Calcified apoptotic vesicles from PROCR+ fibroblasts initiate heterotopic ossification.

Heterotopic ossification (HO) comprises the abnormal formation of ectopic bone in extraskeletal soft tissue. The factors that initiate HO remain elusive. Herein, we found that calcified apoptotic vesicles (apoVs) led to increased calcification and stiffness of tendon extracellular matrix (ECM), which initiated M2 macrophage polarization and HO progression. Specifically, single-cell transcriptome analyses of different stages of HO revealed that calcified apoVs were primarily secreted by a PROCR+ fibroblast population. In addition, calcified apoVs enriched calcium by annexin channels, absorbed to collagen I via electrostatic interaction, and aggregated to produce calcifying nodules in the ECM, leading to tendon calcification and stiffening. More importantly, apoV-releasing inhibition or macrophage deletion both successfully reversed HO development. Thus, we are the first to identify calcified apoVs from PROCR+ fibroblasts as the initiating factor of HO, and might serve as the therapeutic target for inhibiting pathological calcification.

Identification of Endothelial Cell Protein C Receptor by Urinary Proteomics as Novel Prognostic Marker in Non-Recovery Kidney Injury.

Acute kidney injury is a common and complex complication that has high morality and the risk for chronic kidney disease among survivors. The accuracy of current AKI biomarkers can be affected by water retention and diuretics. Therefore, we aimed to identify a urinary non-recovery marker of acute kidney injury in patients with acute decompensated heart failure. We used the isobaric tag for relative and absolute quantification technology to find a relevant marker protein that could divide patients into control, acute kidney injury with recovery, and acute kidney injury without recovery groups. An enzyme-linked immunosorbent assay of the endothelial cell protein C receptor (EPCR) was used to verify the results. We found that the EPCR was a usable marker for non-recovery renal failure in our setting with the area under the receiver operating characteristics 0.776 ± 0.065; 95%CI: 0.648-0.905, (p < 0.001). Further validation is needed to explore this possibility in different situations.

Structural vulnerability in EPCR suggests functional modulation.

The endothelial protein C receptor (EPCR) is a fundamental component of the vascular system in mammals due to its contribution in maintaining blood in a non-prothrombotic state, which is crucial for overall life development. It accomplishes this by enhancing the conversion of protein C (PC) into the anticoagulant activated protein C (APC), with this property being dependent on a known EPCR conformation that enables direct interaction with PC/APC. In this study, we report a previously unidentified conformation of EPCR whereby Tyr154, critical for PC/APC binding, shows a striking non-canonical configuration. This unconventional form is incompatible with PC/APC binding, and reveals, for the first time, a region of structural vulnerability and potential modulation in EPCR. The identification of this malleability enhances our understanding of this receptor, prompting inquiries into the interplay between its plasticity and function, as well as its significance within the broader framework of EPCR's biology, which extends to immune conditions.

Endothelial Protein C Receptor and 3K3A-Activated Protein C Protect Mice from Allergic Contact Dermatitis in a Contact Hypersensitivity Model.

Endothelial protein C receptor (EPCR) is a receptor for the natural anti-coagulant activated protein C (aPC). It mediates the anti-inflammatory and barrier-protective functions of aPC through the cleavage of protease-activated receptor (PAR)1/2. Allergic contact dermatitis is a common skin disease characterized by inflammation and defective skin barrier. This study investigated the effect of EPCR and 3K3A-aPC on allergic contact dermatitis using a contact hypersensitivity (CHS) model. CHS was induced using 1-Fluoro-2,4-dinitrobenzene in EPCR-deficient (KO) and matched wild-type mice and mice treated with 3K3A-aPC, a mutant form of aPC with diminished anti-coagulant activity. Changes in clinical and histological features, cytokines, and immune cells were examined. EPCRKO mice displayed more severe CHS, with increased immune cell infiltration in the skin and higher levels of inflammatory cytokines and IgE than wild-type mice. EPCR, aPC, and PAR1/2 were expressed by the skin epidermis, with EPCR presenting almost exclusively in the basal layer. EPCRKO increased the epidermal expression of aPC and PAR1, whereas in CHS, their expression was reduced compared to wild-type mice. 3K3A-aPC reduced CHS severity in wild-type and EPCRKO mice by suppressing immune cell infiltration/activation and inflammatory cytokines. In summary, EPCRKO exacerbated CHS, whereas 3K3A-aPC could reduce the severity of CHS in both EPCRKO and wild-type mice.

Thrombomodulin Switches Signaling and Protease-Activated Receptor 1 Cleavage Specificity of Thrombin.

BACKGROUND: Cleavage of the extracellular domain of PAR1 (protease-activated receptor 1) by thrombin at Arg41 and by APC (activated protein C) at Arg46 initiates paradoxical cytopathic and cytoprotective signaling in endothelial cells. In the latter case, the ligand-dependent coreceptor signaling by EPCR (endothelial protein C receptor) is required for the protective PAR1 signaling by APC. Here, we investigated the role of thrombomodulin in determining the specificity of PAR1 signaling by thrombin. METHODS: We prepared a PAR1 knockout (PAR1-/-) EA.hy926 endothelial cell line by CRISPR/Cas9 and transduced PAR1-/- cells with lentivirus vectors expressing PAR1 mutants in which either Arg41 or Arg46 was replaced with an Ala. Furthermore, human embryonic kidney 293 cells were transfected with wild-type or mutant PAR1 cleavage reporter constructs carrying N-terminal Nluc (NanoLuc luciferase) and C-terminal enhanced yellow fluorescent protein tags. RESULTS: Characterization of transfected cells in signaling and receptor cleavage assays revealed that, upon interaction with thrombomodulin, thrombin cleaves Arg46 to elicit cytoprotective effects by a ß-arrestin-2 biased signaling mechanism. Analysis of functional data and cleavage rates indicated that thrombin-thrombomodulin cleaves Arg46>10-fold faster than APC. Upon interaction with thrombin, the cytoplasmic domain of thrombomodulin recruited both ß-arrestin-1 and -2 to the plasma membrane. Thus, the thrombin cleavage of Arg41 was also cytoprotective in thrombomodulin-expressing cells by ß-arrestin-1-biased signaling. APC in the absence of EPCR cleaved Arg41 to initiate disruptive signaling responses like thrombin. CONCLUSIONS: These results suggest that coreceptor signaling by thrombomodulin and EPCR determines the PAR1 cleavage and signaling specificity of thrombin and APC, respectively.

EPCR deficiency ameliorates inflammatory arthritis in mice by suppressing the activation and migration of T cells and dendritic cells.

OBJECTIVES: Endothelial protein C receptor (EPCR) is highly expressed in synovial tissues of patients with RA, but the function of this receptor remains unknown in RA. This study investigated the effect of EPCR on the onset and development of inflammatory arthritis and its underlying mechanisms. METHODS: CIA was induced in EPCR gene knockout (KO) and matched wild-type (WT) mice. The onset and development of arthritis was monitored clinically and histologically. T cells, dendritic cells (DCs), EPCR and cytokines from EPCR KO and WT mice, RA patients and healthy controls (HCs) were detected by flow cytometry and ELISA. RESULTS: EPCR KO mice displayed >40% lower arthritis incidence and 50% less disease severity than WT mice. EPCR KO mice also had significantly fewer Th1/Th17 cells in synovial tissues with more DCs in circulation. Lymph nodes and synovial CD4 T cells from EPCR KO mice expressed fewer chemokine receptors CXCR3, CXCR5 and CCR6 than WT mice. In vitro, EPCR KO spleen cells contained fewer Th1 and more Th2 and Th17 cells than WT and, in concordance, blocking EPCR in WT cells stimulated Th2 and Th17 cells. DCs generated from EPCR KO bone marrow were less mature and produced less MMP-9. Circulating T cells from RA patients expressed higher levels of EPCR than HC cells; blocking EPCR stimulated Th2 and Treg cells in vitro. CONCLUSION: Deficiency of EPCR ameliorates arthritis in CIA via inhibition of the activation and migration of pathogenic Th cells and DCs. Targeting EPCR may constitute a novel strategy for future RA treatment.

Bioinformatics analysis of differentially expressed genes related to ischemia and hypoxia in spinal cord injury and construction of miRNA-mRNA or mRNA-transcription factor interaction network.

BACKGROUND: Previous studies show that spinal cord ischemia and hypoxia is an important cause of spinal cord necrosis and neurological loss. Therefore, the study aimed to identify genes related to ischemia and hypoxia after spinal cord injury (SCI) and analyze their functions, regulatory mechanism, and potential in regulating immune infiltration. METHODS: The expression profiles of GSE5296, GSE47681, and GSE217797 were downloaded from the Gene Expression Omnibus database. Gene ontology and Kyoto Encyclopedia of Genes and Genomes analyses were performed to determine the function and pathway enrichment of ischemia- and hypoxia-related differentially expressed genes (IAHRDEGs) in SCI. LASSO model was constructed, and support vector machine analysis was used to identify key genes. The diagnostic values of key genes were evaluated using decision curve analysis and receiver operating characteristic curve analysis. The interaction networks of miRNAs-IAHRDEGs and IAHRDEGs-transcription factors were predicted and constructed with the ENCORI database and Cytoscape software. CIBERSORT algorithm was utilized to analyze the correlation between key gene expression and immune cell infiltration. RESULTS: There were 27 IAHRDEGs identified to be significantly expressed in SCI at first. These genes were mostly significantly enriched in wound healing function and the pathway associated with lipid and atherosclerosis. Next, five key IAHRDEGs (Abca1, Casp1, Lpl, Procr, Tnfrsf1a) were identified and predicted to have diagnostic value. Moreover, the five key genes are closely related to immune cell infiltration. CONCLUSION: Abca1, Casp1, Lpl, Procr, and Tnfrsf1a may promote the pathogenesis of ischemic or hypoxic SCI by regulating vascular damage, inflammation, and immune infiltration.

Glycolytic reprogramming fuels myeloid cell-driven hypercoagulability.

BACKGROUND: Myeloid cell metabolic reprogramming is a hallmark of inflammatory disease; however, its role in inflammation-induced hypercoagulability is poorly understood. OBJECTIVES: We aimed to evaluate the role of inflammation-associated metabolic reprogramming in regulating blood coagulation. METHODS: We used novel myeloid cell-based global hemostasis assays and murine models of immunometabolic disease. RESULTS: Glycolysis was essential for enhanced activated myeloid cell tissue factor expression and decryption, driving increased cell-dependent thrombin generation in response to inflammatory challenge. Similarly, inhibition of glycolysis enhanced activated macrophage fibrinolytic activity through reduced plasminogen activator inhibitor 1 activity. Macrophage polarization or activation markedly increased endothelial protein C receptor (EPCR) expression on monocytes and macrophages, leading to increased myeloid cell-dependent protein C activation. Importantly, inflammation-dependent EPCR expression on tissue-resident macrophages was also observed in vivo. Adipose tissue macrophages from obese mice fed a high-fat diet exhibited significantly enhanced EPCR expression and activated protein C generation compared with macrophages isolated from the adipose tissue of healthy mice. Similarly, the induction of colitis in mice prompted infiltration of EPCR+ innate myeloid cells within inflamed colonic tissue that were absent from the intestinal tissue of healthy mice. CONCLUSION: Collectively, this study identifies immunometabolic regulation of myeloid cell hypercoagulability, opening new therapeutic possibilities for targeted mitigation of thromboinflammatory disease.

CD201+ fascia progenitors choreograph injury repair.

Optimal tissue recovery and organismal survival are achieved by spatiotemporal tuning of tissue inflammation, contraction and scar formation1. Here we identify a multipotent fibroblast progenitor marked by CD201 expression in the fascia, the deepest connective tissue layer of the skin. Using skin injury models in mice, single-cell transcriptomics and genetic lineage tracing, ablation and gene deletion models, we demonstrate that CD201+ progenitors control the pace of wound healing by generating multiple specialized cell types, from proinflammatory fibroblasts to myofibroblasts, in a spatiotemporally tuned sequence. We identified retinoic acid and hypoxia signalling as the entry checkpoints into proinflammatory and myofibroblast states. Modulating CD201+ progenitor differentiation impaired the spatiotemporal appearances of fibroblasts and chronically delayed wound healing. The discovery of proinflammatory and myofibroblast progenitors and their differentiation pathways provide a new roadmap to understand and clinically treat impaired wound healing.

Reprogramming tumor-associated macrophages to outcompete endovascular endothelial progenitor cells and suppress tumor neoangiogenesis.

Tumors develop by invoking a supportive environment characterized by aberrant angiogenesis and infiltration of tumor-associated macrophages (TAMs). In a transgenic model of breast cancer, we found that TAMs localized to the tumor parenchyma and were smaller than mammary tissue macrophages. TAMs had low activity of the metabolic regulator mammalian/mechanistic target of rapamycin complex 1 (mTORC1), and depletion of negative regulator of mTORC1 signaling, tuberous sclerosis complex 1 (TSC1), in TAMs inhibited tumor growth in a manner independent of adaptive lymphocytes. Whereas wild-type TAMs exhibited inflammatory and angiogenic gene expression profiles, TSC1-deficient TAMs had a pro-resolving phenotype. TSC1-deficient TAMs relocated to a perivascular niche, depleted protein C receptor (PROCR)-expressing endovascular endothelial progenitor cells, and rectified the hyperpermeable blood vasculature, causing tumor tissue hypoxia and cancer cell death. TSC1-deficient TAMs were metabolically active and effectively eliminated PROCR-expressing endothelial cells in cell competition experiments. Thus, TAMs exhibit a TSC1-dependent mTORC1-low state, and increasing mTORC1 signaling promotes a pro-resolving state that suppresses tumor growth, defining an innate immune tumor suppression pathway that may be exploited for cancer immunotherapy.

Investigating mortality and morbidity associated with UrINary incontinence during Older Womens Secondary Care Admissions and exploring nurses experiences of delivering related care (U-INconti): a mixed methods research protocol.

INTRODUCTION: Urinary incontinence (UI) is associated with increasing age and is more frequently experienced by women. Despite 40% prevalence in the community, little is known about the prevalence/incidence of UI in older women during hospital admission. UI during hospital admissions, within this group, has also been under-researched in terms of its relationship to specific clinical conditions and mortality rates. Given that UI has serious implications for both patient care and women's general health and well-being on discharge, this protocol describes a planned research project which aims to determine mortality, morbidity, prevalence and incidence of UI in older women (≥55 years) during hospital admission to inform nursing practice. Additionally, it aims to explore the experience of nurses who deliver women's care. METHODS AND ANALYSIS: This is an explanatory mixed-methods study consisting of two phases: (1) retrospecitive analysis of electronic patient care records (EPCR) to determine prevalence/incidence of UI, clinical conditions most likely associated with UI and any associations between UI and death, (2) nurse interviews to explore views, knowledge and perceptions of performing the nursing assessment and providing care for older women (≥55 years) with UI during admission. EPCR will be gained from a National Health Service (NHS) teaching hospital. Nurse interviews will be conducted with nurses from an alternative but similar-sized NHS hospital. ETHICS AND DISSEMINATION: Ethical approval is provided by the University of Salford Ethics Committee and regulatory approval by the NHS Health Research Authority (Integrated Research Application System project ID: 303118). Local NHS trust approval to access electronic care records for the purposes of analysis of anonymised data has been provided by one of the two collaborating NHS hospitals. Findings will be disseminated through open-access geriatric or urogynaecology journals and presented to relevant stakeholders at local, national and international meetings including scientific meetings such as the UK Continence Society and International Continence Society.

Natural immunity to malaria preferentially targets the endothelial protein C receptor-binding regions of PfEMP1s.

Antibody responses to variant surface antigens (VSAs) produced by the malaria parasite Plasmodium falciparum may contribute to age-related natural immunity to severe malaria. One VSA family, P. falciparum erythrocyte membrane protein-1 (PfEMP1), includes a subset of proteins that binds endothelial protein C receptor (EPCR) in human hosts and potentially disrupts the regulation of inflammatory responses, which may lead to the development of severe malaria. We probed peptide microarrays containing segments spanning five PfEMP1 EPCR-binding domain variants with sera from 10 Malian adults and 10 children to determine the differences between adult and pediatric immune responses. We defined serorecognized peptides and amino acid residues as those that elicited a significantly higher antibody response than malaria-naïve controls. We aimed to identify regions consistently serorecognized among adults but not among children across PfEMP1 variants, potentially indicating regions that drive the development of immunity to severe malaria. Adult sera consistently demonstrated broader and more intense serologic responses to constitutive PfEMP1 peptides than pediatric sera, including peptides in EPCR-binding domains. Both adults and children serorecognized a significantly higher proportion of EPCR-binding peptides than peptides that do not directly participate in receptor binding, indicating a preferential development of serologic responses at functional residues. Over the course of a single malaria transmission season, pediatric serological responses increased between the start and the peak of the season, but waned as the transmission season ended. IMPORTANCE Severe malaria and death related to malaria disproportionately affect sub-Saharan children under 5 years of age, commonly manifesting as cerebral malaria and/or severe malarial anemia. In contrast, adults in malaria-endemic regions tend to experience asymptomatic or mild disease. Our findings indicate that natural immunity to malaria targets specific regions within the EPCR-binding domain, particularly peptides containing EPCR-binding residues. Epitopes containing these residues may be promising targets for vaccines or therapeutics directed against severe malaria. Our approach provides insight into the development of natural immunity to a binding target linked to severe malaria by characterizing an "adult-like" response as recognizing a proportion of epitopes within the PfEMP1 protein, particularly regions that mediate EPCR binding. This "adult-like" response likely requires multiple years of malaria exposure, as increases in pediatric serologic response over a single malaria transmission season do not appear significant.

Dual roles of the Arabidopsis PEAT complex in histone H2A deubiquitination and H4K5 acetylation.

Histone H2A monoubiquitination is associated with transcriptional repression and needs to be removed by deubiquitinases to facilitate gene transcription in eukaryotes. However, the deubiquitinase responsible for genome-wide H2A deubiquitination in plants has yet to be identified. In this study, we found that the previously identified PWWP-EPCR-ARID-TRB (PEAT) complex components interact with both the ubiquitin-specific protease UBP5 and the redundant histone acetyltransferases HAM1 and HAM2 (HAM1/2) to form a larger version of PEAT complex in Arabidopsis thaliana. UBP5 functions as an H2A deubiquitinase in a nucleosome substrate-dependent manner in vitro and mediates H2A deubiquitination at the whole-genome level in vivo. HAM1/2 are shared subunits of the PEAT complex and the conserved NuA4 histone acetyltransferase complex, and are responsible for histone H4K5 acetylation. Within the PEAT complex, the PWWP components (PWWP1, PWWP2, and PWWP3) directly interact with UBP5 and are necessary for UBP5-mediated H2A deubiquitination, while the EPCR components (EPCR1 and EPCR2) directly interact with HAM1/2 and are required for HAM1/2-mediated H4K5 acetylation. Collectively, our study not only identifies dual roles of the PEAT complex in H2A deubiquitination and H4K5 acetylation but also illustrates how these processes collaborate at the whole-genome level to regulate the transcription and development in plants.

Stromal heterogeneity may explain increased incidence of metaplastic breast cancer in women of African descent.

The biologic basis of genetic ancestry-dependent variability in disease incidence and outcome is just beginning to be explored. We recently reported enrichment of a population of ZEB1-expressing cells located adjacent to ductal epithelial cells in normal breasts of women of African ancestry compared to those of European ancestry. In this study, we demonstrate that these cells have properties of fibroadipogenic/mesenchymal stromal cells that express PROCR and PDGFRα and transdifferentiate into adipogenic and osteogenic lineages. PROCR + /ZEB1 + /PDGFRα+ (PZP) cells are enriched in normal breast tissues of women of African compared to European ancestry. PZP: epithelial cell communication results in luminal epithelial cells acquiring basal cell characteristics and IL-6-dependent increase in STAT3 phosphorylation. Furthermore, level of phospho-STAT3 is higher in normal and cancerous breast tissues of women of African ancestry. PZP cells transformed with HRasG12V ± SV40-T/t antigens generate metaplastic carcinoma suggesting that these cells are one of the cells-of-origin of metaplastic breast cancers.

Comprehensive genome-wide identification and transferability of chromosome-specific highly variable microsatellite markers from citrus species.

Citrus species among the most important and widely consumed fruit in the world due to Vitamin C, essential oil glands, and flavonoids. Highly variable simple sequence repeats (SSR) markers are one of the most informative and versatile molecular markers used in perennial tree genetic research. SSR survey of Citrus sinensis and Citrus maxima were identified perfect SSRs spanning nine chromosomes. Furthermore, we categorized all SSR motifs into three major classes based on their tract lengths. We designed and validated a class I SSRs in the C. sinensis and C. maxima genome through electronic polymerase chain reaction (ePCR) and found 83.89% in C. sinensis and 78.52% in C. maxima SSRs producing a single amplicon. Then, we selected extremely variable SSRs (> 40 nt) from the ePCR-verified class I SSRs and in silico validated across seven draft genomes of citrus, which provided us a subset of 84.74% in C. sinensis and 77.53% in C. maxima highly polymorphic SSRs. Out of these, 129 primers were validated on 24 citrus genotypes through wet-lab experiment. We found 127 (98.45%) polymorphic HvSSRs on 24 genotypes. The utility of the developed HvSSRs was demonstrated by analysing genetic diversity of 181 citrus genotypes using 17 HvSSRs spanning nine citrus chromosomes and were divided into 11 main groups through 17 HvSSRs. These chromosome-specific SSRs will serve as a powerful genomic tool used for future QTL mapping, molecular breeding, investigation of population genetic diversity, comparative mapping, and evolutionary studies among citrus and other relative genera/species.

Reduced Production of JAK2V617F-positive Microparticles at Mild Hypothermia.

BACKGROUND/AIM: The Philadelphia chromosome-negative (Ph-) myeloproliferative neoplasms (MPNs) are a group of blood cancers that arise from abnormal growth of blood cells in the bone marrow. Patients with MPNs are at increased risk for life-threatening thromboembolic complications. The detection of JAK2V617F in endothelial cells (ECs) brought a new perspective to the research of thromboembolic events. However, the mechanisms by which the mutation contributes to risk have yet to be entirely understood. Consequently, the objective of this study was to investigate how JAK2V617F impacts endothelial cells by considering thermoregulation. MATERIALS AND METHODS: We applied our previously created model for EC that was genetically modified with JAK2 wild type (WT)-GFP and JAK2V617F-GFP lentiviruses; the cells were cultured for 48 h at 37°C for normothermia and 32°C for mild hypothermia. We examined the effect of thermoregulation on infection efficiency and the expression of cell surface markers, including endothelial protein C receptor (EPCR), thrombomodulin (TM), and tissue factor (TF), which are related to the coagulation pathways. Furthermore, the microparticle production from the genetically modified EC (EMPs) was analyzed. RESULTS: We found suppression of the expression of coagulation factors, including EPCR, TM, and TF in JAK2V617F positive ECs under mild hypothermia. JAK2V617F-positive ECs showed slightly higher EMP production under mild hypothermia. CONCLUSION: Although the molecular mechanisms of the thermal effects on the tumor microenvironment with JAK2V617F and its effect on EMP production and coagulation are not known yet, the therapy-oriented effect of thermoregulation might be considered in future studies.

Soluble endothelial protein C receptor is an independent predictor of venous thromboembolism after severe injury: Secondary analysis of a prospective cohort study.

BACKGROUND: Venous thromboembolism is a leading cause of morbidity after trauma. Endothelial cells are essential regulators of coagulation. Although endothelial cell dysregulation is widely reported after trauma, the link between endothelial injury and venous thromboembolism has not been reported. METHODS: We conducted a secondary analysis of the Pragmatic Randomized Optimal Platelets and Plasma Ratios study. Deaths from hemorrhage or within 24 hours were excluded. Venous thromboembolism was diagnosed by duplex ultrasound or chest computed tomography. Endothelial markers soluble endothelial protein c receptor, thrombomodulin, and syndecan-1 were measured in plasma by enzyme-linked immunosorbent assay and compared over the first 72 hours from admission using the Mann-Whitney test. Multivariable logistic regression assessed the adjusted effects of endothelial markers on venous thromboembolism risk. RESULTS: Of 575 patients enrolled, 86 developed venous thromboembolism (15%). The median time to venous thromboembolism was 6 days ([Q1, Q3], [4, 13]). No differences were identified in demographics or injury severity. Soluble endothelial protein c receptor, thrombomodulin, and syndecan-1 showed significant increases over time among patients who developed venous thromboembolism compared to those who did not. Using the last available values, patients were stratified into high and low-soluble endothelial protein c receptor, thrombomodulin, and syndecan-1 groups. Multivariable analyses revealed an independent association between elevated soluble endothelial protein c receptor and venous thromboembolism risk (odds ratio 1.63; 95% confidence interval 1.01, 2.63; P = .04). Cox proportional hazards modeling demonstrated a strong yet nonsignificant trend between elevated soluble endothelial protein c receptor and time to venous thromboembolism. CONCLUSION: Plasma markers of endothelial injury, particularly soluble endothelial protein c receptor, are strongly associated with trauma-related venous thromboembolism. Therapeutics targeting endothelial function could mitigate the incidence of venous thromboembolism after trauma.

Management of Postpartum Extensive Venous Thrombosis after Second Pregnancy.

Background: Pregnancy induces a physiological prothrombotic state. The highest risk period for venous thromboembolism and pulmonary embolism in pregnant women is during the postpartum period. Materials and Methods: We present the case of a young woman who gave birth 2 weeks before admission and was transferred to our clinic for edema. She had an increased temperature in her right limb, and a venous Doppler of the limb confirmed thrombosis of the right femoral vein. From the paraclinical examination, we obtained a CBC with leukocytosis, neutrophilia, and thrombocytosis, and a positive D-dimer test. Thrombophilic tests were negative for AT III, lupus anticoagulant negative, and protein S and C, but were positive for heterozygous PAI-1, heterozygous MTHFR A1298C, and EPCR with A1/A2 alleles. After 2 days of UFH with therapeutic APTT, the patient had pain in her left thigh. We performed a venous Doppler, which revealed bilateral femoral and iliac venous thrombosis. During the computed tomography examination, we assessed the venous thrombosis extension on the inferior cava, common iliac, and bilateral common femoral veins. Thrombolysis was initiated with 100 mg of Alteplase given at a rate of 2 mg/h; however, this did not lead to a considerable reduction in the thrombus. Additionally, the treatment with UFH was continued under therapeutic APTT. After 7 days of UFH and triple antibiotic therapy for genital sepsis, the patient had a favorable evolution with remission of venous thrombosis. Results: Alteplase is a thrombolytic agent that is created with recombinant DNA technology, and it was successfully used to treat thrombosis that occurred in the postpartum period. Conclusions: Thrombophilias are associated with a high VTE risk but also with adverse pregnancy outcomes, including recurrent miscarriages and gestational vascular complications. In addition, the postpartum period is associated with a higher VTE risk. A thrombophilic status with heterozygous PAI-1, heterozygous MTHFR A1298C, and EPCR with A1/A2 positive alleles is associated with a high risk of thrombosis and cardiovascular events. Thrombolysis can be successfully used postpartum to treat VTEs. Thrombolysis can be used successfully in VTE developed in the postpartum period.

Microvascular Environment Influences Brain Microvascular Heterogeneity: Relative Roles of Astrocytes and Oligodendrocytes for the EPCR Expression in the Brain Endothelium.

Postmortem neuropathology shows clear regional differences in many brain diseases. For example, brains from cerebral malaria (CM) patients show more hemorrhagic punctae in the brain's white matter (WM) than grey matter (GM). The underlying reason for these differential pathologies is unknown. Here, we assessed the effect of the vascular microenvironment on brain endothelial phenotype, focusing endothelial protein C receptor (EPCR). We demonstrate that the basal level of EPCR expression in cerebral microvessels is heterogeneous in the WM compared to the GM. We used in vitro brain endothelial cell cultures and showed that the upregulation of EPCR expression was associated with exposure to oligodendrocyte conditioned media (OCM) compared to astrocyte conditioned media (ACM). Our findings shed light on the origin of the heterogeneity of molecular phenotypes at the microvascular level and might help better understand the variation in pathology seen in CM and other neuropathologies associated with vasculature in various brain regions.

Differential expression of endothelial protein C receptor (EPCR) in hematopoietic stem and multipotent progenitor cells in young and old mice.

Endothelial protein C receptor (EPCR) has emerged as one of the most conserved and reliable surface markers for the prospective identification and isolation of hematopoietic stem cells (HSCs). Prior studies have consistently demonstrated that EPCR expression enriches HSCs capable of long-term multilineage repopulation in both mouse and human across different hematopoietic tissues, including bone marrow (BM), fetal liver and ex vivo HSC expansion cultures. However, little is known about the expression profiles of EPCR in multipotent progenitor (MPP) populations located immediately downstream of HSCs in the hematopoietic hierarchy and which play a major role in sustaining lifelong blood cell production. Here, we incorporate EPCR antibody detection into a multi-parameter flow cytometric panel, which allows accurate identification of HSCs and five MPP subsets (MPP1-5) in mouse BM. Our data reveal that all MPP populations contain EPCR-expressing cells. Multipotent MPP1 and MPP5 contain higher proportion of EPCR+ cells compared to the more lineage-biased MPP2-4. Notably, high expression of EPCR enriches phenotypic HSC and MPP5, but not MPP1. Comparison of EPCR expression profiles between young and old BM reveals ageing mediated expansion of EPCR-expressing cells only in HSCs, but not in any of the MPP populations. Collectively, our study provides a comprehensive characterization of the surface expression pattern of EPCR in mouse HSC and MPP1-5 cells during normal and aged hematopoiesis.