Early post-transplant urinary IP-10 expression after kidney transplantation is predictive of short- and long-term graft function

The early identification of renal transplant recipients at enhanced risk of developing acute and subclinical rejection would allow individualized adjustment of immunosuppression before functional graft injury occurs and would exclude these patients from drug-weaning studies. Protein and reverse transcriptase-polymerase chain reaction-based analyses of candidate markers in urine open the opportunity to closely monitor kidney-transplanted patients non-invasively. The chemokine interferon-inducible protein 10 (IP-10; CXCL10) might be an interesting candidate to uncover ongoing immune processes within the graft. Urine samples from kidney-transplanted recipients were retrospectively analyzed for IP-10 mRNA and protein expression. IP-10 levels were correlated with the incidence of acute rejection episodes proven by histology and long-term graft function assessed by the glomerular filtration rate 6 months post transplantation. IP-10 expression in urine identified patients with ongoing acute rejection episodes several days before a biopsy was indicated by rising serum creatinine levels. Most importantly, elevated levels of urinary IP-10 protein within the first four postoperative weeks were predictive of graft function at 6 months even in the absence of acute rejection. These data reveal a correlation between elevated IP-10 expression in urine at early time points post-transplantation and intragraft immune activation that leads to acute rejection and compromised long-term graft function

Mechanical forces switch blood vessel subtypes to arrest adolescent bone growth

Bone growth requires a specialised, highly angiogenic blood vessel subtype, so-called type H vessels, which pave the way for osteoblasts surrounding these vessels. At the end of adolescence, type H endothelial cells differentiate into quiescent type L endothelium lacking the capacity to promote bone growth. Until now, the signals that switch off type H vessel identity and thus arrest adolescent bone growth have remained ill defined. Here we show that mechanical forces, associated with increased body weight at the end of adolescence, trigger the mechanoreceptor PIEZO1 and thereby mediate enhanced production of the kinase FAM20C in osteoblasts. FAM20C phosphorylates dentin matrix protein 1 (DMP1), previously identified as a key factor in bone mineralization. This phosphorylation elicits a burst in DMP1 secretion from osteoblasts. Extracellular DMP1 inhibits vascular endothelial growth factor (VEGF) signalling by preventing VEGFR2 phosphorylation and VEGFR3 expression on the tip cells of type H endothelium. DMP1-mediated VEGF inhibition transforms bone growth-promoting type H vessels into quiescent type L vasculature to arrest bone growth and enhance bone mineralization. This molecular mechanism links mechanical forces and the termination of bone growth via accumulation of an extracellular matrix protein and its regulation of vascular subtypes. It suggests new options for the treatment of diseases characterised by inappropriate turnover or invasion of bone such as osteoarthritis, osteoporosis and osteosarcoma

Mechanical forces couple bone matrix mineralization with inhibition of angiogenesis to limit adolescent bone growth

Bone growth requires a specialised, highly angiogenic blood vessel subtype, so-called type H vessels, which pave the way for osteoblasts surrounding these vessels. At the end of adolescence, type H vessels differentiate into quiescent type L endothelium lacking the capacity to promote bone growth. Until now, the signals that switch off type H vessel identity and thus limit adolescent bone growth have remained ill defined. Here we show that mechanical forces, associated with increased body weight at the end of adolescence, trigger the mechanoreceptor PIEZO1 and thereby mediate enhanced production of the kinase FAM20C in osteoblasts. FAM20C, the major kinase of the secreted phosphoproteome, phosphorylates dentin matrix protein 1, previously identified as a key factor in bone mineralization. Thereupon, dentin matrix protein 1 is secreted from osteoblasts in a burst-like manner. Extracellular dentin matrix protein 1 inhibits vascular endothelial growth factor signalling by preventing phosphorylation of vascular endothelial growth factor receptor 2. Hence, secreted dentin matrix protein 1 transforms type H vessels into type L to limit bone growth activity and enhance bone mineralization. The discovered mechanism may suggest new options for the treatment of diseases characterised by aberrant activity of bone and vessels such as osteoarthritis, osteoporosis and osteosarcoma

Evaluation of NPP1 as a novel biomarker of coronary artery disease: A pilot study in human beings

Purpose: Coronary artery calcification (CAC) is utilized as an important tool for global risk assessment of cardiovascular events in individuals with intermediate risk. Ecto phosphodiesterase/nucleotide phosphohydrolase-1(ENPP1) converts extracellular nucleotides into inorganic pyrophosphate and it is a key regulator of tissue calcification that adjusts calcification in tissues like vascular smooth muscle cells. The main purpose of this clinical study was to find out the correlation between ENPP1 serum concentration and CAC in human for the first time. Methods: In this study 83 patients (16 diabetic patients and 67 non-diabetic patients) with coronary artery disease who fulfilled inclusion and exclusion criteria, entered the study. For all patients a questionnaire consisting demographic data and traditional cardiovascular risk factors were completed. Computed tomography (CT)-Angiography was carried out to determine coronary artery calcium score and enzyme-linked immunosorbent assay (ELISA) method was used for measuring ENPP1 serum concentrations. Results: There was a reverse significant correlation between ENPP1 serum concentration and total CAC score and also CAC of right coronary artery (RCA) (P < 0.05) in non-diabetic patients. Conclusion: On the basis of our results, ENPP1 serum concentration may be a suitable biomarker for coronary artery disease at least in non-diabetic patients. However, more studies with higher sample size are necessary for its confirmation. © 2018 The Authors

Discrete populations of isotype-switched memory B lymphocytes are maintained in murine spleen and bone marrow

At present, it is not clear how memory B lymphocytes are maintained over time, and whether only as circulating cells or also residing in particular tissues. Here we describe distinct populations of isotype-switched memory B lymphocytes (Bsm) of murine spleen and bone marrow, identified according to individual transcriptional signature and B cell receptor repertoire. A population of marginal zone-like cells is located exclusively in the spleen, while a population of quiescent Bsm is found only in the bone marrow. Three further resident populations, present in spleen and bone marrow, represent transitional and follicular B cells and B1 cells, respectively. A population representing 10-20% of spleen and bone marrow memory B cells is the only one qualifying as circulating. In the bone marrow, all cells individually dock onto VCAM1+ stromal cells and, reminiscent of resident memory T and plasma cells, are void of activation, proliferation and mobility

Clonally expanded PD-1-expressing T cells are enriched in synovial fluid of juvenile idiopathic arthritis patients

Juvenile idiopathic arthritis (JIA) is the most common chronic rheumatic condition in childhood. Disease etiology remains largely unknown, however a key role in JIA pathogenesis is surely mediated by T cells. T lymphocytes activity is controlled via signals, known as immune-checkpoints (IC). Delivering an inhibitory signal or blocking a stimulatory signal to achieve immune suppression is critical in autoimmune diseases. However, the role of IC in chronic inflammation and autoimmunity must still be deciphered. In this study, we investigated at single cell level the feature of T cells in JIA chronic inflammation both at transcriptome level via single-cell RNA sequencing and at protein level by flow cytometry. We found that despite the heterogeneity in the composition of synovial CD4+ and CD8+ T cells, those characterized by PD-1 expression were clonally expanded Trm-like cells and displayed the highest pro-inflammatory capacity, suggesting their active contribution in sustaining chronic inflammation in situ. Our data support the concept that novel therapeutic strategies targeting PD-1 may be effective in the treatment of JIA. With this approach, it may become possible to target overactive T regardless of their cytokine production profile

Guidelines for the use of flow cytometry and cell sorting in immunological studies (third edition)

The third edition of Flow Cytometry Guidelines provides the key aspects to consider when performing flow cytometry experiments and includes comprehensive sections describing phenotypes and functional assays of all major human and murine immune cell subsets. Notably, the Guidelines contain helpful tables highlighting phenotypes and key differences between human and murine cells. Another useful feature of this edition is the flow cytometry analysis of clinical samples with examples of flow cytometry applications in the context of autoimmune diseases, cancers as well as acute and chronic infectious diseases. Furthermore, there are sections detailing tips, tricks and pitfalls to avoid. All sections are written and peer-reviewed by leading flow cytometry experts and immunologists, making this edition an essential and state-of-the-art handbook for basic and clinical researchers

Cobalt protoporphyrin represses osteoclastogenesis through blocking multiple signaling pathways

Cobalt protoporphyrin (CoPP) is a metallo-protoporphyrin that works as a powerful inducer of heme oxigenase-1 (HO-1) in various tissues and cells. Our recent studies have demonstrated that induction of HO-1 by several reagents inhibited differentiation and activation of osteoclasts (OCLs), which are multinucleated bone resorbing cells. However, the effects of CoPP on osteoclastogenesis remain to be elucidated. In this study, we report that CoPP inhibits receptor activator of nuclear factor κB ligand (RANKL)-induced OCL formation in a dose dependent manner. Importantly, CoPP had little cytotoxicity, but rather enhanced cell proliferation of OCLs. CoPP suppressed the protein levels of nuclear factor of activated T cells cytoplasmic-1 (NFATc1) as well as those of OCLs markers such as Src and cathepsin K, which are transcriptionally regulated by NFATc1 in mature OCLs. Western blot analyses also showed that CoPP abolished RANKL-stimulated phosphorylation of several major signaling pathways such as IκB, Akt, ERK, JNK and p38 MAPKs in OCL precursor cells. Thus, our results show that CoPP represses osteoclastogenesis through blocking multiple signaling pathways

Distinct tissue niches direct lung immunopathology via CCL18 and CCL21 in severe COVID-19

Prolonged lung pathology has been associated with COVID-19, yet the cellular and molecular mechanisms behind this chronic inflammatory disease are poorly understood. In this study, we combine advanced imaging and spatial transcriptomics to shed light on the local immune response in severe COVID-19. We show that activated adventitial niches are crucial microenvironments contributing to the orchestration of prolonged lung immunopathology. Up-regulation of the chemokines CCL21 and CCL18 associates to endothelial-to-mesenchymal transition and tissue fibrosis within these niches. CCL21 over-expression additionally links to the local accumulation of T cells expressing the cognate receptor CCR7. These T cells are imprinted with an exhausted phenotype and form lymphoid aggregates that can organize in ectopic lymphoid structures. Our work proposes immune-stromal interaction mechanisms promoting a self-sustained and non-resolving local immune response that extends beyond active viral infection and perpetuates tissue remodeling

Guidelines for the use of flow cytometry and cell sorting in immunological studies (third edition)

The third edition of Flow Cytometry Guidelines provides the key aspects to consider when performing flow cytometry experiments and includes comprehensive sections describing phenotypes and functional assays of all major human and murine immune cell subsets. Notably, the Guidelines contain helpful tables highlighting phenotypes and key differences between human and murine cells. Another useful feature of this edition is the flow cytometry analysis of clinical samples with examples of flow cytometry applications in the context of autoimmune diseases, cancers as well as acute and chronic infectious diseases. Furthermore, there are sections detailing tips, tricks and pitfalls to avoid. All sections are written and peer-reviewed by leading flow cytometry experts and immunologists, making this edition an essential and state-of-the-art handbook for basic and clinical researchers