after the onset of and then for several days, masses of polymorphonuclear neutrophils (PMNs) enter the ICH-affected brain. After releasing various cytotoxic moieties, the PMNs undergo rapid apoptosis and then are removed by brain microglia/macrophages (M?). Without fast clearance by M?, apoptotic PMNs (ANs) proceed to secondary necrosis and spillage of wealth of uniquely toxic moieties, causing harm to brain and augment brain injury caused by hematoma. However, given the biological properties of individual moieties released by PMNs, it is likely that PMNs can also add to brain repair. The PMN's chemical composition is set at the precursor stage of PMNs (pPMN) in bone marrow (BM)(BM-pPMN). The mature PMNs retain the pre-established phenotype and act as shuttles, delivering the content (packed in granules) to the sites of PMNs' infiltration (here ICH-injured brain). Here, we propose to study a novel pathway where the signal, specifically IL-27, modifies BM-pPMN phenotype, so that the mature PMNs upon entering the ICH-affected brain would cause less damage, or even provide benefit, including by promoting phagocytosis-mediated cleanup of hematoma and ANs. Lactoferrin (LTF) is a pleiotropic iron-binding/immunoregulatory protein that is synthetized/distributed by PMNs. Our pilot studies show that ICH-induced IL-27 is a signal for BM-pPMN to express more LTF. The LTF- enriched PMNs upon entering the hematoma could release more LTF. This LTF is capable of: (1) stimulating ANs engulfment by M?, (2) upregulating M?' CD91, a scavenger receptor for engulfment of LTF-coated ANs and other hemolysis products, (3) sequestrating the free iron, and (4) polarizing M? to beneficial (M2-like) phenotype. In addition to LTF, IL-27 promotes expression of hemoglobin neutralizing protein haptoglobin, help-me-signal molecule lipocalin-2, and suppresses the expression of MMP-9 and NADPH-oxidase by PMNs. Ultimately, treating animals with recombinant IL-27 (rIL-27) or with recombinant LTF (rLTF), robustly, with a 24h therapeutic window (for rLTF), reduces brain damage and functional loss caused by ICH. Our hypothesis is that after ICH, lL-27 polarizes PMNs to a beneficial type, which upon reaching ICH- injured brain is less cytotoxic and even enhances (in part through LTF) M?-mediated cleanup (in part through upregulating CD91/CD36 scavenger receptor) and lessens secondary brain damage.
Specific aims are: (1) characterize the IL-27-induced PMN phenotype within BM and blood; (2) with primary cell culture, to define the beneficial role of the PMN phenotype and LTF in clearance/detoxification of ANs/heme/iron by the PMNs/LTF- modified M?; and (3) to assess the therapeutic role for rLTF and to define the role of CD91in LTF-assisted cleanup, in mouse ICH model. Our goal is to develop new therapeutic targets for ICH, based on the ability of IL-27-modified PMNs to release more LTF to promote M?-mediated cleanup of hematoma. This approach should not only limit PMNs-mediated CNS damage, but also allow PMNs to act in a beneficial fashion.
Soon after the onset of intracerebral hemorrhage (ICH) and then for several days, millions of neutrophils (a type of white blood cells) enter the injured brai, releasing various toxic moieties that could cause harm to brain and augment brain injury caused by blood components within hematoma. Our novel studies suggest that neutrophils are modifiable at their developing stages in bone marrow. Here, we propose to study a novel pathway where the signal, specifically interleukin 27 (IL-27), modifies developing neutrophils in bone marrow, so that the mature PMNs upon entering the ICH-affected brain would cause less damage, or even provide benefit. Modified neutrophils release lactoferrin, a protein that activates local brain phagocytes (called microglia) and promotes microglia-mediated cleanup of hematoma. Cleanup of blood containing hematoma is essential to allow for better recovery after ICH. Our long-term goal is to develop new effective treatment strategy for patients with ICH.
|Zhao, Xiurong; Ting, Shun-Ming; Sun, Guanghua et al. (2018) Beneficial Role of Neutrophils Through Function of Lactoferrin After Intracerebral Hemorrhage. Stroke 49:1241-1247|
|Zhao, Xiurong; Ting, Shun-Ming; Liu, Chin-Hsuan et al. (2017) Neutrophil polarization by IL-27 as a therapeutic target for intracerebral hemorrhage. Nat Commun 8:602|
|Zhao, Xiurong; Zhang, Liyan; Ting, Shun-Ming et al. (2016) Phagocytosis Assay of Microglia for Dead Neurons in Primary Rat Brain Cell Cultures. Bio Protoc 6:|