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P16INK4a upregulation mediated by TBK1 induces retinal ganglion cell senescence in ischemic injury.

ABSTRACT: Glaucoma is a leading cause of irreversible blindness worldwide that is characterized by progressive retinal ganglion cell (RGC) death. However, RGC senescence as a phase before RGC death, and the mechanism of RGC senescence remains unclear. Here, we demonstrate that TANK-binding protein 1 (TBK1) is upregulated in acute IOP elevation-induced ischemic retinas mouse model. Moreover, pre-treatment with the TBK1 inhibitor BX-795 reduced p16INK4a (p16) expression and RGC senescence. Upregulation of TBK1 via plasmid transfection increased Akt phosphorylation at Ser473 and Bmi1 phosphorylation. The Akt inhibitor MK-2206 decreased the expression of p16 and Bmi1 serine phosphorylation. A Bmi1 Ser316 mutation also attenuated TBK1-induced p16 upregulation. Finally, silencing of TBK1 via shRNA knockdown reduced the expression of p16 as well as Akt and Bmi1 phosphorylation, reducing RGC senescence in vivo. These data suggest that acute IOP elevation-induced ischemia increases TBK1 expression, which then increases p16 expression through the Akt- Bmi1 phosphorylation pathway. This study therefore elucidates a novel mechanism whereby TBK1 regulates p16 expression and RGC senescence, suggesting a potential novel treatment strategy for minimizing RGC senescence in retinal ischemia and glaucoma.

SUBMITTER: Li LU

PROVIDER: S-EPMC5477587 | biostudies-literature | 2017 Apr

REPOSITORIES: biostudies-literature

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P16INK4a upregulation mediated by TBK1 induces retinal ganglion cell senescence in ischemic injury.

Li L U LU Zhao Yin Y Zhang Hong H

Cell death & disease 20170420 4

Glaucoma is a leading cause of irreversible blindness worldwide that is characterized by progressive retinal ganglion cell (RGC) death. However, RGC senescence as a phase before RGC death, and the mechanism of RGC senescence remains unclear. Here, we demonstrate that TANK-binding protein 1 (TBK1) is upregulated in acute IOP elevation-induced ischemic retinas mouse model. Moreover, pre-treatment with the TBK1 inhibitor BX-795 reduced p16INK4a (p16) expression and RGC senescence. Upregulation of T ...[more]

PMID: 28425986

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Project description:BACKGROUND:The preferential use of aerobic glycolysis by tumor cells lead to high accumulation of lactate in tumor microenvironment. Clinical evidence has linked elevated lactate concentration with cancer outcomes. However, the role and molecular mechanisms of lactate in cellular senescence and tumor progression remain elusive. METHODS:The function of Snail in lactate-induced EMT in lung cancer cells was explored by wound healing assay and cell invasion assay. The qRT-PCR and dual luciferase reporter assay were performed to investigate how lactate regulates Snail expression. The level of TGF-?1 in culture supernatant of cells was measured by ELISA for its correlation with extracellular levels of lactate. Ras activity assay and SA-?-gal activity assay were established to determine the effect of lactate on oncogene-induced senescence in human lung epithelial cells. ChIP assays were conducted to determine the binding of snail to p16INK4a promoter. Two TCGA data sets (TCGA-LUAD and TCGA-LUSC) were used to explore the correlations between SNAI1 and CDKN2A expression. RESULTS:In this study, we showed the invasive and migratory potential of lung cancer cells was significantly enhanced by lactate and was directly linked to snail activity. We also demonstrated that extracellular acidification itself is a direct cause of the increased snail expression and physiologically coupled to LDHA-dependent conversion of pyruvate to lactate. Mechanistically, lactate exerts its central function in induction of snail and EMT by directly remodeling ECM and releasing activated TGF-?1. We also demonstrated that Snail help premalignant cells to escape the oncogene-induced senescence by directly targeting and inhibiting p16INK4a expression. CONCLUSIONS:Our study extends the understanding of EMT in tumorigenesis by uncovering the role of snail in cellular senescence. This study also reveals lactate may be a potent tumor-promoting factor and provides the basis for the development of lactate-targeted therapy.

Project description:Cyclosporin A (CsA) inhibits the opening of the mitochondrial permeability transition pore (MPTP) by interacting with cyclophilin D (CypD) and ameliorates neuronal cell death in the central nervous system against ischemic injury. However, the molecular mechanisms underlying CypD/MPTP opening-mediated cell death in ischemic retinal injury induced by acute intraocular pressure (IOP) elevation remain unknown. We observed the first direct evidence that acute IOP elevation significantly upregulated CypD protein expression in ischemic retina at 12?h. However, CsA prevented the upregulation of CypD protein expression and promoted retinal ganglion cell (RGC) survival against ischemic injury. Moreover, CsA blocked apoptotic cell death by decreasing cleaved caspase-3 protein expression in ischemic retina. Of interest, although the expression level of Bcl-xL protein did not show a significant change in ischemic retina treated with vehicle or CsA at 12?h, ischemic damage induced the reduction of Bcl-xL immunoreactivity in RGCs. More importantly, CsA preserved Bcl-xL immunoreactivity in RGCs of ischemic retina. In parallel, acute IOP elevation significantly increased phosphorylated Bad (pBad) at Ser112 protein expression in ischemic retina at 12?h. However, CsA significantly preserved pBad protein expression in ischemic retina. Finally, acute IOP elevation significantly increased mitochondrial transcription factor A (Tfam) protein expression in ischemic retina at 12?h. However, CsA significantly preserved Tfam protein expression in ischemic retina. Studies on mitochondrial DNA (mtDNA) content in ischemic retina showed that there were no statistically significant differences in mtDNA content among control and ischemic groups treated with vehicle or CsA. Therefore, these results provide evidence that the activation of CypD-mediated MPTP opening is associated with the apoptotic pathway and the mitochondrial alteration in RGC death of ischemic retinal injury. On the basis of these observations, our findings suggest that CsA-mediated CypD inhibition may provide a promising therapeutic potential for protecting RGCs against ischemic injury-mediated mitochondrial dysfunction.

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P16INK4a upregulation mediated by TBK1 induces retinal ganglion cell senescence in ischemic injury.

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P16INK4a upregulation mediated by TBK1 induces retinal ganglion cell senescence in ischemic injury.

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