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P279. Mobilization of various types of pluripotent progenitor and stem cells is surrogate prognostic marker of mucosal healing in early clinical remission in Crohn's disease

W. Marlicz1, E. Zuba-Surma2, W. Blogowski3, M. Kucia4, T. Starzynska1, M. Ratajczak4

1Pomeranian Medical University, Department of Gastroenterology, Szczecin, Poland; 2Jagiellonian University, Department of Biochemistry, Cracow, Poland; 3Pomeranian Medical University, Department of Laboratory Diagnostics and Molecular Medicine, Szczecin, Poland; 4University of Louisville, Stem Cell Graham Brown Cancer Center, Louisville, United States

Background: Our group has reported that the number of very small embryonic/epiblast like stem cells (VSEL-SCs) mobilized into peripheral blood (PB) increases in active inflammatory bowel disease (IBD) (JCC 2010;4.1/Mo1815, DDW 2011). Currently we investigated the correlation of various types of stem/progeniotor cells mobilization phenomenon in Crohn's disease (CD) with clinical and endoscopic scores (ES). The activation of the complement cascade (CC) and factors involved in stem cell trafficking and their association with SCs mobilization into PB in IBD were studied.

Methods: 35 active/inactive CD and 30 healthy individuals were enrolled into the study. CD patients were classified according to clinical (Crohns Deasease Activity Index – CDAI) and endoscopic (Simple Endoscopic Score for Crohn's Disease – SES-CD) criteria. CDAI/SES-CD were assessed at week 0 and 10 of treatment with 5‑ASA/AZA and/or mo-anti-TNF-alfa. PB samples were collected, and by employing fluorescence-activated cell sorting (FACS) the numbers of (i) VSELs-SCs (CD45-CXCR4+/CD133+/Lin−), (ii) Mesenchymal SCs (MSC) (CD45−/STRO 1+/CD105+/CD29+), (iii) Endothelial Progenitor Cells (EPC) (CD45−/CD31+/CD34+/KDR+) and iv) Hematopoietic SCs (HSC) (Lin−/CD45+/CD133+) were counted and/or isolated. Immunofluorescence staining of sorted cells and RT-qPCR analysis of expression of mRNA for embryonic SCs markers (Oct‑4, SSEA‑4, Nanog), and markers of intestinal differentiation: Lgr‑5, ASCL2, MSI1, DCLK1, BMI1 was evaluated in sorted VSEL-SCs. Plasma levels of C3a, C5a, membrane attack complex (C5b-9/MAC), VEGF, HGF and SDF‑1 by ELISA were measured.

Results: In IBD patients we found an increase in the number of circulating MSC, EPC and small primitive cells expressing the VSEL phenotype. The number of circulating progenitor/stem cells correlated with SES-CD (p < 0.05) but not CDAI. SES-CD also better correlated with plasma VEGF concentration (R=0.49; p < 0.03) and HGF (R = 0.51; p < 0.02). Active disease was associated with strong mRNA up-regulation of intestinal early differentiation markers (lgr‑5 and ASCL‑2) in circulating PB mononuclear cells. Higher plasma C5a levels in IBD in comparison to controls were noted (61.7±19.7 vs 45.8±11.2; p = 0.024 respectively). No correlation between SDF‑1 and the number of VSEL-SC were found.

Conclusions: Enumeration of circulating stem/progenitor cells could serve as surrogate parameter in accessing treatment efficacy in IBD.

Acknowledgement: Study supported by European Union grant (POIG.01.01.02–00–109/09).