POS0424 DETERMINING CIRCULATING ENDOTHELIAL CELLS USING CELLSEARCH SYSTEM IN SYSTEMIC SCLEROSIS PATIENTS
Authors: F. Pignataro, L. Zorzino, W. Maglione, A. Minniti, G. Clericuzio, M. Picozzi, C. Simonelli, F. Picardo, R. Caporali, N. Del Papa
Journal: Annals of the Rheumatic Diseases (2021)

Background: Endothelial damage and fibroproliferative vasculopathy of small vessels are pathological hallmarks of Systemic Sclerosis (SSc). Detection and analysis of circulating endothelial cells (CECs) detached from affected blood vessels may be an informative tool to study vascular dysfunction and could be considered a novel biomarker of scleroderma vasculopathy. Our group first showed the presence of CECs in SSc by fluorescence-activated cell sorting (FACS), demonstrating that a raised counts of active CECs may represent direct evidence of active vascular disease in SSc. Despite these interesting data, issues related to difficulties in CEC counting through FACS analysis, due their very low concentration in peripheral blood, prevented further investigations in this field. Recently, a specific kit for the detection of CECs has been developed through the CellSearch System (CS), a semi-automated device for the standardized analysis of rare cells, such as CECs, in peripheral blood. Objectives: To assess the counts of CECs determined by the CS in SSc patients and to evaluate their clinical implication and potential as vascular biomarker in SSc. Methods: 10mL of blood samples were collected from 29 subjects (19 SSc patients and 10 healthy donors - HDs) and stored in tubes containing a specific preservative, to allow the analysis of 4mL of blood within 72 hours, according to manufacturer instructions. Out of 19 SSc patients, 18 were female, 10 had the limited form and 9 the diffuse cutaneous variant of SSc. CS uses a proprietary kit containing a ferrofluid-based reagent, that target CD146 to magnetically capture CECs, and the immunofluorescent reagents to stain the CECs, defined as CD146+, CD105-PE+, DAPI+ and CD45-APC-. Clinical, laboratoristic and demographic data were also collected. Results: The mean number of CECs in patients with SSc was significantly higher in comparison to HDs (554/4mL vs. 53.5/4 mL, p=0.0042). When analyzed according to disease subset, both lcSSc and dcSSc showed significantly increased levels of CECs in comparison with HDs (p=0.003 and p=0.005, respectively). No statistical difference was observed in the mean number of CECs in patients with lcSSc compared to those with dcSSc. Regarding vascular involvement, the CECs counts strictly correlated with the presence of digital ulcers (DUs) (p=0.0001) showing a median of 863cells/4mL for the SSc patients with DUs versus a median of 276.2/4mL for the SSc patients without DUs. No statistical correlation was found between CECs and serological autoantibody pattern, skin parameters, or joint and muscle involvement. Patients with active disease, according to the EUSTAR Activity Index, showed a higher CECs value than those with inactive disease (p=0.0012). Conclusion: The amount of CECs detectable in peripheral blood has been recently proposed as a marker of endothelial damage in different vascular diseases, including SSc. However, currently no standardized method is available to determine CEC counts, which makes reported data on CECs reliable and suitable. The CS system is a commercially available semi-automated system that enables standardized determination of CECs. Thus, we examined clinical utility of CECs count by this system in SSc patients. Our results confirm that baseline CEC counts, evaluated by a new standardized method, may represent direct evidence of endothelial damage in SSc and could be a promising tool for monitoring active disease and evaluating therapeutic responses to vascular and immunosuppressive treatments. References: [1]Del Papa N, Pignataro F. Front Immunol. 2018 Jun 18;9:1383 [2]De Simone C et al. J Eur Acad Dermatol Venereol. 2014 May;28(5):590-6 [3]Del Papa N et al. Arthritis Rheum. 2004 Apr;50(4):1296-304 Disclosure of Interests: Francesca Pignataro: None declared, Laura Zorzino: None declared, Wanda Maglione: None declared, Antonina Minniti: None declared, Giulia Clericuzio: None declared, Marco Picozzi: None declared, Cecilia Simonelli Employee of: Menarini Silicon Biosystems, Francesco Picardo Employee of: Menarini Silicon Biosystems, Roberto Caporali: None declared, Nicoletta Del Papa: None declared