stat1 signaling
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2022 ◽  
Vol 12 ◽  
Author(s):  
Sajid Ur Rahman ◽  
Haiyan Gong ◽  
Rongsheng Mi ◽  
Yan Huang ◽  
Xiangan Han ◽  
...  

Cryptosporidium parvum infection is very common in infants, immunocompromised patients, or in young ruminants, and chitosan supplementation exhibits beneficial effects against the infection caused by C. parvum. This study investigated whether chitosan supplementation modulates the gut microbiota and mediates the TLR4/STAT1 signaling pathways and related cytokines to attenuate C. parvum infection in immunosuppressed mice. Immunosuppressed C57BL/6 mice were divided into five treatment groups. The unchallenged mice received a basal diet (control), and three groups of mice challenged with 1 × 106 C. parvum received a basal diet, a diet supplemented with 50 mg/kg/day paromomycin, and 1 mg/kg/day chitosan, and unchallenged mice treated with 1 mg/kg/day chitosan. Chitosan supplementation regulated serum biochemical indices and significantly (p < 0.01) reduced C. parvum oocyst excretion in infected mice treated with chitosan compared with the infected mice that received no treatment. Chitosan-fed infected mice showed significantly (p < 0.01) decreased mRNA expression levels of interferon-gamma (IFN-γ) and tumor necrosis factor-α (TNF-α) compared to infected mice that received no treatment. Chitosan significantly inhibited TLR4 and upregulated STAT1 protein expression (p < 0.01) in C. parvum-infected mice. 16S rRNA sequencing analysis revealed that chitosan supplementation increased the relative abundance of Bacteroidetes/Bacteroides, while that of Proteobacteria, Tenericutes, Defferribacteres, and Firmicutes decreased (p < 0.05). Overall, the findings revealed that chitosan supplementation can ameliorate C. parvum infection by remodeling the composition of the gut microbiota of mice, leading to mediated STAT1/TLR4 up- and downregulation and decreased production of IFN-γ and TNF-α, and these changes resulted in better resolution and control of C. parvum infection.


2021 ◽  
Author(s):  
Michael Schubert ◽  
Christy Hong ◽  
Laura J. Jilderda ◽  
Marta Requesens Rueda ◽  
Andréa E. Tijhuis ◽  
...  

AbstractChromosomal instability is a hallmark of cancer, but also an instigator of aneuploidy-induced stress, reducing cellular fitness. To better understand how cells with CIN adjust to aneuploidy and adopt a malignant fate in vivo, we performed a genome-wide mutagenesis screen in mice. We find that specifically aneuploid tumors inactivate Stat1 signaling in combination with increased Myc activity. By contrast, loss of p53 is common, but not enriched in CIN tumors. Validation in another tissue type confirmed that CIN promotes immune cell infiltration, which is alleviated by Stat1 loss combined with Myc activation, but not with p53 inactivation, or Myc activation alone. Importantly, we find that this mechanism is preserved in human aneuploid cancers. We conclude that aneuploid cancers inactivate Stat1 signaling to circumvent immune surveillance.


2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Dan Qiu ◽  
Xianxin Yan ◽  
Xinqin Xiao ◽  
Guijuan Zhang ◽  
Yanqiu Wang ◽  
...  

Abstract Background The precancerous disease of breast cancer is an inevitable stage in the tumorigenesis and development of breast neoplasms. Quercetin (Que) has shown great potential in breast cancer treatment by inhibiting cell proliferation and regulating T cell function. γδ T cells are a class of nontraditional T cells that have long attracted attention due to their potential in immunotherapy. In this study, we revealed the immunomodulatory function of Que through regulation of the JAK/STAT1 signaling pathway, which was followed by the synergistic killing of breast cancer cells. Methods In the experimental design, we first screened target genes with or without Que treatment, and we intersected the Que target with the disease target by functional enrichment analysis. Second, MCF-10A, MCF-10AT, MCF-7 and MDA-MB-231 breast cancer cell lines were treated with Que for 0 h, 24 h and 48 h. Then, we observed the expression of its subsets by coculturing Que and γδ T cells and coculturing Que and γδ T cells with breast tumor cells to investigate their synergistic killing effect on tumor cells. Finally, Western blotting was used to reveal the changes in proteins related to the JAK/STAT1 signaling pathway after Que treatment in MCF-10AT and MCF-7 cells for 48 h. Results The pathway affected by Que treatment was the JAK/STAT1 signaling pathway and was associated with precancerous breast cancer, as shown by network pharmacology analysis. Que induced apoptosis of MCF-10AT, MCF-7 and MDA-MB-231 cells in a time- and concentration-dependent manner (P < 0.05). Most importantly, Que promoted the differentiation of γδ T cells into the Vδ2 T cell subpopulation. The best ratio of effector cells to target cells (E/T) was 10:1, the killing percentages of γδ T cells against MCF-10A, MCF-10AT, MCF-7, and MDA-MB-231 were 61.44 ± 4.70, 55.52 ± 3.10, 53.94 ± 2.74, and 53.28 ± 1.73 (P = 0.114, P = 0.486, and P = 0.343, respectively), and the strongest killing effect on precancerous breast cancer cells and breast cancer cells was found when the Que concentration was 5 μM and the E/T ratio was 10:1 (64.94 ± 3.61, 64.96 ± 5.45, 55.59 ± 5.98, and 59.04 ± 5.67, respectively). In addition, our results showed that Que increased the protein levels of IFNγ-R, p-JAK2 and p-STAT1 while decreasing the protein levels of PD-L1 (P < 0.0001). Conclusions In conclusion, Que plays a synergistic role in killing breast cancer cells and promoting apoptosis by regulating the expression of IFNγ-R, p-JAK2, p-STAT1 and PD-L1 in the JAK/STAT1 signaling pathway and promoting the regulation of γδ T cells. Que may be a potential drug for the prevention of precancerous breast cancer and adjuvant treatment of breast cancer.


Author(s):  
Zhenwei Li ◽  
Qingqing Xu ◽  
Ning Huangfu ◽  
Xiaomin Chen ◽  
Jianhua Zhu
Keyword(s):  

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 1080-1080
Author(s):  
Hannah Yan ◽  
Forrest C Walker ◽  
Hyojeong Han ◽  
Megan T Baldridge ◽  
Katherine Y. King

Abstract Long-term antibiotic therapy is associated with hematological side effects such as neutropenia and anemia. Our lab and others have shown that long-term antibiotic treatment in mice leads to bone marrow suppression and agranulocytosis in mice through depletion of the commensal bacteria. Our work further showed that Stat1-deficient mice phenocopy the bone marrow suppression phenotype of antibiotic-treated mice, suggesting that commensal microbiota mediate hematopoiesis via Stat1 signaling. However, the upstream actors of this pathway and the bacterial mediators required for commensal microbiota regulation of normal hematopoiesis still remain poorly understood. Such knowledge will be essential for understanding how to treat antibiotic-associated cytopenias. We hypothesize that microbial products detected by host cells trigger STAT1 signaling to potentiate normal hematopoiesis. To identify the host cells that require Stat1 for microbiota-promoted hematopoiesis, we treated conditional Stat1 knock-out mice with two weeks of antibiotic therapy. Of the four conditional knock-out mice we evaluated (LepR-Cre, Villin-Cre, Vav-iCre, LysM-Cre), only the mice deficient in STAT1 in hematopoietic cells (Vav-iCre Stat1 fl/fl) phenocopied the bone marrow suppression of antibiotic-treated mice. Our data suggest that STAT1 signaling is necessary in non-myeloid hematopoietic cells, but not intestinal epithelial cells or mesenchymal stromal cells for microbiota-promoted hematopoiesis. Non-competitive transplantation of Stat1 -/- bone marrow into wild type mice validated these findings; mice lacking STAT1 only in hematopoietic tissues phenocopied the Vav-iCre mice, consistent with a specific role for microbiota-mediated STAT1 signaling in the hematopoietic compartment. To assess the upstream mediator of STAT1 signaling in this biological context, we treated interferon (IFN) receptor knock out mice with two weeks of antibiotics. Of the three types of IFN receptor knock-outs evaluated, only mice deficient in type I IFN signaling phenocopied the bone marrow suppression of antibiotic-treated mice. These findings suggest that type I IFN signaling, and not type II or III, was required for microbiota-dependent hematopoiesis. These results were validated by showing that the administration of pegylated-IFNα was sufficient to rescue the depletion of type I IFN-STAT1 signaling in antibiotic-treated mice. To determine the microbial signals that may potentiate hematopoiesis, we evaluated two microbial products that were previously shown to alter hematopoiesis and to activate type I IFN signaling (Iwamura et al. Blood 2017 & Steed et al. Science 2017). We discovered that oral administration of these commensal-derived products, the metabolite desaminotyrosine (DAT) or NOD1 ligand (NOD1L), a motif of peptidoglycan, were each sufficient to rescue the hematopoietic defects induced by antibiotics in mice. To test whether these products rescue hematopoiesis by activating STAT1 signaling, we attempted to rescue the hematopoietic defects in Stat1 -/- mice. These studies showed that NOD1L rescues granulocyte but not progenitor counts in Stat1 -/- mice, suggesting that NOD1 and type I IFN signaling work together at the progenitor level, but independently at the downstream myeloid progenitor level to promote granulopoiesis. Overall, our studies expand our understanding of the signaling pathways by which the microbiota promotes normal hematopoiesis and identify novel therapeutic agents that can be used to ameliorate antibiotic-induced BM suppression. Disclosures No relevant conflicts of interest to declare.


Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 86-86
Author(s):  
Shuichiro Takashima ◽  
Roshan Sharma ◽  
Anastasiya Egorova ◽  
Jason Kuttiyara ◽  
Takahiro Ito ◽  
...  

Abstract Crypt apoptosis and regeneration are characteristic findings in GI-GVHD. Intestinal stem cells (ISCs) are critical for maintaining the intestinal epithelium, but their frequencies are reduced in experimental GVHD, and the mechanisms driving crypt regeneration in this context are poorly understood. To better understand the impact of GVHD on individual epithelial components, we performed single cell RNA sequencing (scRNAseq) of purified small intestine crypts from B6 mice during homeostasis and five days after B6 into B6 syngeneic (syn) or B10.Br into B6 allogeneic (allo) BMT. Sequenced cells were first partitioned into distinct clusters using PhenoGraph, and the cluster identities were annotated based on their gene expression profiles. Unsupervised clustering indicated multiple subpopulations within the various recognized crypt components (Fig 1A). While secretory lineages clustered similarly across experimental conditions, there was highly distinct clustering among ISC populations and striking dissimilarity between allo and syn ISCs. Quantification by computing the subpopulation's phenotypic distance, a composite score integrating both the number and amplitude of differentially expressed genes, indicated that ISCs demonstrated the greatest transcriptional change in response to GVHD among all crypt lineages (Fig 1B). Gene Set Enrichment Analysis (GSEA) highlighted activation of the interferon-γ (IFNγ) pathway in allo ISCs, and differential gene expression indicated that STAT1 was their most highly upregulated transcription factor. We then performed MHC-mismatched allo-BMT into STAT1-floxed x villin-Cre recipients. Consistent with a role in IFNγ-related toxicity, STAT1-deficient recipients initially demonstrated reduced GVHD pathology, as well as a reduction in proliferating Ki67 + cells (Fig 2A). However, the pathology reduction in STAT1-deficient recipients was transient, while reduction in crypt proliferation persisted. STAT1-deficient recipients ultimately demonstrated increased mortality after allo-BMT, indicating a complex response to epithelial STAT1 signaling in GVHD. While IFNγ can induce epithelial apoptosis and kill organoid cultures, organoid exposure to IFNγ augmented size even at IFNγ concentrations that did not impair viability in a STAT1-dependent manner, and co-culture with allo T cells augmented organoid size (Fig 2B, C). Moreover, cell cycle analysis showed augmentation of cell cycle in ISCs after IFNγ treatment in association with upregulation of cyclin D1 (Fig 2D), and human organoids also showed increased size in response to IFNγ treatment, further suggesting that this growth promotion was not simply a secondary response to tissue injury. In addition to the IFNγ signaling, GSEA of allo ISCs indicated activation of the Myc pathway. scRNAseq data showed specific upregulation of Myc in allo ISCs (Fig 3A). Myc expression within individual ISCs indicated that greater STAT1 expression and IFNγ signaling directly correlated with Myc expression in the same ISCs, providing a potential direct link between T-cell-derived IFNγ and ISC-dependent regeneration (Fig 3B). Additionally, intestinal organoid qPCR showed that Myc expression was upregulated after IFNγ treatment, and scRNAseq of IFNγ-treated organoids indicated this Myc upregulation was restricted to the ISC/progenitor compartment. Although Myc is downstream of Wnt signaling, which is critical for ISC maintenance, expression of the representative Wnt target gene Axin2 was downregulated after IFNγ treatment, and Irf1, a representative IFNγ/STAT1 target gene, was upregulated, suggesting that IFNγ/STAT1 signaling could replace Wnt/β-catenin as a driver of ISC Myc expression. We next examined Myc function and found that treatment with the Myc inhibitor 10058-F4 suppressed IFNγ-dependent organoid growth (Fig 3C). Finally, immunofluorescent staining showed Myc protein expression in intestinal crypts after allo-BMT in a STAT1-dependent manner (Fig 3D), and Myc inhibitor treatment in vivo suppressed crypt epithelial proliferation in mice with GVHD. In summary, we found that epithelial STAT1 contributes to crypt regeneration in GVHD by transmitting T-cell-derived JAK/STAT cytokine signaling to activate Myc expression in ISCs. Clinical use of JAK inhibitors in GVHD may inhibit this regenerative response, necessitating concurrent interventions aimed at restoring it. Figure 1 Figure 1. Disclosures Hanash: Evive Biotech: Ended employment in the past 24 months.


2021 ◽  
Vol 23 (Supplement_6) ◽  
pp. vi26-vi26
Author(s):  
Sabbir Khan ◽  
Rajasekaran Mahalingam ◽  
Shayak Sen ◽  
Kaitlin Gandy ◽  
Kristin Alfaro-Munoz ◽  
...  

Abstract Interferon (IFN) signaling contributes to stemness, cell proliferation, cell death, and cytokine signaling in cancer and immune cells; however, the role of IFN signaling in glioblastoma (GBM) and GBM stem-like cells (GSCs) is unclear. This study aimed to investigate the cancer cell-intrinsic IFN signaling in tumorigenesis and malignant phenotype of GBM. We characterized cell-intrinsic IFN signaling in The Cancer Genome Atlas, patient-derived cohorts of GSCs, and published single-cell RNA sequencing datasets by in-silico analyses. The in-silico findings were further validated by evaluating the cytokine secretion and using pharmacological activators and blockers of IFN/transducer and activator of transcription 1 (STAT1) signaling. We found that GSCs and GBM tumors exhibited differential cell-intrinsic IFN signaling, and high IFN/STAT1 signaling is associated with mesenchymal phenotype and poor survival outcomes. Ruxolitinib, a pharmacological inhibitor of IFN/STAT1, abolished the IFN/STAT1 signaling in GSCs with intrinsically high IFN signaling. IFN-γ treatment for 1 week promotes the mesenchymal phenotype in GSCs with low IFN signature. In addition, chronic inhibition of IFN/STAT1 signaling with ruxolitinib decreased cell proliferation and mesenchymal signatures (CD44, YKL40, and TIMP1) in GSCs with intrinsically active IFN/STAT1 signaling. Publicly available human glioma single-cell RNA-seq (scRNA-seq) datasets analyses showed that both tumor and nontumor cells expressed IFN signaling genes, and the mesenchymal signature was highly expressed in the same cluster where IFN signaling genes were upregulated. We demonstrated that cell-intrinsic IFN signaling in GSCs and GBM tumors is associated with mesenchymal signatures and cell proliferation. Our study provides evidence for the possibility of targeting IFN signaling in a specific group of GBM patients.


2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A774-A774
Author(s):  
Kristin DePeaux ◽  
Dayana Rivadeneira ◽  
McLane Watson ◽  
Andrew Hinck ◽  
Stephen Thorne ◽  
...  

BackgroundOncolytic viruses are an underappreciated immunotherapy capable of inflaming the tumor microenvironment (TME), vaccinating a patient against their own tumor, and delivering gene therapy to the TME. However, apart from the oncolytic HSV T-vec, these therapies have not seen widespread use, due in part to incomplete understanding of their immunologic mechanisms of action. We sought to determine features of oncolytic vaccinia virus (VV) response and resistance using subclones of the HPV+ head and neck cancer model MEER rendered sensitive or resistant to VV.MethodsA VV sensitive MEER tumor resisting treatment was serially passaged in mice and treated with VV until a stably resistant line was generated (Fig1). Sensitive or resistant MEER tumors were implanted, treated with a single intratumoral dose of VV, and harvested 4–7 days later for cytometric analysis. A genetically encoded TGFβ inhibitor was recombined into oncolytic VV (VV-TGFβi).ResultsWe used serial in vivo passaging to generate a VV-resistant MEER line (MEERvvR) from one sensitive to VV (MEERvvS, figure 1) and compared their immune infiltrate. While VV promoted acute cytokine production and cytotoxicity in conventional T cells, the major determining factor between sensitivity and resistance was the phenotype of Treg cells. At baseline, Treg cells in MEERvvS had lower Nrp1 expression and higher IFNγ-STAT1 signaling compared to MEERvvR, indicative of Treg 'fragility'. VV treatment induced MEERvvS Treg cells to become immunostimulatory and produce IFNγ (figure 2). RNAseq revealed MEERvvR produced more TGFβ than MEERvvS cells, suggesting these tumors directly stabilize Treg cells. To determine if MEERvvR could be sensitized to VV, we engineered oncolytic vaccinia to produce a genetically-encoded TGFβ inhibitor which binds TGFβRII, preventing TGFβ1-3 binding (VV-TGFβi). When MEERvvR were treated with VV-TGFβi, elite responses were restored, with commensurate increase in survival (figure 3) associated with increased STAT1 signaling in Treg cells.ConclusionsResistance to oncolytic vaccinia is controlled by Treg cell phenotype; tumors harboring more fragile Treg cells respond exquisitely to VV. An oncolytic vaccinia engineered to produce a novel TGFβi could remodel the TME to be less supportive of Tregs, rendering resistant tumors sensitive to VV. Our data highlight the importance of Treg cell status in resistance to oncolytic virus therapy and suggest TGFβ can be effectively targeted through an inhibitor encoded within the virus. Importantly, this TME directed production of the TGFβi carries no toxicity previously associated with systemic TGFβ inhibition, suggesting a viral approach to TGFβ inhibition can be an effective strategy support broader immunotherapy response.Abstract 743 Figure 1Strategy used to generate a vaccinia resistant MEER (MEERvvR) from vaccinia sensitive MEER (MEERvvS)Abstract 743 Figure 2IFNγ production in Treg cells in MEERvvS and MEERvvR after treatment with PBS or control vaccinia (VV-Ctrl)Abstract 743 Figure 3Survival of VV-resistant MEER treated with PBS, control vaccinia (VV-Ctrl), or vaccinia engineered to deliver a potent inhibitor of TGFβ (VV-TGFβi)


2021 ◽  
Vol 27 (5) ◽  
pp. 1286-1292
Author(s):  
Il-Joo Jo ◽  
Mee-Ok Choi

Kyungohkgo (KOG) is an oriental herbal medicine that has been used for its various pharmacological effects, which include anti-oxidant, anti-inflammatory and immuno-regulation activities. But its effects and mechanisms of anti-atopic dermatitis (AD) have not been elucidated. HaCaT cells were pre-treated with KOG for 1 h and stimulated with tumor necrosis factor (TNF)-α and interferon (IFN)-γ (10 ng/ml each). After 24 h, cells were harvested to measure the production of reactive oxygen species (ROS) and chemokines such as regulated upon activation, normal T cell expressed and secreted (RANTES/CCL5), Thymus and activation-regulated chemokine (TARC/CCL17) and macrophage-deraived chemokine (MDC/CCL22). To investigate the regulatory mechanisms of KOG, we also assessed the phosphorylation of signal transducer and activator of transcription (STAT1) signaling pathways in HaCaT cells. Treatment of KOG decreased the ROS production and mRNA levels of RANTES, TARC, MDC with a concentration dependent manner. In addition, KOG significantly inhibited TNF-α and IFN-γ induced phosphorylation of STAT1. This could indicate that the KOG shows anti-AD activity mainly through STAT1. Thus, we propose that KOG may be a promising anti-AD skin protector, which could suggest the clinical basis for cosmetics development.


2021 ◽  
Author(s):  
Jianyuan Li ◽  
Hui Shi ◽  
Xiaoyu Liu ◽  
Yanwei Wang ◽  
Haiyan Wang ◽  
...  

Abstract I. Background: Peroxiredoxin 6 (Prdx6) is widely expressed in mammalian tissues. Our previous study demonstrated that Prdx6 was expressed in human epididymis and spermatozoa, and the protective role of Prdx6 in human spermatozoa was also reported. In this study, we demonstrate the potential role and mechanism of Prdx6 in human epididymis epithelial cells (HEECs).II. Methods and Results: Western blotting was used to measure expression levels of key proteins in the JAK / STAT signaling pathway. Digital gene expression analysis (DGE) was used to identify gene expression patterns in control HECs and in HECs after Prdx6-RNA interference (P6-RNAi). The DGE analysis identified 589 up-regulated and 314 down-regulated genes (including Prdx6) in Prdx6-RNAi (P6-RNAi) HEECs. Thirteen significantly different pathways were identified between the two groups, with the majority different expressed genes belonging to the CCL, CXCL, IL, and IFIT families. In particular, the expression levels of IL6, IL6ST, and eighteen IFN related genes were significantly increased in the condition of the down-regulated expression of Prdx6. Compared to control HEECs, the expression levels of JAK1, STAT1, phosphorylated JAK1 and STAT1 were significantly increased, while the expression levels of SOCS3 was significantly decreased in P6-RNAi HEECs. The Malondialdehyde (MDA) level and total antioxidant capacity in P6-RNAi HEECs were significantly increased and decreased compared to that of control, respectively. III. Conclusions: We speculated that knockdown of Prdx6 resulted in higher levels of ROS in HEECs, which in turn, activated the JAK1 / STAT1 signaling pathway induced by IL-6 receptor and IFN.


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