The human fetal immune system begins to develop early during gestation; however, factors responsible for fetal immune-priming remain elusive. We explored potential exposure to microbial agents in utero and their contribution toward activation of memory T cells in fetal tissues. We profiled microbes across fetal organs using 16S rRNA gene sequencing and detected low but consistent microbial signal in fetal gut, skin, placenta, and lungs in the 2^nd trimester of gestation. We identified several live bacterial strains including Staphylococcus and Lactobacillus in fetal tissues, which induced in vitro activation of memory T cells in fetal mesenteric lymph node, supporting the role of microbial exposure in fetal immune-priming. Finally, using SEM and RNA-ISH, we visualized discrete localization of bacteria-like structures and eubacterial-RNA within 14^th weeks fetal gut lumen. These findings indicate selective presence of live microbes in fetal organs during the 2^nd trimester of gestation and have broader implications toward the establishment of immune competency and priming before birth.

Epileptogenic triggers are multifactorial and not well understood. Here we aimed to address the hypothesis that inappropriate pro-inflammatory mechanisms contribute to the pathogenesis of refractory epilepsy (non-responsiveness to antiepileptic drugs) in human patients. We used single-cell cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq) to reveal the immunotranscriptome of surgically resected epileptic lesion tissues. Our approach uncovered a pro-inflammatory microenvironment, including extensive activation of microglia and infiltration of other pro-inflammatory immune cells. These findings were supported by ligand–receptor (LR) interactome analysis, which demonstrated potential mechanisms of infiltration and evidence of direct physical interactions between microglia and T cells. Together, these data provide insight into the immune microenvironment in epileptic tissue, which may aid the development of new therapeutics.

The clinical relevance of immune landscape intratumoural heterogeneity (immune-ITH) and its role in tumour evolution remain largely unexplored. Here, we uncover significant spatial and phenotypic immune-ITH from multiple tumour sectors and decipher its relationship with tumour evolution and disease progression in hepatocellular carcinomas (HCC). Immune-ITH is associated with tumour transcriptomic-ITH, mutational burden and distinct immune microenvironments. Tumours with low immune-ITH experience higher immunoselective pressure and escape via loss of heterozygosity in human leukocyte antigens and immunoediting. Instead, the tumours with high immune-ITH evolve to a more immunosuppressive/exhausted microenvironment. This gradient of immune pressure along with immune-ITH represents a hallmark of tumour evolution, which is closely linked to the transcriptome-immune networks contributing to disease progression and immune inactivation. Remarkably, high immune-ITH and its transcriptomic signature are predictive for worse clinical outcome in HCC patients. This in-depth investigation of ITH provides evidence on tumour-immune co-evolution along HCC progression.

CyTOF enables the study of the immune system with a complexity, depth, and multidimensionality never achieved before. However, the full potential of using CyTOF can be limited by scarce cell samples. Barcoding strategies developed based on direct labeling of cells using maleimido‐monoamide‐DOTA (m‐DOTA) provide a very useful tool. However, using m‐DOTA has some inherent problems, mainly associated with signal intensity. This may be a source of uncertainty when samples are multiplexed. As an alternative or complementary approach to m‐DOTA, conjugating an antibody, specific for a membrane protein present on most immune cells, with different isotopes could address the issues of stability and signal intensity needed for effective barcoding. We chose for this purpose CD45, and designed experiments to address different types of cultures and the ability to detect extra‐ and intra‐cellular targets. We show here that our approach provides an useful alternative to m‐DOTA in terms of sensitivity, specificity, flexibility, and user‐friendliness. Our manuscript provides details to effectively barcode immune cells, overcoming limitations in current technology and enabling the use of CyTOF with scarce samples (for instance precious clinical samples). © 2015 The Authors. Published by Wiley Periodicals, Inc.

Systemic sclerosis (SSc) is an autoimmune disease associated with increased mortality and poor morbidity, impairing the quality of life in patients. Whilst we know that SSc affects multiple organs via vasculopathy, inflammation, and fibrosis, its exact pathophysiology remains elusive. Microvascular injury and vasculopathy are the initial pathological features of the disease. Clinically, the vasculopathy in SSc is manifested as Raynaud’s phenomenon (reversible vasospasm in reaction to the cold or emotional stress) and digital ulcers due to ischemic injury. There are several reports that medications for vasculopathy, such as bosentan and soluble guanylate cyclase (sGC) modulators, improve not only vasculopathy but also dermal fibrosis, suggesting that vasculopathy is important in SSc. Although vasculopathy is an important initial step of the pathogenesis for SSc, it is still unclear how vasculopathy is related to inflammation and fibrosis. In this review, we focused on the clinical evidence for vasculopathy, the major cellular players for the pathogenesis, including pericytes, adipocytes, endothelial cells (ECs), and myofibroblasts, and their signaling pathway to elucidate the relationship among vasculopathy, inflammation, and fibrosis in SSc.