Background. Sickle cell disease (SCD) is caused by a single mutation of the ?-globin gene and is the most common form of inherited blood disorder. In the US, approximately 100,000 of Americans are affected. Patients with SCD suffer from repeated red cell sickling and vaso-occlusion episodes, which, since early age, lead to acute and chronic pain, stroke, anemia, infections, organ failure and premature death. Vaso-occlusion episodes cause ischemia-reperfusion injury, activation of endothelial cells, immune cells and the coagulation cascade. These processes lead to a state of chronic inflammation in SCD, associated with increased pro- inflammatory cytokines, TLR4 activation and oxidative stress in all organs, including the bone marrow (BM). To date, little is known about how inflammation impacts the BM microenvironment and HSC functions in SCD patients. Notably, few and limited studies have been conducted on human samples. Preliminary results. Our previous studies using different animal models have shown that TLR4 activation and upregulation of miR155 and NF-kB signaling during inflammation results in the remodeling of the BM niche, myeloid lineage skewing and loss of hematopoietic stem cell self-renewal. Preliminary results that we obtained in SCD patient samples enrolled in a HSC transplantation (HSCT) clinical trial at City of Hope demonstrated a decrease in frequency and number of CD34+ progenitor cells, decreased ability to generate colonies and increased miR155 and NF-kB signaling in SCD BM-derived mesenchymal cells (MSC). Hypothesis. We hypothesize that SCD-induced chronic inflammation remodels the BM niche and impairs HSC function in SCD patients, and that these changes impact the engraftment of donor HSC during transplant.
Aims and Strategy. This study will be conducted on BM specimens of SCD patients (n=21) prior HSCT and at day +15, +30 and +1yr after HSCT. Characterization of the cellular components and inflammatory pathways will be carried on BM biopsies and aspirates by using complementary approaches such as multiparametric flow cytometry analysis, immunohistochemistry and nanostring assays; CD34+ cells will be isolated at baseline, used for RNA-seq and tested for colony-forming assay and ability to engraft NSG mice in vivo; BM and peripheral blood serum will be probed for inflammatory cytokines; MSC will be BM derived in vitro and analyzed characterization and persistence of the inflammatory signature. Results will be correlated with HSCT outcomes. Aged matched healthy donors will be used as controls. Relevance. This study represents a unique opportunity to better understand the biology of SCD in human specimens. Important, the dissection of the mechanisms involved in BM niche dysfunction and inflammatory state in severe SCD should offer avenues for therapeutic intervention to improve engraftment and hematopoietic reconstitution in patients with SCD.

Public Health Relevance

Approximately 100,000 Americans have sickle cell disease, the most common form of inherited blood disorder in which abnormal hemoglobin induces the red blood cells to block blood flow, causing pain and infections and numerous complications, including stroke, acute chest syndrome, and organ damage and in some cases premature death. Current treatments for SCD aim at pain management and prevention of complications; up to now the only curative treatment is bone marrow transplant, which presents risks and it is limited by the presence of available matched donors. In this application we propose to study the bone marrow microenvironment of SCD patients prior and after bone marrow transplant to identify determinants of bone marrow engraftment and to identify novel targets of intervention to improve transplant strategies and other therapeutic approaches, including gene therapy.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Research Project (R01)
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Molecular and Cellular Hematology Study Section (MCH)
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Bai, C Brian
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Beckman Research Institute/City of Hope
United States
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