Pan-cancer analysis of immune checkpoint receptors and ligands in various cells in the tumor immune microenvironment.

Drugs that target immune checkpoint have become the most popular weapon in cancer immunotherapy, yet only have practical benefits for a small percentage of patients. Tumor cells constantly interact with their microenvironment, which is made up of a variety of immune cells as well as endothelial cells and fibroblasts. Immune checkpoint expression and blocked signaling of immune cells in the tumor microenvironment (TME) are key to tumor progression. In this study, we perform deliberation convolution on the TCGA database for human lung, breast, and colorectal cancer to infer crosstalk between immune checkpoint receptors (ICRs) and ligands (ICLs) in TME of pan-carcinogenic solid tumor types, validated by flow cytometry. Analysis of immune checkpoints showed that there was little variation between different tumor types. It showed that CD160, LAG3, TIGIT were found to be highly expressed in CD8+ T cells instead of CD4+ T cells, PD-L1, PD-L2, CD86, LGALS9, TNFRSF14, LILRB4 and other ligands were highly expressed on macrophages, FVR, NECTIN2, FGL1 were highly expressed on Epithelial cells, CD200 was highly expressed in Endothelial cells, and CD80 was highly expressed in CD8 High expression on T cells. Overall, our study provides a new resource for the expression of immune checkpoints in TME on various types of cells. Significance: This study provides immune checkpoint expression of immune cells of multiple cancer types to infer immune mechanisms in the tumor microenvironment and provide ideas for the development of new immune checkpoint-blocking drugs.

Chronic stress induces pulmonary epithelial cells to produce acetylcholine that remodels lung pre-metastatic niche of breast cancer by enhancing NETosis.

BACKGROUND: Chronic stress promotes most hallmarks of cancer through impacting the malignant tissues, their microenvironment, immunity, lymphatic flow, etc. Existing studies mainly focused on the roles of stress-induced activation of systemic sympathetic nervous system and other stress-induced hormones, the organ specificity of chronic stress in shaping the pre-metastatic niche remains largely unknown. This study investigated the role of chronic stress in remodeling lung pre-metastatic niche of breast cancer. METHODS: Breast cancer mouse models with chronic stress were constructed by restraint or unpredictable stress. Expressions of tyrosine hydroxylase, vesicular acetylcholine transporter (VAChT), EpCAM and NETosis were examined by immunofluorescence and confocal microscopy. mRNA and protein levels of choline acetyltransferase (ChAT), VAChT, and peptidylarginine deiminase 4 were detected by qRT-PCR and Western blotting, respectively. Immune cell subsets were analyzed by flow cytometry. Acetylcholine (ACh) and chemokines were detected by ELISA and multi chemokine array, respectively. ChAT in lung tissues from patients was examined by immunohistochemistry. RESULTS: Breast cancer-bearing mice suffered chronic stress metastasized earlier and showed more severe lung metastasis than did mice in control group. VAChT, ChAT and ChAT+ epithelial cells were increased significantly in lung of model mice undergone chronic stress. ACh and chemokines especially CXCL2 in lung culture supernatants from model mice with chronic stress were profoundly increased. Chronic stress remodeled lung immune cell subsets with striking increase of neutrophils, enhanced NETosis in lung and promoted NETotic neutrophils to capture cancer cells. ACh treatment resulted in enhanced NETosis of neutrophils. The expression of ChAT in lung tissues from breast cancer patients with lung metastasis was significantly higher than that in patients with non-tumor pulmonary diseases. CONCLUSIONS: Chronic stress promotes production of CXCL2 that recruits neutrophils into lung, and induces pulmonary epithelial cells to produce ACh that enhances NETosis of neutrophils. Our findings demonstrate for the first time that chronic stress induced epithelial cell derived ACh plays a key role in remodeling lung pre-metastatic niche of breast cancer.

Mature dendritic cells enriched in immunoregulatory molecules (mregDCs): A novel population in the tumour microenvironment and immunotherapy target.

BACKGROUND: Dendritic cells (DCs) mediate divergent immune effects by activating T cells or negatively regulating the immune response to promote immune tolerance. They perform specific functions determined by their tissue distribution and maturation state. Traditionally, immature and semimature DCs were described to have immunosuppressive effects, leading to immune tolerance. Nonetheless, recent research has demonstrated that mature DCs can also suppress the immune response under certain circumstances. MAIN BODY: Mature DCs enriched in immunoregulatory molecules (mregDCs) have emerged as a regulatory module across species and tumour types. Indeed, the distinct roles of mregDCs in tumour immunotherapy have sparked the interest of researchers in the field of single-cell omics. In particular, these regulatory cells were found to be associated with a positive response to immunotherapy and a favourable prognosis. CONCLUSION: Here, we provide a general overview of the latest and most notable advances and recent findings regarding the basic features and complex roles of mregDCs in nonmalignant diseases and the tumour microenvironment. We also emphasise the important clinical implications of mregDCs in tumours.

γδ T cell exhaustion: Opportunities for intervention.

T lymphocytes are the key protective contributors in chronic infection and tumor, but experience exhaustion by persistent antigen stimulation. As an unconventional lineage of T cells, γδ T cells can rapidly response to varied infectious and tumor challenges in a non-MHC-restricted manner and play key roles in immune surveillance via pleiotropic effector functions, showing promising as candidates for cellular tumor immunotherapy. Activated γδ T cells can also acquire exhaustion signature with elevated expression of immune checkpoints, such as PD-1, decreased cytokine production, and functional impairment. However, the exhaustion features of γδ T cells are distinct from conventional αß T cells. Here, we review the researches regarding the characteristics, heterogeneity, and mechanisms of γδ T cell exhaustion. These studies provide insights into the combined strategies to overcome the exhaustion of γδ T cells and enhance antitumor immunity. Summary sentence: Review of the characteristics, heterogeneity, and mechanisms of γδ T cell exhaustion provides insights into the combined strategies to enhance γδ T cell-based antitumor immunotherapy.

Multi-Omics Profiling Suggesting Intratumoral Mast Cells as Predictive Index of Breast Cancer Lung Metastasis.

Breast cancer lung metastasis has a high mortality rate and lacks effective treatments, for the factors that determine breast cancer lung metastasis are not yet well understood. In this study, data from 1067 primary tumors in four public datasets revealed the distinct microenvironments and immune composition among patients with or without lung metastasis. We used multi-omics data of the TCGA cohort to emphasize the following characteristics that may lead to lung metastasis: more aggressive tumor malignant behaviors, severer genomic instability, higher immunogenicity but showed generalized inhibition of effector functions of immune cells. Furthermore, we found that mast cell fraction can be used as an index for individual lung metastasis status prediction and verified in the 20 human breast cancer samples. The lower mast cell infiltrations correlated with tumors that were more malignant and prone to have lung metastasis. This study is the first comprehensive analysis of the molecular and cellular characteristics and mutation profiles of breast cancer lung metastasis, which may be applicable for prognostic prediction and aid in choosing appropriate medical examinations and therapeutic regimens.

The potential of aptamers for cancer research.

Aptamers are promising alternatives to antibodies and can be used as high affinity agents for the cancer detection and the targeted drug transportation. In this manuscript, we highlight the advantages of aptamers, such as high affinities, specificity and excellent chemical stabilities, which are likely to benefit for the diagnosis of cancer in its early stages and then achieve molecular-level treatment. Also, we discuss the challenges and problems in the current application of aptamers.

Combined effects of tumor necrosis factor-α and interleukin-1ß on lysyl oxidase and matrix metalloproteinase expression in human knee synovial fibroblasts in vitro.

Previous studies have demonstrated that inflammatory cytokines are associated with matrix metalloproteinases (MMPs) and/or lysyl oxidases (LOXs) produced by anterior cruciate ligament (ACL) fibroblasts, which may contribute to the poor healing ability of the ACL. To evaluate whether the synovium also participates in ACL healing, the inflammatory microenvironment of the knee joint cavity was mimicked following ACL injury, and the combined effects of tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß) on the expression of MMPs and LOXs in synovial fibroblasts were studied. Cell viability was evaluated using trypan blue staining in the presence of TNF-α and IL-1ß, and the expression of LOXs and MMPs was measured by reverse transcription-quantitative polymerase chain reaction. MMP-2 activity was also measured by zymography. The results indicated that the combined effects of TNF-α and IL-1ß inhibited LOX expression, while promoting MMP-1, -2 and -3 expression and MMP-2 activity in synovial fibroblasts. These changes may impede healing by altering the balance between the degradative and biosynthetic arms of the ligament tissue remodeling process. Collectively, the present results suggest that the poor healing ability of cruciate ligaments may be due to the sensitivity of the synovium to inflammatory factors. Therefore, the synovium potentially serves a key regulatory role in the joint cavity microenvironment and in the healing process of the ACL, and thus should be considered as a therapeutic target to aid in the treatment of patients with ACL trauma.

Synthesis of cell composite alginate microfibers by microfluidics with the application potential of small diameter vascular grafts.

Fabrication of small diameter vascular grafts (SDVGs) with appropriate responses for clinical application is still challenging. In the present work, the production and characterization of solid alginate based microfibers as potential SDVG candidates through the method of microfluidics were considered original. A simple glass microfluidic device with a 'L-shape' cylindrical-flow channel in the microfluidic platform was developed. The gelation of microfibers occurred when the alginate solution and a CaCl2 solution were introduced as a core flow and as a sheath flow, respectively. The diameters of the microfibers could be controlled by varying the flow rates and the glass capillary tubes diameters at their tips. The generated microfibers had somewhat rough and porous surfaces, their suture retention strengths were comparable to the strength of other tissue engineered grafts. The encapsulated mesenchymal stem cells proliferated well in the microfibers, and showed a stable endothelialization under the angiogenesis effects of vascular endothelial growth factor and fibroblastic growth factor. The in vivo implant into the mice abdomens indicated that cell composite microfibers caused a mild host reaction. These encouraging results suggest great promise of the application of microfluidics as a future alternative in SDVGs engineering.

TNF-α induced down-regulation of lysyl oxidase family in anterior cruciate ligament and medial collateral ligament fibroblasts.

BACKGROUND: The lysyl oxidase (LOX) family has the capacity to catalyze the cross-linking of collagen and elastin, implicating its important fundamental role in injury healing. Tumor necrosis factor alpha (TNF-α) is considered to be an important chemical mediator in the acute inflammatory phase of the ligament injury. The role of the lysyl oxidase family induced by TNF-α in the knee ligaments' wound healing process is poorly understood. Our purpose was to determine the different expressions of the LOXs in poorly self-healing anterior cruciate ligament (ACL) and well functionally self-healing medial collateral ligament (MCL) induced by TNF-α. METHODS: Semi-quantitative PCR, quantitative real-time PCR and western blot were performed for original research. RESULTS: The results showed that all LOX family members were expressed at higher levels in MCL than those in ACL fibroblasts; the significant differences existed in the down-regulations of the LOXs induced by TNF-α; and the TNF-α-mediated down-regulations of the LOXs were more prominent in ACL than those in MCL fibroblasts. 1-20 ng/ml TNF-α down-regulated mRNA levels in ACL and MCL fibroblasts by up to 76% and 58% in LOX; 90% and 45% in LOXL-1; 97.5% and 90% in LOXL-2; 89% and 68% in LOXL-3; 52% and 25% in LOXL-4, respectively. Protein assay also showed LOXs had lower expressions in ACL than those in MCL. CLINICAL RELEVANCE: Based on these results, the differential expressions of the LOXs might help to explain the intrinsic differences between the poorly self-healing ACL and well functionally self-healing MCL.

Targeting eradication of malignant cells derived from human bone marrow mesenchymal stromal cells.

Human bone marrow mesenchymal stromal cells (hBMSC) have been shown to participate in malignant transformation. However, hampered by the low frequency of malignant transformation of hBMSC, we do not yet know how to prevent malignant transformation of implanted hBMSC. In this study, in order to establish a model for the eradication of hBMSC-derived malignant cells, a gene fusion consisting of a human telomerase (hTERT) promoter modified with both c-Myc and myeloid zinc finger protein2 (MZF-2) binding elements and followed by the E. coli cytosine deaminase (CD) and luciferase genes was stably transferred into hBMSC via lentiviral transduction; n-phosphonacelyl-L-aspartic acid (PALA) selection was used to generate malignant cell colonies derived from transduced hBMSC after treatment with the carcinogenic reagent BPDE. Cells that were amplified after PALA selection were used for transplantation and 5-FC pro-drug cytotoxicity tests. The results showed that PALA-resistant malignant cells could be generated from hBMSC co-induced with lentiviral transduction and treatment with Benzo(a)pyrene Diol Epoxide (BPDE); the modification of c-Myc and MZF-2 binding elements could remarkably enhance the transcriptional activities of the hTERT promoter in malignant cells, whereas transcriptional activity was depressed in normal hBMSC; malignant cells stably expressing CD under the control of the modified hTERT promoter could be eliminated by 5-FC administration. This study has provided a method for targeted eradication of malignant cells derived from hBMSC.