Urolithin a alleviates oxidative stress-induced senescence in nucleus pulposus-derived mesenchymal stem cells through SIRT1/PGC-1α pathway

BACKGROUND In degenerative intervertebral disc(IVD),an unfavorable IVD environment leads to increased senescence of nucleus pulposus(NP)-derived mesenchymal stem cells(NPMSCs)and the inability to complete the differentiation from NPMSCs to NP cells,leading to further aggravation of IVD degeneration(IDD).Urolithin A(UA)has been proven to have obvious effects in delaying cell senescence and resisting oxidative stress.AIM To explore whether UA can alleviate NPMSCs senescence and to elucidate the underlying mechanism.METHODS In vitro,we harvested NPMSCs from rat tails,and divided NPMSCs into four groups:the control group,H2O2 group,H2O2+UA group,and H2O2+UA+SR-18292 group.Senescence-associatedβ-Galactosidase(SA-β-Gal)activity,cell cycle,cell proliferation ability,and the expression of senescence-related and silent information regulator of transcription 1/PPAR gamma coactivator-1α(SIRT1/PGC-1α)pathway-related proteins and mRNA were used to evaluate the protective effects of UA.In vivo,an animal model of IDD was constructed,and Xrays,magnetic resonance imaging,and histological analysis were used to assess whether UA could alleviate IDD in vivo.RESULTS We found that H2O2 can cause NPMSCs senescence changes,such as cell cycle arrest,reduced cell proliferation ability,increased SA-β-Gal activity,and increased expression of senescence-related proteins and mRNA.After UA pretreatment,the abovementioned senescence indicators were significantly alleviated.To further demonstrate the mechanism of UA,we evaluated the mitochondrial membrane potential and the SIRT1/PGC-1αpathway that regulates mitochondrial function.UA protected mitochondrial function and delayed NPMSCs senescence by activating the SIRT1/PGC-1αpathway.In vivo,we found that UA treatment alleviated an animal model of IDD by assessing the disc height index,Pfirrmann grade and the histological score.CONCLUSION In summary,UA could activate the SIRT1/PGC-1αsignaling pathway to protect mitochondrial function and alleviate cell senescence and IDD in vivo and vitro.

Hepatocellular carcinoma-derived exosomal miRNA-761 regulates the tumor microenvironment by targeting the SOCS2/JAK2/STAT3 pathway

BACKGROUND:Exosomes and exosomal microRNAs have been implicated in tumor occurrence and metastasis.Our previous study showed that microRNA-761(miR-761)is overexpressed in hepatocellular carcinoma(HCC)tissues and that its inhibition affects mitochondrial function and inhibits HCC metastasis.The mechanism by which exosomal miR-761 modulates the tumor microenvironment has not been elucidated.METHODS:Exosomal miR-761 was detected in six cell lines.Cell counting kit-8(CCK-8)and transwell migration assays were performed to determine the function of exosomal miR-761 in HCC cells.The luciferase reporter assay was used to analyze miR-761 target genes in normal fi broblasts(NFs).The inhibitors AZD1480 and C188-9 were employed to determine the role of the Janus kinase 2/signal transducer and activator of transcription 3(JAK2/STAT3)signaling pathway in the transformation of cancer-associated fi broblasts(CAFs).RESULTS:In this study,we characterized the mechanism by which miR-761 reprogrammed the tumor microenvironment.We found that HCC-derived exosomal miR-761 was taken up by NFs.Moreover,HCC exosomes aff ected the tumor microenvironment by activating NFs via suppressor of cytokine signaling 2(SOCS2)and the JAK2/STAT3 signaling pathway.CONCLUSIONS:These results demonstrated that exosomal miR-761 modulated the tumor microenvironment via SOCS2/JAK2/STAT3 pathway-dependent activation of CAFs.Our fi ndings may inspire new strategies for HCC prevention and therapy.

Liuwei Dihuang Pill(六味地黄丸)Treats Postmenopausal Osteoporosis with Shen(Kidney) Yin Deficiency via Janus Kinase/Signal Transducer and Activator of Transcription Signal Pathway by Up-regulating Cardiotrophin-Like Cytokine Factor 1 Expression

Objectives： To investigate the mechanism of Liuwei Dihuang Pill （六味地黄丸, LDP） in treating postmenopausal osteoporosis （PMOP） with Shen （Kidney） yin deficiency. Methods： In this study, 205 cases of PMOP were divided into the PMOP Shen-yin deficiency group （Group A）, PMOP Shen-yang deficiency group （Group B）, PMOP without Shen deficiency group （Group C）, and control group （Group N）. Real-time polymerase chain reaction （RT-PCR） and Western blot techniques were used to observe the effects of LDP treatment on the cardiotrophin-like cytokine factor 1 （CLCF1）, ankyrin repeat and SOCS box containing 1 （ASB1）, and proldneticin 2 （PROK2） genes and the Janus kinase/signal transducer and activator of transcription （JAK/STAT） signaling pathway. Results： The mRNA （P〈0.05） and protein （P〈0.01） expression levels of the CLCF1 gone in Group A were significantly lower than the corresponding levels in Group N. After LDP treatment for 3 months, the mRNA expression levels of the CLCF1 gone were obviously up-regulated （P〈0.01）. After 6-month treatment, the expression levels of CLCF1 mRNA and protein were significantly up-regulated （both P〈0.01）, and the average bone density of the top femur had significantly increased （P〈0.05）. In vitro, CLCF1 overexpression resulted in a significant increase in the total protein and phosphorylated protein levels of JAK2 and STAT3. Conclusions： The CLCF1 gone is an important gone associated with PMOP Shen-yin deficiency and the therapeutic effects of LDP may be mediated by up-regulation of CLCF1 gone expression and activation of the JAK/STAT signaling pathway.

Interleukin-10 Contributes to Therapeutic Effect of Mesenchymal Stem Cells for Acute Liver Failure via Signal Transducer and Activator of Transcription 3 Signaling Pathway

Background：Mesenchymal stem cells （MSCs） transplantation has been proven to have therapeutic potential for acute liver failure （ALF）.However,the mechanism remains controversial.Recently,modulation of inflammation by MSCs has been regarded as a crucial mechanism.The aim of the present study was to explore the soluble cytokines secreted by MSCs and their therapeutic effects in ALF.Methods：MSCs isolated from Sprague-Dawley rats were identified by fluorescence-activated cell sorting analysis.Conditioned medium derived from MSCs （MSCs-CM） was collected and analyzed by a cytokine microarray.MSCs and MSCs-CM were transplanted into rats with D-galactosamine-induced ALF.Liver function,survival rate,histology,and inflammatory factors were determined.Exogenous recombinant rat interleukin （IL）-10,anti-rat IL-10 antibody,and AG490 （signal transducer and activator of transcription 3 [STAT3] signaling pathway inhibitor） were administered to explore the therapeutic mechanism of MSCs-CM.Statistical analysis was performed with SPSS version 19.0,and all data were analyzed by the independent-sample t-test.Results：There are statistical differences of the survival curve between ALF＋MSCs group and ALF＋Dulbecco＆#39;s modified Eagle＆#39;s medium （DMEM） group,as well as ALF＋MSCs-CM group and ALF＋DMEM group （all P 〈 0.05）.Serum alanine aminotransferase （ALT） level in the ALF＋MSCs and ALF＋MSCs-CM groups was lower than that in the ALF＋DMEM group （865.53＆#177;52.80 vs.1709.75＆#177;372.12 U/L and 964.72＆#177;414.59 vs.1709.75＆#177;372.12 U/L,respectively,all P 〈 0.05）;meanwhile,serum aspartate aminotransferase （AST） level in the ALF＋MSCs and ALF＋MSCs-CM groups was lower than that in the ALF＋DMEM group （2440.83＆#177;511.94 vs.4234.35＆#177;807.30 U/L and 2739.83＆#177;587.33 vs.4234.35＆#177;807.30 U/L,respectively,all P 〈 0.05）.Furthermore,MSCs or MSCs-CM treatment significantly reduced serum interferon-γ （IFN-γ）,IL-1β,IL-6 levels and increased serum IL-10 level compared with DMEM （all P 〈 0.05）.Proteome profile analysis of MSCs-CM indicated the presence of anti-inflammatory factors and IL-l 0 was the most distinct.Blocking of IL-10 confirmed the therapeutic significance of this cytokine.Phosphorylated STAT3 was upregulated after IL-l 0 infusion and inhibition of STAT3 by AG490 reversed the therapeutic effect of IL-10.Conclusions：The factors released by MSCs,especially IL-10,have the potential for therapeutic recovery of ALF,and the STAT3 signaling pathway may mediate the anti-inflammatory effect of IL-10.

Profilin-1 is involved in macroangiopathy induced by advanced glycation end products via vascular remodeling and inflammation

BACKGROUND The accumulation of advanced glycation end products(AGEs)have been implicated in the development and progression of diabetic vasculopathy.However,the role of profilin-1 as a multifunctional actin-binding protein in AGEs-induced atherosclerosis(AS)is largely unknown.AIM To explore the potential role of profilin-1 in the pathogenesis of AS induced by AGEs,particularly in relation to the Janus kinase 2(JAK2)and signal transducer and activator of transcription 3(STAT3)signaling pathway.METHODS Eighty-nine individuals undergoing coronary angiography were enrolled in the study.Plasma cytokine levels were detected using ELISA kits.Rat aortic vascular smooth muscle cells(RASMCs)were incubated with different compounds for different times.Cell proliferation was determined by performing the MTT assay and EdU staining.An AGEs-induced vascular remodeling model was established in rats and histological and immunohistochemical analyses were performed.The mRNA and protein levels were detected using real-time PCR and Western blot analysis,respectively.In vivo,shRNA transfection was performed to verify the role of profilin-1 in AGEs-induced proatherogenic mediator release and aortic remodeling.Statistical analyses were performed using SPSS 22.0 software.RESULTS Compared with the control group,plasma levels of profilin-1 and receptor for AGEs(RAGE)were significantly increased in patients with coronary artery disease,especially in those complicated with diabetes mellitus(P<0.01).The levels of profilin-1 were positively correlated with the levels of RAGE(P<0.01);additionally,the levels of both molecules were positively associated with the degree of coronary artery stenosis(P<0.01).In vivo,tail vein injections of AGEs induced the release of proatherogenic mediators,such as asymmetric dimethylarginine,intercellular adhesion molecule-1,and the N-terminus of procollagen III peptide,concomitant with apparent aortic morphological changes and significantly upregulated expression of the profilin-1 mRNA and protein in the thoracic aorta(P<0.05 or P<0.01).Downregulation of profilin-1 expression with an shRNA significantly attenuated AGEs-induced proatherogenic mediator release(P<0.05)and aortic remodeling.In vitro,incubation of vascular smooth muscle cells(VSMCs)with AGEs significantly promoted cell proliferation and upregulated the expression of the profilin-1 mRNA and protein(P<0.05).AGEs(200μg/mL,24 h)significantly upregulated the expression of the STAT3 mRNA and protein and JAK2 protein,which was blocked by a JAK2 inhibitor(T3042-1)and/or STAT3 inhibitor(T6308-1)(P<0.05).In addition,pretreatment with T3042-1 or T6308-1 significantly inhibited AGEs-induced RASMC proliferation(P<0.05).CONCLUSION AGEs induce proatherogenic events such as VSMC proliferation,proatherogenic mediator release,and vascular remodeling,changes that can be attenuated by silencing profilin-1 expression.These results suggest a crucial role for profilin-1 in AGEs-induced vasculopathy.

Effect of interleukin-6 / signal transducer and activator of transcription 3 pathway on cyclooxygenase- 2 expression in THP- 1 monocyte

Objective To investigate the relationship between the interleukin-6(IL-6)/signal transducer and activator of transcription 3(STAT3)signaling pathway and cyclooxygenase-2(COX-2)expression in THP-1 monocytes.Methods Human THP-1 monocyte was used as the research cell,and the time-dependent expressions of STAT3 phosphorylation and COX-2 were detected

Targeting the JAK/STAT pathway in solid tumors

Aberrant activation of signal transducer and activator of transcription(STAT)proteins is associated with the development and progression of solid tumors.However,as transcription factors,these proteins are difficult to target directly.In this review,we summarize the role of targeting Janus kinases(JAKs),upstream activators of STATs,as a strategy for decreasing STAT activation in solid tumors.Preclinical studies in solid tumor cell line models show that JAK inhibitors decrease STAT activation,cell proliferation,and cell survival;in in vivo models,they also inhibit tumor growth.JAK inhibitors,particularly the JAK1/2 inhibitor ruxolitinib,sensitize cell lines and murine models to chemotherapy,immunotherapy,and oncolytic viral therapy.Ten JAK inhibitors have been or are actively being tested in clinical trials as monotherapy or in combination with other agents in patients with solid tumors;two of these inhibitors are already Food and Drug Administration(FDA)approved for the treatment of myeloproliferative disorders and rheumatoid arthritis,making them attractive agents for use in patients with solid tumors as they are known to be well-tolerated.Four JAK inhibitors(two of which are FDA approved for other indications)have exhibited promising anti-cancer effects in preclinical studies;however,clinical studies specifically assessing their activity against the JAK/STAT pathway in solid tumors have not yet been conducted.In summary,JAK inhibition is a viable option for targeting the JAK/STAT pathway in solid tumors and merits further testing in clinical trials.

Effect of electroacupuncture on JAK2/STAT3 pathway in synovial tissues of rats with rheumatoid arthritis

Objective: To observe the effect of electroacupuncture (EA) on Janus kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) pathway in knee joint synovial tissues of rats with rheumatoid arthritis (RA) and to explore the action mechanism of EA on RA. Methods: Twelve of the 48 SPF male Sprague-Dawley (SD) rats were assigned to a normal group by the random number table method. The remaining 36 rats were subjected to RA model preparation by intradermal injection of the Freund's complete adjuvant into the right hind foot pad of each rat under sterile conditions. After the model was successfully prepared, rats were then divided into a model group, a drug group and an EA group according to a random number table method (n=12). Rats in the drug group were treated with 2 mL aqueous solution of tripterygium glycosides [8.1 mg/(kg?bw)];rats in the EA group were treated with EA at bilateral Yanglingquan (GB 34) and Zusanli (ST 36), for 30 min each time;rats in the normal group and the model group were placed in a special rat fixation tank for 30 min each time, and received the same dose of normal saline as those in the drug group. Rats in all groups received intervention once a day for 4 weeks. Diameter of rat ankle joint and rat arthritis index were measured before and after the intervention. At the end of the experiment, the expressions of phospho-JAK2 and phospho-STAT3 were determined by immunohistochemistry. Quantitative real-time polymerase chain reaction (RT-qPCR) was used to detect JAK2 and STAT3 mRNAs expressions. Results: After the model was produced, the arthritis index >2 was considered successful in model preparation. Compared with the model group, the ankle joint diameters and arthritis indexes of rats in the drug group and the EA group were significantly lower (all P<0.01);immunohistochemical staining cells with phospho-JAK2 and phospho-STAT3 were significantly decreased (all P<0.01);the expression levels of JAK2 and STAT3 mRNAs were decreased with statistical differences (all P<0.01). There were no significant differences between the EA group and the drug group (all P>0.05). Conclusion: EA can alleviate the inflammatory response of RA rats, improve their pathological conditions, reduce the expressions of phospho-JAK2 and phospho-STAT3 in the synovial tissue of knee joint, and decrease the expressions of JAK2 and STAT3 mRNAs. The therapeutic effect of EA is comparable to that of the tripterygium glycosides. The mechanism of EA treatment may be related to the inactivation of the JAK2/STAT3 pathway.