In this project, we have discovered using intravital microcopy imaging in a humanized mouse model of sickle cell disease (SCD) that activated neutrophils play a direct role in vaso-occlusive crisis (VOC) by interacting with circulating erythrocytes (sRBC). We have described heterogeneity in the ability of neutrophils to capture sRBC and recently ascribed it to their chronological aging in the circulation and the exposure to the microbiota. Indeed, microbiota depletion markedly reduces aged neutrophil counts, and improves the acute and chronic SCD complications. Preliminary data using a model of psychological stress, a known VOC trigger, suggest that stress- induced exacerbation of VOC also depends on the microbiota and aged neutrophil generation promoted by the IL-17A/G-CSF pathway. In this funding period, we propose to investigate the innovative hypothesis that the microbiota critically regulates SCD activity.
In Specific Aim 1, we will evaluate how the microbiota modulates psychological stress-induced sickle cell VOC. We will assess the mechanisms of stress signals linking the brain to the immune response, focusing on neural (sympathetic nervous system) and stress hormones (glucocorticoids). We will identify the source of IL-17A elicited by stress, evaluate its function using IL-17A- deficient mice, and the mechanisms by which the microbiota activates the IL-17A/G-CSF pathway.
In Specific Aim 2, we will define the role of neutrophils and microbiota in chronic sickle cell-induced end-organ damage. We have found that depletion of the microbiota markedly improved the chronic organ damage in SCD mice. Whether the microbiota mediates organ damage through interactions with leukocytes or other targets is unclear. We will investigate the role of neutrophils in organ damage in G-CSF-deficient mice which are neutropenic. We will investigate whether the microbiota signals to hematopoietic or non-hematopoietic cells using conditional Myd88- deletion. Since our preliminary data suggest that microbiota depletion significantly reduces iron deposition in tissues, we will investigate the role of iron chelation therapy in SCD mice. In collaboration with Dr. Craig Branch, we will monitor the impact of microbiota depletion by T2* magnetic resonance imaging and test the potential of siderophore probiotics.
In Specific Aim 3, we will develop new strategies to harness the microbiota for SCD treatment. We will characterize the differential effect of antibiotics using 16S rDNA sequencing. We will manipulate the microbiota with probiotics and investigate specifically the role of segmented filamentous bacteria (SFB), as they are known to induce IL-17A/G-CSF, using fecal transplantations in vancomycin-treated or germ- free SCD mice. Since hydroxyurea (HU) has antimicrobial activity that can significantly alter the microbiome, we will evaluate its effect on gut microbiota by 16S rDNA sequencing and determine using fecal transplant the contribution of HU-modified microbiota in disease activity. These studies will provide insight on a new mechanism regulating SCD manifestations and lead to novel ways to target inflammation in this disease.

Public Health Relevance

Investigations conducted under this project have uncovered an important role for aged neutrophils and the microbiota in the pathogenesis of sickle cell disease. Here, we will study the mechanisms by which the microbiota influences disease activity, and develop new approaches to target the responsible bacterial strain(s) for prevention or treatment of the disease.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
2R01HL069438-16
Application #
9383680
Study Section
Molecular and Cellular Hematology Study Section (MCH)
Program Officer
Qasba, Pankaj
Project Start
2001-09-30
Project End
2021-05-31
Budget Start
2017-08-01
Budget End
2018-05-31
Support Year
16
Fiscal Year
2017
Total Cost
Indirect Cost
Name
Albert Einstein College of Medicine, Inc
Department
Type
DUNS #
079783367
City
Bronx
State
NY
Country
United States
Zip Code
10461
Xu, Chunliang; Gao, Xin; Wei, Qiaozhi et al. (2018) Stem cell factor is selectively secreted by arterial endothelial cells in bone marrow. Nat Commun 9:2449
Wei, Qiaozhi; Frenette, Paul S (2018) Niches for Hematopoietic Stem Cells and Their Progeny. Immunity 48:632-648
Boulais, Philip E; Mizoguchi, Toshihide; Zimmerman, Samuel et al. (2018) The Majority of CD45- Ter119- CD31- Bone Marrow Cell Fraction Is of Hematopoietic Origin and Contains Erythroid and Lymphoid Progenitors. Immunity 49:627-639.e6
Gao, Xin; Xu, Chunliang; Asada, Noboru et al. (2018) The hematopoietic stem cell niche: from embryo to adult. Development 145:
Pierce, Halley; Zhang, Dachuan; Magnon, Claire et al. (2017) Cholinergic Signals from the CNS Regulate G-CSF-Mediated HSC Mobilization from Bone Marrow via a Glucocorticoid Signaling Relay. Cell Stem Cell 20:648-658.e4
Zahalka, Ali H; Arnal-Estapé, Anna; Maryanovich, Maria et al. (2017) Adrenergic nerves activate an angio-metabolic switch in prostate cancer. Science 358:321-326
Chen, Grace; Chang, Jungshan; Zhang, Dachuan et al. (2016) Targeting Mac-1-mediated leukocyte-RBC interactions uncouples the benefits for acute vaso-occlusion and chronic organ damage. Exp Hematol 44:940-6
Zhang, Dachuan; Xu, Chunliang; Manwani, Deepa et al. (2016) Neutrophils, platelets, and inflammatory pathways at the nexus of sickle cell disease pathophysiology. Blood 127:801-9
Khan, Jalal A; Mendelson, Avital; Kunisaki, Yuya et al. (2016) Fetal liver hematopoietic stem cell niches associate with portal vessels. Science 351:176-80
Birbrair, Alexander; Frenette, Paul S (2016) Niche heterogeneity in the bone marrow. Ann N Y Acad Sci 1370:82-96

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