Untoward effector CD4+ T cell responses are kept in check by immune regulatory mechanisms mediated by CD4+ and CD8+ T cells. CD4+ T helper 17 (Th17) cells, characterized by IL-17 production, play important roles in the pathogenesis of autoimmune diseases (such as arthritis, multiple sclerosis, psoriasis, inflammatory bowel disease, among others) and in the host response to infection and cancer. Here, we demonstrate that human CD4+ T cells cells exposed to a Th17-differentiating milieu are significantly more resistant to immune suppression by CD8+ T cells compared to control Th0 cells. This resistance is mediated, in part, through the action of IL-17A, IL-17F, and IL-17AF heterodimer through their receptors (IL-17RA and IL-17RC) on CD4+ T cells themselves, but not through their action on CD8+ T cells or APC. We further show that IL-17 can directly act on non-Th17 effector CD4+ T cells to induce suppressive resistance, and this resistance can be reversed by blockade of IL-1β, IL-6, or STAT3. These studies reveal a role for IL-17 cytokines in mediating CD4-intrinsic immune resistance. The pathways induced in this process may serve as a critical target for future investigation and immunotherapeutic intervention.

Resident macrophages in the tumor microenvironment exert a dual role in tumor progression. So far, the mechanism of intratumoral macrophage generation is still largely unknown. In the present study, the importance of macrophages in the pro-tumor role of gastric cancer-derived mesenchymal stromal cells (GC-MSCs) was observed in a mouse xenograft model with macrophage depletion. In gastric cancer tissues, high expression levels of Ym-1, Fizz-1, arginase-1, and CCR-2, as well as a low expression level of iNOS, were verified, and co-localization of GC-MSCs and tumor-associated macrophages (TAMs) was observed by dual immunofluorescence histochemistry. TAMs isolated from gastric cancer tissues predominantly displayed an M2 phenotype. In a co-culture system, the contribution of GC-MSCs to M2 polarization of macrophages was confirmed by the M2-related protein expression, M2-like immunophenotype and cytokine profile of GC-MSC-primed macrophages in vitro. Blockade of IL-6/IL-8 by neutralizing antibodies significantly attenuated the promoting effect of GC-MSCs on M2-like macrophage polarization via the JAK2/STAT3 signaling pathway. In addition, GC-MSC-primed macrophages promoted the migration and invasion of gastric cancer cells, and the process of EMT in gastric cancer cells was significantly enhanced by GC-MSC-primed macrophage treatment. Our study showed that tumor-promoting GC-MSCs contribute to M2 macrophage polarization within the gastric cancer niche through considerable secretion of IL-6 and IL-8. These GC-MSC-primed macrophages can subsequently prompt gastric cancer metastasis via EMT promotion in gastric cancer cells.

This study aimed to investigate the relationship between interleukin-6 (IL-6) and NS5ATP9 in autophagy of liver cancer cells. Autophagy is one of the important regulators of the replication of hepatitis C virus and the survival of tumors. IL-6 is a multifunctional cytokine that plays an important role in autophagy and development of many kinds of tumors. However, the role of IL-6 in autophagy has not been fully explored. A previous study had shown that a novel gene, NS5ATP9, could modulate autophagy. The present study demonstrated that human IL-6 recombinant protein induced autophagy of HepG2 cells. Conversely, autophagy decreased after IL-6 was silenced or neutralized with monoclonal antibody against human IL-6. In addition, NS5ATP9 was upregulated by IL-6 via nuclear factor-kappaB activation, as detected by Western blot. Further studies indicated that the induction of autophagy by IL-6 could be attenuated by silencing NS5ATP9. Interestingly, the expression of NS5ATP9, in turn, resulted in the upregulation of IL-6. In conclusion, IL-6 could induce autophagy by expressing NS5ATP9, while NS5ATP9 upregulated IL-6 levels in turn, which further induced autophagy.
© 2017 Wiley Periodicals, Inc.

Medical research is developing an ever greater need for comprehensive high-quality data generation to realize the promises of personalized health care based on molecular biomarkers. The nucleic acid proximity-based methods proximity ligation and proximity extension assays have, with their dual reporters, shown potential to relieve the shortcomings of antibodies and their inherent cross-reactivity in multiplex protein quantification applications. The aim of the present study was to develop a robust 96-plex immunoassay based on the proximity extension assay (PEA) for improved high throughput detection of protein biomarkers. This was enabled by: (1) a modified design leading to a reduced number of pipetting steps compared to the existing PEA protocol, as well as improved intra-assay precision; (2) a new enzymatic system that uses a hyper-thermostabile enzyme, Pwo, for uniting the two probes allowing for room temperature addition of all reagents and improved the sensitivity; (3) introduction of an inter-plate control and a new normalization procedure leading to improved inter-assay precision (reproducibility). The multiplex proximity extension assay was found to perform well in complex samples, such as serum and plasma, and also in xenografted mice and resuspended dried blood spots, consuming only 1 µL sample per test. All-in-all, the development of the current multiplex technique is a step toward robust high throughput protein marker discovery and research.