SIRT1 mRNA Research Articles

Role of SIRT-1 Gene in The Development of Diabetic Retinopathy

Background: Diabetes mellitus (DM) has had a consistent increase in global prevalence. A common microvascular disease associated with diabetes is diabetic retinopathy (DR). As a complex, progressive, as well as asymptomatic neurovascular consequence of DM, DR is the leading cause of vision impairment and blindness in working-age people. The presence of retinal neovascularization classifies diabetic retinopathy (DR) as proliferative, non-proliferative, or diabetic macular edema. Aim of the Study: to evaluate the level of expression of mRNA sirt1- gene in cases of type 2 DM that have retinopathy or vulnerable to it. Patients and Methods: In this research, 60 cases (15 patients with diabetic retinopathy, 15 diabetic cases without retinopathy (PDR), and 30 non-diabetic controls) were recruited. SIRT1 mRNA expression from the two groups was analyzed. Notably, patients with DR exhibited a decrease in the expression of SIRT1 mRNA. ELISA was also used to measure the levels of IL-17 expression in the serum. The results showed that the sera of people with DR had higher levels of IL-17 expression than the sera of healthy people. Results: The findings of this research indicate that imbalanced IL-17 & SIRT1 expression levels may play a role in the etiology of DR, with SIRT1 potentially exerting a protective effect by suppressing IL-17 production.

LncRNA MALAT1 to Enhance Pyroptosis in Viral Myocarditis Through UPF1-Mediated SIRT6 mRNA Decay and Wnt-β-Catenin Signal Pathway.

Viral myocarditis (VMC) is an inflammatory disease of the myocardium caused by cardioviral infection, especially coxsackievirus B3 (CVB3), and is a major contributor to acute heart failure and sudden cardiac death in children and adolescents. LncRNA MALAT1 knockdown reportedly inhibits the differentiation of Th17 cells to attenuate CVB3-induced VMC in mice. Moreover, long non-coding RNAs (lncRNAs) interact with RNA-binding proteins (RBPs) to regulate UPF1-mediated mRNA decay. However, it remains unclear whether MALAT1 can bind to UPF1 to mediate the mRNA decay of its target genes in VMC. Herein, we aimed to explore the effect of lncRNA MALAT1 on UPF1-mediated SIRT6 mRNA decay in VMC using in vivo and in vitro experiments. CVB3-infected BABL/C mice were used as VMC models, and MALAT1 interfering adenovirus was injected to achieve MALAT1 knockdown. The heart function of the VMC mice was assessed using echocardiography. Pathological changes in myocardial tissues were assessed after hematoxylin-eosin staining. Myocardial injury and inflammation were evaluated by measuring creatine kinase isoenzyme B, cardiac troponin T, interleukin (IL)-1β, and IL-18. TUNEL staining was performed to assess apoptosis in myocardial tissues. In vitro experiments were performed using H9c2 cells after transfection and CVB3 infection. The lactic dehydrogenase release, caspase-1 activity, and IL-1β and IL-18 levels in the cellular supernatant were detected. Western blotting was performed to determine the expression of pyroptosis-related proteins (GSDMD-N, NLRP3, ASC, and Cleaved-Caspase-1) and Wnt/β-catenin signal pathway-related proteins (Wnt1, β-catenin, and p-GSK-3β). RNA immunoprecipitation and RNA stability assays assessed the relationship between MALAT1, UPF1, and SIRT6. CVB3-infected mice and H9c2 cells exhibited elevated MALAT1 and reduced SIRT6 expression. MALAT1 knockdown or SIRT6 overexpression suppressed inflammation and pyroptosis and inhibited the activation of the Wnt/β-catenin signal pathway in myocardial tissues and cells. MALAT1 enhanced the enrichment of SIRT6 mRNA by UPF1 and disturbed the stability of SIRT6 mRNA to promote the development of VMC. MALAT1 can bind UPF1 to mediate SIRT6 mRNA decay and activate the Wnt/β-catenin signal pathway in VMC.

METTL3-mediated m6A modification of SIRT1 mRNA affects the progression of diabetic cataracts through cellular autophagy and senescence

BackgroundThe increasing incidence of diabetes mellitus has established diabetic cataracts (DC) as a significant worldwide public health issue. The mechanisms underlying DC remain unknown, and effective prevention and treatment strategies are lacking. Accordingly, we aimed to explore the role and mechanism behind N6-methyladenosine (m6A) in DC progression.MethodsMethyltransferase-like 3 (METTL3), p21, Beclin1, LC3, and p62 expression levels were measured in human tissues. This study assessed total m6A levels and common m6A-regulated biomarkers in both in vitro and in vivo DC models. Autophagy flux was detected in vitro through Ad-mCherry-GFP-LC3B and Monodansylcadaverine (MDC) staining. Cellular senescence was assessed utilizing the senescence-associated β-galactosidase (SA-β-Gal) assay. Furthermore, the effect of METTL3 on SIRT1 mRNA modification was demonstrated, and its mechanism was elucidated using RT-qPCR, western blot, RNA stability assays, and RIP analysis.ResultsMETTL3, p21, and p62 expression levels were elevated in lens epithelial cells (LECs) from DC patients, while Beclin1 and LC3 levels were reduced. Silencing METTL3-mediated m6A modifications restored high-glucose-induced autophagy inhibition and prevented premature senescence in LECs. Notably, SIRT1720 and Metformin significantly enhanced autophagosome generation and delayed cellular senescence. The m6A-reading protein YTHDF2 bound to m6A modifications, and YTHDF2 silencing significantly reduced METTL3-mediated SIRT1 inactivation.ConclusionsMETTL3 induces senescence in DC by destabilizing SIRT1 mRNA in an m6A-YTHDF2-dependent manner. The METTL3-YTHDF2-SIRT1 axis is a key target and potential pathogenic mechanism in DC.

MiR-9-5p regulates Sirt1 involved in testicular development and spermatogenesis in mouse

Testicular development and spermatogenesis are critical for male reproduction, with histone (de)acetylation playing a key role in chromatin remodeling within germ cells. Sirt1, a key histone deacetylase, is implicated in chromatin remodeling, but its expression pattern and specific role in testicular development and spermatogenesis need further study. This study comprehensively analyzed Sirt1 expression in adult and juvenile mouse testicular tissues and across various male germ cells, utilizing RT-qPCR, Western blot, immunofluorescence, and cell transfection. GO and KEGG enrichment analyses were performed to elucidate the biological functions and pathways associated with Sirt1 and its related genes. Multiple miRNA databases were utilized to predict miRNAs targeting Sirt1, and their expression levels were validated using RT-qPCR. Lentiviral transfection was used to knockdown candidate miRNAs to assess their functional roles. The results revealed a significant downregulation of Sirt1 expression in adult mouse testicular tissues compared to juvenile tissues, with pronounced variation across diverse male germ cells. Sirt1 was highly expressed in spermatogonia and mature sperm, but comparatively lower in spermatocytes and spermatids. GO and KEGG enrichment analyses highlighted Sirt1's role in key biological processes, including chromatin organization, regulation of cell proliferation, and energy homeostasis, as well as its association with signaling pathways like cellular senescence, the FoxO signaling pathway, and the AMPK signaling pathway. Bioinformatic analysis and subsequent RT-qPCR validation identified miR-9-5p as a miRNA targeting Sirt1. The expression of miR-9-5p was significantly higher in adult mouse testicular tissues compared to juvenile tissues, inversely correlating with Sirt1 levels. Moreover, the knockdown of miR-9-5p led to a notable increase in Sirt1 mRNA and protein expression. In conclusion, Sirt1 is a key player in mouse testicular development and spermatogenesis. The discovery that miR-9-5p negatively regulates Sirt1 suggests a critical regulatory axis that may govern these processes, providing novel insights into male fertility and potential targets for therapeutic intervention.

Elucidating microRNA-34a organisation within human Argonaute-2 by dynamic nuclear polarisation-enhanced magic angle spinning NMR.

Understanding mRNA regulation by microRNA (miR) relies on the structural understanding of the RNA-induced silencing complex (RISC). Here, we elucidate the structural organisation of miR-34a, which is de-regulated in various cancers, in human Argonaute-2 (hAgo2), the effector protein in RISC. This analysis employs guanosine-specific isotopic labelling and dynamic nuclear polarisation (DNP)-enhanced Magic Angle Spinning (MAS) NMR. Homonuclear correlation experiments revealed that the non-A-form helical conformation of miR-34a increases when incorporated into hAgo2 and subsequently bound to SIRT1 mRNA compared to the free miR-34a or the free mRNA:miR duplex. The C8-C1' correlation provided a nucleotide-specific distribution of C2'- and C3'-endo sugar puckering, revealing the capture of diverse dynamic conformations upon freezing. Predominantly C3'-endo puckering was observed for the seed region, while C2'-endo conformation was found in the central region, with a mixture of both conformations elsewhere. These observations provide insights into the molecular dynamics underlying miR-mediated mRNA regulation and demonstrate that experiments conducted under cryogenic conditions, such as at 90K, can capture and reveal frozen dynamic states, using methods like DNP-enhanced MAS NMR or Cryo-ElectronMicroscopy.

S-nitrosoglutathione (GSNO) induces necroptotic cell death in K562 cells: Involvement of p73, TSC2 and SIRT1

BackgroundNitric oxide and Reactive Nitrogen Species are known to effect tumorigenicity. GSNO is one of the main NO carrying signalling moiety in cell. In the current study, we tried to delve into the effect of GSNO induced nitrosative stress in three different myelogenous leukemic K562, U937 and THP-1 cell lines. MethodWST-8 assay was performed to investigate cell viability. RT-PCR and western-blot analysis were done to investigate mRNA and protein expression. Spectrophotometric and fluorimetric assays were done to investigate enzyme activities. ResultWe found that GSNO exposure led to reduced cell viability and the mode of cell death in K562 was non apoptotic in nature. GSNO promoted impaired autophagic flux and necroptosis. GSNO treatment heightened phosphorylation of AMPK and TSC2 and inhibited mTOR pathway. We observed increase in NAD+/ NADH ratio following GSNO treatment. Increase in both SIRT1 m-RNA and protein expression was observed. While total SIRT activity remained unaltered. GSNO increased tumor suppressor TAp73/ oncogenic ∆Np73 ratio in K562 cells which was correlated with cell mortality. Surprisingly, GSNO did not alter cellular redox status or redox associated protein expression. However, steep increase in total SNO and PSNO content was observed. Furthermore, inhibition of autophagy, AMPK phosphorylation or SIRT1 exacerbated the effect of GSNO. Altogether our work gives insights into GSNO mediated necroptotic event in K562 cells which can be excavated to develop NO based anticancer therapeutics. ConclusionOur data suggests that GSNO could induce necroptotic cell death in K562 through mitochondrial dysfunctionality and PTM of different cellular proteins.

Sirt1 Mitigates Hepatic Lipotoxic Injury Induced by High-Fat-Diet in Fish Through Ire1α Deacetylation

BackgroundSilent information regulator protein 1 (Sirt1) is crucial in regulating lipid metabolism, but its specific role and mechanism in fish hepatic lipotoxic injury remain undefined. ObjectivesThis study aimed to elucidate the regulatory role of Sirt1 and the underlying mechanisms in dietary lipid-induced hepatic lipotoxic injury in a marine teleost black seabream. MethodsBlack seabream were fed a control diet (12% lipid level), high-fat diet (HFD) [18% lipid level, oleic acid (OA)-rich], or HFD supplemented with 0.25%, 0.50%, or 1.00% resveratrol (RSV) for 8 wk. The cultured hepatocytes were stimulated by OA (200 μM), OA supplemented with RSV (20 μM), or transfection with sirt1-small interfering RNA (sisirt1). Biochemical indices, gene expression (qPCR), histology, transmission electron microscope, immunofluorescence, Western blot, flow cytometry, and immunoprecipitation assays were conducted to evaluate hepatic lipid deposition, lipid metabolism, endoplasmic reticulum stress, inflammation and apoptosis, and determine protein interactions between Sirt1 and Ire1α. ResultsIn vivo, RSV supplementation increased mRNA and protein expression levels of sirt1 (236.2% ± 16.1% and 53.1% ± 14.3%) and downregulated the mRNA and phosphorylated protein expression levels of ire1α/Ire1α (46.0% ± 7.6% and 38.6% ± 7.0%), jnk/Jnk (57.6% ± 7.3% and 122.1%), and nuclear factor κ B (nf-κb/Nf-κb) p65 (41.7% ± 7.1% and 24.6% ± 0.8%) compared with the HFD group. Similar patterns were found in the in vitro experiments; however, after knockdown of sirt1, although the cells were incubated with RSV, the expression levels of ire1α/ Ire1α, jnk/Jnk, and nf-κb/Nf-κb p65 showed no significant differences compared with the OA treatment. Moreover, we found that mutation of K61 to arginine to mimic Ire1α deacetylation confers protection against Ire1α-mediated OA-rich HFD-induced inflammation and apoptosis. ConclusionsThe findings revealed that Sirt1 protects against OA-rich HFD-induced hepatic lipotoxic injury via the deacetylation of Ire1α on K61, hence reducing Ire1α autophosphorylation level, and suppressing Jnk and Nf-κb p65 activation. This mechanism is elucidated for the first time in fish.

Sirt1 m6A modification-evoked Leydig cell senescence promotes Cd-induced testosterone decline

Diminished testosterone levels have been documented as a key factor in numerous male health disorders. Both human and animal studies have consistently demonstrated that cadmium (Cd), a pervasive environmental heavy metal, results in decreased testosterone levels. However, the exact mechanism through which Cd interferes with testosterone synthesis remains incompletely elucidated. This research sought to examine the impact of cellular senescence on Cd-suppressed testosterone synthesis. We also investigated the related m6A modification mechanism. The results demonstrated that Cd (100 mg/L) led to a decrease in testosterone levels, along with downregulated expression of testosterone synthase in C57BL/6 N male mice. Furthermore, Cd significantly increased β-galactosidase staining intensity, senescence-related proteins, and senescence-related secretory phenotypes in mouse testicular Leydig cells. Subsequent investigations revealed that Cd decreased the mRNA and protein levels of NAD-dependent deacetylase Sirtuin-1 (SIRT1) in Leydig cells. Mechanistically, mice treated with resveratrol (50 mg/kg), a specific SIRT1 activator, mitigated Leydig cell senescence and reversed Cd-reduced testosterone levels in mouse testes. These effects were also restored by SIRT1 overexpression in Leydig cells. Additionally, we found that Cd increased the level of methyltransferase enzyme METTL3 and Sirt1 m6A modification in Leydig cells. Mettl3 siRNA effectively restored Cd-enhanced Sirt1 m6A level and reversed Cd-downregulated Sirt1 mRNA expression in Leydig cells. Overall, our findings suggest that Cd exposure inhibits testosterone synthesis via Sirt1 m6A modification-mediated senescence in mouse testes. These results offer an experimental basis for investigating the causes and potential treatments of hypotestosteronemia induced by environmental factors.

SIRT1 and FOXO1 role on MASLD risk: effects of DHA-rich n-3 PUFA supplementation and exercise in aged obese female mice and in post-menopausal overweight/obese women

Sirtuins 1 (SIRT1) and Forkhead box protein O1 (FOXO1) expression have been associated with obesity and metabolic dysfunction-associated steatotic liver disease (MASLD). Exercise and/or docosahexaenoic acid (DHA) supplementation have shown beneficial effects on MASLD. The current study aims to assess the relationships between Sirt1, Foxo1 mRNA levels and several MASLD biomarkers, as well as the effects of DHA-rich n-3 PUFA supplementation and/or exercise in the steatotic liver of aged obese female mice, and in peripheral blood mononuclear cells (PBMCs) of postmenopausal women with overweight/obesity. In the liver of 18-month-old mice, Sirt1 levels positively correlated with the expression of genes related to fatty acid oxidation, and negatively correlated with lipogenic and proinflammatory genes. Exercise (long-term treadmill training), especially when combined with DHA, upregulated hepatic Sirt1 mRNA levels. Liver Foxo1 mRNA levels positively associated with hepatic triglycerides (TG) content and the expression of lipogenic and pro-inflammatory genes, while negatively correlated with the lipolytic gene Hsl. In PBMCs of postmenopausal women with overweight/obesity, FOXO1 mRNA expression negatively correlated with the hepatic steatosis index (HSI) and the Zhejiang University index (ZJU). After 16-weeks of DHA-rich PUFA supplementation and/or progressive resistance training (RT), most groups exhibited reduced MASLD biomarkers and risk indexes accompanying with body fat mass reduction, but no significant changes were found between the intervention groups. However, in PBMCs n-3 supplementation upregulated FOXO1 expression, and the RT groups exhibited higher SIRT1 expression. In summary, SIRT1 and FOXO1 could be involved in the beneficial mechanisms of exercise and n-3 PUFA supplementation related to MASLD manifestation.

Hyperglycemia-induced Sirt3 downregulation increases microglial aerobic glycolysis and inflammation in diabetic neuropathic pain pathogenesis.

Hyperglycemia-induced neuroinflammation significantly contributes to diabetic neuropathic pain (DNP), but the underlying mechanisms remain unclear. To investigate the role of Sirt3, a mitochondrial deacetylase, in hyperglycemia-induced neuroinflammation and DNP and to explore potential therapeutic interventions. Here, we found that Sirt3 was downregulated in spinal dorsal horn (SDH) of diabetic mice by RNA-sequencing, which was further confirmed at the mRNA and protein level. Sirt3 deficiency exacerbated hyperglycemia-induced neuroinflammation and DNP by enhancing microglial aerobic glycolysis invivo and invitro. Overexpression of Sirt3 in microglia alleviated inflammation by reducing aerobic glycolysis. Mechanistically, high-glucose stimulation activated Akt, which phosphorylates and inactivates FoxO1. The inactivation of FoxO1 diminished the transcription of Sirt3. Besides that, we also found that hyperglycemia induced Sirt3 degradation via the mitophagy-lysosomal pathway. Blocking Akt activation by GSK69093 or metformin rescued the degradation of Sirt3 protein and transcription inhibition of Sirt3 mRNA, which substantially diminished hyperglycemia-induced inflammation. Metformin invivo treatment alleviated neuroinflammation and diabetic neuropathic pain by rescuing hyperglycemia-induced Sirt3 downregulation. Hyperglycemia induces metabolic reprogramming and inflammatory activation in microglia through the regulation of Sirt3 transcription and degradation. This novel mechanism identifies Sirt3 as a potential drug target for treating DNP.

Skin anti-aging and wound healing effects of a passion fruit seed extract rich in piceatannol

BACKGROUND: UVB radiation triggers skin photoaging by inducing excess cellular oxidants, contributing to senescence, and overproducing matrix metalloproteinase-1 (MMP-1), causing skin wrinkles. Senescence also impedes skin wound closure. Passion fruit seeds (PFS) boast abundant piceatannol (PCT), recognized for its antioxidant and anti-aging properties. OBJECTIVE: To study potential of PFS extract in diminishing photoaging, accelerating wound healing, and enhancing SIRT1 production in human keratinocytes (HaCaT cells). METHODS: The PFS extract was assessed for PCT using HPLC analysis. The antioxidant effects, reduced senescence-associated β-galactosidase (SA-β-gal) activity, and MMP-1 production were measured in HaCaT cells exposed to UVB using a DCFA-DA assay, SA-β-gal activity, and flow cytometry/immunofluorescent including MMP-1 mRNA expression, respectively. The effect of wound healing acceleration was evaluated, including EGFR mRNA expression. Additionally, SIRT1 enhancement was investigated using flow cytometry and immunofluorescent techniques, which also included the assessment of SIRT1 mRNA expression. RESULTS: The PFS extract, which is rich in PCT, effectively diminished cellular oxidants and aging. It decreased UVB-induced cellular oxidants, senescence, and MMP-1 production at both protein and mRNA levels. Moreover, the extract enhanced wound closure by boosting cell migration and increasing EGFR mRNA expression. Additionally, SIRT1 production and expression, associated with cellular rejuvenation, increased in extract-treated cells. CONCLUSIONS: PFS extract exhibits potential for skincare and wound healing applications, offering a natural strategy to combat skin aging effects and support tissue repair.

Vitamin D alleviation of oxidative stress in human retinal pigment epithelial cells.

Oxidative stress-induced retinal pigment epithelium (RPE) cell damage is a major factor in age-related macular degeneration (AMD). Vitamin D3 (VD3) is a powerful antioxidant and it has been suggested to have anti-aging properties and potential for treating AMD. This study aimed to investigate the effect of VD3 on RPE cell oxidative apoptosis of RPE cells in order to provide experimental evidence for the treatment of AMD. Human retinal pigment epithelial cell 19 (ARPE-19) cells were divided into four groups: blank group (untreated), model group (incubated in medium with 400μmol/L H2O2 for 1h), VD3 group (incubated in medium with 100μmol/L VD3 for 24h), and treatment group (incubated in medium with 400μmol/L H2O2 for 1h and 100μmol/L VD3 for 24h). Cell viability, cell senescence, ROS content, expression levels of vitamin D specific receptors, Akt, Sirt1, NAMPT, and JNK mRNA expression levels, SOD activity, and MDA, GSH, and GPX levels were measured. We first established an ARPE-19 cell stress model with H2O2. Our control experiment showed that VD3 treatment had no significant effect on ARPE-19 cell viability within 6-48h. Treating the stressed ARPE-19 cells with VD3 showed mixed results; caspase-3 expression was decreased, Bcl-2 expression was increased, MDA level of ARPE-19 cells was decreased, GSH-PX, GPX and SOD levels were increased, the relative mRNA expression levels of Akt, Sirt1, NAMPT were increased (P < 0.05), and the relative mRNA expression level of JNK was decreased (P < 0.05). VD3 can potentially slow the development of AMD.

Uncovering SIRT3 and SHMT2-dependent pathways as novel targets for apigenin in modulating colorectal cancer: In vitro and in vivo studies

Despite significant advances in the treatment of colorectal cancer (CRC), identification of novel targets and treatment options are imperative for improving its prognosis and survival rates. The mitochondrial SIRT3 and SHMT2 have key roles in metabolic reprogramming and cell proliferation. This study investigated the potential use of the natural product apigenin in CRC treatment employing both in vivo and in vitro models and explored the role of SIRT3 and SHMT2 in apigenin-induced CRC apoptosis. The role of SHMT2 in CRC patients’ survival was verified using TCGA database. In vivo, apigenin treatment restored the normal colon appearance. On the molecular level, apigenin augmented the immunohistochemical expression of cleaved caspase-3 and attenuated SIRT3 and SHMT2 mRNA expression CRC patients with decreased SHMT2 expression had improved overall and disease-free survival rates. In vitro, apigenin reduced the cell viability in a time-dependent manner, induced G0/G1 cell cycle arrest, and increased the apoptotic cell population compared to the untreated control. Mechanistically, apigenin treatment mitigated the expression of SHMT2, SIRT3, and its upstream long intergenic noncoding RNA LINC01234 in CRC cells. Conclusively, apigenin induces caspase-3-dependent apoptosis in CRC through modulation of SIRT3-triggered mitochondrial pathway suggesting it as a promising therapeutic agent to improve patient outcomes.

ACTIVATION OF KLOTHO/SIRT1 SIGNALING PATHWAY ATTENUATES MYOCARDIAL ISCHEMIA REPERFUSION INJURY IN DIABETIC RATS.

Diabetes and myocardial ischemia reperfusion (MIR) injury are characterized by oxidative stress, inflammation, autophagy disorders, and cardiac contractile dysfunction. Klotho and SIRT1 regulate the level of oxidative stress to participate in the regulation of many physiological functions such as cell survival, aging, apoptosis, autophagy, mitochondrial biogenesis, and inflammation. We hypothesized that the activation of Klotho/SIRT1 signaling pathway could attenuate MIR in diabetic rats. Type 1 diabetes and MIR injury model were established to examine this hypothesis in vivo . Primary rat cardiomyocytes and H9c2 cells were exposed to high glucose conditions and hypoxia/reoxygenation (H/R) insult in vitro . Hemodynamic parameters of heart function, myocardial infarct size, oxidative stress, markers of MIR injury or cell viability, and the mRNA and protein expression of Klotho and SIRT1 were measured. There was lower expression of Klotho and SIRT1 in diabetic MIR hearts than in nondiabetic rats, as well as significantly increased oxidative stress levels and decreased autophagy levels. Recombinant Klotho (rKlotho) protein and the SIRT1 agonist SRT1720 could significantly attenuate MIR injury in diabetes by activating Klotho/SIRT1 signaling pathway to reduce oxidative stress and restore autophagy levels. These findings suggest that the Klotho/SIRT1 pathway plays an important role in MIR injury in diabetic rats, and rKlotho protein and agonist SRT1720 have therapeutic potential for alleviating diabetic myocardial IR injury by activating Klotho/SIRT1 to reduce oxidative stress and restore autophagy levels.

SIRT2 as a Potential Biomarker in Lung Adenocarcinoma: Implications for Immune Infiltration.

SIRT2 play important roles in cell cycle and cellular metabolism in the development of non-small cell lung cancer (NSCLC), and SIRT2 exhibits its therapeutic effect on NSCLC tumors with high expression of SIRT2. Nevertheless, the clinical relevance of SIRT2 in lung adenocarcinoma (LUAD), particularly its impact on tumor growth and prognostic implications, remains obscure. This investigation entailed a comprehensive analysis of SIRT2 mRNA and protein expression levels in diverse tumor and corresponding healthy tissues, utilizing databases such as TIMER 2.0, UALCAN, and HPA. Prognostic correlations of SIRT2 expression in LUAD patients, stratified by distinct clinicopathological characteristics, were evaluated using the KM Plotter database. Additionally, the TCGA and TIMER 2.0 databases were employed to assess the relationship between SIRT2 and immune infiltration, as well as to calculate immunity, stromal, and estimation scores, thus elucidating the role of SIRT2 in modulating tumor immunotherapy responses. Furthermore, Gene Set Enrichment Analysis (GSEA) was utilized to elucidate SIRT2's biological functions in pan-cancer cells. Our findings revealed a marked reduction in both mRNA and protein levels of SIRT2 in LUAD tumors relative to healthy tissue. Survival analysis indicated that diminished SIRT2 expression correlates with adverse prognostic outcomes in LUAD. Furthermore, SIRT2 expression demonstrated a significant association with various clinicopathologic attributes of LUAD patients, influencing survival outcomes across different clinicopathologic states. Functional enrichment analyses highlighted SIRT2's involvement in cell cycle regulation and immune response. Notably, SIRT2 exhibited a positive correlation with immune cell infiltration, including natural killer (NK) cells, macrophages, and dendritic cells (DCs). In summary, SIRT2 was a potential prognostic biomarker for LUAD and and a new immunotherapy target.

MiR-34a ameliorates arterial blood flow in rats with lower limb arteriosclerosis obliterans via Sirt1 signaling pathway.

In this study, we investigated the impact of microRNA-34a (miR-34a) on lower limb arteriosclerosis obliterans in rats through the Sirtuin 1 (Sirt1) signaling pathway. Thirty-six Sprague-Dawley rats were divided into normal, model, and miR-34a mimics groups. Rats in the normal group were raised normally, while the model group underwent lower limb arteriosclerosis obliterans induction and received saline injections. The miR-34a mimics group also underwent arteriosclerosis obliterans modeling but received miR-34a mimics injections. Immunohistochemistry revealed significantly increased vascular endothelial growth factor (VEGF) expression in both model and miR-34a mimics groups compared to the normal group, with the miR-34a mimics group showing higher levels. Western blotting indicated elevated Sirt1 protein expression in both non-normal groups, with the miR-34a mimics group exhibiting significantly higher levels. Quantitative polymerase chain reaction (qPCR) demonstrated higher levels of miR-34a, VEGF mRNA, and Sirt1 mRNA in the model group compared to the normal group, but significantly lower levels than the miR-34a mimics group. Enzyme-linked immunosorbent assay (ELISA) showed increased VEGF content in the model group compared to the normal group but decreased compared to the miR-34a mimics group. Hemorrheological detection revealed a reduced PU index in both non-normal groups compared to the normal group, with a significant increase in the miR-34a mimics group compared to the model group. Overall, miR-34a upregulation enhanced VEGF expression in rat blood vessels, ameliorating arterial blood flow in lower limb arteriosclerosis obliterans through the Sirt1 signaling pathway.

Abstract 2451: Investigation of SIRT6 usefulness as a novel biomarker for oral cancer

Abstract Introduction: SIRT6 is one of the seven human sirtuin genes and is known to function as an onco-suppressor gene in colorectal and ovarian cancers, although it is up-regulated in other cancers. Thus, SIRT6 is considered performing both tumor-suppressing and promoting roles. However, the association of SIRT6 with oral squamous cell carcinoma (OSCC) and its role in OSCC pathogenesis are currently unclear. Currently, SCC or CYFRA are used as a tumor marker to assist in the diagnosis of OSCC in a clinical setting whereas its sensitivity is low. Therefor the development of a novel biomarker for early diagnosis and identification of new therapeutic targets is critical to resolve the pressing clinical issues related to the management of OSCC. This study aimed to investigate the expression and the relevance of SIRT6 in patients with OSCC and its potential as a biomarker for early detection and prognosis prediction. Materials and Methods: This study enrolled 78 patients with OSCC and 9 patients with carcinoma in situ (CIS). The OSCC patients had undergone surgery or chemoradiotherapy (51 surgery and 27 chemoradiotherapy). The CIS patients had undergone surgery. Samples were also obtained from 10 patients with oral potentially malignant disorders (OPMDs). Biopsied or surgical resected samples were used for the study. We obtained 21 noncancerous tissues as normal tissues from the adjacent noncancerous tissues after tumor resection. Immunohistochemistry, quantitative real-time RT-PCR, and microarray analyses were performed to determine SIRT6 expression and its association with clinicopathological features in OSCC, CIS, and OPMDs using clinical paraffin embedded specimens. Results: SIRT6 mRNA levels were higher in OSCC tissues than those in noncancerous tissues (p&amp;lt;0.05). And SIRT6 protein expression levels were higher in OSCC, CIS and OPMDs tissues than those in noncancerous tissues (p&amp;lt;0.05). SIRT6 expression was predominant in patients aged ≥65 years and significantly correlated with shorten overall survival. In the microarray analysis, some SIRT6-associated genes, such as ANXA2 was significantly up-regulated, and PIGC and RGPD4 were down-regulated in OSCC. Conclusion: SIRT6 plays a role in tumor homeostasis, leading to a poor prognosis in OSCC. SIRT6 may represent a novel target not only for the treatment, but also as a prognostic marker in OSCC. Citation Format: Haruka Yoshii, Ami Shimoda, Kenji Mitsudo, Mitomu Kioi. Investigation of SIRT6 usefulness as a novel biomarker for oral cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2451.

Identification of Osteoarthritis Inflamm-Aging Biomarkers by Integrating Bioinformatic Analysis and Machine Learning Strategies and the Clinical Validation

To identify inflamm-aging related biomarkers in osteoarthritis (OA). Microarray gene profiles of young and aging OA patients were obtained from the Gene Expression Omnibus (GEO) database and aging-related genes (ARGs) were obtained from the Human Aging Genome Resource (HAGR) database. The differentially expressed genes of young OA and older OA patients were screened and then intersected with ARGs to obtain the aging-related genes of OA. Enrichment analysis was performed to reveal the potential mechanisms of aging-related markers in OA. Three machine learning methods were used to identify core senescence markers of OA and the receiver operating characteristic (ROC) curve was used to assess their diagnostic performance. Peripheral blood mononuclear cells were collected from clinical OA patients to verify the expression of senescence-associated secretory phenotype (SASP) factors and senescence markers. A total of 45 senescence-related markers were obtained, which were mainly involved in the regulation of cellular senescence, the cell cycle, inflammatory response, etc. Through the screening with the three machine learning methods, 5 core senescence biomarkers, including FOXO3, MCL1, SIRT3, STAG1, and S100A13, were obtained. A total of 20 cases of normal controls and 40 cases of OA patients, including 20 cases in the young patient group and 20 in the elderly patient group, were enrolled. Compared with those of the young patient group, C-reactive protein (CRP), interleukin (IL)-6, and IL-1β levels increased and IL-4 levels decreased in the elderly OA patient group (P<0.01); FOXO3, MCL1, and SIRT3 mRNA expression decreased and STAG1 and S100A13 mRNA expression increased (P<0.01). Pearson correlation analysis demonstrated that the selected markers were associated with some indicators, including erythrocyte sedimentation rate (ESR), IL-1β, IL-4, CRP, and IL-6. The area under the ROC curve of the 5 core aging genes was always greater than 0.8 and the C-index of the calibration curve in the nomogram prediction model was 0.755, which suggested the good calibration ability of the model. FOXO3, MCL1, SIRT3, STAG1, and S100A13 may serve as novel diagnostic biomolecular markers and potential therapeutic targets for OA inflamm-aging.

Exploring the anti-ferroptosis mechanism of Kai-Xin-San against Alzheimer's disease through integrating network pharmacology, bioinformatics, and experimental validation strategy in vivo and in vitro

Ethnopharmacological relevanceKai Xin San (KXS), first proposed by Sun Simiao during the Tang Dynasty, has been utilized to treat dementia by tonifying qi and dispersing phlegm. Aim of the studyThis study aimed to elucidate the mechanism by which KXS exerts its therapeutic effects on Alzheimer's disease (AD) by targeting ferroptosis, using a combination of network pharmacology, bioinformatics, and experimental validation strategies. Materials and methodsThe active target sites and the further potential mechanisms of KXS in protecting against AD were investigated through molecular docking, molecular dynamics simulation, and network pharmacology, and combined with the validation of animal experiments. ResultsComputational and experimental findings provide the first indication that KXS significantly improves learning and memory defects and inhibits neuronal ferroptosis by repairing mitochondria damage and upregulating the protein expression of ferroptosis suppressor protein 1 (FSP1) in vivo APP/PS1 mice AD model. According to bioinformatics analysis, the mechanism by which KXS inhibits ferroptosis may involve SIRT1. KXS notably upregulated the mRNA and protein expression of SIRT1 in both vivo APP/PS1 mice and in vitro APP-overexpressed HT22 cells. Additionally, KXS inhibited ferroptosis induced by APP-overexpression in HT22 cells through activating the SIRT1-FSP1 signal pathway. ConclusionsCollectively, our findings suggest that KXS may inhibit neuronal ferroptosis through activating the SIRT1/FSP1 signaling pathway. This study reveals the scientific basis and underlying modern theory of replenishing qi and eliminating phlegm, which involves the inhibition of ferroptosis. Moreover, it highlights the potential application of SIRT1 or FSP1 activators in the treatment of AD and other ferroptosis-related diseases.

LncRNA SOX2OT enhances 5-fluorouracil resistance of cholangiocarcinoma cells by promoting autophagy via up-regulating SIRT1 expression

To investigate the role of SIRT1/autophagy pathway in mediating the regulatory effect of lncRNA SOX2OT on 5-fluorouracil (5-FU) resistance in cholangiocarcinoma cells. HCCC-9810 cells were used to construct a 5-FU-resistant cell model (HCCC-9810/5-FU cells), and the expression levels of lncRNA SOX2OT and SIRT1 mRNA and the protein expressions of SIRT1, Beclin1, LC3 and P62 were detected with qRT-PCR and Western blotting. The effects of transfection with a SOX2OT mimic on drug resistance and cell migration of HCCC-9810/5-FU cells were detected using CCK-8 assay and wound healing assay, and the changes in expressions of SOX2OT, SIRT1, Beclin1, LC3 and P62 were detected. Rescue experiment was performed by co-transfection of HCCC-9810/5-FU cells with both a SOX2OT-overexpressing plasmid and si-SIRT1 to confirm the role of SIRT1 in SOX2OT-mediated regulation of 5-FU resistance. A RNA pulldown assay was used to verify the targeted binding between SOX2OT and SIRT1. The proliferation of HCCC-9810 cells was significantly inhibited after treatment with different concentrations of 5-FU (P < 0.05). The 5-FU-resistant cells showed significantly increased protein expressions of SIRT1, Beclin1 and p62, an increased LC3 Ⅱ/LC3 Ⅰ ratio, and enhanced expressions of SIRT1 mRNA and SOX2OT (P < 0.05). Transfection of the resistant cells with SOX2OT mimic significantly enhanced cell migration and increased the protein expressions of SIRT1, Beclin1 and p62, the LC3Ⅱ/LC3Ⅰ ratio, and expression levels of SIRT1 mRNA and SOX2OT (P < 0.05), and these changes were obviously attenuated by SIRT1 knockdown, which also resulted in lowered 5-FU resistance of the cells without significantly affecting the expression level of SOX2OT (P > 0.05). RNA pulldown assay suggested that SOX2OT could directly bind to SIRT1. LncRNA SOX2OT enhances 5-FU resistance in HCCC-9810 cells by promoting autophagy through up-regulating SIRT1 expression.