed so far. METHODS: We utilized short hairpin RNA (shRNA) and clustered regularly interspaced short palindromic repeats (CRISPR) to verify the role of peroxisomes in WMI following SAH. We evaluated short- and long-term neurobehavior after SAH. Western blotting, immunofluorescence, and Golgi staining techniques were performed to assess the changes in protein levels. RESULTS: Catalase (CAT) CRISPR treatment significantly attenuated neurological deficits and reduced long-term spatial learning and memory impairments after SAH by increasing the level of myelin basic protein (MBP) while decreasing the levels of amyloid precursor protein (APP), interleukin 6 (IL-6), and tumor necrosis factor (TNF)-α. The use of thioredoxin-interacting protein (TXNIP) shRNA significantly offset the effects of CAT shRNA, and the use of glycerone phosphate acyl transferase (GNPAT) shRNA significantly reversed the effects of CAT CRISPR by decreasing the levels of plasmalogens and reactive oxidative species (ROS). CONCLUSION: Peroxisomal dysfunction induced by SAH reversely exacerbated cerebral WMI following SAH, which was at least partly mediated by TXNIP and GNPAT pathways.

An unusual case report of brain abscess caused by prevotella loescheii identified using the metagenomic next-generation sequencing.

OBJECTIVES: Brain abscess (BA) is a rare and life-threatening disease which remains to be a challenge for physicians despite recent advances in both the diagnosis and treatment strategies. Appropriate management of brain abscesses requires a combined surgical and medical approach to prevent associated life-threatening complications. In clinical practice, new diagnostic techniques, such as metagenomic next-generation sequencing (mNGS) and quantitative electroencephalogram (qEEG), can help physicians identify the causative pathogens of brain abscesses in order to provide early pathogen-targeted therapy. CLINICAL PRESENTATION: Here, we report a case of a 27-year-old Chinese woman with a Prevotella loescheii brain abscess into ventricular system. The diagnosis of the causative pathogen was identified by mNGS of abscess drainage fluid, leading to precise and targeted antimicrobial therapy and successful management of the abscess. We, therefor, avoided devastating consequences of the intraventricular rupture of the brain abscess. CONCLUSION: Appropriate management of BA requires a combination of both surgical and medical approaches. The increasing availability of molecular diagnostic techniques, such as the mNGS that is used to elucidate the microbial composition of the abscesses for definitive diagnosis, is likely to drive the future development of optimal treatment strategies for BA.

Overexpression of Mfsd2a attenuates blood brain barrier dysfunction via Cav-1/Keap-1/Nrf-2/HO-1 pathway in a rat model of surgical brain injury.

INTRODUCTION: Disruption of the blood brain barrier (BBB) and subsequent cerebral edema formation is one of the major adverse effects of brain surgery, leading to postoperative neurological dysfunction. Recently, Mfsd2a has been shown to have a crucial role for the maintenance of BBB functions. In this study, we aimed to evaluate the role of Mfsd2a on BBB disruption following surgical brain injury (SBI) in rats. MATERIALS AND METHODS: Rats were subjected to SBI by partial resection of the right frontal lobe. To evaluate the effect of Mfsd2a on BBB permeability and neurobehavior outcome following SBI, Mfsd2a was either overexpressed or downregulated in the brain by administering Mfsd2a CRISPR activation or knockout plasmids, respectively. The potential mechanism of Mfsd2a-mediated BBB protection through the cav-1/Nrf-2/HO-1 signaling pathway was evaluated. RESULTS: Mfsd2a levels were significantly decreased while cav-1, Nrf-2 and HO-1 levels were increased in the right frontal perisurgical area following SBI. When overexpressed, Mfsd2a attenuated brain edema and abolished neurologic impairment caused by SBI while downregulation of Mfsd2a expression further deteriorated BBB functions and worsened neurologic performance following SBI. The beneficial effect of Mfsd2a overexpression on BBB functions was associated with diminished expression of cav-1, increased Keap-1/Nrf-2 dissociation and further augmented levels of Nrf-2 and HO-1 in the right frontal perisurgical area, leading to enhanced levels of tight junction proteins following SBI. The BBB protective effect of Mfsd2a was blocked by selective inhibitors of Nrf-2 and HO-1. CONCLUSIONS: Mfsd2a attenuates BBB disruption through cav-1/Nrf-2/HO-1 signaling pathway in rats subjected to experimental SBI.

cGAS/STING Pathway Activation Contributes to Delayed Neurodegeneration in Neonatal Hypoxia-Ischemia Rat Model: Possible Involvement of LINE-1.

cGAS is a sensor of cytosolic DNA and responds equally to exogenous and endogenous DNA. After recognition of cytosolic dsDNA or ssDNA, cGAS synthesizes the second messenger 2'3'-cGAMP, which then binds to and activates stimulator of interferon genes (STING). STING plays an essential role in responding to pathogenic DNA and self-DNA in the context of autoimmunity. In pathologic conditions, such as stroke or hypoxia-ischemia (HI), DNA can gain access into the cytoplasm of the cell and leak from the dying cells into the extracellular environment, which potentially activates cGAS/STING. Recent in vivo studies of myocardial ischemia, traumatic brain injury, and liver damage models suggest that activation of cGAS/STING is not only a side-effect of the injury, but it can also actively contribute to cell death and apoptosis. We found, for the first time, that cGAS/STING pathway becomes activated between 24 and 48 h after HI in a 10-day-old rat model. Silencing STING with siRNA resulted in decreased infarction area, reduced cortical neurodegeneration, and improved neurobehavior at 48 h, suggesting that STING can contribute to injury progression after HI. STING colocalized with lysosomal marker LAMP-1 and blocking STING reduced the expression of cathepsin B and decreased the expression of Bax and caspase 3 cleavage. We observed similar protective effects after intranasal treatment with cGAS inhibitor RU.521, which were reversed by administration of STING agonist 2'3'-cGAMP. Additionally, we showed that long interspersed element 1 (LINE-1) retrotransposon, a potential upstream activator of cGAS/STING pathway was induced at 48 h after HI, which was evidenced by increased expression of ORF1p and ORF2p proteins and increased LINE-1 DNA content in the cytosol. Blocking LINE-1 with the nucleoside analog reverse-transcriptase inhibitor (NRTI) stavudine reduced infarction area, neuronal degeneration in the cerebral cortex, and reduced the expression of Bax and cleaved caspase 3. Thus, our results identify the cGAS/STING pathway as a potential therapeutic target to inhibit delayed neuronal death after HI.

Inhibition of PAR-2 Attenuates Neuroinflammation and Improves Short-Term Neurocognitive Functions Via ERK1/2 Signaling Following Asphyxia-Induced Cardiac Arrest in Rats.

OBJECTIVE: Global cerebral ischemia-induced neuroinflammation causes neurofunctional impairment following cardiac arrest. Previous studies have demonstrated that the activation of protease-activated receptor-2 (PAR-2) contributes to neuroinflammation. In the present study, we aimed to determine the potential treatment effect of PAR-2 inhibition against neuroinflammation in the setting of asphyxial CA (ACA) in rats. METHODS: A total of 116 adult, male Sprague-Dawley rats were randomly divided into Sham (n = 18) and ACA (n = 98) groups. Time course, short-term outcome, and mechanism studies were conducted. All drugs were delivered intranasally. The effect of PAR-2 inhibitor FSLLRY-NH2 on neurocognitive functions was assessed by neurologic deficit score, number of seizures, and T-maze test, while hippocampal neuronal degeneration was evaluated by Fluoro-Jade C staining after ACA. Western blotting was performed for the mechanism study at 24 h following ACA. Selective PAR-2 agonist (AC55541) and ERK1/2 inhibitor (PD98059) were used for intervention. RESULTS: Inhibition of PAR-2 decreased neuroinflammation, reduced the number of degenerating hippocampal neurons and improved neurocognitive functions following ACA. PAR-2 activator alone exerted opposite effects to PAR-2 inhibitor. PAR-2 mediated the augmented brain levels of proinflammatory cytokines by promoting the phosphorylation of ERK1/2. CONCLUSIONS: PAR-2 inhibition diminished neuroinflammation and thereby reduced hippocampal neuronal degeneration and neurocognitive impairment following ACA. This effect was at least partly mediated via the PAR-2/ERK1/2 signaling.

CCR1 Activation Promotes Neuroinflammation Through CCR1/TPR1/ERK1/2 Signaling Pathway After Intracerebral Hemorrhage in Mice.

The activation of C-C chemokine receptor type 1 (CCR1) has been shown to be pro-inflammatory in several animal models of neurological diseases. The objective of this study was to investigate the activation of CCR1 on neuroinflammation in a mouse model of intracerebral hemorrhage (ICH) and the mechanism of CCR1/tetratricopeptide repeat 1 (TPR1)/extracellular signal-regulated kinase 1/2 (ERK1/2) signaling pathway in CCR1-mediated neuroinflammation. Adult male CD1 mice (n = 210) were used in the study. The selective CCR1 antagonist Met-RANTES was administered intranasally at 1 h after autologous blood injection. To elucidate potential mechanism, a specific ERK1/2 activator (ceramide C6) was administered prior to Met-RANTES treatment; CCR1 activator (recombinant CCL5, rCCL5) and TPR1 CRISPR were administered in naïve mouse. Neurobehavioral assessments, brain water content, immunofluorescence staining, and western blot were performed. The endogenous expressions of CCR1, CCL5, TPR1, and p-ERK1/2 were increased in the brain after ICH. CCR1 were expressed on microglia, neurons, and astrocytes. The inhibition of CCR1 with Met-RANTES improved neurologic function, decreased brain edema, and suppressed microglia/macrophage activations and neutrophil infiltration after ICH. Met-RANTES treatment decreased expressions of CCR1, TPR1, p-ERK, TNF-α, and IL-1ß, which was reversed by ceramide C6. The brain CCR1 activation by rCCL5 injection in naïve mouse resulted in neurological deficits and increased expressions of CCR1, TPR1, p-ERK, TNF-α, and IL-1ß. These detrimental effects of rCCL5 were reversed by TPR1 knockdown using TPR1 CRISPR. Our study demonstrated that CCR1 activation promoted neuroinflammation through CCR1/TPR1/ERK1/2 signaling pathway after ICH in mice. CCR1 inhibition with Met-RANTES attenuated neuroinflammation, thereby reducing brain edema and improving neurobehavioral functions. Targeting CCR1 activation may provide a promising therapeutic approach in the management of ICH patients.