[Total saponins of Panax japonicus alleviates CCl4-induced acute liver injury in rats by regulating the PI3K/AktNF-κB signaling pathway].

OBJECTIVE: To investigate the protective effect of total saponins of Panax japonicus (TSPJ) against CCl4-induced acute liver injury (ALI) in rats and explore the underlying pharmacological mechanisms. METHODS: Male SD rat models of CCl4-induced ALI were given intraperitoneal injections of distilled water, 100 mg/kg biphenyl bisabololol, or 50, 100, and 200 mg/kg TSPJ during modeling (n=8). Liver functions (AST, ALT, TBil and ALP) of the rats were assessed and liver pathologies were observed with HE staining. Immunohistochemistry was used to detect the expressions of PI3K/Akt/NF-κB signaling pathway molecules in liver tissue; ELISA was used to determine the levels of T-SOD, GSH-Px, and MDA. Western blotting was performed to detect the expression levels of PI3K-Akt and SIRT6-NF-κB pathways in the liver tissue. RESULTS: Network pharmacological analysis indicated that the key pathways including PI3K/Akt mediated the therapeutic effect of TSPJ on ALI. In the rat models of ALI, treatments with biphenyl bisabololol and TSPJ significantly ameliorated CCl4-induced increase of serum levels AST, ALT, ALP, TBil and MDA and decrease of T-SOD and GSH-Px levels (all P < 0.01). The rat models of ALI showed significantly increased expression of p-NF-κB (P < 0.01), decreased expressions of PI3K, p-Akt and SIRT6 proteins, and elevated expression levels of p-NF-κB, TNF-α and IL-6 proteins in the liver, which were all significantly improved in the treatment groups (P < 0.05 or 0.01). CONCLUSION: TSPJ can effectively alleviate CCl4-induced ALI in rats by suppressing inflammatory responses and oxidative stress in the liver via regulating the PI3K/Akt and SIRT6/NF-κB pathways.

SIRT3 and SIRT4 double-genes remodeled the mitochondrial network to induce hepatocellular carcinoma cell line differentiation and suppress malignant phenotypes.

Tumor cells with damaged mitochondria undergo metabolic reprogramming, but gene therapy targeting mitochondria has not been comprehensively reported. In this study, plasmids targeting the normal hepatocyte cell line (L-O2) and hepatocellular carcinoma cell line were generated using three genes SIRT3, SIRT4, and SIRT5. These deacetylases play a variety of regulatory roles in cancer and are related to mitochondrial function. Compared with L-O2, SIRT3 and SIRT4 significantly ameliorated mitochondrial damage in HCCLM3, Hep3B and HepG2 cell lines and regulated mitochondrial biogenesis and mitophagy, respectively. We constructed double-gene plasmid for co-express SIRT3 and SIRT4 using the internal ribosome entry site (IRES). The results indicated that the double-gene plasmid effectively expressed SIRT3 and SIRT4, significantly improved mitochondrial quality and function, and reduced mtDNA level and oxidative stress in HCC cells. MitoTracker analysis revealed that the mitochondrial network was restored. The proliferation, migration capabilities of HCC cells were reduced, whereas their differentiation abilities were enhanced. This study demonstrated that the use of IRES-linked SIRT3 and SIRT4 double-gene vectors induced the differentiation of HCC cells and inhibited their development by ameliorating mitochondrial dysfunction. This intervention helped reverse metabolic reprogramming, and may provide a groundbreaking new framework for HCC treatment.

Rosmarinic Acid Activates the Nrf2/ARE Signaling Pathway via the miR-25-3p/SIRT6 Axis to Inhibit Vascular Remodeling.

The vital pathological processes in intimal hyperplasia include aberrant vascular smooth muscle cells (VSMCs) proliferation, migration, and phenotypic switching. Rosmarinic acid (RA) is a natural phenolic acid compound. Nevertheless, the underlying mechanism of RA in neointimal hyperplasia is still unclear. Our analysis illustrated that miR-25-3p mimics significantly enhanced PDGF-BB-mediated VSMCs proliferation, migration, and phenotypic switching while RA partially weakened the effect of miR-25-3p. Mechanistically, we found that miR-25-3p directly targets sirtuin (SIRT6). The suppressive effect of the miR-25-3p inhibitor on PDGF-BB-induced VSMCs proliferation, migration, and phenotypic switch was partially eliminated by SIRT6 knockdown. The suppression of the PDGF-BB-stimulated Nrf2/ARE signaling pathway that was activated by the miR-25-3p inhibitor was exacerbated by the SIRT6 knockdown. In in vivo experiments, RA reduced the degree of intimal hyperplasia while miR-25-3p agomir partially reversed the suppressive effect of RA in vascular remodeling. Our results indicate that RA activates the Nrf2/ARE signaling pathway via the miR-25-3p/SIRT6 axis to inhibit vascular remodeling.

SIRT7 promotes the proliferation and migration of anaplastic thyroid cancer cells by regulating the desuccinylation of KIF23.

OBJECTIVE: This study was designed to investigate the regulatory effects of kinesin family member (KIF) 23 on anaplastic thyroid cancer (ATC) cell viability and migration and the underlying mechanism. METHODS: Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used to analyze the levels of KIF23 in ATC cells. Besides, the effects of KIF23 and sirtuin (SIRT) 7 on the viability and migration of ATC cells were detected using cell counting kit-8, transwell and wound healing assays. The interaction between SIRT7 and KIF23 was evaluated by co-immunoprecipitation (Co-IP) assay. The succinylation (succ) of KIF23 was analyzed by western blot. RESULTS: The KIF23 expression was upregulated in ATC cells. Silencing of KIF23 suppressed the viability and migration of 8505C and BCPAP cells. The KIF23-succ level was decreased in ATC cells. SIRT7 interacted with KIF23 to inhibit the succinylation of KIF23 at K537 site in human embryonic kidney (HEK)-293T cells. Overexpression of SIRT7 enhanced the protein stability of KIF23 in HEK-293T cells. Besides, overexpression of KIF23 promoted the viability and migration of 8505C and BCPAP cells, which was partly blocked by silenced SIRT7. CONCLUSIONS: SIRT7 promoted the proliferation and migration of ATC cells by regulating the desuccinylation of KIF23.

SIRT6 Reduces Rheumatoid Arthritis Injury by Inhibiting MyD88-ERK Signaling Pathway.

BACKGROUND: Rheumatoid arthritis (RA) is an autoimmune disease characterized by destruction of synovial joints, abnormal immune responses and chronic inflammatory manifestations, which seriously affects patients' well-being. We explored this study to ascertain the effect and mechanism of silent information regulator 6 (SIRT6) on RA. METHODS: Genes of RA patients and normal volunteers were analyzed using Gene Expression Omnibus (GEO), Kyoto-Encyclopedia of Genes and Genomes (KEGG) and Disconet databases. Serum samples of RA patients and normal subjects were collected before detection of myeloid differentiation factor-88 (MyD88)-extracellular signal-regulated kinase (ERK) pathway proteins expression with Western blot. In vitro RA fibroblast-like synoviocytes (FLS) cell model (RA-FLS) was established by treating RSC-364 with recombinant rat IL-1ß (10 ng/mL) after which SIRT6 and MyD88 adenoviruses treatment was carried out. The enzyme linked immunoassay (ELISA), real time polymerase chain reaction (RT-PCR) and Western blot were respectively used to measure inflammatory factors, related messenger ribonucleic acid (mRNA) and protein expressions. Also, we constructed RA rat model with bovine type II collagen (BIIC) and complete Freund's adjuvant, before treatment with SIRT6 and MyD88 adenoviruses. RESULTS: Low expression of SIRT6 gene were detected in RA patients. Also, levels of MyD88, ERK and phosphorylated extracellular signal-regulated protein kinase (p-ERK) protein expressions in RA patients were increased, whilst that of SIRT6 protein decreased. Compared to FLS cells in Control group, inflammatory factors levels of rats in Model batch increased significantly. SIRT6 adenovirus treatment potentially and significantly inhibited inflammation including suppression of increased inflammatory factors induced by MyD88. In comparison with FLS cells in Control group, Model batch cells' MyD88, interleukin (IL)-1ß, IL-21, IL-22, IL-6, IL-17, tumor necrosis factor-alpha (TNF-α) and monocyte chemo-attractant protein-1 (MCP-1) mRNA expressions increased but SIRT6 gene treatment could reduce mRNA expression of the aforesaid factors, even after MyD88 adenovirus treatment. Besides, overpressed SIRT6 negatively regulated levels of MyD88, ERK and p-ERK proteins expressions. SIRT6 demonstrated anti-RA effect by regulating MyD88-ERK pathway and inhibiting inflammatory response in RA rats. CONCLUSIONS: SIRT6 could potentially inhibit the inflammatory response of RA via a regulatory mechanism mainly relating to MyD88-ERK signal pathway. Thus, SIRT6 and its agonists may serve as new targets for developing drugs that can potentially treat RA.

Inhibition of SIRT7 overcomes sorafenib acquired resistance by suppressing ERK1/2 phosphorylation via the DDX3X-mediated NLRP3 inflammasome in hepatocellular carcinoma.

AIMS: Sirtuin 7 (SIRT7) plays an important role in tumor development, and has been characterized as a potent regulator of cellular stress. However, the effect of SIRT7 on sorafenib acquired resistance remains unclear and a possible anti-tumor mechanism beyond this process in HCC has not been clarified. We examined the therapeutic potential of SIRT7 and determined whether it functions synergistically with sorafenib to overcome chemoresistance. METHODS: Cancer Genome Atlas-liver HCC data and unbiased gene set enrichment analyses were used to identify SIRT7 as a potential effector molecule in sorafenib acquired resistance. Two types of SIRT7 chemical inhibitors were developed to evaluate its therapeutic properties when synergized with sorafenib. Mass spectrometry was performed to discover a direct target of SIRT7, DDX3X, and DDX3X deacetylation levels and protein stability were explored. Moreover, an in vivo xenograft model was used to confirm anti-tumor effect of SIRT7 and DDX3X chemical inhibitors combined with sorafenib. RESULTS: SIRT7 inhibition mediated DDX3X depletion can re-sensitize acquired sorafenib resistance by disrupting NLRP3 inflammasome assembly, finally suppressing hyperactive ERK1/2 signaling in response to NLRP3 inflammasome-mediated IL-1ß inhibition. CONCLUSIONS: SIRT7 is responsible for sorafenib acquired resistance, and its inhibition would be beneficial when combined with sorafenib by suppressing hyperactive pro-cell survival ERK1/2 signaling.

Sirtuin 6 ameliorates arthritis through modulating cyclic AMP-responsive element binding protein/CCN1/cyclooxygenase 2 pathway in osteoblasts.

INTRODUCTION: CCN1 is an immediate-early gene product pivotal for arthritis progression. We have previously shown that sirtuin 6 (SIRT6) inhibited hypoxia-induced CCN1 expression in osteoblasts. Herein we examined the contribution of cyclic AMP-responsive element binding protein (CREB)/CRE to this suppressive action and the influence of CCN1 on cyclooxygenase (COX) 2 synthesis. MATERIALS AND METHODS: MC3T3-E1 murine osteoblasts were cultured under normoxia (21% oxygen) or hypoxia (2% oxygen). Expressions of CCN1, phospho-CREB (Ser133), COX2 and relevant kinases were assessed by Western blot. SIRT6 was overexpressed in cultured osteoblasts and arthritic joints by a lentiviral-based technique. Activities of CCN1 gene promoter constructs were examined by luciferase reporter assay. Interaction between CREB and CCN1 promoter was assessed by chromatin immunoprecipitation (ChIP). Collagen-induced arthritis (CIA) was established in 20 rats to evaluate the effects of SIRT6 therapy on osteoblastic expressions of phospho-CREB, CCN1 and COX2. RESULTS: SIRT6 suppressed hypoxia-enhanced CCN1 expression and CREB phosphorylation. Attenuation of calcium/calmodulin-dependent protein kinase II (CaMKII) may be responsible for SIRT6-induced CREB inhibition. CRE at - 286 bp upstream of the ATG start codon was essential for CCN1 expression under hypoxia and SIRT6 reduced hypoxia-stimulated CREB/CRE interaction. Forced expression of CREB rescued SIRT6-suppressed CCN1 synthesis. CCN1 induced COX2 expression in osteoblasts. In rat CIA, the therapeutic effect of SIRT6 was accompanied by decreases in osteoblastic expressions of phospho-CREB, CCN1 and COX2. CONCLUSION: Our study indicated that the benefits of SIRT6 to inflammatory arthritis and bone resorption are at least partially derived from its modulation of CREB/CCN1/COX2 pathway in osteoblasts.

Overexpression of SIRT6 regulates NRF2/HO-1 and NF-κB signaling pathways to alleviate UVA-induced photoaging in skin fibroblasts.

Skin photoaging, resulting from prolonged exposure to sunlight, especially UVA rays, has been identified as a key contributor to age-related skin degeneration. However, the mechanism by which UVA radiation induces skin cell senescence has not been fully elucidated. In this investigation, bioinformatics technology was employed to identify SIRT6 as the core hub gene involved in the progression of skin photoaging. The study evinced that prolonged exposure of cutaneous fibroblasts to UVA radiation results in a marked reduction in the expression of SIRT6, both in vivo and in vitro. Knockdown of SIRT6 in skin fibroblasts resulted in the upregulation of genes associated with cellular aging, thereby exacerbating the effects of UVA radiation-induced photoaging. Conversely, overexpression of SIRT6 decreased the expression of cell aging-related genes, indicating that SIRT6 plays a role in the regulation of senescence in skin fibroblasts induced by UVA radiation. We proffer substantiation that overexpression of SIRT6 protects skin fibroblasts from UVA-induced oxidative stress by activating the NRF2/HO-1 signaling cascade. Moreover, SIRT6 overexpression also reduced UVA-induced type I collagen degradation by inhibiting NF-κB signaling cascade. In summary, our findings showed that overexpression of SIRT6 inhibits UVA-induced senescence phenotype and type I collagen degradation in skin fibroblasts by modulating the NRF2/HO-1 and NF-κB signaling pathways. And the regulation of these signaling pathways by SIRT6 may be achieved through its deacetylase activity. Therefore, SIRT6 is a novel and promising therapeutic target for skin aging related to age and UV.

Oridonin ameliorates doxorubicin induced-cardiotoxicity via the E2F1/Sirt6/PGC1α pathway in mice.

Doxorubicin induced cardiotoxicity (DIC) arises from mitochondrial dysfunction and oxidative stress. Oridonin (Ori), a natural tetracycline diterpenoid, has shown cardiac protective effect; however, its role in DIC remains unclear. This study investigates the protective effect of Ori against DIC and elucidates its underlying molecular mechanisms. The results demonstrate that Ori significantly alleviated DIC by improving myocardial structure, reducing the proportion of apoptotic cells, and alleviating the myocardial oxidative damage and mitochondrial dysfunction both in vivo and in vitro. Doxorubicin significantly decreased Sirt6 and PGC1α levels in cardiac tissues, which was reversed by Ori. Furthermore, Sirt6 overexpression significantly improved myocardial structure and reduced the proportion of apoptotic cells by reducing oxidative stress and improving mitochondrial function. The protective effect of Ori is neutralized by the Sirt6 inhibitor OSS_128167, evidenced by downregulated mRNA and protein expression of PGC1α. The transcription factor E2F1 was upregulated by doxorubicin, leading to decreased Sirt6 expression-an effect mitigated by Ori. Molecular docking simulations indicate direct binding between Ori and specific amino acid residues on E2F1 through hydroxyl bonds. These findings uncover a novel mechanism whereby Ori attenuates DIC by modulating the E2F1/Sirt6/PGC1α pathway.

Orientin inhibits inflammation in chondrocytes and attenuates osteoarthritis through Nrf2/NF-κB and SIRT6/NF-κB pathway.

Effects of Orientin on murine chondrocytes treated with interleukin-1ß (IL-1ß) were evaluated using qPCR, western blot analysis, ELISA, and immunofluorescent staining in vitro. In vivo, We established a standard OA model by performing the destabilized medial meniscus (DMM) surgery on C57BL/6 mice, and assessed healing effect of Orientin by X-ray imaging, histopathological analysis, immunohistochemical staining. Osteoarthritis (OA) is the most common form of degenerative joint disease in clinic and the chondrocyte inflammation plays the most important role in OA development. The natural flavonoid compound (Orientin) has anti-inflammatory bioactive properties in the treatment of various diseases. But studies have not explored whether Orientin modulates OA progression. In this study, a significant suppression in IL-1ß-mediated pro-inflammatory mediators and the degradation of cartilage extracellular matrix (ECM) was observed in vitro through qPCR, western blot analysis, ELISA, and immunofluorescent staining after the treatment with Orientin. In addition, Orientin abrogated DMM surgery induced cartilage degradation in mice, which was assessed by X-ray imaging, histopathological analysis, immunohistochemical staining. Mechanistic studies showed that Orientin suppressed OA development by downregulating activation of NF-κB by activating Nrf2/HO-1 axis and SIRT6 signaling pathway. These results provide evidence that Orientin serves as a potentially viable compound for the treatment of OA.

Effects of atorvastatin on the Sirtuin/PXR signaling pathway in Mugilogobius chulae.

Atorvastatin (ATV) is a hypolipidemic drug widely detected in the aquatic environment. Nevertheless, limited information is provided about the toxic effects of ATV on estuary or coastal species and the underlying mechanisms. In the present study, the responses of genes expression in pregnane X receptor (PXR) signaling pathway and enzymatic activities in the liver of the estuarine benthic fish (Mugilogobius chulae) were investigated under acute and sub-chronic ATV exposure. Results showed that PXR was significantly inhibited in the highest exposure concentration of ATV for a shorter time (24 h, 500 µg L-1) but induced in a lower concentration (72 h, 5 µg L-1). The downstream genes in PXR signaling pathway such as CYP3A, SULT, UGT, and GST showed similar trends to PXR. P-gp and MRP1 were repressed in most treatments. GCLC associated with GSH synthesis was mostly induced under ATV exposure for a long time (168 h), suggesting that reactive oxygen species (ROS) were generated under ATV exposure. Similarly, GST and SOD enzymatic activities significantly increased in most exposure treatments. Under ATV exposure, SIRT1 and SIRT2 displayed induction to some extent in most treatments, suggesting that SIRTs may affect PXR expression by regulating the acetylation levels of PXR. The investigation demonstrated that ATV exposure affected the expression of the Sirtuin/PXR signaling pathway, thus further interfered adaption of M. chulae to the environment.

SIRT6 ameliorates LPS-induced apoptosis and tight junction injury in ARDS through the ERK1/2 pathway and autophagy.

Sirtuin6 (SIRT6) has been demonstrated to be involved in a range of physiological processes and diseases, while its role in acute respiratory distress syndrome (ARDS) remains unclear. Therefore, this study focused on the role and underlying mechanism of SIRT6 in ARDS with the aim of identifying potential therapeutic targets. In this study, we found that SIRT6 was significantly decreased in lipopolysaccharide (LPS)-induced A549 cells and a murine model. In vitro overexpression of SIRT6 restored the expression of tight junction proteins (ZO-1 and occludin) and alleviated cell apoptosis and inflammation, while knockdown of SIRT6 aggravated the loss of tight junction proteins (ZO-1 and occludin) and promoted cell apoptosis and inflammation in LPS-induced A549 cells. Furthermore, the overexpression of SIRT6 enhanced autophagy and inhibited the ERK1/2 pathway, while the knockdown of SIRT6 inhibited autophagy and activated the ERK1/2 pathway. The autophagy activator rapamycin and the ERK1/2 inhibitor PD98059 rescued the effects of SIRT6 knockdown on tight junction proteins, apoptosis, and inflammation. Mechanistically, SIRT6 deacetylated histone 3 at Lys9 to negatively regulate the ERK1/2 pathway. In vivo, the SIRT6-specific inhibitor OSS_128167 also significantly accelerated LPS-induced loss of tight junction proteins, lung inflammation, and apoptosis. Meanwhile, the SIRT6-specific inhibitor OSS_128167 also activated the ERK1/2 pathway and inhibited lung autophagy. These results suggested that SIRT6 could ameliorate the loss of tight junction proteins, inflammation, and apoptosis in LPS-induced ARDS by inhibiting the ERK1/ 2 pathway and enhancing autophagy, indicating that SIRT6 plays a beneficial role in ARDS and might be a potential therapeutic target for ARDS.

Metabolic Syndrome and Cardiac Remodeling Due to Mitochondrial Oxidative Stress Involving Gliflozins and Sirtuins.

PURPOSE OF REVIEW: To address the mechanistic pathways focusing on mitochondria dysfunction, oxidative stress, sirtuins imbalance, and other contributors in patient with metabolic syndrome and cardiovascular disease. Sodium glucose co-transporter type 2 (SGLT-2) inhibitors deeply influence these mechanisms. Recent randomized clinical trials have shown impressive results in improving cardiac function and reducing cardiovascular and renal events. These unexpected results generate the need to deepen our understanding of the molecular mechanisms able to generate these effects to help explain such significant clinical outcomes. RECENT FINDINGS: Cardiovascular disease is highly prevalent among individuals with metabolic syndrome and diabetes. Furthermore, mitochondrial dysfunction is a principal player in its development and persistence, including the consequent cardiac remodeling and events. Another central protagonist is the renin-angiotensin system; the high angiotensin II (Ang II) activity fuel oxidative stress and local inflammatory responses. Additionally, sirtuins decline plays a pivotal role in the process; they enhance oxidative stress by regulating adaptive responses to the cellular environment and interacting with Ang II in many circumstances, including cardiac and vascular remodeling, inflammation, and fibrosis. Fasting and lower mitochondrial energy generation are conditions that substantially reduce most of the mentioned cardiometabolic syndrome disarrangements. In addition, it increases sirtuins levels, and adenosine monophosphate-activated protein kinase (AMPK) signaling stimulates hypoxia-inducible factor-1ß (HIF-1 beta) and favors ketosis. All these effects favor autophagy and mitophagy, clean the cardiac cells with damaged organelles, and reduce oxidative stress and inflammatory response, giving cardiac tissue protection. In this sense, SGLT-2 inhibitors enhance the level of at least four sirtuins, some located in the mitochondria. Moreover, late evidence shows that SLGT-2 inhibitors mimic this protective process, improving mitochondria function, oxidative stress, and inflammation. Considering the previously described protection at the cardiovascular level is necessary to go deeper in the knowledge of the effects of SGLT-2 inhibitors on the mitochondria function. Various of the protective effects these drugs clearly had shown in the trials, and we briefly describe it could depend on sirtuins enhance activity, oxidative stress reduction, inflammatory process attenuation, less interstitial fibrosis, and a consequent better cardiac function. This information could encourage investigating new therapeutic strategies for metabolic syndrome, diabetes, heart and renal failure, and other diseases.

A Comprehensive Study on the Anti-cancer Effects of Quercetin and Its Epigenetic Modifications in Arresting Progression of Colon Cancer Cell Proliferation.

Colon cancer etiology involves a wide spectrum of genetic and epigenetic alterations, finding it challenging to find effective therapeutic strategies. Quercetin exhibits potent anti-proliferative/apoptotic properties. In the present study, we aimed to elucidate the anti-cancer and anti-aging effect of quercetin in colon cancer cell lines. The anti-proliferative effect of quercetin was assessed in vitro by CCK-8 in normal and colon cancer cell lines. To check the anti-aging potential of quercetin, collagenase, elastase, and hyaluronidase inhibitory activity assays were performed. The epigenetic and DNA damage assays were performed using the human NAD-dependent deacetylase Sirtuin-6, proteasome 20S, Klotho, Cytochrome-C, and telomerase ELISA kits. Furthermore, the aging-associated miRNA expression profiling was performed on colon cancer cells. The treatment with quercetin inhibited cell proliferation of colon cancer cells in a dose-dependent manner. Quercetin arrested colon cancer cell growth by modulating expression of aging proteins including Sirtuin-6 and Klotho and also by inhibiting telomerase activity to restrict the telomere length which is evident from qPCR analysis. Quercetin also exhibited DNA damage protection by reducing proteasome 20S levels. The miRNA expression profiling results displayed differential expression of miRNA in colon cancer cell, and in addition, the highly upregulated miRNA was involved in the regulation of cell cycle, proliferation, and transcription. Our data suggest that quercetin treatment inhibited cell proliferation in colon cancer cells through regulating the anti-aging protein expression and provides better understanding for quercetin's potential use in colon cancer treatment.

Chrysophanol prevents IL-1ß-Induced inflammation and ECM degradation in osteoarthritis via the Sirt6/NF-κB and Nrf2/NF-κB axis.

Osteoarthritis (OA) is a common joint illness that negatively impacts people's lives. The main active ingredient of cassia seed or rhubarb is chrysophanol. It has various pharmacological effects including anticancer, anti-diabetes and blood lipid regulation. Previous evidence suggests that chrysophanol has anti-inflammatory properties in various diseases, but its effect on OA has not been investigated yet. In this study, chrysophanol inhibited IL-1ß -induced expression of ADAMTS-4, MMP13, COX-2 and iNOS. Meanwhile, it can inhibit aggrecan and collagen degradation in osteoarthritic chondrocytes induced by IL-1ß.Further studies depicted that SIRT6 silencing eliminated the chrysophanol effect on IL-1ß. The results demonstrated that chrysophanol could stimulate SIRT6 activation and, more importantly, increase SIRT6 levels. We also discovered that chrysophanol might impede the NF-κB pathway of OA mice's chondrocytes induced by IL-1ß, which could be because it depends on SIRT6 activation to some extent. It had also been previously covered that chrysophanol could produce a marked effect on Nrf2/NF-κB axis [1]. Therefore, we can infer that chrysophanol may benefit chondrocytes by regulating the SIRT6/NF-κB and Nrf2/NF-κB signaling axis.We examined the anti-inflammatory mechanism and the impact of chrysophanol on mice in vitro and in vivo. In summary, we declare that chrysophanol diminishes the inflammatory reaction of OA in mice in vitro by regulating SIRT6/NF-κB and Nrf2/NF-κB signaling pathway and protects articular cartilage from degradation in vivo. We can infer that chrysophanol could be an efficient therapy for OA.

In vivo exposure to a new 2-cyano-2-p-nitrophenyl-N-benzylthioamide decreases doxorubicin-triggered structural damages in the mature testis.

The use of doxorubicin (DOX) in clinical practice continues to be challenged by its severe toxicity. DOX cytotoxic activity is not only directed against malignant tumours, given that the treatment will damage healthy tissues as well leading to irreversible injuries. This study aimed to address the in vivo effects of DOX and its co-administration with a new analog of thioamide; thiocyanoacetamide (TA) on the germinal epithelium. Thus, male rats received either intravenous injection (iv) of 0.03 mg/kg of body weight/week, 0.9% NaCl and were regarded as the control group (CTR), treated with DOX (3.7 mg/kg/week iv), TA [10 mg/kg/day intragastrically (ig)] or a co-supplementation of DOX and TA. After 50 days, the left testes were dissected and used for toluidine blue, periodic acid-Schiff (PAS) staining (to evaluate the change in polysaccharides/glycoproteins content), and transmission electron microscopy (TEM) (to assess the morphological damages). To estimate the impact of the test compounds on mitochondrial biogenesis, the expression of NAD-dependent deacetylase sirtuin-3 (SIRT-3) and proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) were evaluated by immunofluorescence. Apoptotic cells were observed using Hoechst 33324 fluorescent staining. Data showed testicular injuries in the DOX-treated group, manifested by a significant decrease in total germ cell (GC) number, alteration of Sertoli cell (SC) nucleolus, anchoring junction, along with modifications of the basement membrane (BM) regularity and increase in apoptotic cell count. Mitochondrial aspect and SIRT-3 and PGC-1α expression in the testis were unaffected by the DOX. Co-therapy increased GC number, decreased apoptotic cell count, and restored the BM and anchoring junction regular aspects. This study provides novel insights into understanding DOX-mediated impairment in rats' testis and might offer some basis for the emerging new alternative therapeutic schemes in male patients undergoing chemotherapy.

Ginsenoside Rc Modulates SIRT6-NRF2 Interaction to Alleviate Alcoholic Liver Disease.

Alcoholic liver disease (ALD) is a serious worldwide health problem. Ginsenoside Rc is a major active ingredient isolated from Panax ginseng, whose pharmacological effects counteract oxidative stress, inflammation, and lipid accumulation. However, it is still unclear whether ginsenoside Rc might exert beneficial effects on alcohol-induced liver injury. To this aim, mice primary hepatocytes (MPHs) were challenged with alcohol to test ginsenoside Rc's effects on their intracellular alcohol metabolism. C57BL/6J mice or SIRT6alb-/- mice were chronically fed a diet with added alcohol or given a single gavage of alcohol with or without ginsenoside Rc. Analyses of alcohol metabolism, oxidative stress, inflammation, lipid metabolism, and RNaseq expression were conducted to explore potential targets exploited by ginsenoside Rc to protect against ALD. Our results showed that ginsenoside Rc attenuated alcohol-induced liver injury by regulating oxidative stress, inflammation, and lipid accumulation both in vivo and in vitro. Ginsenoside Rc did increase the deacetylase activity of SIRT6, thereby lowering acetylated NRF2 levels, which elevated NRF2's stability, and subsequently exerting an antioxidant effect. In keeping with this, the hepatic knockout of SIRT6 almost abolished the hepatoprotective effects of ginsenoside Rc against ALD. Therefore, our results suggest that ginsenoside Rc attenuated hepatocytes' damage and oxidative stress in ALD by up-regulating the SIRT6/NRF2 pathway. Hence, ginsenoside Rc may be a promising drug to treat or relieve ALD.

Responses of bitter melon saponins to oxidative stress and aging via the IIS pathway linked with sir-2.1 and hlh-30.

Saponins from bitter melon (BMS) exert potential bioactivities and pharmacological activities, including anti-oxidation and lifespan extension. However, the exact mechanisms of BMS in response to oxidative stress remain unknown. Results demonstrated that bitter melon saponins could strengthen locomotive activities (body bend and head thrashing) accompanied by delaying the muscle fiber damage with age in Caenorhabditis elegans. In addition, BMS inhibited the ROS accumulation, improved the activities of antioxidant enzymes like SOD (by 57.90% and 94.34% for 100 µg/ml and 200 µg/ml BMS, respectively) and CAT (by 51.45% and 56.91% for 100 µg/ml and 200 µg/ml BMS, respectively), and extend the lifespan of N2 and CL2006 worms under paraquat-induced oxidative stress. Mechanism study suggested that BMS modulated the mRNA expressions of oxidation-related regulators, like the upregulation of cat-1, hsf-1, sir-2.1, and hlh-30. Furthermore, gene-deficient mutants verified that IIS (insulin/insulin-like growth factor-1 signaling) pathway linked with sir-2.1 and hlh-30 factors were involved in the BMS's lifespan-extension effects under oxidative stress. In general, this study supplemented the explanation of BMS in promoting oxidation-resistance and lifespan-extension activities, which could be served as a potential candidate for anti-aging. PRACTICAL APPLICATIONS: Our previous studies have suggested that saponins from bitter melon exhibited fat-lowering activity in C. elegans. However, little was known about the mechanism underlying the anti-oxidation effects of BMS in C. elegans. Current results indicated that the IIS pathway linked with sir-2.1 and hlh-30 transcriptional factors jointly to increase the lifespan in BMS' responses to oxidative stress. Our findings are beneficial to understand the main nutritional ingredients in bitter melon, which are ideal and expected in functional foods for aging.

Critical Reanalysis of the Mechanisms Underlying the Cardiorenal Benefits of SGLT2 Inhibitors and Reaffirmation of the Nutrient Deprivation Signaling/Autophagy Hypothesis.

SGLT2 (sodium-glucose cotransporter 2) inhibitors produce a distinctive pattern of benefits on the evolution and progression of cardiomyopathy and nephropathy, which is characterized by a reduction in oxidative and endoplasmic reticulum stress, restoration of mitochondrial health and enhanced mitochondrial biogenesis, a decrease in proinflammatory and profibrotic pathways, and preservation of cellular and organ integrity and viability. A substantial body of evidence indicates that this characteristic pattern of responses can be explained by the action of SGLT2 inhibitors to promote cellular housekeeping by enhancing autophagic flux, an effect that may be related to the action of these drugs to produce simultaneous upregulation of nutrient deprivation signaling and downregulation of nutrient surplus signaling, as manifested by an increase in the expression and activity of AMPK (adenosine monophosphate-activated protein kinase), SIRT1 (sirtuin 1), SIRT3 (sirtuin 3), SIRT6 (sirtuin 6), and PGC1-α (peroxisome proliferator-activated receptor γ coactivator 1-α) and decreased activation of mTOR (mammalian target of rapamycin). The distinctive pattern of cardioprotective and renoprotective effects of SGLT2 inhibitors is abolished by specific inhibition or knockdown of autophagy, AMPK, and sirtuins. In the clinical setting, the pattern of differentially increased proteins identified in proteomics analyses of blood collected in randomized trials is consistent with these findings. Clinical studies have also shown that SGLT2 inhibitors promote gluconeogenesis, ketogenesis, and erythrocytosis and reduce uricemia, the hallmarks of nutrient deprivation signaling and the principal statistical mediators of the ability of SGLT2 inhibitors to reduce the risk of heart failure and serious renal events. The action of SGLT2 inhibitors to augment autophagic flux is seen in isolated cells and tissues that do not express SGLT2 and are not exposed to changes in environmental glucose or ketones and may be related to an ability of these drugs to bind directly to sirtuins or mTOR. Changes in renal or cardiovascular physiology or metabolism cannot explain the benefits of SGLT2 inhibitors either experimentally or clinically. The direct molecular effects of SGLT2 inhibitors in isolated cells are consistent with the concept that SGLT2 acts as a nutrient surplus sensor, and thus, its inhibition causes enhanced nutrient deprivation signaling and its attendant cytoprotective effects, which can be abolished by specific inhibition or knockdown of AMPK, sirtuins, and autophagic flux.

Fasudil Protects Against Adriamycin-induced Acute Heart Injury by Inhibiting Oxidative Stress, Apoptosis, and Cellular Senescence.

BACKGROUND: The clinical utility of Adriamycin (ADR) is limited due to its toxicity, particularly cardiotoxicity. Therefore, effective cardioprotective adjuvants to minimize ADR-induced acute cardiotoxicity are urgently needed. Our previous studies have demonstrated the protective roles of fasudil on tissue injury. Here, we further explore whether inhibition of Rho-kinase could alleviate the acute heart injury induced by ADR. METHODS: C57BL6 mice were randomly divided into the following four groups: ① ADR group; ② low-dose fasudil (ADR+L); ③ high-dose fasudil (ADR+H); and ④ control group (CON). Animals were injected i.p 20 mg/kg ADR once in group ①~③. Animals were injected i.p fasudil (2 or 10 mg/kg/day) daily for consecutive 6 days in groups ② and ③, respectively. Blood samples and heart tissues were collected for assays. H9C2 cells were treated with fasudil for 30 mins and then incubated with ADR for 24 hours. Cells were collected for immunohistochemistry and western blot study, respectively. RESULTS: In the mouse model, administration of fasudil significantly ameliorated ADR-induced cardiac damage, suppressed cell apoptosis and senescence, and ameliorated redox imbalance and DNA damage. In vitro, fasudil treatment ameliorated ADR-induced immunofluorescence reaction of 8-OHdG, decreased the expression of TUNEL cells and proteins of Bax, Caspase-3 and p53, and increased the expression of proteins of Bcl-2 and SIRT 1. CONCLUSION: Fasudil has a protective effect on ADR induced acute cardiotoxicity, which is partially attributed to its antioxidant, anti-senescence, and anti-apoptotic effects.