This might result from the fact that E-selectin is expressed exclusively on activated ECs and is cleaved from ECs that are activated but not damaged; thus the increase in activated E-selectin positive EMPs might reflect early ECs dysfunction in contrast to activated CECs, whose detection requires severe injury and sloughing of ECs [8, 11C13, 20]
This might result from the fact that E-selectin is expressed exclusively on activated ECs and is cleaved from ECs that are activated but not damaged; thus the increase in activated E-selectin positive EMPs might reflect early ECs dysfunction in contrast to activated CECs, whose detection requires severe injury and sloughing of ECs [8, 11C13, 20]. or Tie2 receptors. Conclusions We found an association between AECAs and the severity of endothelial damage in SSc based on higher levels of total EMPs and CECs. In our study, AECAs were associated with apoptosis of ECs rather than their activation. We also identified a possible role of AECAs in the impairment of vascular repair in SSc as evidenced by significantly fewer angiogenic EPCs. by endothelium-derived biomarkers in the blood. These include circulating endothelial cells (CECs) and endothelial microparticles (EMPs) [8C12]. The CECs are mature endothelial cells that Alpelisib hydrochloride are injured and detached from the vessel wall into the bloodstream. The EMPs are microvesicles shed from the cellular membrane during Alpelisib hydrochloride ECs activation or apoptosis, and their release closely reflects the degree of microvascular damage. They expose specific surface markers identifying their cellular origin and are phenotypically distinct when released from either activated or apoptotic ECs [11C14]. In response to vascular injury, repair mechanisms have been shown to involve the recruitment of circulating bone marrow-derived endothelial progenitor cells (EPCs) that home into sites of insulted endothelium, differentiate into the mature endothelial phenotype and restore its integrity as well as promote neovascularization [10, 15, 16]. Aim Based on the hypothesis that vascular damage in SSc might be associated with AECAs, we aimed to provide a combined assessment of their possible inter-relationship with the number and phenotype of EMPs and CECs. A further goal was to Alpelisib hydrochloride determine whether AECAs could affect endothelial repair capacity by counting circulating EPCs in AECA-positive versus AECA-negative SSc patients. Material and methods The study enrolled 47 SSc patients, referred to the Department of Dermatology, Medical University of Lublin. All patients were women and fulfilled the American College of Rheumatology (ACR)/European League Against Rheumatism (EULAR) 2013 classification criteria . Forty of them were diagnosed with limited SSc (lcSSc) and 7 with diffuse SSc (dcSSc) in accordance with LeRoy = 47) (%):Limited (lcSSc)40 (85)Diffuse (dcSSc)7 (15)Disease duration, mean (SD), range [years]9.99 (6.96), 2C34Antinuclear antibodies, (%):Positive45 (96)Anti-topoisomerase I positive26 (55)Anti-centromere positive17 (36)Disease activity (EScSG-AI), (%):Active disease11 (23.4)Inactive disease36 (76.6)Active digital ulcers, (%):Present9 (19)Absent38 (81)Capillaroscopic pattern:Early14 (30)Active14 (30)Late19 (40) Open in a separate window All patients were on stable treatment for at least 6 months, including low dose prednisone ( 10 mg/day), vasodilators (calcium channel blockers or angiotensin-converting enzyme inhibitors) and pentoxifylline. None of the patients was on endothelin receptor blockers or treated with prostacyclin. Six patients received intravenous cyclophosphamide; however, in those patients blood samples were drawn at least 3 months after the last pulse. The study protocol was in accordance with the Helsinki Declaration of 1975 as revised in 2000 and was approved by the institutional review board for human research at the Medical University of Lublin. All participants signed informed consent for experimentation. AECAs identification To measure the presence of AECAs, 5 ml blood samples were collected. An indirect immunofluorescence procedure with human umbilical vein endothelial cells (HUVECs) and monkey skeletal muscles was performed using TITERPLANE technique (EUROIMMUN). The procedure has been described previously . Briefly, HUVECs cultures and frozen monkey skeletal muscles were incubated for 30 min with diluted Rabbit polyclonal to ABCG5 1 : 100 sera from SSc patients on biochip platforms, then rinsed with phosphate-buffered saline (PBS) Tween wash buffer and incubated with goat anti-human IgG antibodies marked with fluorescein to identify AECAs bound to the substrate. Positive reaction was assessed qualitatively using a Nikon TS100 fluorescent microscope (Figure 1). Open in a separate window Figure 1 Positive anti-endothelial cell reaction on human umbilical vein endothelial cells (HUVECs) and monkey skeletal muscles manifesting with cytoplasmic, granular, yellowish-green fluorescence concentrated around nuclei Preparation of platelet-poor-plasma for EMPs The EMPs were identified in platelet-poor plasma (PPP) according to the procedures described by Jimenez 0.05. All results are presented as mean standard deviation (SD). Results Patients were classified as either having (44.7%, = 21) or not having (55.3%, = 26) AECAs. Observed associations between the presence of AECAs and endothelial markers were as follows: i) AECA-positive patients had significantly higher values of total plasma EMPs [CD31+/CD41(C)] compared to those who were negative for these autoantibodies (= 0.037) (Figure 4 A), ii) SSc patients with AECAs had significantly higher plasma levels.