Publications

A 55-year-old man with alcoholism required sigmoid resection for complicated diverticulitis.A few weeks after starting parenteral nutrition, he developed multidirectional gaze nystagmus, gait ataxia, and confusion, leading to the diagnosis of Wernicke encephalopathy.Thiamine treatment was started and his symptoms improved, helping to confirm the diagnosis.However, the patient developed memory loss consistent with Korsakoff syndrome due to hemorrhage involving the fornix and the bilateral anterior thalami (figure).More typical fluid-attenuated inversion recovery lesions (mamillary bodies, tectal plate, and periaqueductal area), which may be reversible, 1 were not seen.Macroscopic hemorrhage is rarely observed in Wernicke-Korsakoff syndrome 2,3 and is associated with poor prognosis. 3

Purpose: Aortic valve stenosis (AVS) is an inflammatory atherosclerosis-like process characterized by valve interstitial cell (VIC) proliferation and commitment to fibrocalcification. No effective medical treatment is currently able to alter AVS clinical course. Tissue factor (TF) and thrombin expressions were previously shown to be significantly associated with fibrocalcification in human stenosed aortic valves. We investigated differential TF expression in normal and pathological valves and VIC and further described its signalling pathways in profibrosis commitment of normal VIC. Methods: We collected 50 fibrocalcific and 6 normal human aortic valves from which tissue lysates and VIC were isolated. VIC phenotype was analysed by flow cytometry. Normal VIC were stimulated or not with IL-1beta (major inflammatory cytokine involved in AVS at 1 ng/mL). In valve lysates and in VIC, TF and PAR-2 were quantified by qPCR for relative mRNA expression, and by flow cytometry/ELISA for protein and activity quantification. Downstream signalling pathways ERK (cell activation and proliferation) and Smad2 (profibrosis transcription factor) of VIIa/TF in TF-overexpressing VIC were assessed by westernblotting (incubation with FVIIa at 50 ng/mL for 30 min) in presence or absence of anti-TF antibody. Results are expressed as median (interquartile range). Results: VIC were defined as CD31(-) and CD68(-). TF and PAR-2 relative mRNA expressions were significantly higher in fibrocalcific vs normal valves (3.2 (1.5-.8) vs 0.18 (0.02-0.29), p<0.05; 1.15 (0.48-1.62) vs 0.16 (0.02-0.22), p<0.05; respectively). Normal VIC constitutively expressed TF (relative mRNA 4.1 (2.1-21.2); activity 36.2 (13.2-101.3) pg/mL) as well as PAR-2 (mRNA and protein by flow cytometry). TF expression was significantly increased in fibrocalcific VIC (relative mRNA 22.8 (9.8-33.8), p=0.05; activity 153 (71.5-247.0) pg/mL, p<0.05) when compared to normal VIC. Following IL-1beta stimulation of VIC, TF expression was significantly upregulated compared to unstimulated cells (relative mRNA 15.4 (12.9-28.2) fold increase, p<0.05; activity 2.7 (2.2-5.5) fold increase, p<0.05). ERK and Smad2 signalling pathways were upregulated in VIIa stimulated TF-overexpressing VIC (pERK/ERK 3.5 (2.9-4.8) fold increase; pSmad2/Smad2/3 1.8 (1.2-1.8) fold increase), an upregulation blunted by VIC preincubation with an anti-TF antibody. Conclusions: Our results demonstrate the implication of TF/VIIa/PAR-2 axis in VIC commitment to fibrocalcific aortic valve disease. Modulation of this pathway may represent a new therapeutic target for early medical treatment of AVS.

The effect of cannabidiol (CBD), a high-affinity agonist of the transient receptor potential vanilloid-2 (TRPV2) channel, has been poorly investigated in human brain microvessel endothelial cells (BMEC) forming the blood-brain barrier (BBB). TRPV2 expression and its role on Ca²⁺ cellular dynamics, trans-endothelial electrical resistance (TEER), cell viability and growth, migration, and tubulogenesis were evaluated in human primary cultures of BMEC (hPBMEC) or in the human cerebral microvessel endothelial hCMEC/D3 cell line. Abundant TRPV2 expression was measured in hCMEC/D3 and hPBMEC by qRT-PCR, Western blotting, nontargeted proteomics, and cellular immunofluorescence studies. Intracellular Ca²⁺ levels were increased by heat and CBD and blocked by the nonspecific TRP antagonist ruthenium red (RR) and the selective TRPV2 inhibitor tranilast (TNL) or by silencing cells with TRPV2 siRNA. CBD dose-dependently induced the hCMEC/D3 cell number (EC₅₀ 0.3 ± 0.1 μM), and this effect was fully abolished by TNL or TRPV2 siRNA. A wound healing assay showed that CBD induced cell migration, which was also inhibited by TNL or TRPV2 siRNA. Tubulogenesis of hCMEC/D3 cells in 3D matrigel cultures was significantly increased by 41 and 73% after a 7 or 24 h CBD treatment, respectively, and abolished by TNL. CBD also increased the TEER of hPBMEC monolayers cultured in transwell, and this was blocked by TNL. Our results show that CBD, at extracellular concentrations close to those observed in plasma of patients treated by CBD, induces proliferation, migration, tubulogenesis, and TEER increase in human brain endothelial cells, suggesting CBD might be a potent target for modulating the human BBB.

Endoglin (ENG) is a transmembrane glycoprotein expressed on endothelial cells that functions as a co-receptor for several ligands of the transforming growth factor beta (TGF-β) family. ENG is also a recognized marker of angiogenesis and mutations in the endoglin gene are responsible for Hereditary Hemorrhagic Telangiectasia (HHT) type 1, a vascular disease characterized by defective angiogenesis, arteriovenous malformations, telangiectasia, and epistaxis. In addition to its involvement in the TGF-β family signaling pathways, several lines of evidence suggest that the extracellular domain of ENG has a role in integrin-mediated cell adhesion via its RGD motif. Indeed, we have described a role for endothelial ENG in leukocyte trafficking and extravasation <i>via</i> its binding to leukocyte integrins. We have also found that ENG is involved in vasculogenic properties of endothelial progenitor cells known as endothelial colony forming cells (ECFCs). Moreover, the binding of endothelial ENG to platelet integrins regulate the resistance to shear during platelet-endothelium interactions under inflammatory conditions. Because of the need for more effective treatments in HHT and the involvement of ENG in angiogenesis, current studies are aimed at identifying novel biological functions of ENG which could serve as a therapeutic target. This review focuses on the interaction between ENG and integrins with the aim to better understand the role of this protein in blood vessel formation driven by progenitor and mature endothelial cells.

Background Pulmonary arterial hypertension (PAH) results from pulmonary vascular disease and may eventually lead to right heart failure and death. Vasodilator therapy has greatly improved PAH prognosis. Circulating microvesicles are considered as surrogate markers of endothelial and hematopoietic cell activation. Aim Thus, our purpose was to determine if MVs are upregulated in pediatric PAH such as reported in adult patients, and to analyze the impact of vasodilator therapies on MV count and function. Patients Population study consisted of 26 patients of median age 6.09 years, with Congenital Heart Disease (CHD) and elevated pulmonary vascular resistance (CHD‐PAH) or idiopathic PAH (iPAH). Results Compared to healthy controls, all circulating MV subpopulations were found higher in untreated PAH patients. No significant differences of annexin‐V+ total MV, endothelial, or leukocyte derived‐MV counts were found between untreated patients and those receiving oral vasodilator therapies. Conversely, platelet MVs were significantly lower in the group treated with SC‐treprostinil compared with both untreated PAH and oral therapy groups ( P = 0.01), and exhibited a significant decrease of phospholipid procoagulant activity. Control samples treated in vitro with treprostinil at therapeutic concentrations showed as expected a significant decrease of platelet aggregation but also a reduced spontaneous MV generation. Conclusion Our results suggest that treprostinil, besides vasodilation, might exert its beneficial effect through an inhibition of platelet activation, resulting in a decreased number and procoagulant activity of circulating MVs.

To investigate the mechanisms underlying the SARS-CoV-2 infection severity observed in patients with obesity, we performed a prospective study of 51 patients evaluating the impact of multiple immune parameters during 2 weeks after admission, on vital organs’ functions according to body mass index (BMI) categories. High-dimensional flow cytometric characterization of immune cell subsets was performed at admission, 30 systemic cytokines/chemokines levels were sequentially measured, thirteen endothelial markers were determined at admission and at the zenith of the cytokines. Computed tomography scans on admission were quantified for lung damage and hepatic steatosis (n = 23). Abnormal BMI (&gt; 25) observed in 72.6% of patients, was associated with a higher rate of intensive care unit hospitalization (p = 0.044). SARS-CoV-2 RNAaemia, peripheral immune cell subsets and cytokines/chemokines were similar among BMI groups. A significant association between inflammatory cytokines and liver, renal, and endothelial dysfunctions was observed only in patients with obesity (BMI &gt; 30). In contrast, early signs of lung damage (ground-glass opacity) correlated with Th1/M1/inflammatory cytokines only in normal weight patients. Later lesions of pulmonary consolidation correlated with BMI but were independent of cytokine levels. Our study reveals distinct physiopathological mechanisms associated with SARS-CoV-2 infection in patients with obesity that may have important clinical implications.