Mechanisms of Radix Tetrastigma on anti rheumatoid arthritis via regulating the balance of Th17/Treg / 中国临床药理学与治疗学

AIM: To explore the possible mechanism of Radix Tetrastigma (RT) on anti-rheumatoid arthritis (RA). METHODS: The rat model of RA was established by intradermal injection of complete Freund]s adjuvant into the right hind foot of SD rats. RT Extract with different dosage was continuous intragastric administration for 3 weeks, then, the degree of foot swelling, arthritis index score, joint heat and grip of each rat was measured respectively. ELISA was used to detect the expression levels of Interleukin (IL) 6, IL-17, IL-10, tumor necrosis factor-α, and rheumatoid factor. Fully automatic hemorheometer was applied to measure hemorheology indexes. The number of CD4

Influence of matrine on Th17/Treg balance in rats with inflammatory bowel disease by regulating IL-6/STAT3/NF-κB signaling pathway / 西安交通大学学报(医学版)

【Objective】 To investigate the influence of matrine (MT) on the balance of T helper cell 17 (Th17)/regulatory T cells (Treg) in rats with inflammatory bowel disease by regulating interleukin-6 (IL-6)/signal transducer and activator of transcription 3 (STAT3)/nuclear transcription factor-κB (NF-κB) pathway. 【Methods】 SD rats were grouped into control check group (CK group), model group, low-dose MT group (MT-L group, 50 mg/kg), medium-dose MT group (MT-M group, 100 mg/kg), high-dose MT group (MT-H group, 200 mg/kg), mesalazine group (MSLM group, 0.42 g/kg), and MT-H+rIL-6 (IL-6 activator) group (200 mg/kg+0.05 mg/kg) according to the random number table method, with 18 in each group. Except for the CK group, the rats in other groups all received with 5% trinitrobenzenesulfonic acid (20 mg/kg) buffer solution mixed with 50% ethanol at a ratio of 1∶1 and then enema to construct a rat model of inflammatory bowel disease. After the successful modeling, they were treated with drug administration once a day for 7 weeks. The body weight of rats was measured at 1, 3, 5, and 7 weeks of administration; the changes of colon length of rats in each group were compared; HE staining was used to detect the pathological damage of rat colon tissue; the levels of tumor necrosis factor-α (TNF-α), interleukin (IL)-17 and IL-10 in serum of rats were detected by ELISA; the proportions of Th17 and Treg cells in peripheral blood of rats were detected by flow cytometry; Western blottingt was performed to detect the protein expression of retinoic acid-related orphan receptor γt (RORγt), forkhead box protein P3 (Foxp3), IL-6, p-STAT3, and p-NF-κB p65 in rat colon tissue. 【Results】 Compared with the CK group, the colon tissue of the model group was severely damaged by pathology, and the body weight (at 3, 5, and 7 weeks), the level of IL-10, the proportion of Treg cell, and the expression of Foxp3 protein were decreased, the colon length shortened, the levels of TNF-α, IL-17, the proportions of Th17 cell, Th17/Treg ratio, and the protein expression of RORγt, IL-6, p-STAT3, and p-NF-κB p65 increased (P<0.05). Compared with the model group, the corresponding indicators of the MT-L group, MT-M group, MT-H group, and MSLM group had the opposite trends (P<0.05); rIL-6 attenuated the promoting effect of high-dose MT on Th17/Treg balance in inflammatory bowel disease rats. 【Conclusion】 MT may promote Th17/Treg balance in inflammatory bowel disease rats by inhibiting IL-6/STAT3/NF-κB signaling pathway.

Knocking-down Synaptotagmin 1 Attenuated Colitis via Suppressing the IL-6/STAT3 Signaling Pathway / 中国生物化学与分子生物学报

Synaptotagmin 1 (Sytl) is a member of the Synaptotagmin family and plays a role in neurotransmitter vesicle transport and exoeytosis. It has been reported that Sytl appears to be expressed in the intestinal epithelium, but the biological function of Sytl in colitis remains poorly understood. This study aimed to investigate the effects of Sytl in the inflammatory response and intestinal epithelial regeneration in colitis using Sytl transgenic mice and dextran sulfate sodium (DSS)-induced ulcerative colitis mode. qRT-PCR and Western blotting were employed to analyze the dynamic changes of Sytl in colitis. H&E staining, immunostaining and Western blotting were used to explore the roles of Sytl in the inflammatory response and in the regeneration and repair of intestinal epithelium in colitis. The results showed that the expression level of Sytl was indeed high in the colonic epithelium of wild-type mice and the intestinal epithelial cells of the adjacent tissues of colorectal cancer patients. Consistently, DSS-induced inflammation progressively resulted in marked upregulation of Sytl in the colon (P<0.01). In DSS-induced colitis, both the body weight loss and colonic shortening were dampened in Sytl loss-of-function mice compared with the control group (P < 0.05), while the number of regenerated crypts and Ki67 proliferating cells were also increased (P<0.01). Additionally, there were less infiltration of CD45 immune cells and F 4/80 macrophages and the expression of pro-inflammatory cytokines IL-6, TNFα and I L l-β, which were related with the severity of inflammation in the inflammatory bowel disease (I B D), were significantly decreased after Sytl deletion (P<0.05). Immunohistochemistry staining and Western blotting results further showed that IL-6 and p-STAT3 was significantly downregulated in Sytl knockdown mice (P<0.05). Taken together, these results suggested that knocking-down of Sytl may improve colitis by inhibiting the IL6/STAT3 signaling pathway.

Chaihu Guizhi Decoction plus or minus formula combined with capecitabine inhibits IL-6/STAT3 signaling to suppress triple-negative breast cancer xenografts in nude mice / 南方医科大学学报

OBJECTIVE@#To investigate the effect of Chaihu Guizhi Decoction (CHGZD) combined with capecitabine on growth and apoptosis of subcutaneous triple-negative breast cancer xenografts in nude mice and explore the possible mechanism.@*METHODS@#Nude mouse models bearing subcutaneous triple-negative breast cancer xenografts were randomized into 6 groups (n=10) for treatment with distilled water (model group), low (10.62 g/kg), medium (21.23 g/kg) and high (42.46 g/kg) doses of CHGZD, capecitabine (0.2 mg/kg), or the combination of CHGZD (42.46 g/kg) and capecitabine (0.2 mg/k) once daily for 21 consecutive days. The general condition of mice was observed, and after 21-day treatments, the tumors were dissected for measurement of tumor volume and weight and histopathological examination with HE staining. Serum IL-6 levels of the mice were determined with enzyme-linked immunosorbent assay (ELISA), and the expression levels of IL-6, STAT3, p-STAT3, Bax, Bcl-2 and cyclin D1 in the tumor tissues were detected using real-time PCR and Western blotting.@*RESULTS@#Compared with those in the model group, the tumor-bearing mice receiving treatments with CHGZD showed significantly increased food intake with good general condition, sensitive responses, increased body weight, and lower tumor mass (P < 0.01). Compared with capecitabine treatment alone, treatment with CHGZD alone at the medium and high doses and the combined treatment all resulted in significantly higher tumor inhibition rates (P < 0.01), induced obvious tumor tissue degeneration and reduced the tumor cell density. Treatments with CHGZD, both alone and in combination with capecitabine, significantly decreased serum IL-6 level, lowered the mRNA expression levels of IL-6 and STAT3, the protein expressions of IL-6, STAT3 and P-STAT3 (P < 0.05), and the mRNA and protein expressions of Bcl-2 and cyclin D1 (P < 0.05), and increased the mRNA and protein expressions of Bax in the tumor tissues (P < 0.05).@*CONCLUSION@#CHGZD combined with capecitabine can significantly inhibit tumor growth in nude mice bearing triple-negative breast cancer xenografts, the mechanism of which may involve the inhibition of IL-6/STAT3 signaling pathway and regulation of Bax, Bcl-2 and cyclin D1 expressions to suppress tumor cell proliferation and differentiation and induce cell apoptosis.

The IL-6/STAT3 signaling pathway mediated by radiotherapy regulates the expression of PD-L1 in esophageal cancer cells / 中华肿瘤杂志

Objective: To explore the possible mechanism of radiotherapy regulating the expression of PD-L1 in esophageal carcinoma. Methods: Three esophageal cancer cell lines (Eca109, Kyse150, TE1) were irradiated with different doses of X-rays, and 6 Gy+ AG490 group was set. The mRNA expression of PD-L1 was detected by real-time quantitative polymerase chain reaction (RT-qPCR). The protein expressions of PD-L1, STAT3, p-STAT3 were detected by western blotting and the protein level of IL-6 was detected by ELISA. Results: The mRNA expressions of PD-L1 in Eca109, Kyse150 and TE1 were 2.86±0.30, 960.01±21.27 and 106.78±6.67, higher than 1.07±0.15 in normal esophageal cell line HET-1A (P<0.01). The protein expressions of PD-L1 in Eca109, Kyse150 and TE1 were 0.091±0.036, 1.533±0.079 and 0.914±0.035, higher than 0.063±0.01 in normal esophageal cell line HET-1A (P<0.01). After 48 hours of 6 Gy irradiation, the protein expression levels of PD-L1 in Eca109, Kyse150 and TE1 were 0.135±0.007, 1.66±0.06 and 1.32±0.06, higher than 0.09±0.01, 1.21±0.05 and 0.93±0.03 of the 0 Gy group (P<0.01), while the protein expression levels of p-STAT3 in Eca109, Kyse150 and TE1 were 1.44±0.26, 0.75±0.04 and 1.92±0.17, higher than 0.18±0.05, 0.48±0.02 and 0.36±0.06 of the 0 Gy group (P<0.01). IL-6 protein expression increased significantly after different doses of irradiation (P<0.01). After the IL-6/STAT3 signaling pathway was blocked by the specific inhibitor AG490, the expressions of PD-L1 of Eca109, Kyse150 and TE1 in the 6 Gy+ AG490 groups were 0.11±0.03, 1.07±0.08 and 0.96±0.11, without significant differences of 0.09±0.01, 0.96±0.05 and 0.85±0.09 of the 0 Gy group (P>0.05), while the protein expressions of p-STAT3 were 0.76±0.11, 0.59±0.06 and 0.96±0.12, without significant differences of 0.67±0.08, 0.54±0.06 and 0.84±0.11 of the 0 Gy group (P>0.05). Conclusion: Radiotherapy may regulate the expression of PD-L1 in esophageal cancer cells through IL-6 / STAT3 signaling pathway.

Macrophage M1 polarization mediated via the IL-6/STAT3 pathway contributes to apical periodontitis induced by Porphyromonas gingivalis

Abstract Objective: To investigate the involvement of IL-6/STAT3 signaling pathway activation in macrophage polarization and bone destruction related to apical periodontitis (AP) stimulated by Porphyromonas gingivalis. Methodology: Macrophage polarization, IL-6/STAT3 expression, and the presence of P. gingivalis were detected in human AP tissues via RT-qPCR, western blotting, and immunohistochemistry staining. Murine bone marrow derived macrophages were isolated and cultured with P. gingivalis W83 in vitro, and levels of macrophage IL-6 expression, STAT3 phosphorylation, and macrophage polarization with or without the selective STAT3 phosphorylation inhibitor Stattic (5 μM) were detected via ELISA, western blotting, RT-qPCR, and flow cytometry, respectively. P. gingivalis-induced murine AP models were constructed, and bone destruction and macrophage polarization in the apical region were evaluated. Transwell co-culture systems were used to investigate the effects of macrophages infected with P. gingivalis on osteogenesis and osteoclastogenesis. Results: P. gingivalis was detected in human AP tissues that highly expressed IL-6/STAT3, and the M1 subtype of macrophages was more abundant in these tissues. P. gingivalis infection induced IL-6 expression, STAT3 phosphorylation, and M1 polarization of macrophages, while 5 μM of Stattic partially abolished these activation effects. Systemic STAT3 blockade via oral administration of Stattic at a dose of 25 mg kg-1 alleviated murine periapical bone resorption and apical infiltration of M1 macrophages induced by P. gingivalis infection in vivo. Furthermore, macrophages infected with P. gingivalis promoted bone destruction via secretion of IL-6, TNF-α, and RANKL, which hinder pre-osteoblast expression of Runx2 and accelerate pre-osteoclast expression of NFAT2. Conclusions: The activation of IL-6/STAT3 signaling pathway is involved in mediating macrophages M1 polarization in the P. gingivalis induced apical inflammatory context and may also be intimately involved in the bone loss caused by P. gingivalis infection, directing the M1 macrophage infiltration during the progression of AP.

Curcumin mediates IL-6/STAT3 signaling pathway to repair intestinal mucosal injury induced by 5-FU chemotherapy for colon cancer / 中国中药杂志

This study aims to investigate the potential mechanism of curcumin in mediating interleukin-6(IL-6)/signal transducer and activator of transcription 3(STAT3) signaling pathway to repair intestinal mucosal injury induced by 5-fluorouracil(5-FU) chemotherapy for colon cancer. SD rats were intraperitoneally injected with 60 mg·kg~(-1)·d~(-1) 5-FU for 4 days to establish a model of intestinal mucosal injury. Then the rats were randomly divided into model group(equal volume of normal saline), curcumin low, medium and high dose groups(50, 100, 200 mg·kg~(-1)), and normal SD rats were used as control group(equal volume of normal saline). Each group received gavage administration for 4 consecutive days, and the changes of body weight and feces were recorded every day. After administration, blood was collected from the heart, and jejunum tissues were collected. The levels of serum interleukin-1β(IL-1β) and tumor necrosis factor-α(TNF-α) were detected by ELISA, and at the same time, the concentration of Evans blue(EB) in jejunum was measured. Hematoxylin-eosin(HE) staining was used to observe the pathological state of jejunum, and the length of jejunum villi and the depth of crypt were measured. The positive expression levels of claudin, occludin and ZO-1 were detected by immunohistochemistry. Western blot was used to detect the protein expression of IL-6, p-STAT3, E-cadherin, vimentin and N-cadherin in jejunum tissues. The results showed that, curcumin significantly increased body weight and fecal weight(P<0.05 or P<0.01), decreased fecal score, EB concentration, IL-1β and TNF-α levels(P<0.05 or P<0.01) in rats. In addition, curcumin maintained the integrity of mucosal surface and villi structure of jejunum to a large extent, and reduced pathological changes in a dose-dependent manner. Meanwhile, curcumin could increase the positive expression of occludin, claudin and ZO-1(P<0.05 or P<0.01), repair intestinal barrier function, downregulate the protein expression of IL-6, p-STAT3, vimentin and N-cadherin in jejunum tissues(P<0.05 or P<0.01), and upregulate the protein expression of E-cadherin(P<0.05). Therefore, curcumin could repair the intestinal mucosal injury induced by 5-FU chemotherapy for colon cancer, and the mechanism may be related to the inhibition of IL-6/STAT3 signal and the inhibition of epithelial-mesenchymal transition(EMT) process.

Effect of moxibustion on IL-6/STAT3 signaling in frontal cortex of fatigue rats / 针刺研究

OBJECTIVE: To observe the effect of moxibustion on interleukin-6(IL-6)/signal transduction and transcriptional activator 3(STAT3) signaling pathway in the frontal cortex of fatigue rats, so as to reveal its mechanisms underlying alleviation of fatigue. METHODS: Twenty-one male SD rats were randomly divided into normal control, model, and moxibustion groups (n＝7 rats in each group). The fatigue model was established by forcing the rats to have an exhausted swim under load condition, once daily for 21 days. Moxibustion was applied to bilateral "Zusanli"(ST36) for about 15 min, once every other day for 21 days. The level of IL-6 in the frontal cortex was detected by ELISA, and the expression of Janus kinase 2 (JAK2), phosphorylated JAK2 (p-JAK2), signal transduction and transcriptional activator 3(STAT3) and phosphorylated STAT3 (p-STAT3) proteins in the frontal cortex was detected by Western blot. RESULTS: After modeling, the levels of IL-6 content and p-STAT3 protein expression and ratio of p-STAT3/STAT3 were significantly increased in the model group relevant to the normal control group (P0.05). CONCLUSION: Moxibustion intervention can relieve fatigue in fatigue rats, which is associated with its function in inhibiting IL-6/STAT3 signaling pathway to reduce inflammatory injury.

Relationship between long non-coding RNA metastasis associated lung adenocarcinoma transcript 1 and IL-6/STAT3 signaling pathway in isoniazid-induced rats liver injury / 国际药学研究杂志

Objective To explore the link between the expression of long non-coding RNA(lncRNA)metastasis associated lung adenocarcinoma transcript 1(MALAT1)and IL-6/signal trans ducers and activators of transcription 3(STAT3)signaling pathway in isoniazid induced rats liver injury. Methods Fifty-six specific pathogen-free(SPF)SD rats were randomly divided into experimen?tal group(48 rats)and control group(8 rats),each with half females and half males. The rats in experimental group were given isonia?zid of 63 mg/(kg·d)for 3,7,10,14,21 and 28 d,with 8 rats at the same time point of each day. The rats in control group were giv?en distilled water by intragastric administration. Serum levels of ALT and AST were measured by automatic biochemical analyzer;SYBR green real-time polymerase chain reaction was used to test the expression level of lncRNA MALAT1 and IL-6/STAT3 mRNA in the liver. Results Liver tissue injury occurred after 7 days and worsened with the extention of administration time. Compared with the rats in the control group,the expression level of lncRNA MALAT1 and IL-6/STAT3 mRNA as well as ALT and AST showed a trend of increase(P<0.01). The expression of ALT,AST and lncRNA MALAT1 declined at different degrees on 28(P<0.05). LncRNA MALAT1 and IL-6/STAT3 mRNA expression levels were positively correlated(P<0.01). LncRNA MALAT1 and IL-6/STAT3 mRNA expression levels had a positive correlation with the contents of ALT and AST(P<0.01). Conclusion The expression level of lncRNA MALAT1 in isoniazid induced liver injury rat models showed an abnormal rising trend,and the positive detection time preceded that of ALT and AST. The mechanism may be related to the activation of IL-6/STAT3 signaling pathway.

Effects and values of antisurvivin oligonucleotides on IL-6/STAT3 signaling pathway of ovarian cancer cell line / 西安交通大学学报(医学版)

ABSTRACT:Objective To analyze the effects of antisurvivin oligonucleotide (Survivin ASODN)on IL-6/STAT3 signaling pathway and down-stream cancer genes of ovarian cancer SKOV3 cell line and to detect the changes of invasive ability of cell line in order to explore the role of Survivin ASODN in reducing metastasis and recurrence of ovarian cancer and its potential clinical value.Methods ASODN was transfected into SKOV3 cells by lipofectamineTM2000 in ASODN group,and lipofectamineTM 2000 was introduced in control group.The invasive ability in ASODN and control groups was detected by Transwell chamber.The expressions of interieukin-6 (IL-6 ), signal transducer and activator of transcription 3 (STAT3 ), Survivin, and vascular endothelial growth factor (VEGF)in ovarian cancer cell line at mRNA level were detected by Real-time PCR.The expressions of IL-6 , STAT3 ,signal transducer and activator of transcription 3 phosphorylation (p-STAT3 ),Survivin,and VEGF-A in ovarian cancer cell line at protein level were detected by Western blot.Results The expressions of IL-6,STAT3, Survivin,and VEGF in ovarian cancer cell line at mRNA level were significantly down-regulated in ASODN group (P<0 .0 5 ).IL-6 ,STAT 3 ,p-STAT 3 ,Survivin and VEGF-A protein levels in ovarian cancer cell line were significantly down-regulated in ASODN group (P<0.05).The invasive ability was significantly reduced by ASODN (P<0.01).Conclusion ASODN targeting Survivin could reduce the invasive ability of ovarian cancer cell line. The mechanism may be blocking IL-6/STAT3 signaling pathway and its down-stream cancer genes of ovarian cancer cell line.ASODN targeting Survivin may have clinical value in preventing and treating the metastasis and recurrence of ovarian cancer.

Relationship between long non-coding RNA metastasis associated lung adenocarcinoma transcript 1 and IL-6/STAT3 signaling pathway in isoniazid-induced rats liver injury / 国际药学研究杂志

Objective. To explore the link between the expression of long non-coding RNA (lncRNA) metastasis associated lung adenocarcinoma transcript 1 (MALAT1) and IL-6/signal trans ducers and activators of transcription 3(STAT3) signaling pathway in isoniazid induced rats liver injury. Methods. Fifty-six specific pathogen-free (SPF) SD rats were randomly divided into experimental group (48 rats) and control group (8 rats), each with half females and half males. The rats in experimental group were given isoniazid of 63 mg/ (kg•d) for 3, 7, 10, 14, 21 and 28 d, with 8 rats at the same time point of each day. The rats in control group were given distilled water by intragastric administration. Serum levels of ALT and AST were measured by automatic biochemical analyzer; SYBR green real-time polymerase chain reaction was used to test the expression level of lncRNA MALAT1 and IL-6/STAT3 mRNA in the liver. Results. Liver tissue injury occurred after 7 days and worsened with the extention of administration time. Compared with the rats in the control group, the expression level of lncRNA MALAT1 and IL-6/STAT3 mRNA as well as ALT and AST showed a trend of increase (P<0.01). The expression of ALT, AST and lncRNA MALAT1 declined at different degrees on 28 (P<0.05). LncRNA MALAT1 and IL-6/STAT3 mRNA expression levels were positively correlated (P<0.01). LncRNA MALAT1 and IL-6/STAT3 mRNA expression levels had a positive correlation with the contents of ALT and AST (P<0.01). Conclusion. The expression level of lncRNA MALAT1 in isoniazid induced liver injury rat models showed an abnormal rising trend, and the positive detection time preceded that of ALT and AST. The mechanism may be related to the activation of IL-6/STAT3 signaling pathway.

Activation-status of IL-6-related nuclear factors in acute myeloid leukemia cells and their biological roles / 中国免疫学杂志

Objective:To investigate the relation between the effects of IL 6 upon acute myeloid leukemia (AML) cells and its signal transduction mechanism Methods:Determined the effects of IL 6 on the growth of M1, R2, K562 and TF1 cells and the activation status of IL 6 induced nuclear factors (signal transducer and activator of transcription 3, STAT3 and nuclear factor IL6, NF IL6) by electrophoresis mobility shift assay (EMSA) Based upon these, blocked/reduced the activation of STAT3 or NF IL6 in M1 leukemia cells with STAT3/NF IL6 anti sense oligonucleotides (AS ODNs) and determined the oligonucleotides' effects upon IL 6 induced growth arrest of M1 cells Results:NF IL6 was constitutively activated in all these AML cells, while STAT3 was constitutively activated only in R2 and K562 cells Furthermore, STAT3 AS ODN reduced IL 6 induced growth arrest of M1 cells, while blocking/reducing of NF IL6 activation resulted in enhancing of IL 6 induced growth arrest Conclusion:JAK STATs pathway (especially STAT3) and Ras/MAPK pathway antagonize each other in proliferation regulation of AML cells