Recent studies demonstrate that inflammatory signals regulate hematopoietic stem cells (HSCs). Granulocyte-colony stimulating factor (G-CSF) is often induced with infection and plays a key role in the stress granulopoiesis response. However, its effects on HSCs are less clear. Herein, we show that treatment with G-CSF induces expansion and increased quiescence of phenotypic HSCs, but causes a marked, cell-autonomous HSC repopulating defect associated with induction of toll-like receptor (TLR) expression and signaling. The G-CSF-mediated expansion of HSCs is reduced in mice lacking TLR2, TLR4 or the TLR signaling adaptor MyD88. Induction of HSC quiescence is abrogated in mice lacking MyD88 or in mice treated with antibiotics to suppress intestinal flora. Finally, loss of TLR4 or germ free conditions mitigates the G-CSF-mediated HSC repopulating defect. These data suggest that low level TLR agonist production by commensal flora contributes to the regulation of HSC function and that G-CSF negatively regulates HSCs, in part, by enhancing TLR signaling.
Toll-like receptor 2 (TLR2) is a member of the TLR family of receptors that play a central role in innate immunity. In addition to regulating effector immune cells, where it recognizes a wide variety of pathogen-associated and nonpathogen-associated endogenous ligands, TLR2 is expressed in hematopoietic stem cells (HSCs). Its role in HSCs, however, is not well understood. Furthermore, augmented TLR2 signaling is associated with myelodysplastic syndrome, an HSC disorder characterized by ineffective hematopoiesis and a high risk of transformation to leukemia, suggesting that aberrant signaling through this receptor may have clinically significant effects on HSCs. Herein, we show that systemic exposure of mice to a TLR2 agonist leads to an expansion of bone marrow and spleen phenotypic HSCs and progenitors, but a loss of HSC self-renewal capacity. Treatment of chimeric animals shows that these effects are largely cell non-autonomous, with a minor contribution from cell-autonomous TLR2 signaling, and are in part mediated by granulocyte colony-stimulating factor and tumor necrosis factor-α. Together, these data suggest that TLR2 ligand exposure influences HSC cycling and function via unique mechanisms from TLR4, and support an important role for TLR2 in the regulation of HSCs.
Objectives To assess the impact of aggressive protocol to decrease door-to-balloon (DTB) time on the incidence of false-positive STEMI (FP-STEMI) and in-hospital mortality. Patients Consecutive patients with presumed STEMI with confirmed ST-segment elevation that underwent emergent catheterization. Methods In July 1, 2009 we instituted an aggressive protocol to further reduce DTB time. A quality improvement (QI) initiative was initiated in January 1, 2010 to maintain short DTB while improving outcomes. Outcomes were compared before and after aggressive DTB and similarly before and after the QI initiative. Outcomes were DTB time, the incidence of FP-STEMI and in-hospital mortality. A review of the emergency catheterization database over the last 10 years (January 2001-December 2010) was carried out for historical comparison. Results Between July 1, 2008 and December 1, 2012, 1031 consecutive patients with presumed STEMI were assessed. Of these 170 were considered FP-STEMI. The median DTB time decreased from 76 to 61 minutes with the aggressive DTB protocol (P=. 001), accompanied by an increase of FP-STEMI (7.7% vs. 16.5%, p=.02). While TP-STEMI in-hospital mortality witnessed non-significant reduction, this was associated with a significant increase of FP-STEMI in-hospital mortality. After the QI initiative, a shorter DTB time (59 minutes) was maintained while decreasing FP-STEMI in-hospital mortality. Conclusion Aggressive measures to reduce DTB time were associated with an increased incidence of FP-STEMI and FP-STEMI in-hospital mortality. Efforts to reduce DTB time should be monitored systematically to avoid unnecessary procedures that may delays other appropriate therapies in critically ill patients.
Toll like receptors (TLRs) are a family of pattern recognition receptors that play a central role in pathogen recognition and shaping the innate immune response. While most of the studies of the role of TLRs have focused on mature immune cell populations, recent reports suggest that TLR signaling may regulate the immune response from the level of the hematopoietic stem cell (HSC). In this study, we sought to further elucidate the effects of systemic TLR ligand exposure on HSCs and determine the cell-intrinsic versus extrinsic effects of such exposure. We specifically focused on TLR2 signaling, as although TLR2 is expressed on HSCs, it’s role in their regulation is not clear. Furthermore, enhanced TLR2 signaling is associated with myelodysplastic syndrome (Wei et al, Leukemia 2013), suggesting that aberrant signaling through this receptor may have clinically significant effects on HSC function. To elucidate the role of TLR2 signaling in regulating HSCs, we used mice with genetic loss of TLR2, as well as a synthetic agonist of TLR2 (PAM3CSK4) to determine the effects of TLR2 signaling loss or gain, respectively, on HSC cycling, mobilization and function. While TLR2 expression is not required for normal HSC function, treatment of wild-type mice with PAM3CSK4 leads to expansion of HSCs in the bone marrow and spleen, increased HSC cycling, and loss of HSC function in competitive bone marrow transplantation experiments. As TLR2 is expressed on a variety of stromal and hematopoietic cell types, we used bone marrow chimeras (Tlr2-/- + Tlr2+/+ marrow transplanted into Tlr2+/+ recipients) to determine if the effects of PAM3CSK4 treatment are cell intrinsic or extrinsic. The data suggests that HSC cycling and expansion in the marrow and spleen upon PAM3CSK4 treatment are extrinsic (occurring in both transplanted HSC populations), and are associated with increased serum levels of G-CSF. Indeed, inhibition of G-CSF using either a neutralizing antibody or mice lacking the G-CSF receptor (Csf3r-/-) leads to even further enhanced HSC bone marrow expansion upon G-CSF treatment but significantly reduced numbers of spleen HSCs compared to similarly treated wild-type mice. This suggests mobilization in response to TLR2 signaling is an indirect, G-CSF-mediated process. Ongoing studies are aimed at determining the contribution of G-CSF to the PAM3CSK4- induced loss of HSC function, and determining the source (stromal vs hematopoietic) of G-CSF production upon PAM3CSK4 exposure. Collectively, this data suggest that TLR2 signaling affects HSCs in a largely extrinsic fashion, with G-CSF playing a major role in regulating the effects of TLR2 ligand exposure on HSCs. Disclosures No relevant conflicts of interest to declare.
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