Anti-aging Effects of Alu Antisense RNA on Human Fibroblast Senescence Through the MEK-ERK Pathway Mediated by KIF15.

OBJECTIVE: To investigate whether human short interspersed nuclear element antisense RNA (Alu antisense RNA; Alu asRNA) could delay human fibroblast senescence and explore the underlying mechanisms. METHODS: We transfected Alu asRNA into senescent human fibroblasts and used cell counting kit-8 (CCK-8), reactive oxygen species (ROS), and senescence-associated beta-galactosidase (SA-ß-gal) staining methods to analyze the anti-aging effects of Alu asRNA on the fibroblasts. We also used an RNA-sequencing (RNA-seq) method to investigate the Alu asRNA-specific mechanisms of anti-aging. We examined the effects of KIF15 on the anti-aging role induced by Alu asRNA. We also investigated the mechanisms underlying a KIF15-induced proliferation of senescent human fibroblasts. RESULTS: The CCK-8, ROS and SA-ß-gal results showed that Alu asRNA could delay fibroblast aging. RNA-seq showed 183 differentially expressed genes (DEGs) in Alu asRNA transfected fibroblasts compared with fibroblasts transfected with the calcium phosphate transfection (CPT) reagent. The KEGG analysis showed that the cell cycle pathway was significantly enriched in the DEGs in fibroblasts transfected with Alu asRNA compared with fibroblasts transfected with the CPT reagent. Notably, Alu asRNA promoted the KIF15 expression and activated the MEK-ERK signaling pathway. CONCLUSION: Our results suggest that Alu asRNA could promote senescent fibroblast proliferation via activation of the KIF15-mediated MEK-ERK signaling pathway.

Alu antisense RNA ameliorates methylglyoxal-induced human lens epithelial cell apoptosis by enhancing antioxidant defense.

AIM: To determine whether an antisense RNA corresponding to the human Alu transposable element (Aluas RNA) can protect human lens epithelial cells (HLECs) from methylglyoxal-induced apoptosis. METHODS: Cell counting kit-8 (CCK-8) and 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays were used to assess HLEC viability. HLEC viability/death was detected using a Calcein-AM/PI double staining kit; the annexin V-FITC method was used to detect HLEC apoptosis. The cytosolic reactive oxygen species (ROS) levels in HLECs were determined using a reactive species assay kit. The levels of malondialdehyde (MDA) and the antioxidant activities of total-superoxide dismutase (T-SOD) and glutathione peroxidase (GSH-Px) were assessed in HLECs using their respective kits. RT-qPCR and Western blotting were used to measure mRNA and protein expression levels of the genes. RESULTS: Aluas RNA rescued methylglyoxal-induced apoptosis in HLECs and ameliorated both the methylglyoxal-induced decrease in Bcl-2 mRNA and the methylglyoxal-induced increase in Bax mRNA. In addition, Aluas RNA inhibited the methylglyoxal-induced increase in Alu sense RNA expression. Aluas RNA inhibited the production of ROS induced by methylglyoxal, restored T-SOD and GSH-Px activity, and moderated the increase in MDA content after treatment with methylglyoxal. Aluas RNA significantly restored the methylglyoxal-induced down-regulation of Nrf2 gene and antioxidant defense genes, including glutathione peroxidase, heme oxygenase 1, γ-glutamylcysteine synthetase and quinone oxidoreductase 1. Aluas RNA ameliorated methylglyoxal-induced increases of the mRNA and protein expression of Keap1 that is the negative regulator of Nrf2. CONCLUSION: Aluas RNA reduces apoptosis induced by methylglyoxal by enhancing antioxidant defense.

Activation of AMPK restored impaired autophagy and inhibited inflammation reaction by up-regulating SIRT1 in acute pancreatitis.

AIMS: Acute pancreatitis (AP) is a common inflammatory disorder with high incidence and mortality. AMPK-SIRT1 pathway is involved in a variety of diseases, but its role in AP remains elusive. This study was aimed to explore the role of AMPK-SIRT1 pathway in AP. MAIN METHODS: AP models in vivo and vitro were constructed by intraperitoneal administration of L-arginine and caerulein-stimulated respectively. Rat serum amylase, IL-6 and TNF-α were determined by ELISA. The expression levels of AMPK, SIRT1, Beclin-1, LC3 and p62 were determined by qRT-PCR and western blot. The number of autophagosome was checked by transmission electron microscope. KEY FINDINGS: Compared with NC rats, serum amylase, IL-6 and TNF-α were increased in AP rats. The expressions of AMPK and SIRT1 were decreased, while Beclin-1, LC3II/Iratio and p62 were markedly increased in AP rats. After activation of AMPK by metformin, expressions of p-AMPKα, SIRT1 were significantly raised, while expressions of Beclin-1, LC3 II/I, p62, TNF-α, IL-6 were reduced, and the number of autophagosome was decreased significantly in caerulein-stimulated AR42J cells. The inhibition of AMPK by compound C obtained opposite results. SIGNIFICANCE: During AP occurrence, p-AMPK and SIRT1 were down-regulated, leading to the accumulation of p62, increase of autophagic vacuoles, damage of autophagy, and the occurrence of inflammation. It hinted that activation of AMPK restored impaired autophagy and inhibited inflammation reaction by up-regulating SIRT1. Our findings might provide important theoretical basis for explaining the pathogenesis of AP and investigating therapeutic target to treat and prevent AP.

Negative Regulation of SIRT1 by IRF9 Involved in Hyperlipidemia Acute Pancreatitis Associated with Kidney Injury.

BACKGROUND: Interferon regulatory factor 9 (IRF9) acts as a negative regulator of sirtuin-1 (SIRT1) to participate in many diseases. However, the role of SIRT1 and IRF9 in hyperlipidemia acute pancreatitis associated with kidney injury is unclear. AIMS: To explore the function of SIRT1 and IRF9 in hyperlipidemia acute pancreatitis associated with kidney injury and provide theoretical guidance for disease diagnosis and treatment. METHODS: Model rats were established by intraperitoneal injection of 20% L-arginine. Apoptosis of kidney tissue was determined by TUNEL staining. Expressions of IRF9, SIRT1, p53, and acetylated p53 were detected by qRT-PCR and Western blot. Dual-Luciferase Reporter Assay was carried out to validate the regulation of IRF9 on SIRT1. RESULTS: Pancreatic and renal injury was more serious, and apoptosis of kidney epithelial cells increased in acute pancreatitis (AP) and hyperlipidemia acute pancreatitis (HLAP) group. IRF9, p53, and acetylated p53 were up-regulated, and SIRT1 was down-regulated in AP and HLAP group (p < 0.05). Down-regulation of SIRT1 was negatively correlated with up-regulation of IRF9 in AP and HLAP group (p < 0.05). Pancreatic and renal injury and kidney epithelial cells apoptosis in HLAP group were more obvious than AP group (p < 0.05). The up-regulation of IRF9 and down-regulation of SIRT1 in HLAP group were more than AP group (p < 0.05). The promoter activity of SIRT1 was repressed by IRF9. CONCLUSION: In pancreatitis associated with kidney injury, IRF9 was a negative regulator of SIRT1, down-regulated the expression of SIRT1, increased acetylated p53, and promoted renal cell apoptosis. Hyperlipidemia further aggravated pancreatic and renal injury and renal cell apoptosis.

Ditopic Chiral Pineno-Fused 2,2':6',2″-Terpyridine: Synthesis, Self-Assembly, and Optical Properties.

The syntheses of 4'-substituted chiral 2,2':6',2″-terpyridine (tpy) ligands with predetermined configurations and directionalities are rather limited in the supramolecular chemistry field. In this study, a carbazole-linked ditopic chiral ligand L was synthesized using 4'-bromo-substituted pineno-fused tpy 5 as the precursor. Upon complexation with Cd(NO3)2·4H2O and Zn(NO3)2·6H2O, two enantiomerically pure metallosupramolecules, [Cd3L3] and [Zn4L4], have been self-assembled and characterized by NMR, electrospray ionization-mass spectrometry, traveling wave ion mobility-mass spectrometry, and DOSY analysis. In addition, their optical properties are characterized by UV-vis, fluorescence, circular dichroism, and circularly polarized luminescence, suggesting an efficiency transmission and amplification of chirality from the ligand to metal center via self-assembly.