Role of Toll-like receptor 4 and Janus kinase and signal transducer and activator of transcription signal transduction pathway in sepsis-induced brain damage

The Janus kinase and signal transducer and activator of transcription （JAK/STAT） signal transduction pathway is involved in sepsis-induced functional damage to the heart, liver, kidney, and other organs. However, the cellular and molecular mechanisms underlying sepsis-induced brain damage remain elusive. In the present study, we found severe loss of neurons in the hippocampal CA1 region in rats with sepsis-induced brain damage following intraperitoneal injection of endotoxin, The expression of toll-like receptor 4, tumor necrosis factor a, and interleukin-6 was significantly increased in brain tissues following lipopolysaccharide exposure. AG490 （JAK2 antagonist） and rapamycin （STAT3 antagonist） significantly reduced neuronal loss and suppressed the increased expression of toll-like receptor 4, tumor necrosis factor a, and interleukin-6 in the hippocampal CA1 region in sepsis-induced brain damaged rats. Overall, these data suggest that blockade of the JAK/STAT signal transduction pathway is neuroprotective in sepsis-induced brain damage via the inhibition of toll-like receptor 4, tumor necrosis factor a, and interleukin-6 exoression.

Signal transducer and activator of transcription 3 promotes the Warburg effect possibly by inducing pyruvate kinase M2 phosphorylation in liver precancerous lesions

BACKGROUND Study shows that signal transducer and activator of transcription 3(STAT3) can increase the Warburg effect by stimulating hexokinase 2 in breast cancer and upregulate lactate dehydrogenase A and pyruvate dehydrogenase kinase 1 in myeloma. STAT3 and pyruvate kinase M2(PKM2) can also be activated and enhance the Warburg effect in hepatocellular carcinoma. Precancerous lesions are critical to human and rodent hepatocarcinogenesis. However, the underlying molecular mechanism for the development of liver precancerous lesions remains unknown. We hypothesized that STAT3 promotes the Warburg effect possibly by upregulating p-PKM2 in liver precancerous lesions in rats.AIM To investigate the mechanism of the Warburg effect in liver precancerous lesions in rats.METHODS A model of liver precancerous lesions was established by a modified Solt-Farber method. The liver pathological changes were observed by HE staining and immunohistochemistry. The transformation of WB-F344 cells induced with Nmethyl-N'-nitro-N-nitrosoguanidine and hydrogen peroxide was evaluated by the soft agar assay and aneuploidy. The levels of glucose and lactate in the tissue and culture medium were detected with a spectrophotometer. The protein levels of glutathione S-transferase-π, proliferating cell nuclear antigen(PCNA), STAT3,and PKM2 were examined by Western blot and immunofluorescence.RESULTS We found that the Warburg effect was increased in liver precancerous lesions in rats. PKM2 and p-STAT3 were upregulated in activated oval cells in liverprecancerous lesions in rats. The Warburg effect, p-PKM2, and p-STAT3 expression were also increased in transformed WB-F344 cells. STAT3 activation promoted the clonal formation rate, aneuploidy, alpha-fetoprotein expression,PCNA expression, G1/S phase transition, the Warburg effect, PKM2 phosphorylation, and nuclear translocation in transformed WB-F344 cells.Moreover, the Warburg effect was inhibited by stattic, a specific inhibitor of STAT3, and further reduced in transformed WB-F344 cells after the intervention for PKM2.CONCLUSION The Warburg effect is initiated in liver precancerous lesions in rats. STAT3 activation promotes the Warburg effect by enhancing the phosphorylation of PKM2 in transformed WB-F344 cells.

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.

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.

Gossypol acetic acid regulates leukemia stem cells by degrading LRPPRC via inhibiting IL-6/JAK1/STAT3 signaling or resulting mitochondrial dysfunction

BACKGROUND Leukemia stem cells(LSCs)are found to be one of the main factors contributing to poor therapeutic effects in acute myeloid leukemia(AML),as they are protected by the bone marrow microenvironment(BMM)against conventional therapies.Gossypol acetic acid(GAA),which is extracted from the seeds of cotton plants,exerts anti-tumor roles in several types of cancer and has been reported to induce apoptosis of LSCs by inhibiting Bcl2.AIM To investigate the exact roles of GAA in regulating LSCs under different microenvironments and the exact mechanism.METHODS In this study,LSCs were magnetically sorted from AML cell lines and the CD34+CD38-population was obtained.The expression of leucine-rich pentatricopeptide repeat-containing protein(LRPPRC)and forkhead box M1(FOXM1)was evaluated in LSCs,and the effects of GAA on malignancies and mitochondrial RESULTS LRPPRC was found to be upregulated,and GAA inhibited cell proliferation by degrading LRPPRC.GAA induced LRPPRC degradation and inhibited the activation of interleukin 6(IL-6)/janus kinase(JAK)1/signal transducer and activator of transcription(STAT)3 signaling,enhancing chemosensitivity in LSCs against conventional chemotherapies,including L-Asparaginase,Dexamethasone,and cytarabine.GAA was also found to downregulate FOXM1 indirectly by regulating LRPPRC.Furthermore,GAA induced reactive oxygen species accumulation,disturbed mitochondrial homeostasis,and caused mitochondrial dysfunction.By inhibiting IL-6/JAK1/STAT3 signaling via degrading LRPPRC,GAA resulted in the elimination of LSCs.Meanwhile,GAA induced oxidative stress and subsequent cell damage by causing mitochondrial damage.CONCLUSION Taken together,the results indicate that GAA might overcome the BMM protective effect and be considered as a novel and effective combination therapy for AML.

Caprylic Acid Improves Lipid Metabolism, Suppresses the Inflammatory Response and Activates the ABCA1/p-JAK2/pSTAT3 Signaling Pathway in C57BL/6J Mice and RAW264.7 Cells

Objective This study aimed to investigate the effects of caprylic acid(C8:0)on lipid metabolism and inflammation,and examine the mechanisms underlying these effects in mice and cells.Methods Fifty-six 6-week-old male C57BL/6J mice were randomly allocated to four groups fed a highfat diet(HFD)without or with 2%C8:0,palmitic acid(C16:0)or eicosapentaenoic acid(EPA).RAW246.7 cells were randomly divided into five groups:normal,lipopolysaccharide(LPS),LPS+C8:0,LPS+EPA and LPS+cAMP.The serum lipid profiles,inflammatory biomolecules,and ABCA1 and JAK2/STAT3 mRNA and protein expression were measured.Results C8:0 decreased TC and LDL-C,and increased the HDL-C/LDL-C ratio after injection of LPS.Without LPS,it decreased TC in mice(P<0.05).Moreover,C8:0 decreased the inflammatory response after LPS treatment in both mice and cells(P<0.05).Mechanistic investigations in C57BL/6J mouse aortas after injection of LPS indicated that C8:0 resulted in higher ABCA1 and JAK2/STAT3 expression than that with HFD,C16:0 and EPA,and resulted in lower TNF-α,NF-κB mRNA expression than that with HFD(P<0.05).In RAW 264.7 cells,C8:0 resulted in lower expression of pNF-κBP65 than that in the LPS group,and higher protein expression of ABCA1,p-JAK2 and p-STAT3 than that in the LPS and LPS+cAMP groups(P<0.05).Conclusion Our studies demonstrated that C8:0 may play an important role in lipid metabolism and the inflammatory response,and the mechanism may be associated with ABCA1 and the p-JAK2/p-STAT3 signaling pathway.

白藜芦醇通过阻断JAK激酶2/信号转导及转录活化因子3信号通路抑制食管癌细胞增殖和迁移并诱导其凋亡的作用研究

目的探究白藜芦醇对人食管癌细胞增殖、凋亡、迁移及JAK激酶2/信号转导及转录活化因子3(JAK2/STAT3)信号通路的调控作用。方法2021年7月至2022年7月,体外培养人食管癌OE19细胞,将细胞分为对照组(不做干预)和实验组(15、30、60、90和120μmol/L白藜芦醇干预24 h)进行预实验,根据细胞计数试剂盒(CCK-8)预实验结果,筛选有显著作用且细胞活力高于50%的30、60、90μmol/L白藜芦醇进行后续的实验,后续实验又分为对照组(不做干预)、30、60、90μmol/L白藜芦醇组(30、60和90μmol/L白藜芦醇)和30μmol/L白藜芦醇+抑制剂组(30μmol/L白藜芦醇+10μmol/L JAK2/STAT3通路抑制剂AG490),干预24 h。用5-乙炔基-2'脱氧尿嘧啶核苷(EdU)、Hoechst 33258染色、Transwell、实时荧光定量PCR(RT-qPCR)及蛋白质印迹法对细胞增殖率、凋亡情况、迁移数及细胞周期蛋白D1(cyclin D1)、胱天蛋白酶-3(caspase-3)和JAK2/STAT3相关因子表达水平进行分析。结果不同浓度实验组的细胞活力与对照组相比逐渐降低,其中30、60、90和120μmol/L白藜芦醇差异有统计学意义(P<0.05),但120μmol/L白藜芦醇组细胞活力低于50%,所以本研究选择30、60和90μmol/L白藜芦醇继续后续实验;60μmol/L白藜芦醇组细胞增殖率(41.49±5.06)%、迁移细胞数(36.67±2.52)个显著低于对照组(53.34±1.99)%、(58.00±2.00)个,凋亡细胞数(7.67±1.53)个显著高于对照组(2.50±0.71)个,且cyclin D1、p-JAK2和p-STAT3表达量也低于对照组,caspase-3 mRNA和蛋白表达水平高于对照组(P<0.05);30、90μmol/L白藜芦醇组以上各指标与对照组比较同样如此。与30μmol/L白藜芦醇组相比,30μmol/L白藜芦醇+抑制剂组以上各指标变化更显著(P<0.05)。结论白藜芦醇可通过抑制JAK2/STAT3通路抑制人食管癌OE19细胞的增殖和迁移并诱导凋亡。

基于JAK2/STAT3信号通路探究利咽糖浆对慢性咽炎大鼠咽喉组织损伤及咽黏膜修复的影响

目的:基于Janus激酶2(JAK2)/信号转导和转录激活因子3(STAT3)信号通路探究利咽糖浆对慢性咽炎大鼠咽喉组织损伤及咽黏膜修复的影响。方法:取SD大鼠随机分为对照组、模型组、利咽糖浆组、RO8191(JAK2/STAT3激活剂)组、利咽糖浆+RO8191组,模型组与药物干预组大鼠采用氨水刺激咽部构建慢性咽炎模型,对照组大鼠咽部注射等剂量生理盐水,经利咽糖浆与RO8191干预后,检测大鼠一般情况及咽部表观状态,并进行咽部表观状态评分;HE染色检测大鼠咽部病理形态变化;流式细胞术检测大鼠外周血T淋巴细胞亚群CD4^(+)T与CD8^(+)T表达、CD4^(+)T/CD8^(+)T;试剂盒检测大鼠血清肿瘤坏死因子-α(TNF-α)、IL-6、IL-10、丙二醛(MDA)、活性氧(ROS)、总抗氧化能力(T-AOC)水平;以免疫印记法检测大鼠咽部组织JAK2/STAT3通路相关蛋白表达。结果:与对照组比较,模型组大鼠咽部组织出现明显病理形态损伤,外周血CD4^(+)T与CD4^(+)T/CD8^(+)T、IL-10、血清T-AOC水平降低(P<0.05),咽部表观状态评分、CD8^(+)T、血清TNF-α、IL-6、MDA与ROS水平、咽部组织p-JAK2/JAK2与p-STAT3/STAT3水平升高(P<0.05);与模型组、利咽糖浆+RO8191组相比,利咽糖浆组大鼠咽部组织病理形态损伤减轻,外周血CD4^(+)T表达与CD4^(+)T/CD8^(+)T、IL-10、血清T-AOC水平均升高(P<0.05),咽部表观状态评分、CD8^(+)T、血清TNF-α、IL-6、MDA与ROS水平、咽部组织p-JAK2/JAK2与p-STAT3/STAT3水平均降低(P<0.05);RO8191组大鼠各指标变化趋势与利咽糖浆组相反。结论:利咽糖浆可通过抑制JAK2/STAT3信号激活减轻炎症及氧化应激,增强抗炎及抗氧化能力,缓解慢性咽炎大鼠咽喉组织损伤,修复咽黏膜。

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。

冬凌草甲素调节JAK2/STAT3/SOCS-1信号通路对糖耐量异常大鼠胰岛素抵抗的影响

目的探讨冬凌草甲素(Oridonin,Ori)对糖耐量异常(impaired glucose tolerance,IGT)大鼠胰岛素抵抗(insulin resistance,IR)的影响及作用机制。方法采用高脂饮食喂养联合链脲佐菌素注射法构建IGT大鼠IR模型,大鼠分为正常组(CT组)、IGT模型组(IGT组)、Ori组(10 mg·kg^(-1)·d^(-1))、Ori+Colivelin(COL)组(10 mg·kg^(-1)·d^(-1)Ori+2 mg/kg COL),每组6只。血糖检测仪测定空腹血糖(FPG)、葡萄糖耐量试验(OGTT)2 h血糖(2 hPG),ELISA试剂盒测定空腹胰岛素(FINS)、单核细胞趋化蛋白1(MCP-1)、肿瘤坏死因子α(TNF-α)含量,计算胰岛素抵抗指数(HOMA-IR),血液自动分析仪测定血清胆固醇(TC)、三酰甘油(TG)、低密度脂蛋白胆固醇(LDL-C)和高密度脂蛋白胆固醇(HDL-C)水平,HE染色观察肝脏病理形态,Western blot验证附睾脂肪磷酸化(p)-激活Janus激活激酶2(JAK2)、JAK2、p-信号转导和转录激活因子3(STAT3)、STAT3、p-细胞因子信号传导抑制蛋白1(SOCS-1)、SOCS-1蛋白表达。结果与CT组比较,IGT组大鼠肝脏细胞肿胀,胞浆内可见大量大小不一的脂肪空泡,细胞核被脂肪空泡挤压偏位,且发生炎性细胞浸润,FPG、2hPG、FINS、HOMA-IR、TC、TG、LDL-C、MCP-1、TNF-α以及p-JAK2/JAK2、p-STAT3/STAT3、p-SOCS-1/SOCS-1蛋白表达水平升高,血清HDL-C水平下降(P<0.05);与IGT组相比,Ori组大鼠肝脏细胞胞浆内脂肪滴及空泡数量明显减少,细胞肿胀有所缓解,未见炎性细胞浸润,FPG、2hPG、FINS、HOMA-IR、TC、TG、LDL-C、MCP-1、TNF-α以及p-JAK2/JAK2、p-STAT3/STAT3、p-SOCS-1/SOCS-1蛋白表达水平下降,血清HDL-C水平升高(P<0.05);与Ori组相比,Ori+COL组大鼠肝脏脂肪变状况加剧,细胞肿大,血清FPG、2hPG、FINS、HOMA-IR、TC、TG、LDL-C、MCP-1、TNF-α以及p-JAK2/JAK2、p-STAT3/STAT3、p-SOCS-1/SOCS-1蛋白表达水平升高,血清HDL-C水平下降(P<0.05)。结论Ori对IGT大鼠IR的缓解作用可能与抑制JAK2/STAT3/SOCS-1信号通路激活有关。

PIM1基因对急性髓系白血病U937细胞增殖、凋亡及JAK2/STAT3信号通路的影响

目的:探讨PIM1基因对急性髓系白血病(AML)U937细胞增殖、凋亡的影响,以及对JAK2/STAT3通路的调控作用。方法:收集初诊成人AML患者和单纯缺铁性贫血患者的骨髓单个核细胞,荧光定量PCR检测PIM1 mRNA表达。将AML细胞系U937细胞分为:U937组(U937细胞正常培养)、Si-PIM1组(U937细胞转染含PIM1 mRNA的低表达腺病毒载体)、Si-NC组(U937细胞转染不含PIM1 mRNA的低表达腺病毒载体)、CoA1组(U937细胞中加入浓度为20μmol/L的JAK2激活剂CoA1)、Si-PIM1+CoA1组(U937细胞转染含PIM1 mRNA低表达的腺病毒载体并加入浓度为20μmol/L的CoA1)。培养24 h。荧光定量PCR和蛋白印迹法检测U937细胞PIM1 mRNA和蛋白、JAK2/STAT3通路、细胞周期、凋亡相关蛋白表达;噻唑蓝法检测细胞增殖活性;流式细胞术检测细胞周期变化及凋亡率。结果:AML患者骨髓单个核细胞中PIM1 mRNA表达水平高于单纯缺铁性贫血患者(P<0.05)。与U937组相比,Si-PIM1组细胞PIM1 mRNA和蛋白、p-JAK2/JAK2、p-STAT3/STAT3、Cyclin D1、CDK2蛋白、细胞增殖活性、S期比例、G2/M期比例降低(均P<0.05),p27、Caspase-3蛋白、G0/G1期、凋亡率升高(均P<0.05),而CoA1组上述指标的变化情况与Si-PIM1组正好相反,CoA1可逆转Si-PIM1对U937细胞的作用效果。U937组、Si-PIM1+CoA1组、Si-NC组U937细胞上述指标差异无统计学意义(P>0.05)。结论:敲低PIM1基因表达可抑制U937细胞增殖、促进凋亡,缓解ALM进程,且上述作用可能与抑制JAK2/STAT3通路活化有关。

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.

基于IL-6/JAK2/STAT3信号通路探讨灰树花提取物对溃疡性结肠炎大鼠结肠组织炎症反应的影响

目的:探讨灰树花提取物通过调控白细胞介素6(IL-6)/蛋白酪氨酸激酶2(JAK2)/信号转导和激活因子3(STAT3)信号通路,对溃疡性结肠炎(UC)大鼠结肠组织炎症反应的影响及作用机制。方法:将40只SD大鼠随机分为空白对照组、UC模型组、灰树花治疗组、西药治疗组、联合治疗组,每组8只。采用3%DSS自由饮用法7 d建立UC大鼠模型后,治疗组分别灌胃灰树花提取物10 mg/(kg·d)、柳氮磺吡啶0.3 g/(kg·d)及等量两种药物,连续14 d。实验期间观察大鼠一般状态,计算疾病活动指数(DAI)评分;采用HE染色法观察大鼠结肠组织病理改变;Western blot检测大鼠结肠组织IL-6、JAK2、STAT3、pSTAT3蛋白表达水平;ELISA检测大鼠血清中IL-6含量;免疫组化法测定大鼠结肠组织中IL-6R、MPO蛋白含量及表达情况。结果:与空白对照组比较,UC模型组大鼠一般状态欠佳,DAI评分升高,HE染色可见明显的组织黏膜损伤与炎症细胞浸润;结肠组织IL-6、JAK2、STAT3、p-STAT3蛋白表达水平及IL-6R、MPO含量均显著升高(P<0.01);血清中IL-6含量显著升高(P<0.01),差异有统计学意义。与UC模型组比较,各治疗组大鼠的一般状态均有所改善,DAI评分降低,HE染色可见黏膜损伤有不同程度改善,偶见炎症细胞浸润;结肠组织IL-6、JAK2、STAT3、p-STAT3蛋白表达水平显著降低(P<0.01),结肠组织IL-6R和MPO含量及血清中IL-6含量均明显减少(P<0.01或P<0.05),差异有统计学意义。结论:灰树花提取物可通过调控IL-6/JAK2/STAT3信号通路相关因子表达,减轻UC大鼠结肠组织炎症反应。

升降理肺消瘤汤对Lewis肺癌小鼠免疫炎性反应和JAK2/STAT3信号通路的影响

目的探讨升降理肺消瘤汤对Lewis肺癌小鼠免疫和炎性反应的调控作用,以及对JAK2/STAT3信号通路的影响。方法适应性喂养C57BL/6J雄性小鼠1周,其中空白组8只,荷瘤组60只。荷瘤组小鼠注射Lewis肺癌细胞造模成功后,将瘤体肉眼可见的荷瘤小鼠分为模型组、PD-1抑制剂组、升降理肺消瘤汤(XLT)低剂量组、XLT中剂量组、XLT高剂量组、XLT中剂量联合PD-1抑制剂组(联合用药组),每组8只。空白组和模型组每日0.4 mL/20 g生理盐水灌胃;PD-1抑制剂组每3 d腹腔注射100μg信迪利单抗;XLT低、中、高剂量组每日0.4 mL/20 g相应浓度中药灌胃;联合用药组每日0.4 mL/20 g中浓度中药灌胃,每3 d腹腔注射100μg信迪利单抗,各组连续给药14 d。最后一次给药的24 h后,称量记录小鼠体质量,摘眼球取血,颈椎脱臼法处死小鼠,比较各组小鼠抑瘤率、去瘤体质量、胸腺指数和脾指数;ELISA法检测血清中白细胞介素-2(IL-2)、干扰素-γ(IFN-γ)、肿瘤坏死因子-α(TNF-α)、IL-6浓度水平;Western Blot检测Janus激酶2(JAK2)、p-JAK2、信号转导和转录激活因子3(STAT3)、p-STAT3蛋白表达水平。结果与模型组比较,各用药组小鼠平均瘤重均较少,PD-1抑制剂组、XLT中、高剂量组瘤重明显减少(P<0.01);PD-1抑制剂组和XLT中剂量组去瘤体质量增加(P<0.01);各用药组小鼠胸腺指数和脾指数均显著增加(P<0.01);PD-1抑制剂组、XLT中剂量组及联合用药组IL-2水平显著升高(P<0.05),各用药组小鼠IFN-γ水平均明显升高(P<0.05);除XLT低剂量组,各用药组TNF-α和IL-6水平均明显降低,差异有统计学意义(P<0.05);各用药组小鼠JAK2蛋白相对表达量均降低(P<0.05),XLT高剂量组和联合用药组p-JAK2降低差异有统计学意义(P<0.05);除PD-1抑制剂对p-STAT3蛋白的表达抑制作用不明显外,各用药组小鼠STAT3、p-STAT3蛋白相对表达量均较模型组显著降低(P<0.05)。结论升降理肺消瘤汤能够抑制Lewis肺癌小鼠瘤体生长,可能与其增强瘤鼠抗肿瘤免疫反应,减轻炎性反应,抑制JAK2/STAT3信号通路的激活有关。

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。

乔松素调节JAK2/STAT3信号通路对食管鳞癌细胞恶性进展的影响

目的:探究乔松素(pinocembrin,Pin)对食管鳞癌细胞恶性进展的影响及对Janus酪氨酸激酶2(janus activated kinase 2,JAK2)/信号传导与转录激活子3(signal transducer and activator of transcription 3,STAT3)信号通路的调控机制。方法:将人食管鳞癌细胞KYSE-410随机分为KYSE-410组、Pin低浓度(Pin-L)组、Pin中浓度(Pin-M)组、Pin高浓度(Pin-H)组、Pin-H+JAK2激活剂(Pin-H+Broussonin E)组。采用CCK-8法和克隆平板实验检测各组细胞的增殖活性;采用Transwell实验检测细胞的迁移和侵袭能力;采用流式细胞术检测各组细胞的凋亡情况;采用Western blot检测各组细胞中JAK2/STAT3信号通路相关蛋白的表达水平。结果:与KYSE-410组相比,Pin-L组、Pin-M组和Pin-H组细胞存活率(79.91±2.31、62.33±1.41、51.17±1.05,F=307.400),克隆细胞形成数量(162.82±6.33、144.59±5.09、120.18±3.72,F=209.800),迁移细胞数(77.73±3.26、60.83±2.41、49.28±1.60,F=360.314),侵袭细胞数(62.72±2.42、50.93±2.07、32.47±1.55,F=460.221)以及细胞中p-JAK2/JAK2(0.77±0.06、0.60±0.04、0.42±0.03,F=68.226)和p-STAT3/STAT3值(0.70±0.06、0.58±0.04、0.39±0.03,F=61.160)均降低(P<0.001),而细胞凋亡率(24.72±2.18、39.62±3.66、48.33±4.13,F=235.118)升高(P<0.001),Broussonin E的加入逆转了Pin对KYSE-410细胞恶性进展的抑制作用(P<0.05)。结论:Pin能够对食管鳞癌细胞的恶性进展起到抑制作用,其作用机制可能与JAK2/STAT3信号通路被抑制有关。

Janus激酶抑制剂AG490对人视网膜母细胞瘤HXO-RB_(44)细胞JAK2/STAT3信号通路的影响

目的:研究Janus激酶(Janus kinase,JAK)抑制剂AG490对人视网膜母细胞瘤HXO-RB44细胞株体外抗增殖及细胞周期的作用,探讨其对JAK2/信号转导与转录激活因子3(signal transducer and activator of transcription 3,STAT3)信号通路蛋白表达的影响。方法:本实验分为实验组和对照组,实验组又根据不同浓度(6.25,12.50,25.00,50.00,100.00,200.00μmol/L)的AG490处理分为6个不同浓度的实验组。采用细胞的活力测定法检测各组细胞增殖状态。应用流式细胞术对各组中细胞凋亡及周期进行分析。采用Western印迹检测处理后STAT3,p-STAT3及血管内皮生长因子(vascular endothelial growth factor,VEGF)蛋白的表达。结果:AG490处理HXO-RB44细胞株48 h后,随着药物浓度的增加,细胞抑制率增加,细胞存活率下降(均P<0.05)。除6.25μmol/L实验组外,其余5组与对照组两两比较,差异均有统计学意义(均P<0.05)。流式细胞术显示:随着AG490药物浓度的增加,细胞凋亡率呈逐渐增高趋势,与对照组相比,差异均有统计学意义(均P<0.05)。其中,50.00和100.00μmol/L实验组G1期细胞比例显著增多,相应地处于S期的细胞比例减少。Western印迹显示:随着AG490药物浓度的增加,STAT3和p-STAT3蛋白的表达量逐渐下降,与对照组相比,差异均有统计学意义(均P<0.05);VEGF表达量逐渐下降,与对照组相比,6.25和12.50μmol/L实验组的VEGF差异均无统计学意义(均P>0.05),其余各实验组差异均有统计学意义(均P<0.05)。结论:JAK抑制剂AG490能抑制HXORB44细胞株生长及增殖,促进细胞的凋亡增加;并通过阻断JAK2/STAT3信号通路而下调STAT3,p-STAT3和VEGF的表达,从而抑制HXO-RB44细胞株的增殖,加速其凋亡。

丙泊酚对结肠癌细胞侵袭迁移及Janus激酶2/信号转导与转录激活子3信号通路的影响

目的:探讨丙泊酚对人结肠癌细胞株SW480侵袭、迁移的作用及对Janus激酶2/信号转导与转录激活子3(Janus kinase 2/signal transduction and transcriptional activator 3,JAK2/STAT3)信号通路的调控。方法:人结肠癌细胞株SW480分为空白对照组、丙泊酚处理组(又分为2,4,8μg/mL丙泊酚处理组)和丙泊酚+colivelin组,用乳酸脱氢酶(lactate dehydrogenase, LDH)活性检测试剂盒检测LDH活性,用噻唑蓝(methyl thiazolyl tetrazolium,MTT)法检测细胞增殖,用Transwell实验检测细胞迁移和侵袭,用蛋白质印迹法检测细胞中JAK2,磷酸化JAK2(p-JAK2),STAT3和磷酸化STAT3(p-STAT3)的蛋白表达量。结果:与空白对照组相比,丙泊酚处理组细胞株SW480的LDH活性显著升高(P<0.05),细胞增殖、迁移和侵袭能力以及p-JAK2和p-STAT3的表达均显著下降(均P<0.05)。与丙泊酚处理组相比,丙泊酚+colivelin组SW480细胞p-STAT3的表达水平、细胞增殖、迁移和侵袭能力均显著升高(均P<0.05)。结论:丙泊酚能够通过抑制JAK2/STAT3信号通路而抑制人结肠癌细胞的增殖和转移。

IL-6/IL-6R/JAK2/STAT3通路调控骨髓微环境对多发性骨髓瘤生物学行为影响的研究进展

多发性骨髓瘤(MM)是一种克隆浆细胞异常增殖的恶性疾病,疾病的发展表现出广泛的异质性,这种异质性与MM肿瘤细胞、骨髓微环境之间的相互作用密切相关。IL-6/IL-6R/JAK2/STAT3通路可以调节骨髓微环境中相关可溶性因子的转录,促进MM肿瘤细胞增殖、抗凋亡、产生耐药性及引导相关骨破坏。本文就IL-6/IL-6R/JAK2/STAT3通路调控骨髓微环境对MM生物学行为影响的研究进展进行综述,以期为MM的靶向治疗及精准治疗提供新的研究思路。

灯盏细辛注射液通过调节JAK2/STAT3信号通路抗脑卒中大鼠神经损伤的研究

[目的]探讨灯盏细辛注射液(EBI)对脑卒中大鼠的神经保护作用及在该过程中对Janus激酶2(JAK2)/信号转导子和转录激活子3(STAT3)信号通路的调节机制。[方法]通过大脑中动脉闭塞(MCAO)建立缺血性脑卒中大鼠模型,然后将造模成功的大鼠随机分为模型组、尼莫地平组、EBI组、JAK2抑制剂组(AG490组)和EBI+JAK2激动剂组(EBI+CA1组),每组20只,另外选取20只大鼠只暴露颈动脉不进行结扎作为假手术组。利用神经功能缺损评分评估大鼠神经功能;2,3,5-三苯基氯化四氮唑(TTC)染色检测大鼠脑梗死面积;苏木精-伊红(HE)和尼氏(Nissl)染色检测大鼠脑组织的病理性形态;TUNEL染色检测大鼠脑组织中神经细胞凋亡情况;酶联免疫吸附测定(ELISA)检测大鼠脑组织中超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)和丙二醛(MDA)的水平;蛋白免疫印迹法(Western blot)检测大鼠脑组织中Bax、B淋巴细胞瘤-2(Bcl-2)及JAK2、p-JAK2、STAT3、p-STAT3蛋白的表达。[结果]与假手术组相比,模型组大鼠脑梗死面积增大,神经元细胞凋亡率增加,神经缺损评分、MDA水平、Bax蛋白表达及p-JAK2/JAK2、p-STAT3/STAT3比值均升高(P<0.05),同时脑组织中神经元退化,神经元数量减少,SOD、GSH-Px活性及Bcl-2蛋白表达降低(P<0.05);与模型组相比,EBI组、尼莫地平组和AG490组大鼠脑梗死面积减小,神经元细胞凋亡率减少,神经缺损评分、MDA水平、Bax蛋白表达及p-JAK2/JAK2、p-STAT3/STAT3比值均降低(P<0.05),同时脑组织中神经元损伤减轻,神经元数量增加,SOD、GSH-Px活性及Bcl-2蛋白表达升高(P<0.05);进一步利用JAK2激动剂进行回补实验发现,EBI对大鼠神经损伤的保护作用被逆转(P<0.05)。[结论] EBI能够通过抑制JAK2/STAT3信号通路减轻脑卒中大鼠的神经损伤。