The bone marrow endothelial progenitor cell response to septic infection.

Early increase in the level of endothelial progenitor cells (EPCs) in the systemic circulation occurs in patients with septic infection/sepsis. The significance and underlying mechanisms of this response remain unclear. This study investigated the bone marrow EPC response in adult mice with septic infection induced by intravenous injection (i.v.) of Escherichia coli. For in vitro experiments, sorted marrow stem/progenitor cells (SPCs) including lineage(lin)-stem cell factor receptor (c-kit)+stem cell antigen-1 (Sca-1)-, lin-c-kit+, and lin- cells were cultured with or without lipopolysaccharides (LPSs) and recombinant murine vascular endothelial growth factor (VEGF) in the absence and presence of anti-Sca-1 crosslinking antibodies. In a separate set of experiments, marrow lin-c-kit+ cells from green fluorescence protein (GFP)+ mice, i.v. challenged with heat-inactivated E. coli or saline for 24 h, were subcutaneously implanted in Matrigel plugs for 5 weeks. Marrow lin-c-kit+ cells from Sca-1 knockout (KO) mice challenged with heat-inactivated E. coli for 24 h were cultured in the Matrigel medium for 8 weeks. The marrow pool of EPCs bearing the lin-c-kit+Sca-1+VEGF receptor 2 (VEGFR2)+ (LKS VEGFR2+) and LKS CD133+VEGFR2+ surface markers expanded rapidly following septic infection, which was supported by both proliferative activation and phenotypic conversion of marrow stem/progenitor cells. Increase in marrow EPCs and their reprogramming for enhancing angiogenic activity correlated with cell-marked upregulation of Sca-1 expression. Sca-1 was coupled with Ras-related C3 botulinum toxin substrate 2 (Rac2) in signaling the marrow EPC response. Septic infection caused a substantial increase in plasma levels of IFN-γ, VEGF, G-CSF, and SDF-1. The early increase in circulating EPCs was accompanied by their active homing and incorporation into pulmonary microvasculature. These results demonstrate that the marrow EPC response is a critical component of the host defense system. Sca-1 signaling plays a pivotal role in the regulation of EPC response in mice with septic infection.

Endothelial progenitor cells in the host defense response.

Extensive injury of endothelial cells in blood vasculature, especially in the microcirculatory system, frequently occurs in hosts suffering from sepsis and the accompanied systemic inflammation. Pathological factors, including toxic components derived from invading microbes, oxidative stress associated with tissue ischemia/reperfusion, and vessel active mediators generated during the inflammatory response, are known to play important roles in mediating endothelial injury. Collapse of microcirculation and tissue edema developed from the failure of endothelial barrier function in vital organ systems, including the lung, brain, and kidney, are detrimental, which often predict fatal outcomes. The host body possesses a substantial capacity for maintaining vascular homeostasis and repairing endothelial damage. Bone marrow and vascular wall niches house endothelial progenitor cells (EPCs). In response to septic challenges, EPCs in their niche environment are rapidly activated for proliferation and angiogenic differentiation. In the meantime, release of EPCs from their niches into the blood stream and homing of these vascular precursors to tissue sites of injury are markedly increased. The recruited EPCs actively participate in host defense against endothelial injury and repair of damage in blood vasculature via direct differentiation into endothelial cells for re-endothelialization as well as production of vessel active mediators to exert paracrine and autocrine effects on angiogenesis/vasculogenesis. In recent years, investigations on significance of EPCs in host defense and molecular signaling mechanisms underlying regulation of the EPC response have achieved substantial progress, which promotes exploration of vascular precursor cell-based approaches for effective prevention and treatment of sepsis-induced vascular injury as well as vital organ system failure.

Acute Alcohol Intoxication Impairs Sonic Hedgehog-Gli1 Signaling and Activation of Primitive Hematopoietic Precursor Cells in the Early Stage of Host Response to Bacteremia.

BACKGROUND: Activation of hematopoietic stem cells [HSCs, lineage(lin)- stem cell growth factor receptor (c-kit)+ stem cell antigen-1(Sca-1)+ , or LKS cells in mice] is critical for initiating the granulopoietic response. This study determined the effect of alcohol exposure on sonic hedgehog (SHH) signaling in the regulation of HSC activation during bacteremia. METHODS: Acute alcohol intoxication was induced in mice by intraperitoneal (i.p.) injection of 20% alcohol (5 g alcohol/kg body weight). Control mice received i.p. saline. Thirty minutes later, mice were intravenously (i.v.) injected with Escherichia coli (E. coli, 1 to 5 × 107 CFUs/mouse) or saline. RESULTS: SHH expression by lineage-negative bone marrow cells (BMCs) was significantly increased 24 hours after E. coli infection. Extracellular signal-regulated kinase 1/2 (ERK1/2)-specificity protein 1 (Sp1) signaling promotes SHH expression. ERK1/2 was markedly activated in BMCs 8 hours following E. coli infection. Alcohol suppressed both the activation of ERK1/2 and up-regulation of SHH expression following E. coli infection. E. coli infection up-regulated GLI family zinc finger 1 (Gli1) gene expression by BMCs and increased Gli1 protein content in LKS cells. The extent of Gli1 expression was correlated with the activity of proliferation in LKS cells. Alcohol inhibited up-regulation of Gli1 expression and activation of LKS cells in response to E. coli infection. Alcohol also interrupted the granulopoietic response to bacteremia. CONCLUSION: These data show that alcohol disrupts SHH-Gli1 signaling and HSC activation in the early stage of the granulopoietic response, which may serve as an important mechanism underlying the impairment of immune defense against bacterial infection in host excessively consuming alcohol.

Sonic Hedgehog Signaling Regulates Hematopoietic Stem/Progenitor Cell Activation during the Granulopoietic Response to Systemic Bacterial Infection.

Activation and reprogramming of hematopoietic stem/progenitor cells play a critical role in the granulopoietic response to bacterial infection. Our current study determined the significance of Sonic hedgehog (SHH) signaling in the regulation of hematopoietic precursor cell activity during the host defense response to systemic bacterial infection. Bacteremia was induced in male Balb/c mice via intravenous injection (i.v.) of Escherichia coli (5 × 107 CFUs/mouse). Control mice received i.v. saline. SHH protein level in bone marrow cell (BMC) lysates was markedly increased at both 24 and 48 h of bacteremia. By contrast, the amount of soluble SHH ligand in marrow elutes was significantly reduced. These contrasting alterations suggested that SHH ligand release from BMCs was reduced and/or binding of soluble SHH ligand to BMCs was enhanced. At both 12 and 24 h of bacteremia, SHH mRNA expression by BMCs was significantly upregulated. This upregulation of SHH mRNA expression was followed by a marked increase in SHH protein expression in BMCs. Activation of the ERK1/2-SP1 pathway was involved in mediating the upregulation of SHH gene expression. The major cell type showing the enhancement of SHH expression in the bone marrow was lineage positive cells. Gli1 positioned downstream of the SHH receptor activation serves as a key component of the hedgehog (HH) pathway. Primitive hematopoietic precursor cells exhibited the highest level of baseline Gli1 expression, suggesting that they were active cells responding to SHH ligand stimulation. Along with the increased expression of SHH in the bone marrow, expression of Gli1 by marrow cells was significantly upregulated at both mRNA and protein levels following bacteremia. This enhancement of Gli1 expression was correlated with activation of hematopoietic stem/progenitor cell proliferation. Mice with Gli1 gene deletion showed attenuation in activation of marrow hematopoietic stem/progenitor cell proliferation and inhibition of increase in blood granulocytes following bacteremia. Our results indicate that SHH signaling is critically important in the regulation of hematopoietic stem/progenitor cell activation and reprogramming during the granulopoietic response to serious bacterial infection.