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Substance P induces inflammatory responses involving NF-?B in genetically diabetic mice skin fibroblasts co-cultured with macrophages.

ABSTRACT: PURPOSE:Delayed wound healing is an intractable complex of diabetes and substance P (SP) is proved to benefit wound healing, whose functioning mechanism remains elusive. This study aims at revealing whether the influence of SP on diabetic wound healing is dependent on inflammatory responses, particularly NF-?B. METHODS:Skin fibroblasts of genetically diabetic mice were co-cultured with bone marrow-derived macrophages, and treated with SP, SP + L703,606 (a neurokinin-1 receptor antagonist), or SP + MG132 (an inhibitor of NF-?B). For macrophages, their migration ability was assessed by Transwell experiments, and their M2 polarization was analyzed by flow cytometry and markers for M2 phenotype. Pro-inflammatory factors in the supernatant were detected by enzyme-linked immunosorbent assay. In fibroblasts, the transcription levels of the four pro-inflammatory factors and the protein levels of NF-?B regulators like inhibitor of NF-?B alpha (I?B?) and I?B kinases (IKKs) were monitored by real-time quantitative PCR and western blot, respectively. RESULTS:SP could significantly induce migration to fibroblasts (P<0.01), M2 polarization (P<0.001) and pro-inflammatory factor concentration (P<0.01) in the co-culture system. It also promotes the transcription process of pro-inflammatory factors in fibroblasts (P<0.01), and induce activation of IKK?/? and phosphorylation of I?B?, which caused NF-?B activation. All these effects were reversed if NF-?B was inhibited. CONCLUSION:The promoting effects of SP on diabetic wound healing was dependent on enhanced inflammatory responses, especially the activation of NF-?B. This study provided evidence for the potential usage of SP in accelerating diabetic wound healing.

SUBMITTER: Ni T

PROVIDER: S-EPMC4891430 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Substance P induces inflammatory responses involving NF-κB in genetically diabetic mice skin fibroblasts co-cultured with macrophages.

Ni Tao T Liu Yushu Y Peng Yinbo Y Li Ming M Fang Yong Y Yao Min M

American journal of translational research 20160515 5

<h4>Purpose</h4>Delayed wound healing is an intractable complex of diabetes and substance P (SP) is proved to benefit wound healing, whose functioning mechanism remains elusive. This study aims at revealing whether the influence of SP on diabetic wound healing is dependent on inflammatory responses, particularly NF-κB.<h4>Methods</h4>Skin fibroblasts of genetically diabetic mice were co-cultured with bone marrow-derived macrophages, and treated with SP, SP + L703,606 (a neurokinin-1 receptor ant ...[more]

PMID: 27347325

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Project description:BackgroundOvarian cancer has the highest mortality among gynecological cancers due to late diagnosis and lack of effective targeted therapy. Although the study of interplay between cancer cells with their microenvironment is emerging, how ovarian cancer triggers signaling that coordinates with immune cells to promote metastasis is still elusive.MethodsMicroarray and bioinformatics analysis of low and highly invasive ovarian cancer cell lines were used to reveal periostin (POSTN), a matrix protein with multifunctions in cancer, with elevated expression in the highly invasive cells. Anchorage independent assay, Western blot, RNA interference, confocal analysis and neutralizing antibody treatment were performed to analyze the effects of POSTN on tumor promotion and to explore the underlying mechanism. Chemotaxis, flow cytometry and cytokine array analyses were undertaken to analyze the involvement of POSTN in cancer-associated fibroblast (CAF) and macrophage modulation. Correlations between POSTN expression levels and clinical characteristics were analyzed using the Oncomine, commercial ovarian cancer cDNA and China Medical University Hospital patient cohort. In vivo effect of POSTN on metastasis was studied using a mouse xenograft model.ResultsExpression of POSTN was found to be elevated in highly invasive ovarian cancer cells. We observed that POSTN was co-localized with integrin β3 and integrin β5, which was important for POSTN-mediated activation of ERK and NF-κB. Ectopic expression of POSTN enhanced whereas knockdown of POSTN decreased cancer cell migration and invasion in vitro, as well as tumor growth and metastasis in vivo. POSTN enhanced integrin/ERK/NF-κB signaling through an autocrine effect on cancer cells to produce macrophage attracting and mobilizing cytokines including MIP-1β, MCP-1, TNFα and RANTES resulting in increased chemotaxis of THP-1 monocytes and their polarization to M2 macrophages in vitro. In agreement, tumors derived from POSTN-overexpressing SKOV3 harbored more tumor-associated macrophages than the control tumors. POSTN induced TGF-β2 expression from ovarian cancer cells to promote activation of adipose-derived stromal cells to become CAF-like cells expressing alpha smooth muscle actin and fibroblast activation protein alpha. Consistently, increased CAFs were observed in POSTN overexpressing SKOV3 cells-derived metastatic tumors. In clinical relevance, we found that expression of POSTN was positively correlated with advanced-stage diseases and poor overall survival of patients.ConclusionsOur study revealed a POSTN-integrin-NF-κB-mediated signaling and its involvement in enhancing M2 macrophages and CAFs, which could potentially participate in promoting tumor growth. Our results suggest that POSTN could be a useful prognosis marker and potential therapeutic target.

Dataset Information

Substance P induces inflammatory responses involving NF-?B in genetically diabetic mice skin fibroblasts co-cultured with macrophages.

Publications

Substance P induces inflammatory responses involving NF-κB in genetically diabetic mice skin fibroblasts co-cultured with macrophages.

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