Emerging Role of Ferroptosis in the Pathogenesis of Ischemic Stroke: A New Therapeutic Target?

Ischemic stroke is one of the main causes of high morbidity, mortality, and disability worldwide; however, the treatment methods are limited and do not always achieve satisfactory results. The pathogenesis of ischemic stroke is complex, defined by multiple mechanisms; among them, programmed death of neuronal cells plays a significant role. Ferroptosis is a novel type of regulated cell death characterized by iron redistribution or accumulation and increased lipid peroxidation in the membrane. Ferroptosis is implicated in many pathological conditions, such as cancer, neurodegenerative diseases, and ischemia-reperfusion injury. In this review, we summarize current research findings on ferroptosis, including possible molecular mechanisms and therapeutic applications of ferroptosis regulators, with a focus on the involvement of ferroptosis in the pathogenesis and treatment of ischemic stroke. Understanding the role of ferroptosis in ischemic stroke will throw some light on the development of methods for diagnosis, treatment, and prevention of this devastating disease.

A novel CCM3 mutation associated with cerebral cavernous malformation in a Chinese family.

BACKGROUND: Cerebral cavernous malformation (CCM), especially the familial form, is a relatively rare congenital and occult vascular disease of the central nervous system. The familial form of CCM has been linked to three different genes: KRIT1/CCM1, MGC4607/CCM2, and PDCD10/CCM3; however, the genetic basis of CCM is not well understood. The PDCD10/CCM3 is the most recent gene to be identified that results in worse clinical symptoms. Early diagnosis and treatment is important for patient prognosis. CASE REPORT: The proband is a 38-year-old male who has been suffering from weakness in the limbs for 7 months. Investigation of his family history revealed that his mother also suffered from limbs paralysis and had been bedridden for a long time. His older brother suffered from headache for years, whereas his younger brother was asymptomatic. Brain computed tomography analysis of all family members showed multiple high-density shadows. Subsequently, magnetic resonance imaging analysis identified more prominent and similar multiple intracranial lesions in all family members. The lesions were hypo-intense, or showed mixed signs on T1-weighted imaging, and were significantly more intense on T2-weighted imaging. To understand the genetic basis of the disease in the family, DNA sequencing analysis was performed. A novel deletion mutation in the PDCD10/CCM3 gene was identified in the proband and his relatives. The deletion resulted in a frameshift mutation and premature termination of translation of the protein, and potentially caused the disease in this family. CONCLUSIONS: Our study identified a novel PDCD10/CCM3 heterozygous deletion (c.165delT) associated with CCM. This finding expands the CCM gene mutation profile, which will be beneficial for genetic counseling and clinical therapy.

Constitutive expression of NtabSPL6-1 in tobacco and Arabidopsis could change the structure of leaves and promote the development of trichomes.

The coding sequence of NtabSPL6-1 was cloned by high-fidelity PCR with specific primers and was used in construction of a binary vector for overexpression. Wild-type Col-0 Arabidopsis plants and Qinyan95 tobacco leaves were transformed using floral dip and leaf disc methods, respectively. Phenotypic observation showed that constitutive expression of NtabSPL6-1 in Arabidopsis could promote the development of trichomes on leaf epidermis and influence the growth pattern of cauline leaves. In tobacco, ectopic expression of NtabSPL6-1 led to dwarfism of the plants and alteration of the leaf structure, accompanied by changes of the glandular trichomes in development. At the same time, the self-regulation capability of NtabSPL6-1 was determined by yeast two-hybrid system. The results indicated that SBP-C terminal domain and C terminal domain of NtabSPL6-1 possessed strong transcriptional activation ability; the intact protein, N terminal domain, and the first peptide fragment in N terminal domain possessed weak transcriptional activation ability; and the second and the third peptide fragments in N terminal domain had no transcriptional activation ability, suggesting the N terminal domain of NtabSPL6-1 could block the activity of the C terminal domain. NtabSPL6-1 may affect the resistance of plants to biotic stress factors indirectly by regulation of the trichome growth.

Gerstmann-Sträussler-Scheinker disease: A case report.

BACKGROUND: Gerstmann-Sträussler-Scheinker (GSS) disease is an inherited prion disease that is clinically characterized by the early onset of progressive cerebellar ataxia. The incidence of GSS is extremely low and it is particularly rare in China. Therefore, clinicians may easily confuse this disease with other diseases that also cause ataxia, resulting in its under-diagnosis or misdiagnosis. CASE SUMMARY: Here, we report the first case of genetically diagnosed GSS disease in Northeast China. The patient exhibited typical ataxia and dysarthria 2.5 years after symptom onset. However, magnetic resonance imaging of the brain and spinal cord revealed a normal anatomy. Screening results for the spinocerebellar ataxia gene were also negative. We thus proposed to expand the scope of genetic screening to include over 200 mutations that can cause ataxia. A final diagnosis of GSS was presented and the patient was followed for more than 3.5 years, during which we noted imaging abnormalities. The patient gradually exhibited decorticate posturing and convulsions. We recommended administration of oral sodium valproate, which resolved the convulsions. CONCLUSION: Patients with inherited ataxia should be considered for a diagnosis of GSS via genetic testing at an early disease stage.

Dynamic Expression of CX36 Protein in Kainic Acid Kindling induced Epilepsy.

Connexin (Cx) 36 is known to be a component of gap junctions, and has been suggested to play an important role in epilepsy. In order to determine dynamic changes of Cx36 protein expression in epilepsy and investigate the role of Cx36 in electroencephalographic activity and pathogenesis, we utilized kainic acid (KA) to induce epileptogenesis. We found that epileptic discharges began 71.8 ± 23.7 s after KA administration. Spike frequency and amplitude of epileptiform activity reached maximal levels at 30 ± 5.2 min. The maximum level of spike frequency and amplitude of epileptiform activity was 13.9 ± 0.3 Hz and 198 ± 14.3mV respectively. Employing Western blotting and immunohistochemistry, we demonstrated that hippocampal Cx36 protein expression was significantly increased 6 h after KA kindling compared to control or sham groups, but decreased in 3 d and 7d groups. Our results suggested that the dynamic change of Cx36 expression may play an important role inepilepsy, and the specific manipulation of Cx36 expression may be a potential target for the treatment of epilepsy.

Overview of Experimental and Clinical Findings regarding the Neuroprotective Effects of Cerebral Ischemic Postconditioning.

Research on attenuating the structural and functional deficits observed following ischemia-reperfusion has become increasingly focused on the therapeutic potential of ischemic postconditioning. In recent years, various methods and animal models of ischemic postconditioning have been utilized. The results of these numerous studies have indicated that the mechanisms underlying the neuroprotective effects of ischemic postconditioning may involve reductions in the generation of free radicals and inhibition of calcium overload, as well as the release of endogenous active substances, alterations in membrane channel function, and activation of protein kinases. Here we review the novel discovery, mechanism, key factors, and clinical application of ischemic postconditioning and discuss its implications for future research and problem of clinical practice.

Noninvasive and quantitative intracranial pressure estimation using ultrasonographic measurement of optic nerve sheath diameter.

We aimed to quantitatively assess intracranial pressure (ICP) using optic nerve sheath diameter (ONSD) measurements. We recruited 316 neurology patients in whom ultrasonographic ONSD was measured before lumbar puncture. They were randomly divided into a modeling and a test group at a ratio of 7:3. In the modeling group, we conducted univariate and multivariate analyses to assess associations between ICP and ONSD, age, sex, BMI, mean arterial blood pressure, diastolic blood pressure. We derived the mathematical function "Xing &Wang" from the modelling group to predict ICP and evaluated the function in the test group. In the modeling group, ICP was strongly correlated with ONSD (r = 0.758, p < 0.001), and this association was independent of other factors. The mathematical function was ICP = -111.92 + 77.36 × ONSD (Durbin-Watson value = 1.94). In the test group, a significant correlation was found between the observed and predicted ICP (r = 0.76, p < 0.001). Bland-Altman analysis yielded a mean difference between measurements of -0.07 ± 41.55 mmH2O. The intraclass correlation coefficient and its 95%CIs for noninvasive ICP assessments using our prediction model was 0.86 (0.79-0.90). Ultrasonographic ONSD measurements provide a potential noninvasive method to quantify ICP that can be conducted at the bedside.

A case report: Becker muscular dystrophy presenting with epilepsy and dysgnosia induced by duplication mutation of Dystrophin gene.

BACKGROUND: Becker muscular dystrophy (BMD), a genetic disorder of X-linked recessive inheritance, typically presents with gradually progressive muscle weakness. The condition is caused by mutations of Dystrophin gene located at Xp21.2. Epilepsy is an infrequent manifestation of BMD, while cases of BMD with dysgnosia are extremely rare. CASE PRESENTATION: We describe a 9-year-old boy with BMD, who presented with epilepsy and dysgnosia. Serum creatine kinase level was markedly elevated (3665 U/L). Wechsler intelligence tests showed a low intelligence quotient (IQ = 65). Electromyogram showed slight myogenic changes and skeletal muscle biopsy revealed muscular dystrophy. Immunohistochemical staining showed partial positivity of sarcolemma for dystrophin-N. Multiplex ligation-dependent probe amplification revealed a duplication mutation in exons 37-44 in the Dystrophin gene. CONCLUSIONS: The present case report helps to better understand the clinical and genetic features of BMD.

Neuroprotective Effect of Salvianolic Acids against Cerebral Ischemia/Reperfusion Injury.

This study investigated the neuroprotective effect of salvianolic acids (SA) against ischemia/reperfusion (I/R) injury, and explored whether the neuroprotection was dependent on mitochondrial connexin43 (mtCx43) via the phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT) pathway. In vitro, we measured astrocyte apoptosis, mitochondrial membrane potential, and also evaluated the morphology of astrocyte mitochondria with transmission electron microscopy. In vivo, we determined the cerebral infarction volume and measured superoxide dismutase (SOD) activity and malondialdehyde (MDA) content. Additionally, mtCx43, p-mtCx43, AKT, and p-AKT levels were determined. In vitro, we found that I/R injury induced apoptosis, decreased cell mitochondrial membrane potential (MMP), and damaged mitochondrial morphology in astrocytes. In vivo, we found that I/R injury resulted in a large cerebral infarction, decreased SOD activity, and increased MDA expression. Additionally, I/R injury reduced both the p-mtCx43/mtCx43 and p-AKT/AKT ratios. We reported that both in vivo and in vitro, SA ameliorated the detrimental outcomes of the I/R. Interestingly, co-administering an inhibitor of the PI3K/AKT pathway blunted the effects of SA. SA represents a potential treatment option for cerebral infarction by up-regulating mtCx43 through the PI3K/AKT pathway.

Mechanism of Mitochondrial Connexin43's Protection of the Neurovascular Unit under Acute Cerebral Ischemia-Reperfusion Injury.

We observed mitochondrial connexin43 (mtCx43) expression under cerebral ischemia-reperfusion (I/R) injury, analyzed its regulation, and explored its protective mechanisms. Wistar rats were divided into groups based on injections received before middle cerebral artery occlusion (MCAO). Cerebral infarction volume was detected by 2,3,5-triphenyltetrazolim chloride staining, and cell apoptosis was observed by transferase dUTP nick end labeling. We used transmission electron microscopy to observe mitochondrial morphology and determined superoxide dismutase (SOD) activity and malondialdehyde (MDA) content. MtCx43, p-mtCx43, protein kinase C (PKC), and p-PKC expression were detected by Western blot. Compared with those in the IR group, cerebral infarction volumes in the carbenoxolone (CBX) and diazoxide (DZX) groups were obviously smaller, and the apoptosis indices were down-regulated. Mitochondrial morphology was damaged after I/R, especially in the IR and 5-hydroxydecanoic acid (5-HD) groups. Similarly, decreased SOD activity and increased MDA were observed after MCAO; CBX, DZX, and phorbol-12-myristate-13-acetate (PMA) reduced mitochondrial functional injury. Expression of mtCx43 and p-mtCx43 and the p-Cx43/Cx43 ratio were significantly lower in the IR group than in the sham group. These abnormalities were ameliorated by CBX, DZX, and PMA. MtCx43 may protect the neurovascular unit from acute cerebral IR injury via PKC activation induced by mitoKATP channel agonists.

Cortical spreading depression-induced preconditioning in the brain.

Cortical spreading depression is a technique used to depolarize neurons. During focal or global ischemia, cortical spreading depression-induced preconditioning can enhance tolerance of further injury. However, the underlying mechanism for this phenomenon remains relatively unclear. To date, numerous issues exist regarding the experimental model used to precondition the brain with cortical spreading depression, such as the administration route, concentration of potassium chloride, induction time, duration of the protection provided by the treatment, the regional distribution of the protective effect, and the types of neurons responsible for the greater tolerance. In this review, we focus on the mechanisms underlying cortical spreading depression-induced tolerance in the brain, considering excitatory neurotransmission and metabolism, nitric oxide, genomic reprogramming, inflammation, neurotropic factors, and cellular stress response. Specifically, we clarify the procedures and detailed information regarding cortical spreading depression-induced preconditioning and build a foundation for more comprehensive investigations in the field of neural regeneration and clinical application in the future.

Neuroprotection by Carbenoxolone Against Ischemia Injury Involves PI3K/Akt Pathway.

BACKGROUND: Previous experimental studies have shown some protective effects on brain ischemic injures in vivo and in vitro models. However, cotreatment with carbenoxolone (Cbx) and phosatidylinositol 3-kinase (PI3K) inhibitor LY 294002 to a focal cerebral ischemia and reperfusion rat model has not been studied yet. Here we investigate their protective effects on neural cells and examine the function of PI3K/Akt pathway in this protection. METHODS: Both flow cytometry and western blot were used quantitatively and qualitatively to determine cell apoptosis. RESULTS: The neural cell apoptosis is related with Cx43, and Bcl-2/Bax and caspase 3 pathways. The percentage of apoptosis cells following transient middle cerebral artery occlusion (MCAO) in mice decrease with the treatment of Cbx. Our data demonstrated that treatment with Cbx reduced cerebral injury in rats exposed to transient focal ischemia and reperfusion (I/R), and this was mediated by the activation of the PI3K/Akt pathways as well as by blocking the caspase 3 apoptosis pathway. LY294002 blocked the increase in phospho-AKT evoked by Cbx and abolished the associated protective effect. CONCLUSIONS: Taken together, these findings provide evidence that Cbx protects neurons against ischemia injury and this neuroprotective effect involves the PI3K/Akt pathway.

Connexin: a potential novel target for protecting the central nervous system?

Connexin subunits are proteins that form gap junction channels, and play an important role in communication between adjacent cells. This review article discusses the function of connexins/hemichannels/gap junctions under physiological conditions, and summarizes the findings regarding the role of connexins/hemichannels/gap junctions in the physiological and pathological mechanisms underlying central nervous system diseases such as brain ischemia, traumatic brain and spinal cord injury, epilepsy, brain and spinal cord tumor, migraine, neuroautoimmune disease, Alzheimer's disease, Parkinson's disease, X-linked Charcot-Marie-Tooth disease, Pelizaeus-Merzbacher-like disease, spastic paraplegia and maxillofacial dysplasia. Connexins are considered to be a potential novel target for protecting the central nervous system.

Wnt-5a prevents Aß-induced deficits in long-term potentiation and spatial memory in rats.

Although the neurotoxicity of amyloid ß (Aß) protein in Alzheimer's disease (AD) has been reported widely, the exact molecular mechanism underlying the Aß-induced synaptic dysfunction and memory impairment remains largely unclear. Growing evidence indicates that wingless-type (Wnt) signaling plays an important role in neuronal development, synapse formation and synaptic plasticity. In the present study, we investigated the neuroprotective action of Wnt-5a against the synaptic damage and memory deficit induced by Aß25-35 by using in vivo electrophysiological recording and Morris water maze (MWM) test. We found that intracerebroventricular (i.c.v.) injection of Aß25-35 alone did not affect the baseline field excitatory postsynaptic potentials (fEPSPs) and the paired-pulse facilitation (PPF) in the hippocampal CA1 region of rats, but significantly suppressed high frequency stimulation (HFS) induced long-term potentiation (LTP); pretreatment with Wnt-5a prevented the Aß25-35-induced suppression of hippocampal LTP in a dose-dependent manner; soluble Frizzled-related protein (sFRP), a specific Wnt antagonist, effectively attenuated the protective effects of Wnt-5a. In MWM test, Aß25-35 alone significantly disrupted spatial learning and memory ability of rats, while pretreatment with Wnt-5a effectively prevented the impairments induced by Aß25-35. These results in the present study demonstrated for the first time the neuroprotective effects of Wnt-5a against Aß-induced in vivo synaptic plasticity impairment and memory disorder, suggesting that Wnt signaling pathway is one of the important targets of Aß neurotoxicity and Wnt-5a might be used as one of the putative candidates for the therapeutic intervention of AD.

Effect of connexin 36 blockers on the neuronal cytoskeleton and synaptic plasticity in kainic acid-kindled rats.

In this study we investigated the potential anti-epileptogenic effect of neuronal connexin Cx36 gap junction blockage via inhibition of microtubule-associated protein 2 (MAP-2) and synaptophysin (SYP) overexpression. Thirty adult male Wistar rats were divided into five groups (six animals per group): control, sham, carbenoxolone (CBX), quinine (QN), and quinidine (QND). An epilepsy model was produced by injecting kainic acid (KA) into the rat amygdala. Broad-spectrum and selective blockers of the Cx36 channel (CBX, QN, and QND) were administered via intraperitoneal injection. Expression of MAP-2 and SYP was assessed by immunofluorescent and immunohistochemical examination. Expression of MAP-2 and SYP was significantly increased after KA administration in the sham group compared with the control group. Expression of MAP-2 and SYP was significantly decreased in the CBX, QN, and QND groups compared with the sham group. The results provide new evidence regarding the key role of MAP-2 and SYP overexpression in three important mechanisms: the modulation of neuronal plasticity, hyperexcitability of the hippocampal neuronal network, and persistent seizure discharge. Furthermore, the reversal of MAP-2 and SYP overexpression following administration of Cx36 channel blockers indicates a potential role for Cx36 channel blockers in anti-epileptogenic treatment and in doing so, highlights a critical need for further investigation of these compounds.

Combined occurrence of a novel TOR1A and a THAP1 mutation in primary dystonia.

BACKGROUND: The ΔGAG deletion of the TOR1A gene (DYT1) is responsible for DYT1 dystonia. However, no other TOR1A mutation has been reported in the Chinese population. METHODS: Two hundred one dystonia patients without the ΔGAG deletion were screened for other mutations in TOR1A. Gene function changes were analyzed by subcellular distribution and luciferase reporter assay. RESULTS: A novel TOR1A mutation (c.581A>T, p.Asp194Val) was found in a patient with early-onset segmental dystonia harboring a THAP1 mutation (c.539T>C, p.Leu180Ser). Overexpression of mutant TOR1A Asp194Val protein induces inclusion formation in SK-N-AS cell lines, and the repressive activity of the mutant THAP1 Leu180Ser protein on TOR1A gene expression is decreased compared with wild-type THAP1. CONCLUSIONS: This is the first report about a dystonia patient harboring two distinct dystonia gene mutations. Functional analysis indicated a potential additive effect of these two mutations, which might provoke the occurrence of dystonic symptoms in this patient.

Mutation screening of GNAL gene in patients with primary dystonia from Northeast China.

BACKGROUND: Mutations in GNAL have recently been identified as responsible for primary dystonia, however, GNAL mutations in Chinese patients with primary dystonia are not well characterized. PATIENTS AND METHODS: Fifty-nine unrelated patients with cervical onset or cervical involved primary dystonia and 120 neurologically normal controls from Northeast China without mutations of TOR1A and THAP1 were all screened for mutation of GNAL gene. RESULTS: One subject with adult-onset generalized dystonia was found have a novel nonsense GNAL mutation (c.284C>T, p.Ser95X). Another subject with adult-onset cervical dystonia was found harbor the c.932-7T>G tentative splice site mutation. Although another seventeen sequence variants were identified in both patients and controls, no disease association was found among these sequence variants. CONCLUSIONS: Mutations in GNAL gene can cause adult-onset primary dystonia in Chinese patients, and the mutation frequency is 3.4% in cervical onset or cervical involved primary dystonia. This paper identifies the first case of GNAL dystonia in the Chinese population.

Adult-onset leukoencephalopathy with brain stem and spinal cord involvement in Chinese Han population: a case report and literature review.

Leukoencephalopathy with brainstem and spinal cord involvement and lactate elevation (LBSL) is a recently described disorder with an autosomal recessive mode of inheritance. We report a case of rare adult-onset LBSL with typical magnetic resonance imaging (MRI) features. The DARS2 gene mutation analysis has identified a c. 228-20_21delTTinsC (p.R76SfsX5) mutation and a c. 850G > A (p. 284E > K) mutation. With glucocorticosteroid treatment the patient has had improvement in bladder symptoms. This is the first reported adult-onset LBSL case in the Chinese Han population. A review of the literature suggests that brain lactate elevation in adult-onset LBSL is lower than early-onset cases (P < 0.01), and early-onset cases show mild intelligence and cognition decline. These observations suggest that age of onset and brain lactate levels probably influence the prognosis of LBSL.