thromboplastin and Inflammatory-Bowel-Diseases

Intestinal mesenchymal cells play essential roles in epithelial homeostasis, matrix remodeling, immunity, and inflammation. But the extent of heterogeneity within the colonic mesenchyme in these processes remains unknown. Using unbiased single-cell profiling of over 16,500 colonic mesenchymal cells, we reveal four subsets of fibroblasts expressing divergent transcriptional regulators and functional pathways, in addition to pericytes and myofibroblasts. We identified a niche population located in proximity to epithelial crypts expressing SOX6, F3 (CD142), and WNT genes essential for colonic epithelial stem cell function. In colitis, we observed dysregulation of this niche and emergence of an activated mesenchymal population. This subset expressed TNF superfamily member 14 (TNFSF14), fibroblastic reticular cell-associated genes, IL-33, and Lysyl oxidases. Further, it induced factors that impaired epithelial proliferation and maturation and contributed to oxidative stress and disease severity in vivo. Our work defines how the colonic mesenchyme remodels to fuel inflammation and barrier dysfunction in IBD.

Topics: Animals; Cell Proliferation; Colitis; Colon; Epithelial Cells; Fibroblasts; Genetic Heterogeneity; Homeostasis; Humans; Inflammation; Inflammatory Bowel Diseases; Intestinal Mucosa; Intestines; Mesenchymal Stem Cells; Mesoderm; Mice; Mice, Inbred C57BL; Myofibroblasts; Pericytes; RAW 264.7 Cells; Single-Cell Analysis; SOXD Transcription Factors; Thromboplastin; Tumor Necrosis Factor Ligand Superfamily Member 14; Wnt Signaling Pathway

Circulating procoagulant microparticles (MPs) are thought to be involved in the pathogenesis of venous thromboembolism in patients with inflammatory bowel disease (IBD). However, the exposure of tissue factor, the primary initiator of coagulation activation, on microparticles (TF(+)MPs) and its association with hemostasis activation has not yet been studied in IBD patients.. In this case-control study 49 IBD patients (28 Crohn's disease, 21 ulcerative colitis) and 49 sex- and age-matched, healthy controls were included. Clinical disease activity (Crohn's Disease Activity Index and Clinical Activity Index, respectively) was assessed and IBD-related data were determined by chart review. Numbers, cellular origin and procoagulant activity of TF(+)MPs in plasma were determined using flow cytometry and a chromogenic activity assay. D-dimer and high-sensitive C-reactive protein (CRP) served as markers for coagulation activation and inflammation, respectively. The primary endpoint was the number of TF(+)MPs in IBD patients compared to controls.. Median number (interquartile range) of TF(+)MPs was higher in IBD patients than in controls (14.0 (11.9-22.8)×10(3)/mL vs. 11.9 (11.9-19.1)×10(3)/mL plasma, P=0.029). This finding was due to generally higher plasma levels of MPs from platelets and leukocytes in IBD patients. However, the number of TF(+)MPs was neither correlated with their procoagulant activity and D-dimer nor with disease activity and CRP.. Increased numbers of circulating TF(+)MPs represent a new facet of hemostatic abnormalities in IBD. However, the lack of association with activation of the coagulation system and disease activity questions their pathogenetic role for venous thromboembolism in this patient group.

Topics: Adult; Biomarkers; Blood Coagulation; C-Reactive Protein; Case-Control Studies; Cell-Derived Microparticles; Female; Fibrin Fibrinogen Degradation Products; Flow Cytometry; Humans; Inflammatory Bowel Diseases; Male; Middle Aged; Prospective Studies; Thromboplastin

Although angiogenesis is viewed as a fundamental component of inflammatory bowel disease (IBD) pathogenesis, we presently lack a thorough knowledge of the cell type(s) involved in its induction and maintenance in the inflamed intestinal mucosa. This study aimed to determine whether platelet (PLT) adhesion to inflamed intestinal endothelial cells of human origin may favour angiogenesis. Unstimulated or thrombin-activated human PLT were overlaid on resting or tumour necrosis factor (TNF)-α-treated human intestinal microvascular endothelial cells (HIMEC), in the presence or absence of blocking antibodies to either vascular cell adhesion molecule (VCAM)-1, intercellular adhesion molecule (ICAM)-1, integrin α(v)β(3) , tissue factor (TF) or fractalkine (FKN). PLT adhesion to HIMEC was evaluated by fluorescence microscopy, and release of angiogenic factors (VEGF and soluble CD40L) was measured by ELISA. A matrigel tubule formation assay was used to estimate PLT capacity to induce angiogenesis after co-culturing with HIMEC. TNF-α up-regulated ICAM-1, α(v)β(3) and FKN expression on HIMEC. When thrombin-activated PLT were co-cultured with unstimulated HIMEC, PLT adhesion increased significantly, and this response was further enhanced by HIMEC activation with TNF-α. PLT adhesion to HIMEC was VCAM-1 and TF independent but ICAM-1, FKN and integrin α(v)β(3) dependent. VEGF and sCD40L were undetectable in HIMEC cultures either before or after TNF-α stimulation. By contrast, VEGF and sCD40L release significantly increased when resting or activated PLT were co-cultured with TNF-α-pre-treated HIMEC. These effects were much more pronounced when PLT were derived from IBD patients. Importantly, thrombin-activated PLT promoted tubule formation in HIMEC, a functional estimate of their angiogenic potential. In conclusion, PLT adhesion to TNF-α-pre-treated HIMEC is mediated by ICAM-1, FKN and α(v)β(3) , and is associated with VEGF and sCD40L release. These findings suggest that inflamed HIMEC may recruit PLT which, upon release of pro-angiogenic factors, actively contribute to inflammation-induced angiogenesis.

Topics: Blood Platelets; CD40 Ligand; Cells, Cultured; Chemokine CX3CL1; Coculture Techniques; Endothelial Cells; Enzyme-Linked Immunosorbent Assay; Female; Humans; Inflammation; Inflammatory Bowel Diseases; Integrin alphaVbeta3; Intercellular Adhesion Molecule-1; Intestines; Male; Microscopy, Fluorescence; Microvessels; Neovascularization, Pathologic; Platelet Adhesiveness; Thromboplastin; Tumor Necrosis Factor-alpha; Vascular Cell Adhesion Molecule-1; Vascular Endothelial Growth Factor A

Inflammatory bowel diseases (IBD) are characterized by an increased thrombotic risk of uncertain etiology. Endogenous thrombin potential (ETP), a parameter of the thrombin generation curve, represents a new tool in the evaluation of thrombotic and bleeding disorders.. To study ETP in IBD patients and to correlate the results with clinical and biochemical features.. Seventy-four IBD patients (37 ulcerative colitis and 37 Crohn's disease) and 74 sex- and age-matched healthy individuals. ETP was measured upon activation of coagulation with small amounts of tissue factor and phospholipids in the presence or absence of thrombomodulin; results were expressed as nM thrombin.minutes.. Mean+/-SD ETP values were significantly higher in patients (1,499+/-454) than controls (1,261+/-385) (p<0.001) only when the test was performed in the presence of thrombomodulin. ETP evaluated as ratio (with/without thrombomodulin), taken as an index of hypercoagulability, was significantly higher in patients (0.69+/-0.14) than controls (0.62+/-0.18) (p<0.006). Patients with increased C-reactive protein (CRP) had significantly higher mean ETP (1,721+/-458) than those with normal CRP (1,357+/-394) or controls (1,261+/-385) (p<0.001). Patients who at the time of blood sampling were classified as having a clinically active disease had ETP higher than those who were quiescent (1,655+/-451 versus 1,388+/-427, p<0.001) or controls (1,261+/-385, p<0.001).. ETP measured in the presence of thrombomodulin or as ratio (with/without thrombomodulin) is increased in IBD patients, mainly in those with increased CRP or active disease. It may be considered as a candidate test for prospective studies aimed at assessing the risk of thrombosis in IBD patients.

Topics: Adult; C-Reactive Protein; Case-Control Studies; Colitis, Ulcerative; Crohn Disease; Factor VIII; Female; Humans; Inflammatory Bowel Diseases; Male; Middle Aged; Phospholipids; Prospective Studies; Thrombin; Thrombomodulin; Thromboplastin

Topics: Arthritis, Rheumatoid; Female; Fibrocystic Breast Disease; Humans; Inflammatory Bowel Diseases; Male; Neoplasms; Thromboplastin