Argatroban promotes recovery of spinal cord injury by inhibiting the PAR1/JAK2/STAT3 signaling pathway

Argatroban is a synthetic thrombin inhibitor approved by U.S.Food and Drug Administration for the treatment of thrombosis.However,whether it plays a role in the repair of spinal cord injury is unknown.In this study,we established a rat model of T10 moderate spinal cord injury using an NYU Impactor ModerⅢand performed intraperitoneal injection of argatroban for 3 consecutive days.Our results showed that argatroban effectively promoted neurological function recovery after spinal cord injury and decreased thrombin expression and activity in the local injured spinal cord.RNA sequencing transcriptomic analysis revealed that the differentially expressed genes in the argatroban-treated group were enriched in the JAK2/STAT3 pathway,which is involved in astrogliosis and glial scar formation.Western blotting and immunofluorescence results showed that argatroban downregulated the expression of the thrombin receptor PAR1 in the injured spinal cord and the JAK2/STAT3 signal pathway.Argatroban also inhibited the activation and proliferation of astrocytes and reduced glial scar formation in the spinal cord.Taken together,these findings suggest that argatroban may inhibit astrogliosis by inhibiting the thrombin-mediated PAR1/JAK2/STAT3 signal pathway,thereby promoting the recovery of neurological function after spinal cord injury.

Oleanolic acid inhibits colon cancer cell stemness and reverses chemoresistance by suppressing JAK2/STAT3 signaling pathway

Background:Oleanolic acid(OA),a pentacyclic triterpenoid exhibiting specific anti-cancer properties and highly effective antioxidant activity,was isolated from traditional Chinese medicinal herbs.Conversely,the OA that impacts colon cancer(CC)cells and its underlying mechanisms remain poorly understood.Methods:The cytotoxic effect of OA alone or OA-5-Fluorouracil(5-FU)combination on normal and CC cells was analyzed by methyl thiazolyl diphenyl-tetrazolium bromide(MTT).Then,the impact of OA on CC cell lines(LoVo and HT-29)proliferation and stemness were measured using colon formation and tumorsphere formation assays.Octamer-binding transcription factor 4(Oct4),Prominin-1(CD133),Nanog,and transcription factor SOX-2(SOX2)are cell stemness-related indicators whose expression was assessed usingfluorescence qPCR assay,Western blotting,and immunohistochemistry.The effect of OA on the proliferative potency of CC cells was evaluated using an in vivo model.Results:The stem-like characteristics and clone production of colon cancer cells were markedly reduced by OA alone or in combination with OA-5-FU.Moreover,OA increases the susceptibility of CC cells to 5-FU by blocking the cell stemness-related markers(CD133,Nanog,SOX2,and Oct4)expression levels both in vitro and in vivo,as well as by inactivating the activator of transcription 3(STAT3 signaling)and Janus kinase 2/signal transducer(JAK2).Conclusion:Thesefindings imply that oleanolic acid,both in vitro and in vivo,suppresses the JAK2/STAT3 pathway,which in turn reverses chemoresistance and decreases colon cancer cell stemness.Therefore,by reducing the recommended amount of 5-FU,this strategy may improve chemotherapeutic effectiveness and minimize undesired side effects.

Exploring the mechanism of electroacupuncture at different acupoints on acute colitis rats based on JAK2/STAT3/SOCS1 signaling pathway

Objective:To investigate the mechanism of JAK2/STAT3/SOCS1 signaling pathway in electroacupuncture of different acupoints on acute colitis rats.Methods:36 SPF SD rats were randomly divided into 6 groups,with 6 rats in each group.The rat model of acute colitis was prepared by enema with glacial acetic acid solution.After the model was established,electroacupuncture was given to each acupoint group,with density wave,frequency 2Hz-50 Hz,intensity 2 mA,muscle tremor as the degree 20 min/time,1 time/day,for 3 consecutive days.Observe the general condition of rats;the pathological changes of colonic mucosa in rats were observed by HE method.The contents of serum interleukin-4(IL-4)and interleukin-8(IL-8)were detected by ELISA.Western blot and RT-PCR were used to detect the expression of JAK2,STAT3,SOCS1 protein and mRNA in rat colon tissue.Results:In contrast to the normal group,the overall condition of the model group was worse,the colonic mucosa was severely damaged,even necrotic,and the ulcer surface was obvious.The content of IL-4 in serum was obviously reduced,and the content of IL-8 was obviously go up(P<0.01).The protein content of JAK2,STAT3 and the expression of JAK2,STAT3 mRNA in colon tissue of rats were obviously go up,while the protein content of SOCS1 and the expression of SOCS1 mRNA were obviously reduced(P<0.01).In contrast to the model group,the general condition of rats in each acupoint group was significantly improved,the damage and necrosis of colonic mucosa and ulcer surface were obviously alleviated,the content of IL-4 in serum was obviously go up,and the content of IL-8 was significantly decreased(P<0.01).The protein content of JAK2,STAT3 and the expression of JAK2,STAT3 mRNA in colon tissue of rats were obviously reduced,while the protein content of SOCS1 and the expression of SOCS1 mRNA were obviously go up(P<0.05,P<0.01).Comparison of different acupoint groups,the colonic mucosal injury in the Zusanli group was significantly reduced,the content of serum IL-4 was significantly increased,and the content of IL-8 was significantly decreased(P<0.05,P<0.01).The protein content and mRNA expression of JAK2 and STAT3 in colon tissue were significantly down-regulated,while the protein content and mRNA expression of SOCS1 were significantly go up(P<0.05,P<0.01).Conclusion:Electroacupuncture at each acupoint can improve the damage of colonic mucosa and reduce the inflammatory response.The therapeutic effect of Zusanli(ST36)is better than that of Tianshu(ST25),Dachangshu(BL25)and Shangjuxu(ST37).The mechanism may be related to the regulation of JAK2/STAT3/SOCS1 signaling pathway related proteins and inflammatory cytokines IL-4 and IL-8.

Metochalcone induces senescence-associated secretory phenotype via JAK2/STAT3 pathway in breast cancer

Breast and lung cancers are the leading causes of mortality and most frequently diagnosed cancers in women and men,respectively,worldwide.Although the antitumor activity of chalcones has been extensively studied,the molecular mechanisms of isoliquiritigenin analog 2',4',4-trihydroxychalcone(metochalcone;TEC)against carcinomas remain less well understood.In this study,we found that TEC inhibited cell proliferation of breast cancer BT549 cells and lung cancer A549 cells in a concentration-dependent manner.TEC induced cell cycle arrest in the S-phase,cell migration inhibition in vitro,and reduced tumor growth in vivo.Moreover,transcriptomic analysis revealed that TEC modulated the activity of the JAK2/STAT3 and P53 pathways.TEC triggered the senescence-associated secretory phenotype(SASP)by repressing the JAK2/STAT3 axis.The mechanism of metochalcone against breast cancer depended on the induction of SASP via deactivation of the JAK2/STAT3 pathway,highlighting the potential of chalcone in senescence-inducing therapy against carcinomas.

Mechanism of Yanghe Pingchaun granules on airway remodeling in asthmatic rats based on IL-6/JAK2/STAT3 signaling axis

Objective: To investigate the effects of Yanghe Pingchuan Granules on airway remodeling in asthmatic rats, and to explore the mechanism of Interleukin-6/Janus kinase 2/ Signal transducing activator of transcription 3(IL-6/JAK2/STAT3) signal axis. Methods: We separated 42 healthy male SD rats into two groups, a control group (7) and a model group (35).The model group was sensitized with a combination of ovalbumin (OVA) and aluminum hydroxide for 2 weeks, while the control group was given an equal amount of physiological saline.After 2 weeks, the modeling group was randomly divided into Model group, Yanghe Pingchuan Granules high, medium and low dose groups and Dexamethasone group, each group consisted of 7 animals. After 4 weeks, OVA atomization and gavage were used for stimulation and treatment. Yanghe Pingchuan Granules high, middle and low groups were given 15.48, 7.74, 3.87 g∙kg-1 Yanghe Pingchuan Granules daily, dexamethasone group was given 0.0625 mg∙kg-1 dexamethasone daily, and the other groups were given the same amount of normal saline. HE, PAS and Masson staining were used to observe the lung histopathological changes in rats. The levels of interleukin-6, IL-23 and IL-17A were detected by ELISA. The expression levels of JAK-2, P-JAK2, STAT3 and P-STAT3 in lung tissues were detected by Western blot. Real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the mRNA expression levels of IL-6, JAK2 and STAT3 in rat lung tissue. Results: The lung tissue structure of the model group was severely damaged compared to the control group, accompanied by a great many of inflammatory cell infiltration, goblet cell hyperplasia, subepithelial collagen fiber deposition and airway epithelial thickening were more obvious. The expressions of IL-6, IL- 23 and IL-17A in serum were significantly increased (P<0.01), the protein expression levels of JAK-2, P-JAK2, STAT3 and P-STAT3 and the mRNA expression levels of IL-6, JAK2 and STAT3 in lung tissue were significantly increased (P<0.01);Compared with the model group, inflammatory cell infiltration, goblet cell proliferation, subepithelial collagen fiber deposition and airway epithelial thickening were significantly reduced in each administration group, and the expressions of IL-6, IL-23 and IL-17A in serum were significantly decreased (P< 0.01). The protein expression levels of JAK-2, P-JAK2, STAT3 and P-STAT3 and mRNA expression levels of IL-6, JAK2 and STAT3 in lung tissue were significantly decreased (P<0.01). Conclusion: Yanghe Pingchuan Granules can significantly alleviate airway remodeling in asthmatic rats, and its mechanism may be through inhibiting the IL-6/JAK2/STAT3 signal axis.

Galectin 2 regulates JAK/STAT3 signaling activity to modulate oral squamous cell carcinoma proliferation and migration in vitro

Background:Galectin 2(LGALS2)is a protein previously reported to serve as a mediator of disease progression in a range of cancers.The function of LGALS2 in oral squamous cell carcinoma(OSCC),however,has yet to be explored,prompting the present study to address this literature gap.Methods:Overall,144 paired malignant tumor tissues and paracancerous OSCC patient samples were harvested and the LGALS2 expression levels were examined through qPCR and western immunoblotting.The LGALS2 coding sequence was introduced into the pcDNA3.0 vector,to enable the overexpression of this gene,while an LGALS2-specific shRNA and corresponding controls were also obtained.The functionality of LGALS2 as a regulator of the ability of OSCC cells to grow and undergo apoptotic death in vitro was assessed through EdU uptake and CCK-8 assays,and flow cytometer,whereas a Transwell system was used to assess migratory activity and invasivity.An agonist of the Janus Kinase 2(JAK2)/Signal Transducer and Activator of Transcription 3(STAT3)pathway was also used to assess the role of this pathway in the context of LGALS2 signaling.Results:Here,we found that lower LGALS2 protein and mRNA expression were evident in OSCC tumor tissue samples,and these expression levels were associated with clinicopathological characteristics and patient survival outcomes.Silencing LGALS2 enhanced proliferation in OSCC cells while rendering these cells better able to resist apoptosis.The opposite was instead observed after LGALS2 was overexpressed.Mechanistically,the ability of LGALS2 to suppress the progression of OSCC was related to its ability to activate the JAK/STAT3 signaling axis.Conclusion:Those results suggest a role for LGALS2 as a suppressor of OSCC progression through its ability to modulate JAK/STAT3 signaling,supporting the potential utility of LGALS2 as a target for efforts aimed at treating OSCC patients.

基于Hepcidin和JAK2/STAT3信号通路探讨通痹颗粒 对胶原诱导性关节炎大鼠的影响

目的研究通痹颗粒对胶原诱导性关节炎(collagen-induced arthritis,CIA)大鼠铁调素(hepcidin,Hepc)、Janus激酶(janus kinase,JAK)2/信号转导子和转录激活子(signal transduction and activator of transcription,STAT)3信号通路的影响。方法选取36只雌性SD大鼠随机分成空白组、模型组、阳性对照组和通痹颗粒低、中、高剂量组,每组6只。空白组不予处理,其余组用牛Ⅱ型胶原建立CIA模型。造模完成后,空白组、模型组予生理盐水灌胃,其余各组分别以巴瑞替尼片和低、中、高剂量通痹颗粒灌胃。每天1次,连续4周。HE染色行滑膜组织病理学观察;酶联免疫吸附法测定血清Hepc、白细胞介素6(interleukin 6,IL-6)水平;逆转录-聚合酶链反应法测定滑膜中JAK2、STAT3、细胞信号因子传导抑制体(suppressor of cytokine signaling,SOCS)1、SOCS3的mRNA相对表达量;Western blot法检测滑膜中JAK2、p-JAK2、STAT3、p-STAT3、SOCS1、SOCS3的蛋白表达量。结果模型组见滑膜上皮结构缺损,滑膜重度增生,排列紊乱,并有大量炎症细胞浸润和多个血管翳形成;各给药组滑膜炎症均有所减轻,阳性对照组优于通痹颗粒高剂量组,通痹颗粒中、高剂量组优于低剂量组。与模型组相比,各给药组关节炎指数评分、血清Hepc和IL-6水平均显著降低(P<0.01);与阳性对照组相比,通痹颗粒中、低剂量组关节炎指数评分、血清Hepc和IL-6水平均升高(P<0.05)。与模型组比较,阳性对照组和通痹颗粒低、中、高剂量组JAK2、STAT3 mRNA和蛋白以及p-JAK2、p-STAT3的蛋白表达量均降低(P<0.05),而通路抑制因子SOCS1、SOCS3 mRNA和蛋白的表达均升高(P<0.05);与阳性对照组比较,通痹颗粒各剂量组JAK2、STAT3 mRNA和蛋白以及p-JAK2、p-STAT3的蛋白表达量均升高(P<0.05),而SOCS1、SOCS3 mRNA和蛋白的表达均降低(P<0.05)。结论通痹颗粒能够改善CIA大鼠滑膜炎症,其机制可能与抑制JAK2/STAT3信号通路而减少Hepc的表达有关。

静态磁场通过激活FGFR1/JAK2/STAT3信号通路促进皮肤成纤维细胞的增殖活性和迁移表型

目的:通过体外实验探讨静态磁场(SMF)对皮肤成纤维细胞增殖活性和迁移表型的调控作用与潜在机制。方法:将人皮肤成纤维细胞(HSFs)分为5组,包括对照组、SMF组、SMF+阿魏酸组、SMF+芦可替尼组、SMF+Stattic组。对照组为正常培养的HSFs;SMF组的HSFs细胞暴露于1mT的SMF中。其余三组分别将FGFR1、JAK2、STAT3的抑制剂(3μmoL/L阿魏酸、3nmoL/L芦可替尼、20μmoL/L的Stattic)与HSFs一起预培养,并暴露于1mT的SMF中,所有组均处理24h。用细胞计数试剂盒-8(CCK-8)分析细胞的增殖活性。用ELISA试剂盒法检测人类B细胞淋巴瘤2相关X蛋白(BAX)的水平。用Western blot检测凋亡标志蛋白cleaved-caspase3及FGFR1、JAK2、STAT3、磷酸化的(p)-FGFR1、p-JAK2、p-STAT3的表达以及细胞迁移相关表型高迁移率组蛋白1(HMGB1)、波形蛋白(vimentin)、血管内皮生长因子A(VEGFA)、基质金属蛋白酶-9(MMP-9)、MMP-2的表达。结果:与对照组比,SMF组的细胞增殖活性增加,BAX的水平减少,cleaved-caspase3的蛋白表达水平下调,p-FGFR1、p-JAK2、p-STAT3、HMGB1、vimentin、VEGFA、MMP-9、MMP-2的表达水平均显著上调(均P<0.05)。与SMF组比,SMF+阿魏酸组的细胞增殖活性降低,BAX的水平增加,cleaved-caspase3的蛋白表达水平上调,而p-FGFR1、p-JAK2、p-STAT3、HMGB1、vimentin、VEGFA、MMP-9、MMP-2的表达均显著下调(均P<0.05)。与SMF组比,SMF+芦可替尼组的细胞增殖活性降低,BAX的水平增加,cleaved-caspase3的蛋白表达水平上调,JAK2、p-JAK2、p-STAT3、HMGB1、vimentin、VEGFA、MMP-9、MMP-2的表达水平均显著下调(均P<0.05)。与SMF组比,SMF+Stattic组的细胞增殖活性降低,BAX的水平增加,cleaved-caspase3的蛋白表达水平上调,STAT3、p-STAT3、HMGB1、vimentin、VEGFA、MMP-9、MMP-2的表达水平均显著下调(均P<0.05)。结论:SMF通过激活FGFR1/JAK2/STAT3信号通路促进皮肤成纤维细胞的增殖活性和迁移表型。

藏红花素通过抑制JAK2/STAT3信号通路减轻脑缺血再灌注大鼠海马神经元损伤

目的研究藏红花素(CRO)对脑缺血再灌注(CI/R)大鼠海马神经元损伤的影响并探索其潜在机制。方法将144只雄性SD大鼠按照随机数字表法分为假手术(Sham)组,模型(CI/R)组,CRO低、中、高剂量(CRO-L、CRO-M、CRO-H)组和尼莫地平(NMP)组6组,每组24只。采用线栓法制备CI/R大鼠模型,各组分别于造模前7 d开始1次/d腹腔注射(ip)给药(CRO-L、CRO-M、CRO-H组分别ip给药10、20、40 mg/kg,NMP组ip给药1 mg/kg,Sham组和CI/R组ip给予生理盐水5 mL/kg)。再灌注24 h后,通过Morris水迷宫实验检测大鼠学习记忆能力,TTC染色检测脑梗死率,HE染色法行海马CA1区和CA3区神经元病理学检查,TUNEL染色法行海马CA1区和CA3区神经元凋亡检查,ELISA法检测海马组织白细胞介素-1β(IL-1β)、IL-8、肿瘤坏死因子-α(TNF-α)含量,Western blot法检测海马组织Janus激酶2/信号转导与转录激活子3(JAK2/STAT3)信号通路相关蛋白相对表达量。结果与Sham组比较,CI/R组学习记忆能力明显降低,脑梗死率明显升高(P<0.05);海马CA1区和CA3区神经元呈现数量减少、间隙增大、空泡样变、核膜核仁边界模糊、炎性细胞浸润等病理改变,凋亡率明显升高(P<0.05);海马组织IL-1β、IL-8、TNF-α含量明显升高(P<0.05);p-JAK2、p-STAT3、高迁移率族蛋白B1(HMGB1)、Bcl-2相关X蛋白(Bax)、激活型半胱氨酸蛋白酶-3(cleaved Caspase-3)相对表达量和p-JAK2/JAK2、p-STAT3/STAT3、Bax/Bcl-2表达比值均明显升高,Bcl-2相对表达量明显降低(P<0.05)。与CI/R组比较,CRO-M组、CRO-H组和NMP组大鼠学习记忆能力显著改善、脑梗死率明显降低(P<0.05);海马CA1区和CA3区神经元病理学改变明显改善、凋亡率明显降低(P<0.05);海马组织IL-1β、IL-8、TNF-α含量明显降低(P<0.05);p-JAK2、p-STAT3、HMGB1、Bax、Cleaved Caspase-3相对表达量和p-JAK2/JAK2、p-STAT3/STAT3、Bax/Bcl-2表达比值均明显降低(P<0.05)。CRO上述作用呈现一定的剂量依赖性,且CRO-H组对CI/R大鼠学习记忆能力、海马CA1区和CA3区神经元病理学改变和凋亡率、炎症因子含量、JAK2/STAT3信号通路相关蛋白表达的影响显著优于NMP组(P<0.05)。结论CRO可能通过抑制JAK2/STAT3信号通路活化,减轻炎症和神经元凋亡,从而对CI/R大鼠海马神经元损伤起到保护作用。

基于JAK2/STAT3信号通路探讨黄芪-山茱萸对高糖诱导足细胞损伤的保护作用

目的:探究黄芪-山茱萸对高糖诱导肾足细胞损伤的保护作用及机制。方法:CCK8法检测黄芪-山茱萸(0、175、350、700、1400、2800 mg/L)对足细胞(MPC-5)活性的影响。30 mmol/L葡萄糖诱导MPC-5细胞损伤,350、700、1400 mg/L黄芪-山茱萸进行干预;ELISA测定MPC-5细胞IL-6、IL-1β水平,流式细胞术测定细胞凋亡,免疫荧光染色测定MPC-5肾病蛋白(nephrin)、足细胞素(podocin)表达,qRT-PCR测定MPC-5细胞nephrin、podocin、结蛋白(desmin)、Wilm瘤基因1(WT-1)基因表达,Western blot测定细胞Janus激酶2/信号转导和转录激活因子3(JAK2/STAT3)信号通路磷酸化表达。结果:2800 mg/L黄芪-山茱萸可显著抑制MPC-5细胞活性(P<0.01)。MPC-5细胞经高糖诱导后,IL-6、IL-1β水平升高,凋亡增加,nephrin、podocin、WT-1表达降低,desmin表达升高,JAK2/STAT3信号通路磷酸化表达增加(P<0.01)。350、700、1400 mg/L黄芪-山茱萸可抑制高糖诱导MPC-5细胞IL-6、IL-1β水平升高和凋亡,增强nephrin、podocin、WT-1表达,降低desmin表达,抑制JAK2/STAT3信号通路磷酸化表达(P<0.05)。结论:黄芪-山茱萸可减轻高糖诱导足细胞炎症和凋亡损伤,其机制与抑制JAK2/STAT3通路磷酸化有关。

从肝论治取穴针灸通过调控JAK2/STAT3途径对膝骨关节炎大鼠关节软骨的保护作用

目的:探究从肝论治取穴针灸通过调控JAK2/STAT3途径对膝骨关节炎(OA)大鼠关节软骨的保护作用。方法:60只SD大鼠随机分为对照组、OA组、OA+针灸组,每组20只,通过手术构建右膝OA模型。OA+针灸组大鼠接受从肝论治取穴针灸4周。检测软骨退化情况、关节炎症,以及JAK2/STAT3转录和蛋白水平。结果:OA组大鼠OARSI评分高于对照组(P<0.05);干预后,OA+针灸组大鼠OARSI评分低于OA组(P<0.05)。OA组大鼠透明软骨(HC)厚度低于对照组,钙化软骨(CC)厚度高于对照组(P<0.05);干预后,OA+针灸组大鼠HC厚度高于OA组,CC厚度低于OA组(P<0.05)。OA组大鼠白细胞介素-1β(IL-1β)、IL-6、JAK2 mRNA、STAT3 mRNA、JAK2蛋白、STAT3蛋白水平均高于对照组(P<0.05);干预后,OA+针灸组大鼠IL-1β、IL-6、JAK2 mRNA、STAT3 mRNA、JAK2蛋白、STAT3蛋白水平均低于OA组(P<0.05)。结论:从肝论治取穴针灸可通过调控JAK2/STAT3途径发挥对OA大鼠膝关节软骨的保护作用。

丹参酮ⅡA通过抑制JAK2/STAT3通路减轻慢性萎缩性胃炎大鼠胃黏膜损伤的机制研究

[目的]研究丹参酮ⅡA(TanⅡA)对慢性萎缩性胃炎(CAG)大鼠胃黏膜的影响,并基于Janus激酶2(JAK2)/信号转导与转录激活子3(STAT3)通路探讨其机制。[方法]将80只Wistar大鼠随机分为5组(n=16):正常(Normal)组、模型(Model)组、TanⅡA(5 mg/kg)组、TanⅡA(5 mg/kg)+AG490(JAK2抑制剂,5 mg/kg)组和Tan IIA(5 mg/kg)+C-A1(JAK2激动剂,50 mg/kg)组。除Normal组外,其他4组均采用N-甲基-N'-硝基-N-亚硝基胍溶液(MNNG,0.04 g/mL)自由饮联合雷尼替丁灌胃、饥饱失常饮食的综合法构建CAG大鼠模型。各组分别每日1次连续给药治疗12周后,中性红清除法检测胃黏膜血流量,酶联免疫吸附法(ELISA)法检测血清胃泌素(GAS)、血浆胃动素(MTL)及胃黏膜炎症因子水平,苏木精-伊红(HE)染色法观察胃黏膜组织病理学改变并行萎缩评分,TUNEL染色法观察胃黏膜细胞凋亡状况并计算凋亡指数,逆转录聚合酶链反应(RT-PCR)法、蛋白免疫印迹(Western Blot)法检测胃黏膜JAK2/STAT3通路相关mRNA和蛋白表达。[结果]与Model组相比,TanⅡA组大鼠胃黏膜血流量、血清GAS水平、血浆MTL水平均明显升高(P<0.05);胃黏膜中肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)、IL-6等炎症因子水平明显降低(P<0.05);胃黏膜病理学改变明显改善,萎缩评分明显降低(P<0.05);胃黏膜凋亡细胞数量明显减少,凋亡指数明显降低(P<0.05);胃黏膜JAK2、STAT3 mRNA表达量明显降低(P<0.05);B淋巴细胞瘤2(Bcl-2)蛋白表达量明显升高,Bcl-2相关X蛋白(Bax)、激活型半胱氨酸蛋白酶3(C-Cas-3)、C-Cas-9蛋白表达量及p-JAK2/JAK2、p-STAT3/STAT3、胞核核因子-κB(NF-κB)p65/胞浆NF-κB p65表达比值明显降低(P<0.05)。AG490能够明显增强TanⅡA对CAG大鼠各检测指标的调控作用,C-A1则能够明显逆转TanⅡA对CAG大鼠各检测指标的调控作用(P<0.05)。[结论] TanⅡA可能通过抑制JAK2/STAT3通路活化及NF-κB核转位,减轻炎症反应和细胞凋亡,从而减轻CAG大鼠胃黏膜结构和功能损伤。

缺氧诱导因子-1α通过miR-126/JAK2-STAT3轴调控慢性肾病小鼠肾纤维化

目的:探究缺氧诱导因子-1α(HIF-1α)通过miR-126/JAK2-STAT3轴促进慢性肾病小鼠肾纤维化的分子机制。方法:选取8周龄雄性C57/BL6小鼠,分为假手术组、模型组、HIF-1αmimic组、miR-126 mimic组、HIF-1α+miR mimic组、WP1066组、miR-126 mimic+WP1066组;用全自动生化仪检测小鼠肾24 h尿蛋白、血尿素氮(BUN)、血肌酐(SCr)水平,qRT-PCR检测肾组织内miR-126相对表达量,免疫印迹检测肾组织内HIF-α,肾纤维化关键蛋白fibronectin、collagen-Ⅰ、α-SMA,JAK2/STAT3信号通路关键蛋白表达;Masson染色分析小鼠肾病理形态及间质纤维化程度。结果:模型组模型建立后miR-126表达受到抑制,差异有统计学意义。与假手术组相比,模型组小鼠24 h尿蛋白、BUN、SCr水平均上调,肾组织肾小管、间质结构紊乱,肾纤维化明显,fibronectin、collagen-Ⅰ、α-SMA、p-JAK2、p-STAT3表达上调;HIF-1α过表达进一步上调24 h尿蛋白、BUN、SCr、fibronectin、collagen-Ⅰ、α-SMA、p-JAK2、p-STAT3表达;miR-126过表达下调24 h尿蛋白、BUN、SCr、fibronectin、collagen-Ⅰ、α-SMA、p-JAK2、p-STAT3表达;HIF-1α过表达可以逆转miR-126过表达抑制肾纤维化的趋势,差异有统计学意义。与模型组相比,JAK2/STAT3信号通路被阻断后,p-JAK2、p-STAT3、fibronectin、collagen-Ⅰ、α-SMA表达明显下调,miR-126的过表达可以进一步下调小鼠肾组织内p-JAK2、p-STAT3、fibronectin、collagen-Ⅰ、α-SMA表达,差异有统计学意义。结论:小鼠肾纤维化促进肾组织内HIF-1α高表达,HIF-1α通过抑制miR-126提升JAK2-STAT3信号通路活性,最终促进肾纤维化进展。

胃动蛋白2调控JAK2/STAT3通路在胃癌迁移和侵袭中的作用

目的 阐明胃动蛋白2(gastrokine 2,GKN2)在人胃癌中的作用及相关分子机制。方法 采用免疫组织化学技术检测90例胃癌(gastric cancer,GC)组织、48例癌旁胃组织(adjacent tissue,PT)和22例远端胃黏膜组织(distal gastric mucosa,DGM)中GKN2和TFF1的表达;构建GKN2基因高表达载体,转染人胃癌细胞MKN28和SGC7901,qRT-PCR和Western blot验证转染效率,实验分为3组:GKN2组、NC组、空白组。采用CCK-8、Transwell迁移和侵袭实验测定细胞增殖、迁移和侵袭能力;Western blot观察GKN2转染后JAK2、STAT3、p-JAK2、p-STAT3蛋白表达变化。结果 与癌旁胃组织(GKN2,43.75%;TFF1,62.50%)和远端胃黏膜组织(GKN2,86.36%;TFF1,81.82%)相比,胃癌组织中GKN2(6.67%)、TFF1(21.11%)表达下调(P<0.05);但胃癌组织中GKN2与TFF1的表达无相关性(r≈0.23,P=0.074)。与NC组(0.25±0.03)和空白组(0.24±0.03)相比,GKN2转染MKN28细胞24 h后OD570下降至(0.15±0.02),GKN2转染48 h后的OD570(0.22±0.06)也低于相应的NC组(0.49±0.06)和空白组(0.44±0.02),72 h后的OD570(0.25±0.05)比相应的NC组(0.65±0.21)和空白组(0.63±0.03)减少(P<0.05);SGC7901细胞中GKN2转染24 h后OD570(0.721±0.014)低于NC组(1.352±0.168)和空白组(1.381±0.168),48 h后的OD570(0.674±0.028)也低于相应的NC组(1.459±0.211)和空白组(1.565±0.351),72 h后GKN2的OD570(0.400±0.028)比NC组(1.745±0.194)和空白组(1.792±0.385)减少(P<0.05)。Transwell迁移实验发现,MKN28细胞中GKN2转染组穿过膜的癌细胞数(26±5.01)明显低于NC组(109±7.10)和空白组(110±4.04)(P<0.05);与NC组(94±6.00)和空白组(75+7.05)相比,SGC7901中GKN2高表达显著降低了穿过膜的细胞数(37±3.11)(P<0.05)。Transwell侵袭实验证实,与NC组(67±5.02)和空白组(66±5.16)相比,GKN2高表达显著降低了穿过基质胶的MKN28细胞数(28±4.10)(P<0.05);SGC7901细胞中GKN2转染组穿过基质胶的癌细胞数(10±2.06)远低于NC组(58±5.13)和空白组(55±3.17)(P<0.05)。Western blot结果显示,GKN2高表达显著下调MKN28和SGC7901细胞中总JAK2、STAT3蛋白表达以及磷酸化形式p-JAK2和p-STAT3的表达水平(P<0.05)。结论 GKN2高表达可通过阻断JAK2/STAT3通路抑制胃癌细胞迁移和侵袭。

Elevated retinol binding protein 4 levels are associated with atherosclerosis in diabetic rats via JAK2/STAT3 signaling pathway

BACKGROUND Atherosclerosis is a major cause of mortality worldwide and is driven by multiple risk factors,including diabetes,which results in an increased atherosclerotic burden,but the precise mechanisms for the occurrence and development of diabetic atheroscerosis have not been fully elucidated.AIM To summarize the potential role of retinol binding protein 4(RBP4) in the pathogenesis of diabetic atheroscerosis,particularly in relation to the RBP4-Janus kinase 2/signal transducer and activator of transcription 3(JAK2/STAT3)signaling pathway.METHODS Male Wistar rats were randomly divided into three groups,including a control group(NC group),diabetic rat group(DM group),and diabetic atherosclerotic rat group(DA group).The contents of total cholesterol(TC), high-density lipoprotein cholesterol(HDL-c), triglycerides(TG), low-density lipoprotein cholesterol(LDLc), fasting insulin(FINS),fasting plasma glucose,and hemoglobin A1 c(HbA1 c)were measured.Moreover,the adipose and serum levels of RBP4,along with the expression levels of JAK2, phosphorylated JAK2(p-JAK2), STAT3,phosphorylated STAT3(p-STAT3), B-cell lymphoma-2(Bcl-2), and Cyclin D1 in aortic tissues were also measured.Besides,homeostasis model assessment of insulin resistance(HOMA-IR) and atherogenic indexes(AI) were calculated.RESULTS Compared with the NC and DM groups,the levels LDL-c,TG,TC,FINS,HOMAIR,RBP4,and AI were upregulated,whereas that of HDL-c was downregulated in the DA group(P <0.05);the mRNA levels of JAK2,STAT3,Cyclin D1,and Bcl-2 in the DA group were significantly increased compared with the NC group and the DM group;P-JAK2,p-JAK2/JAK2 ratio,p-STAT3,p-STAT3/STAT3 ratio,Cyclin D1,and Bcl-2 at protein levels were significantly upregulated in the DA group compared with the NC group and DM group.In addition,as shown by Pearson analysis,serum RBP4 had a positive correlation with TG,TC,LDL-c,FINS,HbA1 C,p-JAK2,p-STAT3,Bcl-2,Cyclin D1,AI,and HOMA-IR but a negative correlation with HDL-c.In addition,multivariable logistic regression analysis showed that serum RBP4,p-JAK2,p-STAT3,and LDL-c were predictors of the presence of diabetic atherosclerosis.CONCLUSION RBP4 could be involved in the initiation or progression of diabetic atherosclerosis by regulating the JAK2/STAT3 signaling pathway.

基于IL-6/JAK2/STAT3信号轴研究阳和平喘颗粒调控哮喘大鼠气道重塑作用机制

目的:研究阳和平喘颗粒对哮喘大鼠气道重塑及白细胞介素-6(IL-6)/Janus蛋白酪氨酸激酶2(JAK2)/信号转导和转录活化因子3(STAT3)信号轴在其中的作用机制。方法:选取健康雄性SD大鼠42只,随机数字法分为正常对照组7只与造模组35只,造模组采用卵清蛋白(OVA)联合氢氧化铝腹腔注射2周的方式进行致敏,正常对照组采用等量的生理盐水;2周后将造模组随机分为模型组、阳和平喘高、中、低剂量组和地塞米松组,每组7只;后4周采用OVA雾化+灌胃的方式进行激发和治疗,阳和平喘高中低组每日分别予以15.48、7.74、3.87 g/kg阳和平喘颗粒灌胃,地塞米松组予以0.0625 mg/kg地塞米松进行灌胃,其余组灌胃等量生理盐水。HE、PAS、Masson染色观察大鼠肺组织病理学变化;ELISA检测大鼠血清中IL-6、IL-23、IL-17A水平;Western blot检测肺组织中JAK-2、P-JAK2、STAT3、P-STAT3蛋白表达量;qRT-PCR检测大鼠肺组织中IL-6、JAK2、STAT3的mRNA水平。结果:与正常对照组比较,模型组大鼠肺组织有大量炎性细胞浸润,杯状细胞增生、上皮下胶原纤维沉积、气道上皮增厚较为明显;血清中IL-6、IL-23、IL-17A水平显著升高(P<0.01),肺组织JAK-2、P-JAK2、STAT3、P-STAT3的蛋白表达量和IL-6、JAK2、STAT3的mRNA表达水平显著升高(P<0.01);与模型组比较,各给药组炎性细胞浸润、杯状细胞增生、上皮下胶原纤维沉积、气道上皮增厚程度明显减轻,血清中IL-6、IL-23、IL-17A水平显著降低(P<0.01),肺组织JAK-2、P-JAK2、STAT3、P-STAT3的蛋白表达量和IL-6、JAK2、STAT3的mRNA水平显著降低(P<0.01)。结论:阳和平喘颗粒可明显缓解哮喘大鼠气道重塑,其机制可能是通过抑制IL-6/JAK2/STAT3信号轴发挥作用。

Triptolide Inhibits Expression of Inflammatory Cytokines and Proliferation of Fibroblast-like Synoviocytes Induced by IL-6/sIL-6R-Mediated JAK2/STAT3 Signaling Pathway

Triptolide,a component of the Chinese herb Tripterygium wilfordii Hook F,has been proved to be effective in the treatment of rheumatoid arthritis(RA).However,its underlying mechanisms on RA have not yet been well established.We observed the inhibitory effect of triptolide on the expression of inflammatory cytokines and proliferation of fibroblast-like synoviocytes(FLS)induced by the complex of interleukin-6(IL-6)and the soluble form of the IL-6 receptor(sIL-6R).Furthermore,to clarify the underlying mechanisms,we treated FLS with the Janus-activated kinase 2(JAK2)inhibitor/signal transducer and activator of transcription 3(STAT3)activation blocker AZD1480.In this study,immunohistochemical staining was used to identify vimentin(+)and CD68(−)in FLS.The FLS proliferation was measured by cell proliferation assay,and the cell cycles were analyzed by flow cytometry.Furthermore,ELISA was used to detect the expression of the inflammatory factors in culture solution.The expression levels of p-JAK2,JAK2,p-STAT3 and STAT3 were investigated through Western blotting analysis.The results showed that IL-6/sIL-6R significantly increased the cell proliferation and expression of inflammatory cytokines,including IL-6,interleukin-1β(IL-1β)and vascular endothelial growth factor(VEGF).Triptolide or AZD1480 inhibited the cell proliferation and inflammatory cytokine expression in IL-6/sIL-6R-stimulated FLS by suppressing JAK2/STAT3.The study suggested that the physiological effects of triptolide on RA were due to its contribution to the inhibition of the inflammatory cytokine expression and FLS proliferation by suppressing the JAK2/STAT3 signaling pathway.It may provide an innovative insight into the effect of triptolide in preventing RA pathogenesis.

Germacrone Induces Apoptosis in Human Hepatoma HepG2 Cells through Inhibition of the JAK2/STAT3 Signalling Pathway

Summary： Previous studies have shown that STAT3 plays a vital role in the genesis and progression of cancer. In this study, we investigated the relationship between the JAK2/STAT3 signalling pathway and germacrone-induced apoptosis in HepG2 cells. HepG2 cells were incubated with germacrone for 24 h, the protein expression of p-STAT3, STAT3, p-JAK2 and JAK2 was detected by Westem Blotting, and RT-PCR was used to determine the expression of STAT3, p53, Bcl-2 and Bax at transcriptional levels. Besides that, HepG2 cells were pre-treated with AG490 or IL-6 for 2 h, and then incubated with ger- macrone for 24 h. The expression ofp-JAK2, JAK2, p-STAT3, STAT3, p53, Bax and Bcl-2 was detected by Western blotting. The activity of HepG2 cells was tested by MTT assay. The apoptosis of HepG2 cells and levels of reactive oxygen species （ROS） were flow cytometrically measured. The results showed that germacrone exposure decreased p-STAT3 and p-JAK2 and regulated expression of p53 and Bcl-2 family members at the same time. Moreover, IL-6 enhanced the activation of the JAK2/STAT3 signalling pathway and therefore attenuated the germacrone-induced apoptosis. Suppression of JAK2/STAT3 signalling pathway by AG490, an inhibitor of JAK2, resulted in apoptosis and an increase in ROS in response to germacrone exposure. We therefore conclude that germacrone induces apoptosis through the JAK2/STAT3 signalling pathway.

Pyrrolidine dithiocarbamate alleviates the anti-tuberculosis drug-induced liver injury through JAK2/STAT3 signaling pathway:An experimental study

Objective:To study the effect of pyrrolidine dithiocarbamate(PDTC) on the anti-tuberculosis drug-induced liver injury and the molecular mechanism. Methods:Clean male SD rats were selected as experimental animals and randomly divided into normal group,model group,PDTC group and AG490 group. Animal model of anti-tuberculosis drug-induced liver injury was established by intragastric administration isoniazid + rifampicin. PDTC group received intraperitoneal injection of PDTC,and AG490 group received intraperitoneal injection of AG490. Twenty-eight days after intervention,the rats were executed,and the liver injury indexes,inflammation indexes and oxidative stress indexes in serum as well as JAK2/STAT3 expression,liver injury indexes,inflammation indexes and oxidative stress indexes in liver tissue were determined. Results:p-JAK2,p-STAT3,TNF-α,IL-1β,IL-6,ROS,8-OHdG and MDA expression in liver tissue as well as TBIL,ALT,AST,γ-GT,TNF-α,IL-1β,IL-6,ROS,8-OHdG and MDA levels in serum of model group were significantly higher than those of normal group while p-JAK2,p-STAT3,TNF-α,IL-1β,IL-6,ROS,8-OHdG and MDA expression in liver tissu as well as TBIL,ALT,AST,γ-GT,TNF-α,IL-1β,IL-6,ROS,8-OHdG and MDA levels in serum of PDTC group and AG490 group were significantly lower than those of model group. Conclusions:PDTC can inhibit the inflammation and oxidative stress mediated by JAK2/STAT3 signaling pathway to alleviate the anti-tuberculosis drug-induced liver injury.

IL-6/JAK2/STAT3通路中葛花解酲汤对脾虚湿热型溃疡性结肠炎“炎-癌转化”的预防作用

目的:探讨葛花解酲汤对脾虚湿热型溃疡性结肠炎“炎-癌转化”(UC-UCAC)小鼠结肠组织IL-6/JAK2/STAT3信号通路的影响。方法:80只SPF级C57BL/6雄性小鼠随机选出10只作为空白组,其余70只为造模组。造模组在建立脾虚湿热模型后随机分为模型组(第1、2、3周期)、葛花解酲汤高、中、低剂量组、美沙拉嗪组,10只/组,以氧化偶氮甲烷(AOM)/葡聚糖硫酸钠(DSS)继续建立UC-UCAC转化模型。各组给予相应药物治疗4周。观察小鼠一般状态;统计小鼠疾病活动指数(DAI)评分;HE染色观察小鼠结肠黏膜组织病理;Western blot、IHC和RT-q PCR检测小鼠结肠组织EGFR、IL-6、JAK2、STAT3、 p-STAT3蛋白和基因表达。结果:与空白组相比,模型组(第3周期)小鼠一般状态较差,结肠黏膜组织出现癌变,DAI评分、各目标蛋白及基因表达显著升高(P<0.01);与模型组(第3周期)相比,各治疗组小鼠一般状态有所恢复,结肠组织病理不同程度改善,除葛花解酲汤低剂量组外,其他各治疗组各目标蛋白及基因表达显著下降(P<0.01)。结论:葛花解酲汤可能通过抑制IL-6/JAK2/STAT3信号通路激活破坏肿瘤炎症微环境,修复受损结肠黏膜组织,延缓UC-UCAC进程,预防UCAC。