Abstract
Inflammation shapes the immune response to foreign antigens by influencing effector immune function and is dysregulated in many forms of inflammatory bowel disease and other auto-immune conditions. It is increasingly recognized that inflammation also impacts hematopoietic stem and progenitor cell (HSPC) function. Interleukin-10 (IL-10) is an anti-inflammatory cytokine that regulates inappropriate immune responses by suppressing inflammatory programs in mature immune cells. However, the direct role of IL-10 in modulating HSPC function and in suppressing hematopoietic cell output in the context of inflammation has not previously been demonstrated.
IL10Rß-deficient (Il10rb-/-) mice have an increased inflammatory cytokine profile and develop overt colitis symptoms between 4-6 months of age. We observed an expansion in the relative frequency of immunophenotypically-defined HSPCs (LSK+: Lineage-c-Kit+Sca-1+) and frequency and absolute numbers of granulocyte-monocyte progenitors (GMPs: Lineage-c-Kit+CD34+FcgRII/III+) in pre-colitic Il10rb-/- mice relative to wildtype mice (WT) (see table). Increased GMPs correlated with increased myeloid and reduced lymphoid frequency and number in the bone marrow (BM). Consistent with the expansion in the Il10rb-/-BM HSPC compartment, transplantation of a 1:1 ratio of Il10rb-/-: WT whole BM also resulted in a 2-fold higher short-term reconstitution of Il10rb-/- CD45+ cells in peripheral blood relative to WT cells. Moreover, in vitro we observed suppressed expansion of WT but not Il10rb-/- lineage- cells after IL-10 treatment, suggesting that IL10 suppresses HSPC proliferation. To determine if IL-10 signaling affects the function of hematopoietic stem cells (HSCs), we compared an equal number of purified HSCs (LSK+CD150+CD48-) from Il10rb-/- and WT mice in repopulation assays. Il10rb-/- HSCs demonstrated a 6-fold (P<0.01) reduced lineage reconstitution in peripheral blood relative to WT HSCs in competitive transplantation, thus demonstrating that, while Il10rb-/- mice have an increased pool of committed progenitor cells, HSC function is reduced relative to WT mice. To confirm direct IL-10 signaling in HSPC populations, purified LSK+ and myeloid progenitors (LK+Sca-1-) were exposed to IL-10. Significant STAT3 phosphorylation was observed in HSPCs and myeloid progenitors, with the highest proportion of phosphorylated cells in the GMP fraction. These data suggest that IL-10 directly acts on multiple stem and progenitor populations and that myeloid progenitors robustly respond to IL-10. To determine if the alterations in Il10rb-/- BM are cell intrinsic or a product of underlying inflammation in the absence of IL-10 signaling in vivo, we generated mixed whole BM WT: Il10rb-/- chimeric mice and induced inflammation through E. coli lipopolysaccharide (LPS) injection. LPS exposure led to a 1.5-fold larger depletion of Il10rb-/- GMPs compared to WT GMPs in mixed chimeric mice 24 hours post-injection relative to vehicle treatment. These data suggest a protective role for IL-10 on progenitors during an acute inflammatory insult. Upon transplantation of BM from primary LPS-treated mixed chimeric mice, a short-term increase in peripheral blood myeloid cells was observed when compared to transplanted BM from vehicle-treated mice. Notably, the enhanced peripheral blood myeloid output from LPS-treated donor BM was skewed 2-fold in favor of Il10rb-/- cells and BM myeloid colony-forming-cells were skewed 4-fold in favor of Il10rb-/- cells relative to WT cells. These data demonstrate that IL-10 attenuates inflammation-induced myeloid output upon BM transplantation.
Taken together, this study reveals a previously unrecognized role for IL-10 in suppressing progenitor cell expansion and promoting HSC function at steady state while mitigating myeloid lineage skewing after an inflammatory insult.
Williams:Bluebird Bio: Research Funding.
