Abstract
Fast and efficient homing and engraftment of hematopoietic stem progenitor cells (HSPCs) is crucial for positive clinical outcomes from transplantation. We found that this process depends on activation of the Nlrp3 inflammasome, both in the HSPCs to be transplanted and in the cells in the recipient bone marrow (BM) microenvironment. For the first time we provide evidence that functional deficiency in the Nlrp3 inflammasome in transplanted cells or in the host microenvironment leads to defective homing and engraftment. At the molecular level, functional deficiency of the Nlrp3 inflammasome in HSPCs leads to their defective migration in response to the major BM homing chemoattractant stromal-derived factor 1 (SDF-1) and to other supportive chemoattractants, including sphingosine-1-phosphate (S1P) and extracellular adenosine triphosphate (eATP). We report that activation of the Nlrp3 inflammasome increases autocrine release of eATP, which promotes incorporation of the CXCR4 receptor into membrane lipid rafts at the leading surface of migrating cells. On the other hand, a lack of Nlrp3 inflammasome expression in BM conditioned for transplantation leads to a decrease in expression of SDF-1 and danger-associated molecular pattern molecules (DAMPs), which are responsible for activation of the complement cascade (ComC), which in turn facilitates the homing and engraftment of HSPCs.
Document Type
Article
Publication Date
10-2020
Digital Object Identifier (DOI)
https://doi.org/10.1007/s12015-020-10005-w
Funding Information
This work was supported by NIH grants 2R01 DK074720 and R01HL112788, the Stella and Henry Endowment, and the OPUS grant DEC-2016/23/B/NZ3/03157 to MZR. AT was supported by NIH T32 HL134644 Training Grant to MZR. AAL is supported by the University of Kentucky COBRE Early Career Program (P20 GM103527) and the NIH Grant R01 HL124266.
Related Content
Descriptions of supplementary materials:
Supplementary Figure 1
Panel A. Measurement of MCC950 toxicity. Murine BMMNCs were incubated for 1 h with different doses of the Nlrp3-selective inhibitor MCC950, then resuspended in human methylcellulose base medium, supplemented with GM-CSF (25 ng/ml) and IL-3 (10 ng/ml), for determining the number of CFU-GM colonies, and with thrombopoietin (TPO, 100 ng/ml) and IL-3 (10 ng/ml), for determining the number of burst-forming unit-erythroid (BFU-E) colonies. Cultures were incubated for 7 and 14 days respectively (37 °C, 95% humidity, and 5% CO2), at which time they were scored under an inverted microscope for the number of colonies. Results from three independent experiments plated in duplicate are pooled together. Panel B. The chemotactic responsiveness of mBMMNCs, untreated or treated with MCC950, to medium supplemented with SDF-1, S1P, C1P, or ATP, according to FACS or the number of CFU-GM clonogenic progenitors. Results are combined from two independent experiments. *p > 0.05. (PPTX 65 kb)
Supplementary Figure 2
Defect in short- and long-term engraftment of HSPCs treated with an Nlrp3-selective inhibitor in WT mice. Panel A. Lethally irradiated WT mice (9 per group) were transplanted with bone marrow mononuclear cells (BMMNCs) that had been previously treated with MCC950 and labeled with a PKH67 cell linker. Twenty-four hours after transplantation, femoral BMMNCs were harvested, the number of PKH67+ cells evaluated by FACS, and the CFU-GM clonogenic progenitors enumerated in an in vitro colony assay. Panel B. Lethally irradiated WT mice (9 per group) were transplanted with BMMNCs treated with MCC950, and 12 days after transplantation femoral BMMNCs were harvested and plated to count the number of CFU-GM colonies and the spleens removed for counting the number of CFU-S colonies. No colonies were formed in lethally irradiated, untransplanted mice (irradiation control). *p < 0.05. Panel C. Lethally irradiated WT mice (9 per group) were transplanted with BMMNCs treated with MCC950. White blood cells (above) and platelets (below) were counted at intervals (at 0, 3, 7, 14, 21, and 28 days after transplantation). *p < 0.05. (PPTX 74 kb)
Supplementary Figure 3
Defect in short- and long-term engraftment of HSPCs in WT mice treated with an Nlrp3-selective inhibitor. Panel A. Lethally irradiated WT mice (9 per group), untreated or treated with MCC950, were transplanted with bone marrow mononuclear cells (BMMNCs) from WT mice that had previously been labeled with a PKH67 cell linker. Twenty-four hours after transplantation, femoral BMMNCs were harvested, the number of PKH67+ cells evaluated by FACS, and the CFU-GM clonogenic progenitors enumerated in an in vitro colony assay. Panel B. Lethally irradiated WT mice (9 per group), untreated or treated with MCC950, were transplanted with BMMNCs from WT mice, and 12 days after transplantation femoral BMMNCs were harvested and plated to count the number of CFU-GM colonies and the spleens removed for counting the number of CFU-S colonies. No colonies were formed in lethally irradiated, untransplanted mice (irradiation control). *p < 0.05. Panel C. Lethally irradiated mice (9 per group), untreated or treated with MCC950, were transplanted with BMMNCs from WT mice. White blood cells (left) and platelets (right) were counted at intervals (at 0, 3, 7, 14, 21, and 28 days after transplantation). *p < 0.05. (PPTX 80 kb)
Supplementary Figure 4
A reduced number of HSPCs in the BM of Nlrp3-KO mice. The number of SKL cells in the BM of Nlrp3-KO mice compared with WT control animals was evaluated by FACS and by the number of CFU-GM and BFU-E clonogenic progenitors in in vitro methylcellulose cultures. Results are combined from two independent experiments (4 mice per group per repeat). (PPTX 51 kb)
Repository Citation
Adamiak, Mateusz; Abdel-Latif, Ahmed K.; Bujko, Kamila; Thapa, Arjun; Anusz, Krzysztof; Tracz, Michał; Brzezniakiewicz-Janus, Katarzyna; Ratajczak, Janina; Kucia, Magda; and Ratajczak, Mariusz Z., "Nlrp3 Inflammasome Signaling Regulates the Homing and Engraftment of Hematopoietic Stem Cells (HSPCs) by Enhancing Incorporation of CXCR4 Receptor into Membrane Lipid Rafts" (2020). Internal Medicine Faculty Publications. 213.
https://uknowledge.uky.edu/internalmedicine_facpub/213
Supplementary Figure 1
12015_2020_10005_MOESM2_ESM.pptx (74 kB)
Supplementary Figure 2 Defect in short- and long-term engraftment of HSPCs treated with an Nlrp3-selective inhibitor in WT mice.
12015_2020_10005_MOESM3_ESM.pptx (80 kB)
Supplementary Figure 3 Defect in short- and long-term engraftment of HSPCs in WT mice treated with an Nlrp3-selective inhibitor.
12015_2020_10005_MOESM4_ESM.pptx (51 kB)
Supplementary Figure 4 A reduced number of HSPCs in the BM of Nlrp3-KO mice.
Notes/Citation Information
Published in Stem Cell Reviews and Reports, v. 16, issue 5.
© The Author(s) 2020
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