23 Temporal Expression of TLR2 On Hematopoietic Stem Cells in Murine Fetal Liver.

Abstract

Recently Toll like receptors (TLRs) have been shown to play an essential role in innate immunity, recognizing a specific component of foreign pathogens. TLR2 recognizes gram-positive bacterial cell wall molecules such as peptidoglycan (PG), whereas TLR4 recognizes gram-negative bacterial lipopolysaccharide. TLRs are expressed on various immunological cells such as neutrophils, monocyte/macrophages, dendritic cells and B cells, however, their expression pattern during hematopoietic development/differentiation is not well known. We examined expression of TLR2 or 4 on hematopoietic stem cells (HSCs) in fetal liver. Fetal liver cells at 12.5-16.5 days postcoitum (dpc) were collected and analyzed by flow cytometry. TLR2 and 4 were detected in the murine CD34+c-kit+Sca-1+lineage- (34KSL) population of the12.5 d.p.c fetal liver. 34KSL cells were separated into 3 fractions depending on the expression of TLR2 and 4; TLR2brightTLR4+ cells (7.3%), TLR2dimTLR4- cells (84.3%), and TLR2-TLR4- cells (3.5%). TLR2dimTLR4- and TLR2-TLR4- cells had high colony-forming ability but TLR2brightTLR4+ cells formed very few colonies. In 14.5dpc fetal liver, 90% of 34KSL cells were TLR2dim, however, in 16.5dpc fetal liver, most of the 34KSL cells were TLR2 negative. Interestingly, this TLR2 expression on fetal liver HSCs was synchronized with the expression level of Mac1. When TLR2 bright, TLR2 dim or TLR2- 34KSL cells of the 12.5 dpc fetal liver from Ly5.1 C57BL/6 mice were transplanted into lethally irradiated adult Ly5.2 C57BL/6 mice respectively, TLR2dim 34KSL cells were confirmed to reconstitute the recipient hematopoietic system. To examine the function of TLR2 on fetal liver HSCs, TLR2dim34KSL cells were cultured with/without PG in the presence of SCF and TPO. After 7 days of incubation, cells were collected and analyzed by colony assay and surface markers. Colony forming ability was maintained in both groups but cells differentiated into different lineages. We conclude that TLR2 is temporary expressed on fetal liver HSCs but the role of TLR2 in hematopoiesis is unclear.