A mouse model of yellow fluorescent protein (YFP) expression in hematopoietic cells to assess leukocyte-endothelial interactions in the microcirculation

In this study, we describe the use of intravital microscopy in a transgenic mouse model expressing yellow fluorescent protein (YFP) under the control of a monocyte specific promoter c-fms (CD115) to track and quantify specific leukocyte subsets. Flow cytometry on peripheral and bone marrow leukocytes revealed that YFP was predominantly expressed by CD11a(+), CD11b(+), and CD14(+) monocytes. In the bone marrow, 67 +/- 4% of Ly6C(high) F4/80(+) cells were YFPhigh while 55 +/- 1% of Ly6C(low) F4/80(+) cells were YFPlow supporting the use of c-fms(YFP) expression as a marker of monocyte lineage. 70 +/- 7% of CD11b(+) F4/80(+) Ly6C(+) ("triple positive") cells expressed YFP. To assess leukocyte-endothelial interactions in YFP+ cells in c-fms(YFP+) mice, we evaluated leukocyte adhesion, rolling and local shear stress responses in the cremasteric endothelium 4 h following administration of TNF alpha. TNF alpha resulted in a five-fold increase in adhesion of YFP+ cells to the endothelium and provided superior discriminative ability in assessing rolling and adhesion events when compared with bright field microscopy. Additionally, when compared with Rhodamine-6G labeled leukocytes or GFP(+) cells in mice transplanted with green fluorescent protein (GFP) positive bone marrow, the level of detail observed in the c-fms(YFP+) was greater, with both GFP(+) and YFP+ cells demonstrating superior signal to noise compared to bright field microscopy. A weak positive linear correlation between wall shear stress and YFP+ cell adhesion (r(2) = 0.20, p < 0.05) was seen in the cremasteric microcirculation. Taken together, these data demonstrate the use of c-fms(YFP+) mice in identifying distinct monocyte subsets and highlight the potential of this model for real-time monocyte-endothelial interactions using intravital microscopy. (C) 2009 Elsevier Inc. All rights reserved.