Porcine deltacoronavirus nucleocapsid protein antagonizes JAK-STAT signaling pathway by targeting STAT1 through KPNA2 degradation.

Porcine deltacoronavirus (PDCoV) is an enteric pathogenic coronavirus that causes acute and severe watery diarrhea in piglets and has the ability of cross-species transmission, posing a great threat to swine production and public health. The interferon (IFN)-mediated signal transduction represents an important component of virus-host interactions and plays an essential role in regulating viral infection. Previous studies have suggested that multifunctional viral proteins encoded by coronaviruses antagonize the production of IFN via various means. However, the function of these viral proteins in regulating IFN-mediated signaling pathways is largely unknown. In this study, we demonstrated that PDCoV and its encoded nucleocapsid (N) protein antagonize type I IFN-mediated JAK-STAT signaling pathway. We identified that PDCoV infection stimulated but delayed the production of IFN-stimulated genes (ISGs). In addition, PDCoV inhibited JAK-STAT signal transduction by targeting the nuclear translocation of STAT1 and ISGF3 formation. Further evidence showed that PDCoV N is the essential protein involved in the inhibition of type I IFN signaling by targeting STAT1 nuclear translocation via its C-terminal domain. Mechanistically, PDCoV N targets STAT1 by interacting with it and subsequently inhibiting its nuclear translocation. Furthermore, PDCoV N inhibits STAT1 nuclear translocation by specifically targeting KPNA2 degradation through the lysosomal pathway, thereby inhibiting the activation of downstream sensors in the JAK-STAT signaling pathway. Taken together, our results reveal a novel mechanism by which PDCoV N interferes with the host antiviral response.IMPORTANCEPorcine deltacoronavirus (PDCoV) is a novel enteropathogenic coronavirus that receives increased attention and seriously threatens the pig industry and public health. Understanding the underlying mechanism of PDCoV evading the host defense during infection is essential for developing targeted drugs and effective vaccines against PDCoV. This study demonstrated that PDCoV and its encoded nucleocapsid (N) protein antagonize type I interferon signaling by targeting STAT1, which is a crucial signal sensor in the JAK-STAT signaling pathway. Further experiments suggested that PDCoV N-mediated inhibition of the STAT1 nuclear translocation involves the degradation of KPNA2, and the lysosome plays a role in KPNA2 degradation. This study provides new insights into the regulation of PDCoV N in the JAK-STAT signaling pathway and reveals a novel mechanism by which PDCoV evades the host antiviral response. The novel findings may guide us to discover new therapeutic targets and develop live attenuated vaccines for PDCoV infection.

An extracellular humanized IFNAR immunocompetent mouse model for analyses of human interferon alpha and subtypes.

Type I interferons (IFN-Is) have key roles in immune defense and treatments for various diseases, including chronic hepatitis B virus (HBV) infection. All IFN-Is signal through a shared IFN-I heterodimeric receptor complex comprising IFN-α receptor 1 (IFNAR1) and IFNAR2 subunits, but differences in antiviral and immunomodulatory responses among IFN-I subtypes remain largely unknown. Because the IFN-IFNAR interactions are species-specific, mice exhibit weak responses to human IFN-I. To more fully characterize the actions of human IFN-α and its subtypes in vivo, a gene targeting strategy was employed to generate gene knock-in mice with extracellular-humanized IFNAR1/2 (IFNAR-hEC) in the C57BL/6N strain. IFNAR-hEC mice actively responded to human IFN-I, and endogenous mouse IFN-I signalling remained active in heterozygous mice (IfnarhEC/+). Analyses of IFNAR-hEC mice and isolated cells showed that human IFN-α2 and α14 subtypes exerted differential effect on the activation of JAK-STAT signalling and immune responses. Compared with IFN-α2, IFN-α14 induced greater activation of STAT1/2 and IFN-stimulated genes, synergistically elicited IFN-α and -γ signalling, and induced higher numbers of antigen-specific CD8+ T cells. Moreover, IFNAR-hEC mice with HBV replication displayed long-term viral suppression upon treatment with the clinically-used PEGylated hIFN-α2. These results indicate that IFNAR-hEC mice may be useful for elucidating antiviral and immunomodulatory functions of human IFN-Is and for conducting preclinical studies. A better understanding of the distinct activities of IFN-α subtypes can provide insights concerning the development of improved IFN-based therapy.

JAK-STAT signaling pathway and rosacea / 中华皮肤科杂志

Studies have shown that rosacea is related to inflammatory factors, neurovascular function, micro-ecological environment and other factors. The Janus kinase (JAK) -signal transducer and activator of transcription (STAT) signaling pathway involves a variety of inflammatory cytokines, and plays an important role in cell proliferation, differentiation, apoptosis, angiogenesis and immune regulation. This review summarizes the JAK-STAT signaling pathway and explores its potential role in rosacea.

Immune regulation mediated by JAK/STAT signaling pathway in hemocytes of Pacific white shrimps, Litopenaeus vannamei stimulated by lipopolysaccharide.

To understand the regulatory mechanism of Janus kinase/Signal Transducers and Activators of Transcription (JAK/STAT) signaling pathway on the immune system of the Pacific white shrimp, Litopenaeus vannamei, RNA interference technique was used to investigate the effects of JAK/STAT signaling pathway on the immune response of hemocyte in Litopenaeus vannamei stimulated by lipopolysaccharide (LPS). The results showed that 1) after 6 h of LPS stimulation, the expression levels of immune genes in hemocyte were significantly up-regulated (P < 0.05), the immune defense ability (hemocyte number, phagocytosis rate, hemagglutination activity, bacteriolytic activity, antibacterial activity, prophenoloxidase system activity) and the hemocyte antioxidant ability were significantly higher than the control group, especially at 12 h. 2) After 48 h of STAT gene interference, the expression levels of immune genes in hemocytes were significantly down-regulated, and the immune defense ability (hemocyte count, phagocytosis rate, plasma agglutination activity, lysozyme activity, antibacterial activity, proPO system activity) and the antioxidant ability were reduced and significantly lower than control. Concurrently, after LPS stimulation, the immune indexes were significantly up-regulated at 12 h to the maximum but was still lower the undisturbed LPS group. These results indicate that JAK/STAT signaling pathway is involved in the immune regulation mechanism of L. vannamei against LPS stimulation through positive regulation of cellular immune and humoral immune. These results provide a basis for further research on the role and status of JAK/STAT signaling pathway in the immune defense of crustaceans.

MGF360-9L Is a Major Virulence Factor Associated with the African Swine Fever Virus by Antagonizing the JAK/STAT Signaling Pathway.

African swine fever (ASF)-an aggressive infectious disease caused by the African swine fever virus (ASFV)-is significantly unfavorable for swine production. ASFV has a complex structure and encodes 150-167 proteins; however, the function of most of these proteins is unknown. This study identified ASFV MGF360-9L as a negative regulator of the interferon (IFN)-ß signal. Further evidence showed that MGF360-9L interacts with signal transducer and activator of transcription (STAT) 1 and STAT2 and degrades STAT1 and STAT2 through apoptosis and ubiquitin-proteasome pathways, respectively. Subsequently, the activation of IFN-ß signaling was inhibited. Naturally isolated or genetically manipulated live attenuated viruses are known to protect against the virulent parental ASFV strains. Therefore, through homologous recombination, we deleted MGF360-9L from the virulent ASFV CN/GS/2018 strain to construct a recombinant strain, ASFV-Δ360-9L. Compared with the parent ASFV CN/GS/2018 strain, the replication level of ASFV-Δ360-9L decreased in primary porcine alveolar macrophage cultures at 24 h postinfection, but the difference is unlikely to be biologically relevant. Notably, ASFV-Δ360-9L was partially attenuated in pigs. To our knowledge, this study is the first to uncover the function of MGF360-9L during ASFV infection. MGF360-9L inhibits IFN-ß signaling through the targeted degradation of STAT1 and STAT2. Furthermore, MGF360-9L is a key virulence gene of ASFV. Our findings reveal a new mechanism by which ASFV inhibits host antiviral response; this might facilitate the development of live attenuated ASFV vaccines. IMPORTANCE African swine fever-an acute, febrile, hemorrhagic, highly contacting, and highly lethal disease caused by African swine fever virus (ASFV)-jeopardizes the global pig industry. Understanding the mechanism ASFV employs to evade host defense during infection is essential for developing targeted drugs and vaccines against ASFV. To our knowledge, this study identifies the mechanism of innate immunity against by MGF360-9L and the effect of MGF360-9L on ASFV pathogenicity. The results showed that MGF360-9L may help ASFV escape the host immunity by degrading STAT1 and STAT2 and thus inhibiting IFN-ß signaling. MGF360-9L is also an important virulence factor of ASFV. The deletion of MGF360-9L reduces ASFV virulence in pigs. This study explored a new mechanism of ASFV against innate immunity and identified a new ASFV virulence factor; these findings may guide the development of live attenuated ASFV vaccines.

Pentagalloylglucose reduces AGE-induced inflammation by activating Nrf2/HO-1 and inhibiting the JAK2/STAT3 pathway in mesangial cells.

Pentagalloylglucose (PGG), a gallotannin polyphenolic compound, has been found to possess a host of beneficial pharmacologic activities, such as anti-inflammatory and antioxidative activities. We previously demonstrated that PGG is capable of binding to the cell membrane of renal mesangial cells, but the pharmacological effect of PGG on diabetic renal injury and the underlying mechanisms are still not yet clear. In this study, the effects of PGG on Nrf2/HO-1 and JAK2/STAT3 signaling were explored in AGE-stimulated mesangial cells. Furthermore, the Nrf2 transcriptional inhibitor ML385 was used to verify the involvement of Nrf2 in the PGG-mediated inhibition of the JAK2/STAT3 cascade. Our results showed that PGG significantly inhibited AGE-induced ROS generation and activated AGE-inhibited Nrf2/HO-1 signaling. Moreover, AGE-induced inflammatory cytokines (IL-1ß and TNF-α) and their signaling through JAK2/STAT3 were blocked by PGG. Furthermore, ML385 suppressed Nrf2/HO-1 signaling, elevated ROS and cytokine production, and activated JAK2/STAT3 cascade were reversed by PGG. These findings indicate that PGG inhibits the JAK2/STAT3 cascade by activating Nrf2/HO-1 signaling.

Grass carp (Ctenopharyngodon idellus) PIAS1 inhibits innate immune response via interacting with STAT1.

Protein inhibitor of activated signal transducer and activator of transcription (PIAS) family protein involved in gene transcriptional regulation acts as negative regulator in Janus kinase-signal transducer and activator of transcription (JAK/STAT) signaling pathway. But until now, the roles of PIAS in fish are not clear. In this study, we identified the two mammalian PIAS1 orthologs from Ctenopharyngodon idellus, namely CiPIAS1a and CiPIAS1b, respectively. They can respond to the stimulation from Polyribocytidylic acid (Poly I:C), Grass Carp Reovirus (GCRV) and Lipopolysaccharides (LPS) respectively, so we suggested that they could participate in interferon (IFN)-mediated antiviral and antibacterial immune response. The subcellular localization and nuclear cytoplasm extraction showed that CiPIAS1a and CiPIAS1b were mainly distributed in the nucleus. In addition, Co-IP showed that they separately inhibited the phosphorylation of STAT1 via interacting with it, which leads to the reduction of IFN1 expression.

Studies on the anti-psoriasis effects and its mechanism of a dual JAK2/FLT3 inhibitor flonoltinib maleate.

Psoriasis is a chronic, inflammatory autoimmune disease mediated by T cells, and characterized with abnormal proliferation and differentiation of keratinocytes, and inflammatory infiltration. The Janus kinase-signal transducer and activator of transcription (JAK-STAT) pathway has been identified to play essential roles in mediating various of biological processes, and is closely related to autoimmune diseases. Dendritic cells (DCs) are important antigen presenting cells and play an important regulatory role in T cells. The proliferation, differentiation and function of DCs are regulated by JAK and FMS-like tyrosine kinase 3 (FLT3) signal pathways. Flonoltinib maleate (FM), a high selectivity dual JAK2/FLT3 inhibitor with IC50 values of 0.8 nM and 15 nM for JAK2 and FLT3, respectively, was developed by our laboratory. Moreover, FM was a potent JAK2 inhibitor with 863-fold and 696-fold selectivity over JAK1 and JAK3, respectively. In this study, the anti-psoriasis activity of FM was evaluated both in vitro and in vivo. FM effectively inhibited the proliferation of HaCaT, the inflammatory keratinocyte induced by M5 and markedly suppressed the generation and differentiation of DCs from bone marrow (BM), and inhibited the expression of FLT3 in DCs in vitro. FM effectively inhibited the ear thickening and improved the pathological changes of the ear in interleukin (IL)-23-induced psoriasis-like acanthosis mouse model. Further in keratin 14-vascular endothelial growth factor (K14-VEGF) transgenic homozygous mice model, FM could obviously improve the psoriatic symptom and pathological changes, significantly inhibit the generations of Th1 and Th17 cells in the spleen, and the accumulations of DCs in the ears. FM could also significantly reduce the expression of various inflammatory factors both in C57BL/6 and K14-VEGF mice ears, and the serum of K14-VEGF mice. Mechanism revealed that FM effectively suppressed the phosphorylation of JAK2, STAT3 and STAT5 in inflammatory keratinocytes and the mice ears of C57BL/6 and K14-VEGF, as well as the phosphorylation of FLT3 in K14-VEGF mice ears. In conclusion, FM plays an excellent anti-psoriasis activity, including inhibiting keratinocyte proliferation and regulating inflammatory response through inhibiting JAK2 and FLT3 signaling pathway.

MicroRNA-144-3p enhances LPS induced septic acute lung injury in mice through downregulating Caveolin-2.

OBJECTIVE: The emphasis of this study focused on the possible implication and the mechanism of miR-144-3p in septic acute lung injury (ALI) condition. METHODS: Mice were pre-injected with miR-144-3p agomir, miR-144-3p antagomir, sh-Caveolin-2 or PBS before 10 mg/kg LPS induced sepsis model establishment. The ratio of wet weight of lung tissues and body weight (W/W) was calculated. The pathological changes on lung tissues were observed by H&E staining. Secretions of inflammatory cytokines (TNF-α, IL-1ß and IL-6) in both mouse serum and lung tissues were determined by ELISA. Cell apoptosis and cell morphology were measured by TUNEL staining and H&E staining. The expressions of miR-144-3p, Caveolin-2, apoptotic related proteins and JAK/STAT pathway related proteins were measured by qRT-PCR or/and Western blot. Dual luciferase reporter assay was applied to detect the binding of miR-144-3p with Caveolin-2. RESULTS: LPS resulted in increased W/W, disrupted lung tissue, enhanced inflammatory response and cell apoptosis. miR-144-3p was upregulated while Caveolin-2 was downregulated in response to LPS treatment. Inflammation and cell apoptosis induced by LPS can be alleviated by miR-144-3p antagomir injection, but enhanced by miR-144-3p agomir or sh-Caveolin-2 treatment. miR-144-3p can negatively target Caveolin-2. miR-144-3p can activate the JAK/STAT signal pathway through Caveolin-2 in septic ALI mouse. CONCLUSION: miR-144-3 can promote LPS induced septic ALI through downregulating Caveolin-2 to activate the JAK/STAT signal pathway.

Dexmedetomidine inhibits apoptosis of astrocytes induced by oxygen-glucose deprivation via targeting JAK/STAT3 signal pathway.

OBJECTIVE: There is an increasing interest concerning the contribution of astrocytes to the intrinsic bioremediation of ischemic brain injury. The aim of this work was to disclose the effects and mechanism of dexmedetomidine (DEX) on the apoptosis of astrocytes under oxygen glucose deprivation (OGD) condition. METHODS: Primary cultured astrocytes separated from Sprague-Dawley (SD) rats were subjected to OGD treatment. Astrocytes were transfected with si-JMJD3 or pcDNA3.1-JMJD3 and then treated with DEX or JAK/STAT inhibitor (WP1066) before cell apoptosis was detected by TUNEL apoptosis kit. Western blot was applied to assess the level of apoptosis-related proteins Caspase-3, Bax and Bcl-2. Astrocyte cell viability was assessed by measuring the lactate dehydrogenase (LDH) level using a LDH assay kit. RESULTS: Astrocytes received OGD treatment had increased LDH and elevated apoptotic rate (P < 0.05). DEX could suppress OGD induced cytotoxic effect on astrocytes, as evidenced by decreased LDH release and suppressed cell apoptosis rate (P < 0.05). Meanwhile, DEX and WP1066 treatment were also found to inhibit the phosphorylation level of STAT1 and STAT3 (P < 0.05), indicating the DEX could suppress the activation of JAK/STAT signal pathway. JMJD3 overexpression in astrocytes could suppress the anti-apoptotic function of WP1066 in OGD treated astrocytes and hamper the protective effect of DEX in cell apoptosis (P < 0.05), suggesting that DEX and JAK/STAT signal pathway inhibits OGD induced apoptosis in astrocytes by down-regulating JMJD3. CONCLUSION: DEX protects astrocytes against apoptosis via inhibiting JAK2/STAT3 signal pathway and downregulating JMJD3 expression in vitro.

Antiviral activity of a polysaccharide from Radix Isatidis (Isatis indigotica Fortune) against hepatitis B virus (HBV) in vitro via activation of JAK/STAT signal pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Hepatitis B virus (HBV) infection frequently results in both acute and chronic hepatitis and poses serious threats to human health worldwide. Despite the availability of effective HBV vaccine and anti-HBV drugs, apparently inevitable side effects and resistance have limited its efficiency, thus prompt the search for new anti-HBV agents. The traditional Chinese medicine Radix Isatidis has been used for thousands of years, mainly for the treatment of viral and bacterial infection diseases including hepatitis. AIM OF THE STUDY: In this study, antiviral activities of a Radix Isatidis (Isatis indigotica Fortune) polysaccharide (RIP) were evaluated in vitro model using the HepG2.2.15 cell line and the underlying mechanism was elucidated with the aim of developing a novel anti-HBV therapeutic agent. MATERIALS AND METHODS: Structure features of the purified polysaccharide RIP were investigated by a combination of chemical and instrumental analysis. Drug cytotoxicity was assessed using the MTT assay. The contents of HBsAg, HBeAg, intracellular and extracellular IFN-α level were measured using respective commercially available ELISA kit. The HBV DNA expression was evaluated by real-time quantitative polymerase chain reaction (PCR) and the relevant proteins involved in TFN/JAK/STAT signaling pathways were examined by western blot assay. RESULTS: MTT assay showed that RIP had no toxicity on HepG2.2.15 cell line below the concentration 400 µg/ml at Day 3, 6 and 9. Furthermore, RIP at the concentration of 50, 100 and 200 µg/ml significantly reduced extracellular and intracellular level of HBsAg, HBeAg and HBV DNA in HepG2.2.15 cells in a time and dose-dependent manner. Moreover, RIP also enhanced the production of IFN-α in HepG2.2.15 cell via activation of JAK/STAT signal pathway and induction of antiviral proteins, as evidenced by the increased protein expression of p-STAT-1, p-STAT-2, p-JAK1, p-TYK2, OAS1, and Mx in HepG2.2.15 cells. In addition, the over expression of SOCS-1 and SOCS-3 was significantly abolished under same conditions. CONCLUSIONS: These results suggested that the HBV inhibitory effect of RIP was possibly due to the activation of IFN-α-dependent JAK/STAT signal pathway and induction of the anti-HBV protein expression.

The mechanism and tumor inhibitory study of Lagopsis supine ethanol extract on colorectal cancer in nude mice.

BACKGROUND: This study was aimed to determination the tumor inhibitory effect and explore the potential mechanisms of Lagopsis supine ethanol extract (Ls) on colorectal cancer. METHODS: The cell growth inhibition experiment of Ls in colorectal cancer cell lines was determined by MTT method in the time course of 24, 48 and 72 h in four gradient drug concentrations. The protein expression levels of pSTAT3, pJAK2, STAT3, JAK2, Bcl-2 and caspase 3 were measured by Western blot method. The mRNA levels of the downstream genes of STAT3 were detected through semi-quantitative RT PCR. Sixty Balb/c-nude mice were xenograft with HCT116 colorectal cancer cells through subcutaneously. The xenografts were divided into five groups: model group, positive group (capecitabine 300 mg/kg) and three dosages of Ls treated groups (75, 150 and 300 mg/kg). Tumor size and tumor weight were calculated for evaluation the anti-tumor effects. H & E staining and immunohistochemical analysis were used to determine the histopathological changes and the levels of pSTAT3 and pJAK2 in the tumor tissues. RESULTS: Ls exhibited a significant anti-proliferation effect in HCT116 and SW480 cells in vitro. The protein levels of pSTAT3, pJAK2 and Bcl-2, and the mRNA levels of Bcl-2 and Bak notably reduced with a dose-dependent manner. While the protein levels of caspase 3, and mRNA levels of Bax and caspase-3 remarkably increased in the gradient dosage of Ls in HCT116 cells. HCT116 in vivo xenografts experiment showed that the growth of the tumors significantly inhibited by Ls administration, which with no any significant body weight changes in each experiment group. The histopathology analysis displayed that Ls significantly reduced the inflammatory cells in tumor tissue. Furthermore, Ls also significantly down-regulate the protein levels of pSTAT3 and pJAK2 in the tumor tissues, compared with the model group. CONCLUSIONS: This work shows that Ls inhibited the cell proliferation of colorectal cancer in vitro and significantly reduced the tumor growth in HCT116 xenografts in vivo, which is probably related with the JAK/STAT signal pathway.

Tetramethylpyrazine attenuates blood-brain barrier disruption in ischemia/reperfusion injury through the JAK/STAT signaling pathway.

Tetramethylpyrazine (TMP) has been studied in depth and is widely used in the treatment of many kinds of diseases in China. However, whether it has neuroprotective effects on cerebral ischemia remains unclear. An ischemia/reperfusion (I/R) injury animal model was established via middle cerebral artery occlusion in this study. We set several different groups in which the rats were performed in different ways to explore the effects of TMP on blood-brainbarrier (BBB) disruption and determine whether TMP relieved BBB disruption through blocking the JAK/STAT signaling pathway. Our results showed that TMP could reduce the neurological functional loss, decrease the brain edema and BBB permeability, as well as increase the expression of tight junction proteins via inhibiting the activation of JAK/STAT signaling pathway. Overall, we demonstrated that TMP promoted neurological recovery after I/R injury via restoring the integrity and function of BBB.

Adeno-associated virus 9-mediated RNA interference targeting SOCS3 alleviates diastolic heart failure in rats.

OBJECTIVE: To explore the effect of adeno-associated virus 9-mediated RNA interference targeting SOCS3 (AAV9-SOCS3 siRNA) on the treatment of diastolic heart failure (DHF). METHOD: A rat DHF model was established, and cardiac function and hemodynamic changes were measured. HE, Sirius red and TUNEL staining were applied to observe the pathological changes in the myocardium. Immunoblotting and immunohistochemical staining were utilized to detect SOCS3 expression. The expression levels of various factors, including fibrosis-related factors (collagen I, collagen II, α-SMA and TGF-ß), inflammatory-related factors (IL-1ß, IL-6, TNF-α, p-p65 and ICAM-1) and factors related to the JAK/STAT signal pathway were analyzed by immunoblotting and/or qPCR. The serum levels of IL-1ß, IL-6, and TNF-α were measured using ELISA. RESULTS: SOCS3 expression was significantly downregulated in the DHF rat model by SOCS3 siRNA delivery. In the successfully established DHF rat model, cardiac function was clearly decreased, and cardiomyocyte apoptosis and myocardial fibrosis were significantly increased. These changes were ameliorated by treatment with AAV9-SOCS3 siRNA. The expression levels of p-JAK2 and p-STAT3 were significantly upregulated in the AAV9-SOCS3 siRNA group compared with the sham and AAV9-siRNA control groups, indicating that SOCS3 is a negative regulator of this signaling pathway. The expression levels of collagen I/III, α-SMA and TGF-ß were also decreased at both the mRNA and protein levels. In addition, the serum and myocardial tissue expression levels of inflammatory-related factors, such as IL-6, IL-1ß, and TNF-α, were also reduced by the administration of AAV9-SOCS3 siRNA compared with the AAV9-siRNA control. CONCLUSIONS: SOCS3 gene silencing by AAV9-SOCS3 siRNA administration in a DHF rat model significantly reduced myocardial fibrosis and the inflammatory response and improved heart function. Therefore, this treatment is a potential therapeutic method for treating DHF.

SOCS3 Suppression Promoted the Recruitment of CD11b+Gr-1-F4/80-MHCII- Early-Stage Myeloid-Derived Suppressor Cells and Accelerated Interleukin-6-Related Tumor Invasion via Affecting Myeloid Differentiation in Breast Cancer.

Interleukin-6 (IL-6) is an important trigger for the expansion and recruitment of myeloid-derived suppressor cells (MDSCs), which are regarded to be major coordinators of the immunosuppressive tumor microenvironment. In this study, we constructed IL-6-knockdown breast cancer mice models to explore the molecular events involved in the IL-6-mediated effects on MDSC development. We defined a subset of early-stage MDSCs (e-MDSCs) with the phenotype of CD11b+Gr-1-F4/80-MHCII- in IL-6 high-expressing 4T1 mice mammary carcinoma models, which were the precursors of CD11b+Gr-1+ conventional MDSCs. Furthermore, sustained suppression of SOCS3 and aberrant hyperactivation of the JAK/STAT signaling pathway was exclusively detected in wide-type 4T1 tumor-bearing mice, which promoted the accumulation of e-MDSCs in situ and their immunosuppressive capability in vitro. After blocking the IL-6/STAT3 signaling pathway with the IL-6 receptor antibody or STAT3 antagonist JSI-124 in tumor-bearing mice, significant shrinkage of primary tumors and decrease in lung metastatic nodules were observed in vivo, accompanied by the dramatic decrease of e-MDSC recruitment and recovery of anti-tumor T cell immunity. Thus, SOCS3 suppression accelerated the IL-6-mediated growth and metastasis of mammary carcinoma via affecting myeloid differentiation in breast cancer. Moreover, the IL-6/STAT3 signaling pathway might be a promising candidate target in developing novel therapeutic strategies to eliminate e-MDSCs and improve breast cancer prognosis.

SOCS3 regulates mucus hypersecretion via JAK/STAT signal pathway / 安徽医科大学学报

Objective To investigate the SOCS3 regulates mucus hypersecretion via JAK/STAT signal pathway in inflammatory cells.Methods Culture 16HBE cells and divide into three groups:control group,inter leukin(IL)-6-exposed group;IL-6-exposed and microRNA-203-transfered group.The protein levels of JAK1/2,SOCS3 and MUC5AC were measured by Western blot.The mRNA expressions of SOCS3,STAT3 and MUC5AC were detected by Real-time PCR.The protein levels of p-JAK1/2,SOCS3 and MUC5AC were analyzed by ELISA.Results Compared with the control group,the mRNA expressions of SOCS3,STAT3,MUC5AC and the protein levels of p-JAK1/2,SOCS3,MUC5AC were both significantly increased in IL-6-exposed group (P ＜ 0.05);in the IL-6-exposed and miR-203-transfered group,the protein levels of p-JAK1/2,MUC5AC and the mRNA expressions of STAT3 were both significantly increased (P ＜ 0.05),but the mRNA expressions of SOCS3 was almost as much as that in IL-6-exposed group,the protein levels of SOCS3 was significantly decreased (P ＜ 0.05).Conclusion SOCS3 over-express when cells are stimulated by IL-6,and negative-feedback regulates MUC5AC secretion via JAK/STAT signal pathway.

Rifampicin attenuates experimental autoimmune encephalomyelitis by inhibiting pathogenic Th17 cells responses.

Rifampicin, a broad-spectrum antibiotic, has neuroprotective, immunosuppressive, and anti-inflammatory properties. However, the effect of rifampicin on autoimmune disorders of the nervous system is not clear. In this study, we investigated whether rifampicin was beneficial to myelin oligodendrocyte glycoprotein peptide (MOG33-35 )-induced female C57BL/6 experimental autoimmune encephalomyelitis (EAE) mice, the well-established animal model of multiple sclerosis. Rifampicin treatment (daily from the first day after EAE immunization) remarkably attenuated clinical signs and loss of body weight, which are associated with suppression of inflammatory infiltration and demyelination in spinal cords of EAE mice. Furthermore, rifampicin dramatically reduced the disruption of blood-brain barrier integrity, down-regulated serum concentration of IL-6 and IL-17A, inhibited pathological Th17 cell differentiation, and modulated the expression of p-STAT3 and p-p65. These results suggest that rifampicin is effective for attenuating the clinical severity of EAE mice, which may be related to its inhibitive ability in differentiation of Th17 cell and secretion of its key effector molecule IL-17A via regulation of excessive activation of the key signaling molecules of JAK/STAT pathway. Our findings may be helpful for developing therapeutic and preventive strategies for multiple sclerosis.

Effect of IFN-CSP on gene of JAK-STAT signaling pathway in HBV transgenic mice / 中国生化药物杂志

Objective To investigate the anti-HBV molecular mechanisms of liver targeting interferon ( IFN-CSP ) in Balb/c-HBV transgenic mice.Methods Balb/c-HBV transgenic mice were randomly divided into 3 groups.Control group (treated with physiological saline), IFN α2b group (treated with 103 U/g IFN α2b), IFN-CSP group (treated with 102 U/g IFN-CSP).Another group of the non-transgenic mice were used as the Normal group.Each mouse was intramuscular injected with 50 μL dose once a day for 4 weeks.Total RNA of mice liver were extracted, and STAT1, STAT2, IRF-9, OAS1 gene expression of JAK-STAT signaling pathway were analyzed by real-time PCR.Results IFN α2b and IFN-CSP can significantly up regulate the expression of STAT1, STAT2, IRF-9, OAS1 gene of JAK-STAT signaling pathway (P<0.01).The induce effects of IFN-CSP on STAT1, STAT2, IRF-9, OAS1 were significantly better than that of IFN α2b (P<0.05).Conclusion The anti-HBV molecular mechanisms of liver targeting interferon (IFN-CSP) in Balb/c-HBV transgenic mice maybe related to regulate the expression of STAT1, STAT2, IRF-9, OAS1 gene of JAK-STAT signaling pathway.These results will lay a basis for the application of recombinant liver-targeting interferon.

Effects of TSA on Cerebral ischemia/reperfusion injury via JAK/STAT signal pathway in rats / 中华急诊医学杂志

Objective To investigate the protective effect of trichostatin-A (TSA) on cerebral ischemia/reperfusion injury via Janus kinase/signal transducer and activator of transcription (JAK/STAT) signal pathway.Methods 36 male SD rats were randomly (random number) divided into 3 groups:shamoperated group,ischemia/reperfusion (I/R) group and TSA group.Rat model of middle cerebral artery occlusion/reperfusion (MCAO) was established using a modified filament method.No occlusion was applicated to the sham-operated group.TSA group was injected with TSA 0.05 mg/kg via penile vein,20 minutes before operation.Reperfusion was carried out 24 hours after modeling.Longa 5 score was used to assess the neurological function,and TTC staining was applied to calculate the percentage of cerebral infarction area,The expression of JAK2 and p-JAK2 proteins was detected by Elisa.Results The low expression of JAK2 was observed in each group,and there was no statistical difference between groups (P =0.266).Compared with I/R group,TSA group had lower score in cerebral ischemia-reperfusion injury assessment (P=0.019),smaller area of cerebral infarction (P ＜0.01),reduced expression of p-JAK2 (P =0.009),all of which were of significant difference.Condusions TSA can reduce the cerebral ischemia/reperfusion injury via JAK/STAT signal pathway by down regulating p-JAK2 expression.

Variable interference with interferon signal transduction by different strains of porcine reproductive and respiratory syndrome virus.

Porcine reproductive and respiratory syndrome virus (PRRSV) is a positive-sense single-strand RNA virus that has contributed to substantial losses to the swine industry. The objective of this study was to examine the interference of interferon (IFN)-activated signaling by PRRSV viral proteins and compare the effects of several PRRSV strains on the IFN signaling. Three non-structural proteins (nsp 1ß, 7 and 12) and two structural proteins (GP3 and N) of VR-2385 were found to significantly inhibit the expression of IFN-stimulated response element (ISRE) luciferase reporter. Nsp1ß and N showed robust inhibition and their-encoding sequences from the six PRRSV strains (VR-2385, Ingelvac PRRS MLV, VR-2332, NVSL97-7895, MN184, and Lelystad) were cloned for further characterization. The nsp1ßs of all the strains except MLV inhibited expression of IFN-induced ISRE reporter, interferon-stimulated gene 56 (ISG56) and signal transducer and activator of transcription 2 (STAT2). The N proteins inhibited the IFN-induced ISRE reporter expression and STAT2 elevation and blocked nuclear translocation of STAT1. In MARC-145 cells, all the six PRRSV strains with the exception of MN184 blocked the activity of exogenous IFN-α. In primary porcine pulmonary alveolar macrophages (PAMs), only MLV and NVSL failed to inhibit the activity of IFN-α. These results indicate that some PRRSV strains, such as NVSL and MN184, have variable effects on IFN-activated signaling in the two types of cells.