Cells from the embryonic pancreas (pooled), liver and adult BM were incubated with ER-MP58-biotin (own culture) and afterwards with streptavidin-allophycocyanin (Becton Dickinson). ER-MP58+ cells were sorted with FACSAria (Becton Dickinson). Subsequently, ER-MP58+ cells were cultured for 8 days on 0.5% gelatin-coated wells (96-well plate) learn more in RPMI 1640 medium supplemented with 10% FCS, 50 μM β-mercaptoethanol and 50 ng/mL GM-CSF (MT Diagnostics, Etten-Leur, The Netherlands). Finally cells were harvested with 2 mM EDTA. To monitor the proliferation

capability, cells from the embryonic pancreas (pooled), liver, adult BM and blood were labeled with 5 μM carboxyfluorescein succinimidyl ester (CFSE) (Sigma Aldrich) and incubated for 10 min at 37°C. Cells were washed and incubated with ER-MP58-biotin and afterwards with streptavidin-allophycocyanin. ER-MP58+ cells were sorted with FACSAria and were cultured for 8 days with 50 ng/mL GM-CSF. Cells were harvested with 2 mM EDTA. Cryostat sections (6 μm) of E15.5 pancreases from C57BL/6 and NOD/LTj mice were prepared and fixed with cold methanol and acetone. Slides were incubated with guinea pig-anti-insulin (DAKO, Glostrup, Denmark) and rat-anti-ER-MP58 followed by rabbit-anti-guinea pig-FITC (Abcam) and goat-anti-rat-TexasRed (Southern Biotechnology

Associates, Birmingham, AL, USA). Finally, slides were mounted in Vectashield with DAPI (Vector Laboratories, Burlingame, CA, USA). Cryostat sections (6 μm) of 5-wk-old pancreases from C57BL/6, NOR/LTj and NOD/LTj mice were prepared and fixed with cold methanol and acetone. Slides EGFR inhibitor were Non-specific serine/threonine protein kinase incubated with Ki-67-FITC and rat-anti-ER-MP58 followed by goat-anti-rat-TexasRed. Finally, slides were mounted in Vectashield with DAPI. Data were analyzed

by Mann–Whitney U test for unpaired data. All analyses were carried out using SPSS software (SPSS, Chicago, IL, USA) and considered statistically significant if p<0.05. The authors thank Pieter Leenen for his expert advice and the Juvenile Diabetes Research Foundation for supporting this study Conflicts of interest: The authors declare no financial or commercial conflict of interest. Detailed facts of importance to specialist readers are published as ”Supporting Information”. Such documents are peer-reviewed, but not copy-edited or typeset. They are made available as submitted by the authors. "
“Human monocytes respond to a variety of stimuli with a complex spectrum of activities ranging from acute defense mechanisms to cell differentiation or cytokine release. However, the individual intracellular signaling pathways related to these functions are not well understood. CXC chemokine ligand 4 (CXCL4) represents a broad activator of monocytes, which induces acute as well as delayed activities in these cells including cell differentiation, survival, or the release of ROS, and cytokines.