GM-CSF confers resistance to influenza by enhancing innate immune mechanisms that depend on alveolar macrophages. Pulmonary delivery of this cytokine has the potential to reduce the morbidity and mortality due to influenza virus.

Smoking is associated with increased susceptibility to tuberculosis and influenza. However, little information is available on the mechanisms underlying this increased susceptibility. Mice were left unexposed or were exposed to cigarette smoke and then infected with Mycobacterium tuberculosis by aerosol or influenza A by intranasal infection. Some mice were given a DNA vaccine encoding an immunogenic M. tuberculosis protein.Gamma interferon (IFN-␥) production by T cells from the lungs and spleens was measured. Cigarette smoke exposure inhibited the lung T-cell production of IFN-␥ during stimulation in vitro with anti-CD3, after vaccination with a construct expressing an immunogenic mycobacterial protein, and during infection with M. tuberculosis and influenza A virus in vivo. Reduced IFN-␥ production was mediated through the decreased phosphorylation of transcription factors that positively regulate IFN-␥ expression. Cigarette smoke exposure increased the bacterial burden in mice infected with M. tuberculosis and increased weight loss and mortality in mice infected with influenza virus. This study provides the first demonstration that cigarette smoke exposure directly inhibits the pulmonary T-cell response to M. tuberculosis and influenza virus in a physiologically relevant animal model, increasing susceptibility to both pathogens.

BackgroundTumor-associated macrophages (TAMs) facilitate tumor progression via establishment of an immunosuppressive tumor microenvironment (TME). However, it is poorly understood how tumor cells could functionally modulate TAMs. Our previous work indicated that tumor cell-released autophagosomes (TRAPs), a type of LC3-II+ double-membrane extracellular vesicles (EVs) was sufficient to suppress anti-tumor immune responses by inducing IL-10-producing B cells and immune suppressive neutrophils. Here, we hypothesized that TRAPs may participate in regulating macrophage polarization.MethodsTRAPs isolated from multiple murine tumor cell lines and pleural effusions or ascites of cancer patients were incubated with bone marrow-derived macrophages (BMDMs) and monocytes, respectively. Cellular phenotypes were examined by flow cytometry, ELISA and quantitative PCR. TRAPs treated BMDMs were tested for the ability to suppress T-cell proliferation in vitro, and for promotion of tumor growth in vivo. Transwell chamber and neutralization antibodies were added to ascertain the inhibitory molecules expressed on BMDMs exposed to TRAPs. Knockout mice were used to identify the receptors responsible for TRAPs-induced BMDMs polarization and the signaling mechanism was examined by western blot. Autophagy-deficient tumors were profiled for phenotypic changes of TAMs and IFN-γ secretion of T cells by flow cytometry. The phenotype of monocytes from pleural effusions or ascites of cancer patients was assessed by flow cytometry.ResultsTRAPs converted macrophages into an immunosuppressive M2-like phenotype characterized by the expression of PD-L1 and IL-10. These macrophages inhibited the proliferation of both CD4+ and CD8+ T cells in vitro, and promoted tumor growth mainly through PD-L1 in vivo. TRAPs-induced macrophage polarization was dependent on TLR4-mediated MyD88-p38-STAT3 signaling. In vivo studies indicated that disruption of autophagosome formation in B16F10 cells by silencing the autophagy gene Beclin1 resulted in a remarkable delay in tumor growth, which was associated with reduced autophagosome secretion, TAMs reprogramming and enhanced T cell activation. Moreover, the levels of LC3B+ EVs appeared to correlate significantly with up-regulation of PD-L1 and IL-10 in matched monocytes from effusions or ascites of cancer patients, and TRAPs isolated from these samples could also polarize monocytes to an M2-like phenotype with increased expression of PD-L1, CD163 and IL-10, decreased expression of HLA-DR, and T cell-suppressive function.ConclusionsThese findings suggest the TRAPs-PD-L1 axis as a major driver of immunosuppression in the TME by eliciting macrophage polarization towards an M2-like phenotype, and highlight the potential novel therapeutic approach of simultaneously targeting autophagy and PD-L1.Electronic supplementary materialThe online version of this article (10.1186/s40425-018-0452-5) contains supplementary material, which is available to authorized users.

Background/Aims: Minimal residual leukemia cells (MRLs) are difficult to eradicate through traditional treatment and therefore remain to be a major threat to the long-term survival of leukemia patients. Tumor-derived exosomes (TEXs), which carry tumor associated antigens (TAA), may be a potential cell-free tumor vaccine for the specific eradication of MRLs. However, TEXs are intended to be less immunogenic due to exosomal TGF-β1. To further optimize the efficacy of TEX-based vaccines, we investigated whether exosomes from TGF-β1 silenced leukemia cells (LEX_TGF-β1si) had an increased potential to induce a specific antitumor effect compared with non-modified exosomes. Methods: Exosomal TGF-β1 was downregulated via lentiviral shRNA silencing of TGF-β1 in leukemia cells. The characteristics of LEX_TGF-β1si were determined via electron microscopy, western blot analysis, and flow cytometry. The antitumor effect of LEX_TGF-β1si was evaluated by detecting the properties of LEX_TGF-β1si-pulsed dendritic cells (DCs), CD4⁺ T-cell proliferation, Th1 cytokine secretion, specific CTL activity, and NK cell function. Moreover, to verify the superiority of LEX_TGF-β1si immunization, LEX_TGF-β1si was subcutaneously injected into DBA/2 mice: either followed by tumor challenge or tumor bearing. Results: The lentiviral shRNA silencing of TGF-β1 in parental leukemia cells successfully downregulated the TGF-β1 level in leukemia cell derived exosomes (LEX). LEX_TGF-β1si was uptaken by DCs and was more potent in promoting DC function by upregulating the surface expression of costimulatory factors and MHC class II molecules, while inducing the secretion of IL-12p70 and TNF-α. Furthermore, immunization with LEX_TGF-β1si facilitated CD4⁺ T-cell proliferation and Th1 cytokine secretion, and stimulated stronger specific cytotoxic lymphocyte (CTL) response and nature killer (NK) cell cytotoxicity more efficiently compared to non-modified LEX. In mice models, immunization with LEX_TGF-β1si resulted in a more potent capability to inhibit tumor growth and to prolong survival, suggesting that LEX_TGF-β1si was more effective in both protective and therapeutic antitumor tests than non-modified LEX. Conclusions: These data suggested that down-regulation of exosomal TGF-β1 effectively induced potent anti-tumor immunity. Our strategy of optimizing exosome vaccine may have promising potential for leukemia immunotherapy.

Tumor-derived exosomes (TEX) can induce a specific antitumor immune response and have been developed as a promising tumor vaccine. Despite promising preclinical data, TEX exhibit relatively low efficacy and limited clinical benefit in clinical trials. In the present study, we investigated whether exosomes from the TGF-β1 silenced L1210 cells (LEX) can enhance the efficacy of DC-based vaccines. We silenced TGF-β1 in L1210 cells with a lentiviral shRNA vector and prepared the LEX. It was shown that LEX can significantly decrease TGF-β1 expression of dendritic cells (DC) and effectively promote their maturation and immune function. In addition, DC pulsed with LEX (DC) more effectively promoted CD4 T cell proliferation in vitro and Th1 cytokine secretion and induced tumor-specific CTL response. This response was higher in potency compared to that noted by the other two formulations. Moreover, DC inhibited tumor growth more efficiently than other formulations did as the preventive or therapeutic tumor vaccine. Accordingly, these findings revealed that DC induced a more potent antigen-specific anti-leukemic immunity than DC pulsed with exosomes from non-manipulated L1210 cells. This indicated that the targeting of DC by LEX may be used as a promising approach for leukemia immunotherapy.

Adolescents with type 1 diabetes face self-management challenges that make it difficult for them to achieve good glycemic control. In our population of adolescents with poorly controlled type 1 diabetes, the use of continuous glucose monitoring (CGM) improved patients’ glycemic time in range (TIR) and identified hypoglycemia more frequently than with intermittent self-monitoring of blood glucose throughout a 4-week interval. However, the adolescents were unable to synthesize this information to problem-solve or reduce the frequency of hypoglycemic events. Setting SMART (specific, measurable, achievable, relevant, and time-bound) diabetes management goals and providing intensive diabetes education and support could increase adolescents’ TIR and prevent hypoglycemia.

Background: CD4 + T cells are critical effectors of anti-tumor immunity, but how tumor cells influence CD4 + T cell effector function is not fully understood. Tumor cell-released autophagosomes (TRAPs) are being recognized as critical modulators of host anti-tumor immunity during tumor progression. Here, we explored the mechanistic aspects of TRAPs in the modulation of CD4 + T cells in the tumor microenvironment. Methods: TRAPs isolated from tumor cell lines and pleural effusions or ascites of cancer patients were incubated with CD4 + T cells to examine the function and mechanism of TRAPs in CD4 + T cell differentiation and function. TRAPs-elicited CD4 + T cells were tested for their suppression of effector T cell function, induction of regulatory B cells, and promotion of tumorigenesis and metastasis in a mouse model. Results: Heat shock protein 90α (HSP90α) on the surface of TRAPs from malignant effusions of cancer patients and tumor cell lines stimulated CD4 + T cell production of IL-6 via a TLR2-MyD88-NF-κB signal cascade. TRAPs-induced autocrine IL-6 further promoted CD4 + T cells secretion of IL-10 and IL-21 via STAT3. Notably, TRAPs-elicited CD4 + T cells inhibited CD4 + and CD8 + effector T cell function in an IL-6-and IL-10-dependent manner and induced IL-10producing regulatory B cells (Bregs) via IL-6, IL-10 and IL-21, thereby promoting tumor growth and metastasis. Consistently, inhibition of tumor autophagosome formation or IL-6 secretion by CD4 + T cells markedly retarded tumor growth. Furthermore, B cell or CD4 + T cell depletion impeded tumor growth by increasing effector T cell function. Conclusions: HSP90α on the surface of TRAPs programs the immunosuppressive functions of CD4 + T cells to promote tumor growth and metastasis. TRAPs or their membrane-bound HSP90α represent important therapeutic targets to reverse cancer-associated immunosuppression and improve immunotherapy.

Nanoparticles composed of galactosylated chitosan oligosaccharide (Gal-CSO) and adenosine triphosphate (ATP) were prepared for hepatocellular carcinoma cell-specific uptake, and the characteristics of Gal-CSO/ATP nanoparticles were evaluated. CSO/ATP nanoparticles were prepared as a control. The average diameter and zeta potential of Gal-CSO/ATP nanoparticles were 51.03 ± 3.26 nm and 30.50 ± 1.25 mV, respectively, suggesting suitable properties for a drug delivery system. Subsequently, the cytotoxicity of Gal-CSO/ATP nanoparticles were examined by the methyl tetrazolium (MTT) assay, and the half maximal inhibitory concentration (IC50) values were calculated with HepG2 (human hepatocellular carcinoma cell line) cells. The results showed that the cytotoxic effect of nanoparticles on HepG2 cells was low. In the meantime, it was also found that the Gal-CSO/ATP nanoparticles could be uptaken by HepG2 cells, due to expression of the asialoglycoprotein receptor (ASGP-R) on their surfaces. The presented results indicate that the Gal-CSO nanoparticles might be very attractive to be used as an intracellular drug delivery carrier for hepatocellular carcinoma cell targeting, thus warranting further in vivo or clinical investigations.