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CPKC?-Modulated Sequential Reactivation of mTOR Inhibited Autophagic Flux in Neurons Exposed to Oxygen Glucose Deprivation/Reperfusion.

ABSTRACT: We have reported that conventional protein kinase Cγ (cPKCγ)-modulated neuron-specific autophagy improved the neurological outcome of mice following ischemic stroke through the Akt-mechanistic target of rapamycin (mTOR) pathway. However, its detailed molecular mechanism remains unclear. In this study, primary cortical neurons from postnatal one-day-old C57BL/6J cPKCγ wild-type (cPKCγ+/+) and knockout (cPKCγ−/−) mice suffering oxygen glucose deprivation/reperfusion (OGD/R) were used to simulate ischemia/reperfusion injury in vitro. A block of autophagic flux was observed in cPKCγ+/+ neurons under OGD/R exposure, characterized by accumulation of p62. Immunofluorescent results showed a decrease in colocalization between LC3 and Atg14 or Stx17 in cPKCγ+/+ neurons when compared with cPKCγ−/− neurons after OGD/R. However, the colocalization between LC3 and Lamp2 was barely decreased, indicating the presence of autolysosomes. The larger lysotracker-positive structures were also significantly increased. These results suggest that cPKCγ-induced inhibition of autophagy occurred at the stages of autophagosome formation, Stx17 anchoring, and the degradation of autolysosomes in particular. In addition, cPKCγ-modulated phosphorylation of mTOR at Ser 2481 was dependent on the site of Ser 2448, which may have blocked autophagic flux. cPKCγ-modulated sequential reactivation of mTOR inhibited autophagic flux in neurons exposed to OGD/R, which may provide endogenous interventional strategies for stroke, especially ischemia/reperfusion injury.

SUBMITTER: Hua R

PROVIDER: S-EPMC5983661 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

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cPKCγ-Modulated Sequential Reactivation of mTOR Inhibited Autophagic Flux in Neurons Exposed to Oxygen Glucose Deprivation/Reperfusion.

Hua Rongrong R Han Song S Zhang Nan N Dai Qingqing Q Liu Ting T Li Junfa J

International journal of molecular sciences 20180506 5

We have reported that conventional protein kinase Cγ (cPKCγ)-modulated neuron-specific autophagy improved the neurological outcome of mice following ischemic stroke through the Akt-mechanistic target of rapamycin (mTOR) pathway. However, its detailed molecular mechanism remains unclear. In this study, primary cortical neurons from postnatal one-day-old C57BL/6J cPKCγ wild-type (cPKCγ+/+) and knockout (cPKCγ−/−) ...[more]

PMID: 29734780

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Project description:Reperfusion injury after extended ischemia accounts for approximately 50% of myocardial infarct size, and there is no standard therapy. HDAC inhibition reduces infarct size and enhances cardiomyocyte autophagy and PGC1α-mediated mitochondrial biogenesis when administered at the time of reperfusion. Furthermore, a specific autophagy-inducing peptide, Tat-Beclin 1 (TB), reduces infarct size when administered at the time of reperfusion. However, since SAHA affects multiple pathways in addition to inducing autophagy, whether autophagic flux induced by TB maintains mitochondrial homeostasis during ischemia/reperfusion (I/R) injury is unknown. We tested whether the augmentation of autophagic flux by TB has cardioprotection by preserving mitochondrial homeostasis both in vitro and in vivo. Wild-type mice were randomized into two groups: Tat-Scrambled (TS) peptide as the control and TB as the experimental group. Mice were subjected to I/R surgery (45 min coronary ligation, 24 h reperfusion). Autophagic flux, mitochondrial DNA (mtDNA), mitochondrial morphology, and mitochondrial dynamic genes were assayed. Cultured neonatal rat ventricular myocytes (NRVMs) were treated with a simulated I/R injury to verify cardiomyocyte specificity. The essential autophagy gene, ATG7, conditional cardiomyocyte-specific knockout (ATG7 cKO) mice, and isolated adult mouse ventricular myocytes (AMVMs) were used to evaluate the dependency of autophagy in adult cardiomyocytes. In NRVMs subjected to I/R, TB increased autophagic flux, mtDNA content, mitochondrial function, reduced reactive oxygen species (ROS), and mtDNA damage. Similarly, in the infarct border zone of the mouse heart, TB induced autophagy, increased mitochondrial size and mtDNA content, and promoted the expression of PGC1α and mitochondrial dynamic genes. Conversely, loss of ATG7 in AMVMs and in the myocardium of ATG7 cKO mice abolished the beneficial effects of TB on mitochondrial homeostasis. Thus, autophagic flux is a sufficient and essential process to mitigate myocardial reperfusion injury by maintaining mitochondrial homeostasis and partly by inducing PGC1α-mediated mitochondrial biogenesis.

Dataset Information

CPKC?-Modulated Sequential Reactivation of mTOR Inhibited Autophagic Flux in Neurons Exposed to Oxygen Glucose Deprivation/Reperfusion.

Publications

cPKCγ-Modulated Sequential Reactivation of mTOR Inhibited Autophagic Flux in Neurons Exposed to Oxygen Glucose Deprivation/Reperfusion.

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