IL-6Cooperates With G-CSF To Augment Pro-angiogenesis Function Of Neutrophils In Tumor By Enhancing The Activation Of STAT3

Tumour growth is tightly related to new blood vessel formation, tissue remodelling and invasiveness capacity. Recent evidence has shown that polymorphonuclear neutrophils (PMNs) produce several proangiogenic cytokines, including MMP-9, VEGF, IL-6, and IL-8, to promote angiogenesis of tumor. Although neutrophils are known to have anti-tumor potential, in recent years, the tumor-promoting effect of neutrophils has been well demonstrated. However, it remains unclear the reason why the conversion of neutrophil function from tumor suppressive to tumor promotional.Here we report that IL-6cooperates with G-CSF to cause this conversion before neutrophils enter tumor milieu. Pre-modulation of neutrophils by G-CSF/IL-6enabled these cells to promote tumor angiogenesis before they enter the tumor milieu. G-CSFR or IL-6R alone could attenuate the angiogenesis promotion effect of neutrophils, whereas their cooperation was required for modulating neutrophils to inhibit tumor angiogenesis.To further prove that the modulation by G-CSF and IL-6resulted in the alteration of neutrophil function, we next analyzed the Mmp9, Bv8and Trail mRNA levels of neutrophils in tissue and in bone marrow. G-CSF and IL-6up-regulated Mmp9and Bv8but down-regulate Trail expression in neutrophils of bone marrow and tissue. Moreover, G-CSF and IL-6influenced production or release of functional molecules by neutrophils in tissue—more pro-angiogenesis factor production and release (e.g MMP-9) but less anti-angiogenesis factor production and release (e.g. TRAIL).Effect of STAT3signaling pathway was important for G-CSF/IL-6-modulated gene expression in neutrophils. STAT3was crucial for up-regulating the expression of Mmp9, Bv8and down-regulating the expression of Trail in neutrophils. Compared with G-CSF, IL-6was indeed a weaker inducer for STAT3activation in bone marrow neutrophils. But IL-6could enhance STAT3activation by increasing STAT3expression in neutrophils as a co-stimulator in vivo. Co-stimulation with G-CSF and IL-6induced higher level of phosphorylated STAT3in neutrophils, which was further augmented by the increase of total STAT3protein in neutrophils due to down-regulation of IFN-P expression in bone marrow macrophages by IL-6. Consistently, IFN-P attenuated or abrogated the modulatory effect of G-CSF/IL-6and tumor on the expressions of Mmp9, Bv8, and Trail genes, and abrogated tumor-promoting effect of bone marrow neutrophils in angiogenesis.Based on the altered activation of signaling pathways, neutrophils in response to complex stimuli in tumor released much less TRAIL, but showed much higher expressions of Mmp9and Bv8genes, thus favoring tumor growth and tumor-angiogenesis. These findings highlight the molecular mechanisms underlying the alteration of proangiogenesis function of neutrophil.