MicroRNA-379-5p targets MAP3K2 to reduce autophagy and alleviate neuronal injury following cerebral ischemia via the JNK/c-Jun signaling pathway.

MicroRNAs (miRNAs) are abundant in neurons and play key roles in the function and development of the nervous system. This study focuses on the function of miR-379-5p in neurological function recovery during ischemic stroke. The expression of miR-379-5p in the serum of patients with ischemic stroke was determined. Human cerebral cortical neuron cells (HCN-2) were subjected to oxygen/glucose deprivation (OGD) to mimic an ischemic stroke in vitro, whereas mice subjected to middle cerebral artery occlusion (MCAO) were used as an animal model. The serum of patients with ischemic stroke and OGD-treated HCN-2 cells displayed a poor expression of miR-379-5p. Upregulation of miR-379-5p reduced the OGD-induced cell damage and decreased the expression of the autophagy marker protein Beclin1 in cells. Rapamycin, an autophagy activator, blocked the protective functions of miR-379-5p. Further, miR-379-5p directly bound to MAP3K2. MAP3K2 activated the JNK/c-Jun signaling pathway and suppressed the neuroprotective events mediated by miR-379-5p. The in vitro results were reproduced in vivo, where upregulation of miR-379-5p reduced neurological impairment and infarct size in MCAO-induced mice. This study suggested that miR-379-5p showed a neuroprotective effect on ischemic stroke and reduced autophagy of neurons through the suppression of MAP3K2 and the JNK/c-Jun axis.

Autophagy and inflammation in ischemic stroke.

Appropriate autophagy has protective effects on ischemic nerve tissue, while excessive autophagy may cause cell death. The inflammatory response plays an important role in the survival of nerve cells and the recovery of neural tissue after ischemia. Many studies have found an interaction between autophagy and inflammation in the pathogenesis of ischemic stroke. This study outlines recent advances regarding the role of autophagy in the post-stroke inflammatory response as follows. (1) Autophagy inhibits inflammatory responses caused by ischemic stimulation through mTOR, the AMPK pathway, and inhibition of inflammasome activation. (2) Activation of inflammation triggers the formation of autophagosomes, and the upregulation of autophagy levels is marked by a significant increase in the autophagy-forming markers LC3-II and Beclin-1. Lipopolysaccharide stimulates microglia and inhibits ULK1 activity by direct phosphorylation of p38 MAPK, reducing the flux and autophagy level, thereby inducing inflammatory activity. (3) By blocking the activation of autophagy, the activation of inflammasomes can alleviate cerebral ischemic injury. Autophagy can also regulate the phenotypic alternation of microglia through the nuclear factor-κB pathway, which is beneficial to the recovery of neural tissue after ischemia. Studies have shown that some drugs such as resveratrol can exert neuroprotective effects by regulating the autophagy-inflammatory pathway. These studies suggest that the autophagy-inflammatory pathway may provide a new direction for the treatment of ischemic stroke.

Ischemic Postconditioning Alleviates Cerebral Ischemia-Reperfusion Injury Through Activating Autophagy During Early Reperfusion in Rats.

This study aimed to investigate whether ischemic postconditioning (IpostC) alleviates cerebral ischemia/reperfusion (I/R) injury involved in autophagy. Adult Sprague-Dawley rats were divided into five groups: sham (sham surgery), I/R (middle cerebral artery occlusion [MCAO] for 100 min, then reperfusion), IpostC (MCAO for 100 min, reperfusion for 10 min, MCAO for 10 min, then reperfusion), IpostC+3MA (3-methyladenine, an autophagy inhibitor, administered 30 min before first reperfusion), and IpostC+Veh (vehicle control for IpostC+3MA group). Infarct volume was measured using cresyl violet staining. Autophagy-related proteins were detected by western blot and immunohistochemistry. Autophagosomes, autophagolysosomes, and mitochondrial damage were identified by transmission electron microscopy. Cortical cell apoptosis was detected by the TUNEL assay. Neurologic function was assessed using the modified Neurologic Severity Score. IpostC improved neurological function and reduced infarct volume after I/R (P < 0.05). These effects of IpostC were inhibited by 3MA (P < 0.05). Autophagosome formation was increased in the I/R and IpostC+Veh groups (P < 0.05), but not in the IpostC+3MA group. The I/R group showed enhanced LC3-II/LC3-I ratio, p62, and Cathepsin B levels and decreased LAMP-2 level (all P < 0.05 vs. sham), indicating dysfunction of autophagic clearance. IpostC reduced p62 and Cathepsin B levels and increased the LC3-II/LC3-I ratio, and nuclear translocation of transcription factor EB (all P < 0.05); these effects of IpostC were reversed by 3MA, suggesting IpostC enhanced autophagic flux. Furthermore, IpostC attenuated I/R-induced mitochondrial translocation of Bax and mitochondrial cytochrome-c release (all P < 0.05); 3MA inhibited these effects of IpostC (P < 0.05). In conclusion, IpostC may alleviate cerebral I/R injury by activating autophagy during early reperfusion.

Oxygen-glucose deprivation regulates BACE1 expression through induction of autophagy in Neuro-2a/APP695 cells.

Our previous findings have demonstrated that autophagy regulation can alleviate the decline of learning and memory by eliminating deposition of extracellular beta-amyloid peptide (Aß) in the brain after stroke, but the exact mechanism is unclear. It is presumed that the regulation of beta-site APP-cleaving enzyme 1 (BACE1), the rate-limiting enzyme in metabolism of Aß, would be a key site. Neuro-2a/amyloid precursor protein 695 (APP695) cell models of cerebral ischemia were established by oxygen-glucose deprivation to investigate the effects of Rapamycin (an autophagy inducer) or 3-methyladenine (an autophagy inhibitor) on the expression of BACE1. Either oxygen-glucose deprivation or Rapamycin down-regulated the expression of BACE1 while 3-methyladenine up-regulated BACE1 expression. These results confirm that oxygen-glucose deprivation down-regulates BACE1 expression in Neuro-2a/APP695 cells through the introduction of autophagy.

Autophagy: a double-edged sword for neuronal survival after cerebral ischemia.

Evidence suggests that autophagy may be a new therapeutic target for stroke, but whether activation of autophagy increases or decreases the rate of neuronal death is still under debate. This review summarizes the potential role and possible signaling pathway of autophagy in neuronal survival after cerebral ischemia and proposes that autophagy has dual effects.

Clinical profile of Parkinson's disease in the Gumei community of Minhang district, Shanghai.

OBJECTIVE: We examined the demographic and clinical profiles of Parkinson's disease in Shanghai, China, to assist in disease management and provide comparative data on Parkinson's disease prevalence, phenotype, and progression among different regions and ethnic groups. METHODS: A door-to-door survey and follow-up clinical examinations identified 180 community-dwelling Han-Chinese Parkinson's disease patients (104 males, 76 females). RESULTS: The average age at onset was 65.16 ± 9.60 years. The most common initial symptom was tremor (112 patients, 62.22%), followed by rigidity (38, 21.11%), bradykinesia (28, 15.56%) and tremor plus rigidity (2, 1.11%). Tremor as the initial symptom usually began in a single limb (83.04% of patients). The average duration from onset to mild Parkinson's disease (Hoehn-Yahr phase 1-2) was 52.74 ± 45.64 months. Progression from mild to moderate/severe Parkinson's disease (phase ≥ 3) was significantly slower (87.07 ± 58.72 months; p<0.001), except for patients presenting initially with bradykinesia (53.83 ± 24.49 months). Most patients (149/180, 82.78%) took levodopa with or without other drugs. The Hamilton Anxiety Scale revealed symptoms of clinical anxiety in 35 patients, and the Hamilton Depression Scale revealed depressive symptoms in 88 patients. The depressed or anxious subgroup (123 patients) demonstrated a significantly younger age at onset (55.54 ± 7.68 years) compared with the overall mean (p<0.05). CONCLUSION: Unilateral limb tremor was the most common initial symptom, and motor function deteriorated slowly over ≅4-9 years. Earlier-onset patients experience greater psychiatric dysfunction.

Clinical profile of Parkinson's disease in the Gumei community of Minhang district, Shanghai

OBJECTIVE: We examined the demographic and clinical profiles of Parkinson's disease in Shanghai, China, to assist in disease management and provide comparative data on Parkinson's disease prevalence, phenotype, and progression among different regions and ethnic groups. METHODS: A door-to-door survey and follow-up clinical examinations identified 180 community-dwelling Han-Chinese Parkinson's disease patients (104 males, 76 females). RESULTS: The average age at onset was 65.16±9.60 years. The most common initial symptom was tremor (112 patients, 62.22%), followed by rigidity (38, 21.11%), bradykinesia (28, 15.56%) and tremor plus rigidity (2, 1.11%). Tremor as the initial symptom usually began in a single limb (83.04% of patients). The average duration from onset to mild Parkinson's disease (Hoehn-Yahr phase 1-2) was 52.74±45.64 months. Progression from mild to moderate/severe Parkinson's disease (phase≥3) was significantly slower (87.07±58.72 months; p<0.001), except for patients presenting initially with bradykinesia (53.83±24.49 months). Most patients (149/180, 82.78%) took levodopa with or without other drugs. The Hamilton Anxiety Scale revealed symptoms of clinical anxiety in 35 patients, and the Hamilton Depression Scale revealed depressive symptoms in 88 patients. The depressed or anxious subgroup (123 patients) demonstrated a significantly younger age at onset (55.54±7.68 years) compared with the overall mean (p<0.05). CONCLUSION: Unilateral limb tremor was the most common initial symptom, and motor function deteriorated slowly over ≅4−9 years. Earlier-onset patients experience greater psychiatric dysfunction. .

Mitophagy in ischaemia/reperfusion induced cerebral injury.

Mitochondrial autophagy (Mitophagy), the specific autophagic elimination of mitochondria, has been related with several forms of degenerative disease and mitochondrial dysfunction. It is involved in multiple cellular processes. In addition to one of its established key roles in the maintenance of normal cellular phenotype and function, there is growing interest in the concept that targeted modulation of mitophagy may reduce cerebral ischaemia/reperfusion injury. Induction of mitophagy results in selective clearance of damaged mitochondria in cells. In response to stress such as ischaemia/reperfusion, prosurvival and prodeath pathways are concomitantly activated in neuronal cells.

The role of autophagic and lysosomal pathways in ischemic brain injury.

Autophagy is involved in neural cell death after cerebral ischemia. Our previous studies showed that rapamycin-induced autophagy decreased the rate of apoptosis, but the rate of apoptosis was creased after the autophagy inhibitor, 3-methyladenine, was used. In this study, a suture-occluded method was performed to generate a rat model of brain ischemia. Under a transmission electron microscope, autophagic bodies and autophagy lysosomes were markedly accumulated in neurons at 4 hours post brain ischemic injury, with their numbers gradually reducing over time. Western blotting demonstrated that protein levels of light chain 3-II and cathepsin B were significantly increased within 4 hours of ischemic injury, but these levels were not persistently upregulated over time. Confocal microscopy showed that autophagy was mainly found in neurons with positive light chain 3 signal. Injection of rapamycin via tail vein promoted the occurrence of autophagy in rat brain tissue after cerebral ischemia and elevated light chain 3 and cathepsin B expression. However, injection of 3-methyladenine significantly diminished light chain 3-II and cathepsin B expression. Results verified that autophagic and lysosomal activity is increased in ischemic neurons. Abnormal components in cells can be eliminated through upregulating cell autophagy or inhibiting autophagy after ischemic brain injury, resulting in a dynamic balance of substances in cells. Moreover, drugs that interfere with autophagy may be potential therapies for the treatment of brain injury.

Inhibition of L-type Ca(2+) channels by curcumin requires a novel protein kinase-theta isoform in rat hippocampal neurons.

Curcumin, a major active compound of Curcuma longa, has been reported to have potent neuroprotective activities. However to date, the relevant mechanisms still remain unclear. In this study, we report that curcumin selectively inhibits L-type Ca(2+) channel currents in cultured rat hippocampal neurons. Whole-cell currents were recorded using 10mM barium as a charge carrier. Curcumin reversibly inhibited high-voltage-gated Ca(2+) channel (HVGCC) currents (IBa) in a concentration-dependent manner but had no apparent effects on the cells treated with nifedipine, a specific L-type Ca(2+) channel blocker. Curcumin did not markedly affect the activation of L-type Ca(2+) channels while significantly shifted the inactivation curve in the hyperpolarizing direction. Pretreatment of cells with the classical and novel PKC antagonists GF109203X and calphostin C completely abolished curcumin-induced IBa inhibition, whereas the classical PKC antagonist Gö6976 or inhibition of PKA activity elicited no such effects. Moreover, the curcumin-induced IBa response was abolished by intracellular application of the PKC-θ inhibitory peptide PKC-θ-IP or by siRNA knockdown of PKC-θ in cultured rat hippocampal neurons. In these neurons, novel isoforms of PKC including delta (PKC-δ), epsilon (PKC-ɛ) and theta (PKC-θ), but not eta (PKC-η), were endogenously expressed. Taken together, these results suggest that curcumin selectively inhibits IBavia a novel PKC-θ-dependent pathway, which could contribute to its neuroprotective effects in rat hippocampal neurons.

Therapeutic effects of rapamycin on MPTP-induced Parkinsonism in mice.

In neurodegenerative disorders such as Parkinson's disease (PD), autophagy is implicated in the process of dopaminergic neuron cell death. The α-synuclein protein is a major component of Lewy bodies and Lewy neurites, and mutations in α-synuclein have been implicated in the etiology of familial PD. The current work investigates the mechanisms underlying the therapeutic effects of the autophagy-stimulating antibiotic rapamycin in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD. Male C57BL/6 mice were treated with intravenous rapamycin or saline control for 7 days following MPTP administration. Immunohistochemistry and western blotting were used to detect alterations in the expression of PD biomarkers, including tyrosine hydroxylase (TH), and the level of autophagy was evaluated by the detection of both microtubule-associated protein light chain 3 (LC3) and α-Synuclein cleavage. In addition, levels of monoamine neurotransmitters were measured in the striatum using high performance liquid chromatography (HPLC). Immunohistochemistry using antibodies against TH indicated that the number of dopaminergic neurons in the substantia nigra following MPTP treatment was significantly higher in rapamycin-treated mice compared with saline-treated controls (p < 0.01). Levels of TH expression in the striatum were similar between the groups. α-synuclein Immunoreactivity was significantly decreased in rapamycin-treated mice compared with controls (p < 0.01). Immunoreactivity for LC3, however, was significantly higher in the rapamycin-treated animals than controls (p < 0.01). The concentrations of both striatal dopamine, and the dopamine metabolite DOPAC, were significantly decreased in both MPTP-treated groups compared with untreated controls. The loss of DOPAC was less severe in rapamycin-treated mice compared with saline-treated mice (p < 0.01) following MPTP treatment. These results demonstrate that treatment with rapamycin is able to prevent the loss of TH-positive neurons and to ameliorate the loss of DOPAC following MPTP treatment, likely via activation of autophagy/lysosome pathways. Thus, further investigation into the effectiveness of rapamycin administration in the treatment of PD is warranted.