DNA methylation is a fundamental epigenetic mark that governs gene expression and chromatin organization, thus providing a window into cellular identity and developmental processes1. Current datasets typically include only a fraction of methylation sites and are often based either on cell lines that underwent massive changes in culture or on tissues containing unspecified mixtures of cells2–5. Here we describe a human methylome atlas, based on deep whole-genome bisulfite sequencing, allowing fragment-level analysis across thousands of unique markers for 39 cell types sorted from 205 healthy tissue samples. Replicates of the same cell type are more than 99.5% identical, demonstrating the robustness of cell identity programmes to environmental perturbation. Unsupervised clustering of the atlas recapitulates key elements of tissue ontogeny and identifies methylation patterns retained since embryonic development. Loci uniquely unmethylated in an individual cell type often reside in transcriptional enhancers and contain DNA binding sites for tissue-specific transcriptional regulators. Uniquely hypermethylated loci are rare and are enriched for CpG islands, Polycomb targets and CTCF binding sites, suggesting a new role in shaping cell-type-specific chromatin looping. The atlas provides an essential resource for study of gene regulation and disease-associated genetic variants, and a wealth of potential tissue-specific biomarkers for use in liquid biopsies.

Lung cancer is the second most common malignancy and the leading cause of cancer-related death in the western world. Moreover, despite advances in surgery, chemotherapy and radiotherapy, the death rate from lung cancer remains high and the reported overall five-year survival rate is only 15%. Thus, novel treatments for this devastating disease are urgently needed. Chemokines, a family of 48 chemotactic cytokines interacts with their 7 transmembrane G-protein-coupled receptors, to guide immune cell trafficking in the body under both physiologic and pathologic conditions. Tumor cells, which express a relatively restricted repertoire of chemokine and chemokine receptors, utilize and manipulate the chemokine system in a manner that benefits both local tumor growth and distant dissemination. Among the 19 chemokine receptors, CXCR4 is the receptor most widely expressed by malignant tumors and whose role in tumor biology is most thoroughly studied. The chemokine CXCL12, which is the sole ligand of CXCR4, is highly expressed in primary lung cancer as well as in the bone marrow, liver, adrenal glands and brain, which are all sites for lung cancer metastasis. This review focuses on the pathologic role of the CXCR4/CXCL12 axis in NSCLC and on the potential therapeutic implication of targeting this axis for the treatment of NSCLC.

Mediastinal paraganglioms are rare, highly vascularized tumors arising from chromaffin tissue located in the para-aortic ganglia. Tumors tend to invade bordering structures and may also form metastasis. Up to 50% of patients are asymptomatic and diagnosis is incidental. Presenting symptoms are related to catecholamine hypersecretion or to a mass effect. Complete surgical resection remains the standard of care due to malignant potential of the tumor and poor response to chemotherapy or radiation. Strategic location of the tumor in proximity to great vessels, trachea, and recurrent laryngeal nerve poses challenge for the surgeon. We report a case of a 59-year old asymptomatic female who was incidentally diagnosed with a middle mediastinal mass on a positron-emission tomography (PET-CT) scan performed as part of breast cancer surveillance. Complete resection of the tumor was achieved using cardiopulmonary bypass. The patient recovered uneventfully and in a ten-month follow up there is no evidence of recurrence.

The chemokine receptor CXCR4 is involved in the growth and metastasis of tumor cells. However, the expression of its ligand, the chemokine CXCL12, in tumors and its role in regulating the accumulation of immune cells within the tumors is not clear. Using ELISA and immunohistochemistry we found that CXCL12 is expressed in the majority of nonsmall cell lung cancer tissue sections obtained from stage IA to IIB nonsmall cell lung cancer patients undergoing operation. Histopathologic examination of these sections indicated that high CXCL12 expression correlated with increased tumor inflammation. In addition, disease recurrence rates in a subgroup of adenocarcinoma patients showed a tendency to correlate with high CXCL12 expression in the tumor. Isolation of adenocarcinoma-infiltrating immune cells demonstrated an increase in the percentage of CD4+CD69+CXCR4+ T cells as compared with normal lung tissue. About 30% of these cells expressed the regulatory T cell markers CD25high and FoxP3. The percentage of CD8 T cells within the tumor did not change, however; the percentage of NK and NK T cells was significantly reduced. In correlation with CXCR4 expression, CD4 T cells showed increased migration in response to CXCL12 compared with CD8 T cells and NK cells. Overall, these observations suggest that CXCL12 expression may influence tumor progression by shaping the immune cell population infiltrating lung adenocarcinoma tumors.

Interaction between carcinoma-associated fibroblasts and tumor epithelial cells through the CXCL12/CXCR4 axis plays a role in non-small cell lung cancer tumor proliferation, marking this axis as a target for immune intervention.

ObjectivesAutocrine and paracrine chemokine/chemokine receptor-based interactions promote non-small-cell-lung-cancer (NSCLC) carcinogenesis. CCL20/CCR6 interactions are involved in prostatic and colonic malignancy pathogenesis. The expression and function of CCL20/CCR6 and its related Th-17 type immune response in NSCLC is not yet defined. We sought to characterize the role of the CCL20/CCR6/IL-17 axis in NSCLC tumor growth.MethodsA specialized histopathologist blindly assessed CCL20/CCR6 expression levels in 49 tissue samples of NSCLC patients operated in our department. Results were correlated to disease progression. Colony assays, ERK signaling and chemokine production were measured to assess cancer cell responsiveness to CCL20 and IL-17 stimulation.ResultsCCL20 was highly expressed in the majority (38/49, 77.5%) of tumor samples. Only a minority of samples (8/49, 16.5%) showed high CCR6 expression. High CCR6 expression was associated with a shorter disease-free survival (P = 0.008) and conferred a disease stage-independent 4.87-fold increased risk for disease recurrence (P = 0.0076, CI 95% 1.52–15.563). Cancerous cell colony-forming capacity was increased by CCL20 stimulation; this effect was dependent in part on ERK phosphorylation and signaling. IL-17 expression was detected in NSCLC; IL-17 potentiated the production of CCL20 by cancerous cells.ConclusionOur findings suggest that the CCL20/CCR6 axis promotes NSCLC disease progression. CCR6 is identified as a potential new prognostic marker and the CCL20/CCR6/IL-17 axis as a potential new therapeutic target. Larger scale studies are required to consolidate these observations.

The nasal-mucosa constitutes the primary entry site for respiratory viruses including SARS-CoV-2. While the imbalanced innate immune response of end-stage COVID-19 has been extensively studied, the earliest stages of SARS-CoV-2 infection at the mucosal entry site have remained unexplored. Here we employed SARS-CoV-2 and influenza virus infection in native multi-cell-type human nasal turbinate and lung tissues ex vivo , coupled with genome-wide transcriptional analysis, to investigate viral susceptibility and early patterns of local-mucosal innate immune response in the authentic milieu of the human respiratory tract. SARS-CoV-2 productively infected the nasal turbinate tissues, predominantly targeting respiratory epithelial cells, with rapid increase in tissue-associated viral sub-genomic mRNA, and secretion of infectious viral progeny. Importantly, SARS-CoV-2 infection triggered robust antiviral and inflammatory innate immune responses in the nasal mucosa. The upregulation of interferon stimulated genes, cytokines and chemokines, related to interferon signaling and immune-cell activation pathways, was broader than that triggered by influenza virus infection. Conversely, lung tissues exhibited a restricted innate immune response to SARS-CoV-2, with a conspicuous lack of type I and III interferon upregulation, contrasting with their vigorous innate immune response to influenza virus. Our findings reveal differential tissue-specific innate immune responses in the upper and lower respiratory tract, that are distinct to SARS-CoV-2. The studies shed light on the role of the nasal-mucosa in active viral transmission and immune defense, implying a window of opportunity for early interventions, whereas the restricted innate immune response in early-SARS-CoV-2-infected lung tissues could underlie the unique uncontrolled late-phase lung damage of advanced COVID-19. IMPORTANCE In order to reduce the late-phase morbidity and mortality of COVID-19, there is a need to better understand and target the earliest stages of SARS-CoV-2 infection in the human respiratory tract. Here we have studied the initial steps of SARS-CoV-2 infection and the consequent innate immune responses within the natural multicellular complexity of human nasal-mucosal and lung tissues. Comparing the global innate response patterns of nasal and lung tissues, infected in parallel with SARS-CoV-2 and influenza virus, we have revealed distinct virus-host interactions in the upper and lower respiratory tract, which could determine the outcome and unique pathogenesis of SARS-CoV-2 infection. Studies in the nasal-mucosal infection model can be employed to assess the impact of viral evolutionary changes, and evaluate new therapeutic and preventive measures against SARS-CoV-2 and other human respiratory pathogens.

Both insulin and IGF-I induced endothelial-independent, nitric oxide-independent vasorelaxation of rings from human IMA and saphenous veins, through a mechanism involving activation of potassium channels. This response remained intact in vessels from patients with NIDDM. This result supports the hypothesis that insulin and IGF-I play roles in the regulation of vascular tone in human vessels.