Tat-NTS peptide protects neurons against cerebral ischemia-reperfusion injury via ANXA1 SUMOylation in microglia

Abstract

Rationale: Recent studies indicate that microglial activation and the resulting inflammatory response could be potential targets of adjuvant therapy for ischemic stroke. Many studies have emphasized a well-established function of Annexin-A1 (ANXA1) in the immune system, including the regulation of microglial activation. Nevertheless, few therapeutic interventions targeting ANXA1 in microglia for ischemic stroke have been conducted. In the present study, Tat-NTS, a small peptide developed to prevent ANXA1 from entering the nucleus, was utilized. We discovered the underlying mechanism that Tat-NTS peptide targets microglial ANXA1 to protect against ischemic brain injury. Methods: Preclinical studies of ischemic stroke were performed using an oxygen-glucose deprivation and reperfusion (OGD/R) cell model in vitro and the middle cerebral artery occlusion (MCAO) animal model of ischemic stroke in vivo. Confocal imaging and 3D reconstruction analyses for detecting the protein expression and subcellular localization of microglia in vivo. Co-immunoprecipitation (Co-IP), immunoblotting, ELISA, quantitative real-time PCR (qRT-PCR), Luciferase reporter assay for determining the precise molecular mechanism. Measurement on the cytotoxicity of Tat-NTS peptide for microglia was assessed by CCK-8 and LDH assay. TUNEL staining was used to detect the microglia conditioned medium-mediated neuronal apoptosis. Adeno-associated viruses (AAVs) were injected into the cerebral cortex, striatum and hippocampal CA1 region of adult male Cx3cr1-Cre mice, to further verify the neurofunctional outcome and mechanism of Tat-NTS peptide by TTC staining, the modified Neurological Severity Score (mNSS) test, the open field test (OFT), the novel object recognition task (NORT), the Morris water maze (MWM) test, the long-term potentiation (LTP) and the Transmission electron microscopy (TEM). Results: It was observed that administration of Tat-NTS led to a shift of subcellular localization of ANXA1 in microglia from the nucleus to the cytoplasm in response to ischemic injury. Notably, this shift was accompanied by an increase in ANXA1 SUMOylation in microglia and a transformation of microglia towards an anti-inflammatory phenotype. We confirmed that Tat-NTS-induced ANXA1 SUMOylation in microglia mediated IKKα degradation via NBR1-dependent selective autophagy, then blocking the activation of the NF-κB pathway. As a result, the expression and release of the pro-inflammatory factors IL-1β and TNF-α were reduced in both in vitro and in vivo experiments. Furthermore, we found that Tat-NTS peptide's protective effect on microglia relieved ischemic neuron apoptosis. Finally, we demonstrated that Tat-NTS peptide administration, through induction of ANXA1 SUMOylation in microglia, reduced infarct volume, improved neurological function and facilitated behavioral recovery in MCAO mice. Conclusions: Our study provides evidence for a novel mechanism of Tat-NTS peptide in regu...

A cell‐penetrating peptide exerts therapeutic effects against ischemic stroke by mediating the lysosomal degradation of sirtuin 5

“…Moreover, our previous study reported that the cell‐penetrating peptide Tat‐NTS inhibits the binding of ANXA1 to importin β, significantly inhibiting the nuclear transport of ANXA1 in neurons and thereby protecting against ischemic stroke‐induced neuronal damage. 51 , 52 However, the present study revealed that Tat‐SIRT5‐CTM targets the SIRT5‐ANXA1 complex, resulting in SIRT5 degradation via the lysosomal pathway, which eventually alleviates microglia‐induced neuroinflammation and neuronal injury following ischemic stroke. 20 As the Tat‐NTS and Tat‐SIRT5‐CTM peptides act through different mechanisms in different cells, we supposed that a combination of the two different peptides would have greater efficacy.…”

Melanocortin Derivatives Induced Vascularization and Neuroglial Proliferation in the Rat Brain under Conditions of Cerebral Ischemia

“…Potential neuroprotectors should be aimed at protecting nerve cells from damage and death, restoring vascular function, affecting glial, vascular and inflammatory cells, restoring nervous tissue and brain function after ischemia [ 10 , 11 , 12 ]. Promising candidates include peptide-based drugs possessing antioxidant, anti-inflammatory, regenerative, antiplatelet, and anti-apoptotic properties.…”

Annexin 1 Reduces Dermatitis-Induced Itch and Cholestatic Itch through Inhibiting Neuroinflammation and Iron Overload in the Spinal Dorsal Horn of Mice

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