Absence of PD-L1 signaling hinders macrophage defense against Mycobacterium tuberculosis via upregulating STAT3/IL-6 pathway.

The blockade of programmed death-ligand 1 (PD-L1) pathway has been clinically used in cancer immunotherapy, while its effects on infectious diseases remain elusive. Roles of PD-L1 signaling in the macrophage-mediated innate immune defense against M.tb is unclear. In this study, the outcomes of tuberculosis (TB) in wild-type (WT) mice treated with anti-PD-1/PD-L1 therapy and macrophage-specific Pdl1-knockout (Pdl1ΔΜΦ) mice were compared. Treatment with anti-PD-L1 or anti-PD-1 benefited protection against M.tb infection in WT mice, while Pdl1ΔΜΦ mice exhibited the increased susceptibility to M.tb infection. Mechanistically, the absence of PD-L1 signaling impaired M.tb killing by macrophages. Furthermore, elevated STAT3 activation was found in PD-L1-deficient macrophages, leading to increased interleukin (IL)-6 production and reduced inducible nitric oxide synthase (iNOS) expression. Inhibiting STAT3 phosphorylation partially impeded the increase in IL-6 production and restored iNOS expression in these PD-L1-deficient cells. These findings provide valuable insights into the complexity and mechanisms underlying anti-PD-L1 therapy in the context of tuberculosis.

Transcriptomic Signature of 3D Hierarchical Porous Chip Enriched Exosomes for Early Detection and Progression Monitoring of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is a highly lethal malignant tumor, and the current non-invasive diagnosis method based on serum markers, such as α-fetoprotein (AFP), and des-γ-carboxy-prothrombin (DCP), has limited efficacy in detecting it. Therefore, there is a critical need to develop novel biomarkers for HCC. Recent studies have highlighted the potential of exosomes as biomarkers. To enhance exosome enrichment, a silicon dioxide (SiO2) microsphere-coated three-dimensional (3D) hierarchical porous chip, named a SiO2-chip is designed. The features of the chip, including its continuous porous 3D scaffold, large surface area, and nanopores between the SiO2 microspheres, synergistically improved the exosome capture efficiency. Exosomes from both non-HCC and HCC subjects are enriched using an SiO2-chip and performed RNA sequencing to identify HCC-related long non-coding RNAs (lncRNAs) in the exosomes. This study analysis reveales that LUCAT-1 and EGFR-AS-1 are two HCC-related lncRNAs. To further detect dual lncRNAs in exosomes, quantitative real time polymerase chain reaction (qRT-PCR) is employed. The integration of dual lncRNAs with AFP and DCP significantly improves the diagnostic accuracy. Furthermore, the integration of dual lncRNAs with DCP effectively monitors the prognosis of patients with HCC and detects disease progression. In this study, a liquid biopsy-based approach for noninvasive and reliable HCC detection is developed.

Lycopene suppresses gastric cancer cell growth without affecting normal gastric epithelial cells.

Gastric cancer is one of the leading causes of cancer-related death worldwide. Lycopene, a natural carotenoid, has potent antioxidant activity and anti-cancer effects against several types of cancers. However, the mechanism for the anti-gastric cancer effects of lycopene remains to be fully clarified. Normal gastric epithelial cell line GES-1 and gastric cancer cell line AGS, SGC-7901, Hs746T cells were treated with different concentrations of lycopene and the effects of lycopene were compared. Lycopene specifically suppressed cell growth monitored by Real-Time Cell Analyzer, induced cell cycle arrest and cell apoptosis detected by flow cytometry, and lowered mitochondrial membrane potentials assessed by JC-1 staining of AGS and SGC-7901 cells, while did not affect those of GES-1 cells. Lycopene did not affect the cell growth of Hs746T cells harboring TP53 mutation. Further bioinformatics analysis predicted 57 genes with up-regulated expression levels in gastric cancer and decreased function in cells after lycopene treatment. Quantitative PCR and Western Blot were used to check the critical factors in the cell cycle and apoptosis signaling pathway. Lycopene decreased the high expression levels of CCNE1 and increased the levels of TP53 in AGS and SGC-7901 cells without affecting those in GES-1 cells. In summary, lycopene could effectively suppress gastric cancer cells with CCNE1-amplification, which could be a promising target therapy reagent for gastric cancer.

Engineered exosomes as an in situ DC-primed vaccine to boost antitumor immunity in breast cancer.

BACKGROUND: Dendritic cells (DCs) are central for the initiation and regulation of innate and adaptive immunity in the tumor microenvironment. As such, many kinds of DC-targeted vaccines have been developed to improve cancer immunotherapy in numerous clinical trials. Targeted delivery of antigens and adjuvants to DCs in vivo represents an important approach for the development of DC vaccines. However, nonspecific activation of systemic DCs and the preparation of optimal immunodominant tumor antigens still represent major challenges. METHODS: We loaded the immunogenic cell death (ICD) inducers human neutrophil elastase (ELANE) and Hiltonol (TLR3 agonist) into α-lactalbumin (α-LA)-engineered breast cancer-derived exosomes to form an in situ DC vaccine (HELA-Exos). HELA-Exos were identified by transmission electron microscopy, nanoscale flow cytometry, and Western blot analysis. The targeting, killing, and immune activation effects of HELA-Exos were evaluated in vitro. The tumor suppressor and immune-activating effects of HELA-Exos were explored in immunocompetent mice and patient-derived organoids. RESULTS: HELA-Exos possessed a profound ability to specifically induce ICD in breast cancer cells. Adequate exposure to tumor antigens and Hiltonol following HELA-Exo-induced ICD of cancer cells activated type one conventional DCs (cDC1s) in situ and cross-primed tumor-reactive CD8+ T cell responses, leading to potent tumor inhibition in a poorly immunogenic triple negative breast cancer (TNBC) mouse xenograft model and patient-derived tumor organoids. CONCLUSIONS: HELA-Exos exhibit potent antitumor activity in both a mouse model and human breast cancer organoids by promoting the activation of cDC1s in situ and thus improving the subsequent tumor-reactive CD8+ T cell responses. The strategy proposed here is promising for generating an in situ DC-primed vaccine and can be extended to various types of cancers. Scheme 1. Schematic illustration of HELA-Exos as an in situ DC-primed vaccine for breast cancer. (A) Allogenic breast cancer-derived exosomes isolated from MDA-MB-231 cells were genetically engineered to overexpress α-LA and simultaneously loaded with the ICD inducers ELANE and Hiltonol (TLR3 agonist) to generate HELA-Exos. (B) Mechanism by which HELA-Exos activate DCs in situ in a mouse xenograft model ofTNBC. HELA-Exos specifically homed to the TME and induced ICD in cancer cells, which resulted in the increased release of tumor antigens, Hiltonol, and DAMPs, as well as the uptake of dying tumor cells by cDC1s. The activated cDC1s then cross-primed tumor-reactive CD8+ T cell responses. (C) HELA-Exos activated DCs in situ in the breast cancer patient PBMC-autologous tumor organoid coculture system. ABBREVIATIONS: DCs: dendritic cells; α-LA: α-lactalbumin; HELA-Exos: Hiltonol-ELANE-α-LA-engineered exosomes; ICD: immunogenic cell death; ELANE: human neutrophil elastase; TLR3: Toll-like receptor 3; TNBC: triple-negative breast cancer; TME: tumor microenvironment; DAMPs: damage-associated molecular patterns; cDC1s: type 1 conventional dendritic cells; PBMCs: peripheral blood mononuclear cells.

IL-17F depletion accelerates chitosan conduit guided peripheral nerve regeneration.

Peripheral nerve injury is a serious health problem and repairing long nerve deficits remains a clinical challenge nowadays. Nerve guidance conduit (NGC) serves as the most promising alternative therapy strategy to autografts but its repairing efficiency needs improvement. In this study, we investigated whether modulating the immune microenvironment by Interleukin-17F (IL-17F) could promote NGC mediated peripheral nerve repair. Chitosan conduits were used to bridge sciatic nerve defect in IL-17F knockout mice and wild-type mice with autografts as controls. Our data revealed that IL-17F knockout mice had improved functional recovery and axonal regeneration of sciatic nerve bridged by chitosan conduits comparing to the wild-type mice. Notably, IL-17F knockout mice had enhanced anti-inflammatory macrophages in the NGC repairing microenvironment. In vitro data revealed that IL-17F knockout peritoneal and bone marrow derived macrophages had increased anti-inflammatory markers after treatment with the extracts from chitosan conduits, while higher pro-inflammatory markers were detected in the Raw264.7 macrophage cell line, wild-type peritoneal and bone marrow derived macrophages after the same treatment. The biased anti-inflammatory phenotype of macrophages by IL-17F knockout probably contributed to the improved chitosan conduit guided sciatic nerve regeneration. Additionally, IL-17F could enhance pro-inflammatory factors production in Raw264.7 cells and wild-type peritoneal macrophages. Altogether, IL-17F may partially mediate chitosan conduit induced pro-inflammatory polarization of macrophages during nerve repair. These results not only revealed a role of IL-17F in macrophage function, but also provided a unique and promising target, IL-17F, to modulate the microenvironment and enhance the peripheral nerve regeneration.

Deep learning for COVID-19 chest CT (computed tomography) image analysis: A lesson from lung cancer.

As a recent global health emergency, the quick and reliable diagnosis of COVID-19 is urgently needed. Thus, many artificial intelligence (AI)-base methods are proposed for COVID-19 chest CT (computed tomography) image analysis. However, there are very limited COVID-19 chest CT images publicly available to evaluate those deep neural networks. On the other hand, a huge amount of CT images from lung cancer are publicly available. To build a reliable deep learning model trained and tested with a larger scale dataset, the proposed model builds a public COVID-19 CT dataset, containing 1186 CT images synthesized from lung cancer CT images using CycleGAN. Additionally, various deep learning models are tested with synthesized or real chest CT images for COVID-19 and Non-COVID-19 classification. In comparison, all models achieve excellent results in accuracy, precision, recall and F1 score for both synthesized and real COVID-19 CT images, demonstrating the reliable of the synthesized dataset. The public dataset and deep learning models can facilitate the development of accurate and efficient diagnostic testing for COVID-19.

Fine-Grained Breast Cancer Classification With Bilinear Convolutional Neural Networks (BCNNs).

Classification of histopathological images of cancer is challenging even for well-trained professionals, due to the fine-grained variability of the disease. Deep Convolutional Neural Networks (CNNs) showed great potential for classification of a number of the highly variable fine-grained objects. In this study, we introduce a Bilinear Convolutional Neural Networks (BCNNs) based deep learning method for fine-grained classification of breast cancer histopathological images. We evaluated our model by comparison with several deep learning algorithms for fine-grained classification. We used bilinear pooling to aggregate a large number of orderless features without taking into consideration the disease location. The experimental results on BreaKHis, a publicly available breast cancer dataset, showed that our method is highly accurate with 99.24% and 95.95% accuracy in binary and in fine-grained classification, respectively.

Divergent effects of lycopene on pancreatic alpha and beta cells.

Lycopene is an antioxidant which has potential anti-diabetic activity, but the cellular mechanisms have not been clarified. In this study, different concentrations of lycopene were used to treat pancreatic alpha and beta cell lines, and the changes of cell growth, cell apoptosis, cell cycle, reactive oxygen species (ROS), ATP levels and expression of related cytokines were determined. The results exhibited that lycopene did not affect cell growth, cell apoptosis, cell cycle, ROS and ATP levels of alpha cells, while it promoted the growth of beta cells, increased the ratio of S phase, reduced the ROS levels and increased the ATP levels of beta cells. At the same time, lycopene treatment elevated the mRNA expression levels of tnfα, tgfß and hif1α in beta cells. These findings suggest that lycopene plays cell-specific role and activates pancreatic beta cells, supporting its application in diabetes therapy.

Gastric cancer patients have elevated plasmacytoid and CD1c+ dendritic cells in the peripheral blood.

Dendritic cells (DCs) are important in tumor immunology. Identifying DC subset markers in the peripheral blood, which are informative for gastric cancer stages, is not only useful for prognosis but may also provide mechanistic insights into processes facilitating therapy. The present study investigated plasmacytoid dendritic cells (pDCs) and myeloid CD1c+ dendritic cells (mDC1s) in the peripheral blood of patients with gastric cancer and healthy controls using flow cytometry. Using peripheral DC staining and subset analysis, patients with gastric cancer were identified to have substantially higher numbers of peripheral pDCs and mDC1s. In addition, there was a trend of elevated circulating pDCs with advanced stages and lymph node metastasis in gastric cancer, whereas no differences in circulating mDC1s were observed among the various groups. The results suggested that circulating pDCs are a positive prognostic indicator in patients with gastric cancer of different stages and highlighted the critical role of pDCs immunity in the development of gastric cancer.

Evaluation of Immune Restoration Potential of PD-1 Blockers.

The programmed death 1 (PD-1) receptor and its primary ligand (PD-L1) have crucial roles in tumor-induced immune suppression. PD-1/PD-L1 blockers are designed to restore the immune system and induce potent antitumor effects. In this study we established a direct and reliable method to evaluate the immune restoration potential of human PD-1 blockers. We found anti-human PD-1 antibody could reverse PD-L1 induced suppression of human CD3+ cells proliferation and IL-2 production. This method is suitable for all kinds of PD-1 blockers including antibodies and chemical drugs. This function assay could be easily applied and provide valuable information for drug development.

Interleukin-17F attenuates H2O2-induced cell cycle arrest.

Interleukin IL-17F was expressed in colon epithelial cells and showed multiple functions in colon tumorigenesis. However, the role of IL-17F in colon cancer cell cycle progression remains unclear. In this study, we analyzed the effects of IL-17F on oxidant-induced cell cycle shift in human colon cancer cells. IL-17F overexpressing and wildtype HCT116 cells were challenged with H(2)O(2). Cell cycle distribution analysis showed IL-17F attenuated H(2)O(2)-induced G2/M phase arrest by inhibiting S to G2/M transition. We further checked expression levels of two critical cell cycle regulators p21 and p27. The results showed that IL-17F could inhibit H(2)O(2) induced p27 up-regulation. Meanwhile, IL-17F could increase the phosphorylation of p38 after H(2)O(2) treatment. The regulations of p27 level and p38 activity may contribute to the impaired G2/M phase arrest by IL-17F. Taken together, our findings extend IL-17F as an important factor in colon cancer development and provide new insight into the signaling pathway.

IgG-positive cells surround pancreatic ducts and form multiple layers after streptozotocin treatment.

Although numerous studies of diabetes have focused on cell-mediated immunity to pancreatic islet cells, little is known about immune cells in the pancreatic duct region. In this study, we found that membrane immunoglobulin G (IgG)-positive cells comprised about 1.4% of the total pancreatic cells in mice, forming a thin septum that surrounds large and medium pancreatic ducts. The IgG-positive cells showed low expression of beta-catenin and were amylase-, cytokeratin-, insulin-, and glucagon-negative. Flow cytometric analysis showed that the IgG-positive cells were also positive for CD45, Sca-1, c-Kit, CD49f, and CD133, and negative for Flk-1, suggesting that they were undifferentiated hematopoietic cells. On day 5 after streptozotocin treatment, the percentage of periductal IgG-positive cells increased to 3.37% of total pancreatic cells, and the periductal IgG-positive cells formed multiple layers (beta-catenin-low, and amylase-, cytokeratin-, insulin-, glucagon-negative). These cells were Ki67-negative, suggesting they were recruited from hematopoietic cells. We further found that IgG-positive cells formed multiple layers around large ducts of pancreas from NOD mice. Our findings reveal the existence of periductal IgG-positive cells in the adult mouse pancreas, which were activated during streptozotocin-induced diabetes, adding a new dimension to our understanding of immunity in diabetes.

Progesterone suppresses Th17 cell responses, and enhances the development of regulatory T cells, through thymic stromal lymphopoietin-dependent mechanisms in experimental gonococcal genital tract infection.

In most female patients, the symptoms of genital infection due to Neisseria gonorrhoeae tend to be slight or even absent. Our previous studies suggested that progesterone might play a role in female asymptomatic gonococcal infection. In this study, we demonstrated that progesterone induced the expression of thymic stromal lymphopoietin (TSLP) and regulatory T cells (Treg)-related transcription factor Foxp3, and inhibited the expression of Th17 related transcription factor RORγt, and reduced the influx of neutrophils in murine vaginal gonococcal infection. Blockade of TSLP with antibody partially reversed the effects of progesterone on the murine model of gonococcal vaginal infection. In in vitro experiments, progesterone induced a rapid up-regulation of TSLP in vaginal epithelial cells stimulated with N. gonorrhoeae. Blocking thymic stromal lymphopoietin receptor (TSLPR) with a TSLPR monoclonal antibody partially prevented progesterone suppression of IL-17-producing T cells differentiation, and progesterone promotion of CD4⁺CD25⁺Foxp3⁺ regulatory T cells differentiation. Altogether, our results indicate that the progesterone suppresses Th17 cell responses, and enhances the development of Treg cells, through TSLP-dependent mechanisms, and play a role in female asymptomatic gonococcal infections.

Comparative analysis of dendritic cell numbers and subsets between smoking and control subjects in the peripheral blood.

It has been well known that smoking alters the property and functionality of a wide range of immune cells including dendritic cells (DCs). However, a great deal of effort in the past has been mainly devoted to dissect the effect of smoking on pulmonary DCs, while its exact impact on circulating DCs remains to be fully addressed. Therefore, in the present report we particularly examined the impact of smoking on the number and subset of DCs in the peripheral blood by multi-parametric flow cytometry analysis. A significant increase for peripheral blood mononuclear cells (PBMCs) was noted in the smoking subjects. Subsequent studies revealed that the percentage for plasmacytoid DCs (pDCs) and total DCs in PBMCs was significantly higher in the smoking subjects as compared with that of control subjects, while the percentage for myeloid DCs (mDCs) did not differ between two groups. It was also found that the absolute number for total DCs, mDCs and pDCs were significantly higher in the smoking subjects than that of control subject. However, the mDC/pDC ratio was significantly reduced, suggesting that smoking impairs the balance of DC subsets. Given that pDCs are in favor of tolerogenic function, our data support that smoking could induce the production of pDCs to manifest immunosuppressive properties in the chronic smokers.

A protective role by interleukin-17F in colon tumorigenesis.

Interleukin-17F (IL-17F), produced by Th17 cells and other immune cells, is a member of IL-17 cytokine family with highest homology to IL-17A. IL-17F has been shown to have multiple functions in inflammatory responses. While IL-17A plays important roles in cancer development, the function of IL-17F in tumorigenesis has not yet been elucidated. In the current study, we found that IL-17F is expressed in normal human colonic epithelial cells, but this expression is greatly decreased in colon cancer tissues. To examine the roles of IL-17F in colon cancer, we have used IL-17F over-expressing colon cancer cell lines and IL-17F-deficient mice. Our data showed decreased tumor growth of IL-17F-transfected HCT116 cells comparing to mock transfectants when transplanted in nude mice. Conversely, there were increased colonic tumor numbers and tumor areas in Il-17f(-/-) mice than those from wild-type controls after colon cancer induction. These results indicate that IL-17F plays an inhibitory role in colon tumorigenesis in vivo. In IL-17F over-expressing tumors, there was no significant change in leukocyte infiltration; instead, we found decreased VEGF levels and CD31(+) cells. While the VEGF levels were increased in the colon tissues of Il-17f(-/-) mice with colon cancer. Together, our findings demonstrate a protective role for IL-17F in colon cancer development, possibly via inhibiting tumor angiogenesis.