Sirtuin 4 (SIRT4) is one of seven mammalian sirtuins that possesses ADP-ribosyltransferase, lipoamidase and deacylase activities and plays indispensable role in metabolic regulation. However, the role of SIRT4 in the retina is not clearly understood. The purpose of this study was to explore the location and function of SIRT4 in the retina. Therefore, immunofluorescence was used to analyze the localization of SIRT4 in rat, mouse and human retinas. Western blotting was used to assess SIRT4 and glutamine synthetase (GS) protein expression at different developmental stages in C57BL/6 mice retinas. We further analyzed the retinal structure, electrophysiological function and the expression of GS protein in SIRT4-deficient mice. Excitotoxicity was caused by intravitreal injection of glutamate (50 nmol) in mice with long-term intraperitoneal injection of resveratrol (20 mg/Kg), and then retinas were subjected to Western blotting and paraffin section staining to analyze the effect of SIRT4 on excitotoxicity. We show that SIRT4 co-locates with Müller glial cell markers (GS and vimentin). The protein expression pattern of SIRT4 was similar to that of GS, and both increased with development. There were no significant retinal structure or electrophysiological function changes in 2-month SIRT4-deficient mice, while the expression of GS protein was decreased. Moreover, long-term administration of resveratrol can upregulate the expression of SIRT4 and GS while reducing the retinal injury caused by excessive glutamate. These results suggest that SIRT4 is highly expressed in retinal Müller glial cells and is relevant to the expression of GS. SIRT4 does not appear to be essential in retinal development, but resveratrol, as an activator of SIRT4, can upregulate GS protein expression and protect the retina from excitotoxicity.

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Müller glial cells (MGCs) are a group of glial cells in the retina that provide essential support to retinal neurons; however, the understanding of MGC apoptosis and autophagy remains limited. This study was aimed at investigating the role of autophagy in MGCs under normal and oxidative conditions, and identifying the underlying mechanisms. In addition, the sirtuin 4 (SIRT4)-mediated signaling pathway was observed to regulate the autophagic process in MGCs. To assess the effect of autophagy on MGC mitochondrial function and survival, we treated rMC-1 cells—rat-derived Müller glial cells—with rapamycin and 3-methyladenine (3-MA), and found that MGC death was not induced by such treatment, while autophagic dysfunction could increase MGC apoptosis under oxidative stress, as reflected by the expression level of cleaved caspase 3 and PI staining. In addition, the downregulation of autophagy by 3-MA could influence the morphology of the mitochondrial network structure, the mitochondrial membrane potential, and generation of reactive oxygen species (ROS) under oxidative stress. Moreover, SIRT4 depletion enhanced autophagosome formation, as verified by an increase in the LC3 II/I ratio and a decrease in the expression of SQSTM1/p62, and vice versa. The inhibition of AMPK phosphorylation by compound C could reverse these changes in LC3 II/I and SQSTM1/p62 caused by SIRT4 knockdown. Our research concludes that MGCs can endure autophagic dysfunction in the absence of oxidative stress, while the downregulation of autophagy can cause MGCs to become more sensitized to oxidative stress. Simultaneous exposure to oxidative stress and autophagic dysfunction in MGCs can result in a pronounced impairment of cell survival. Mechanically, SIRT4 depletion can activate the autophagic process in MGCs by regulating the AMPK–mTOR signaling pathway.

First, the constitutive equation of the linear elastic and the basic equations flexibility of the unloading rock mass were introduced, and then using the equations and the boundary conditions the variation principle was contracture. Prior to construct an appropriate minimum potential energy functional, on this basis, pending the introduction of two Lagrange multipliers, and absorb the variation constraints to establish the new functional. Last will 、、、as an independent variable, considering the variation stationary conditions functional of the new functional can identify the Lagrange multipliers 、 to be determined, so can build the generalized variation principle of dual variables