Cobalt protoporphyrin IX increases endogenous G‐ CSF and mobilizes HSC and granulocytes to the blood

• DOI: 10.15252/emmm.201809571
• Publication Date: 2019-11-11T08:20:48Z
• Authors: Agata Szade; Krzysztof Szade; Witold N Nowak; Karolina Bukowska...; Lucie Muchova; Monika Gońka; Monika Żukowska; Maciej Cieśla; Neli Kachamakova‐...; Marzena Rams‐Baron; Alicja Ratuszna; Józef Dulak; Alicja Józkowicz
• Keywords: Male; 0301 basic medicine; Medicine (General); hematopoietic stem and progenitor cells; HO‐1; Hematopoietic stem and progenitor cells; HO-1; 610; Protoporphyrins; Mobilization; QH426-470; Granulocyte colony-stimulating factor; granulocyte colony‐stimulating factor; Mice; 03 medical and health sciences; R5-920; Granulocyte Colony-Stimulating Factor; Genetics; Animals; mobilization; Mice, Knockout; 0303 health sciences; CoPP; Articles; Hematopoietic Stem Cells; Hematopoietic Stem Cell Mobilization; 3. Good health; Mice, Inbred C57BL; Female; granulocyte colony-stimulating factor; Heme Oxygenase-1; Granulocytes

Article11 November 2019Open Access Transparent process Cobalt protoporphyrin IX increases endogenous G-CSF and mobilizes HSC granulocytes to the blood Agata Szade Corresponding Author [email protected] orcid.org/0000-0002-0575-4659 Department of Medical Biotechnology, Faculty Biochemistry, Biophysics Jagiellonian University, Krakow, Poland Search for more papers by this author Krzysztof orcid.org/0000-0002-7227-4276 Witold N Nowak orcid.org/0000-0002-1526-3511 Karolina Bukowska-Strakova orcid.org/0000-0001-9181-439X Clinical Immunology Transplantology, Institute Pediatrics, University College, Lucie Muchova orcid.org/0000-0003-2082-1060 Fourth Internal Medicine Biochemistry Laboratory Medicine, First Charles in Prague, Czech Republic Monika Gońka Żukowska orcid.org/0000-0002-2832-2146 Maciej Cieśla orcid.org/0000-0002-8460-1991 Neli Kachamakova-Trojanowska orcid.org/0000-0002-3226-0726 Malopolska Centre Marzena Rams-Baron A. Chelkowski Physics, Silesia, Chorzow, Silesian Center Education Interdisciplinary Research, Alicja Ratuszna Józef Dulak orcid.org/0000-0001-5687-0839 Józkowicz orcid.org/0000-0002-7317-260X Information *,1, Szade1, Nowak1, Bukowska-Strakova1,2, Muchova3, Gońka1, Żukowska1, Cieśla1,7, Kachamakova-Trojanowska1,4, Rams-Baron5,6, Ratuszna5,6, Dulak1,4 Józkowicz1 1Department 2Department 3Fourth 4Malopolska 5A. 6Silesian 7Present address: Division Molecular Hematology, Lund Lund, Sweden *Corresponding author: Tel: [phone]; E-mail: EMBO Mol Med (2019)11:e09571https://doi.org/10.15252/emmm.201809571 PDFDownload PDF article text main figures. Peer ReviewDownload a summary editorial decision including letters, reviewer comments responses feedback. ToolsAdd favoritesDownload CitationsTrack CitationsPermissions ShareFacebookTwitterLinked InMendeleyWechatReddit Figures & Info Abstract Granulocyte colony-stimulating factor (G-CSF) is used clinical practice mobilize cells from bone marrow blood; however, it not always effective. We show that cobalt (CoPP) plasma concentrations G-CSF, IL-6, MCP-1 mice, triggering mobilization hematopoietic stem progenitor (HSPC). Compared with recombinant CoPP higher number HSPC mature granulocytes. In contrast does increase circulating T cells. Transplantation CoPP-mobilized peripheral mononuclear (PBMC) results chimerism faster reconstitution than transplantation PBMC mobilized G-CSF. Although activate Nrf2/HO-1 axis, observed effects are independent. Concluding, expression mobilization-related cytokines has superior mobilizing efficiency compared This observation could lead development new strategies treatment neutropenia transplantation. Synopsis Recombinant treating prior harvesting describes as efficient upstream concentration IL-6 induces blood. functional exogenous leads recovery donor G-CSF-mobilized neutralization inhibits CoPP-induced mobilization. independent axis. Introduction Porphyrins macrocyclic compounds essential plants, bacteria, animals, found molecules such chlorophylls cytochromes (Chandra et al, 2000). form complexes metals generate metalloporphyrins. Often bound metal ions determine unique properties For example, only Fe-protoporphyrin (heme) substrate heme oxygenase-1 (HO-1), but other protoporphyrins tin (SnPP) can inhibit HO-1 enzymatic activity (Schulz 2012) (Fig 1A B). inducer Nrf-2/HO-1 both vitro vivo. Thus, considered potential where may have therapeutic advantages (Shan 2006). Figure 1. upregulates set cytokinesC3H mice were injected CoPP, SnPP, or solvent controls (NaCl, DMSO) each second day 5 days. Samples collected 24 h after last injection. Chemical structures (HO-1 substrate) vivo inducer). B. Heme degradation reaction catalyzed HO-1. C. oxygenase increased decreased SnPP liver measured gas chromatography. D. Total leukocyte red cell count PB. major WBC types. monocyte percentages increasing, whereas lymphocyte percentage decreasing among PB leukocytes CoPP. E. Heatplot cytokine growth Luminex assay. group (red box). F. Selected MCP-1, IP-10, IL-5. Data information: Results shown mean + SEM, one-way ANOVA Bonferroni post-test, n = per group. *P ≤ 0.05; **P 0.01; ***P 0.001; ****P 0.0001. Download figure PowerPoint an enzyme which degrades into carbon monoxide (CO), iron ions, biliverdin 1B), subsequently reduced bilirubin (Tenhunen 1968). The idea activation purposes based on its broad anti-inflammatory (Ryter also influences maturation myeloid Specific deletion lineage (Lyz-Cre:Hmox1fl/fl) partially blocks differentiation progenitors toward macrophages (Wegiel 2014). showed lack affects granulopoiesis (Bukowska-Strakova 2017). HO-1−/− (PB), what connected myelocyte proliferation (BM). Moreover, dendritic (Chauveau 2005), effect seems be (Mashreghi 2008). Given rationale, we sought study how pharmacological induction influence function lineage. Unexpectedly, efficiently well mice. Because constant need improve strategies, investigated induced approach meet requirement. During mobilization, hematopoiesis enhanced, large numbers released BM blood, immature (Lapidot Petit, 2002). Mechanism complex involves several populations pathways (Duhrsen 1988; Lapidot 2002; Tay 2017), (granulocyte factor) one best characterized factors (Souza 1986; Lapid acts progenitors, driving their (Metcalf Nicola, 1983). Apart many agents, stromal cell-derived 1α (Hattori 2001; Devine 2008), (Andrews 1992), interleukin 6 (IL-6) (Pojda Tsuboi, 1990), IL-8 (Laterveer 1995), Groβ (Pelus Fukuda, 2006; Fukuda 2007), granulocyte-macrophage (GM-CSF) (Gianni 1989) act (reviewed (Lapid 2008)). Recently, Hoggatt al (2018) reported rapid highly engrafting single injection AMD3100 combination. Current progress basic science concerning already been successfully translated (Bronchud 1987; Sheridan 1992). Pharmacological outstanding importance prevention (Kelly Wheatley, 2009; Lyman 2010) (HSC). success depends collection sufficient (Kondo Nowadays, source transplantable necessarily itself, rather circulation (Cashen 2007; To 2011). human widely (Mehta 2015). Despite improvements protocols, some patients application inefficient. Among healthy donors, fails 5–30%, high-risk patients, failure rate reaches even up 60% (Ferraro 2011; Patients who fail response might treated additionally plerixafor (To 2011); price drug sometimes obstacle—single dose costs thousand dollars. Therefore, efficiency, agents additional activities. These include modulating extracellular matrix (Saez 2014), phosphorylation signaling proteins (Wang 2016), inhibition proteasome (Ghobadi (Hoggatt Pelus, Here, describe previously unknown compare standard factor, commonly activator inhibitor HO-1, respectively 1B) As expected, administration C3H resulted 2.6-fold liver, 2.4 times 1C). Along activity, had absolute all types (WBC) visible shift 1D). Erythrocyte parameters unaffected examine whether leukocytosis was linked changes profile plasma, performed screen 32 1E). 1E,F) includes chemoattractant protein 1 (MCP-1, CCL2), interferon γ-induced 10 (IP-10, CXCL10), IL-5, greatest extent 1F). However, did CSFs—M-CSF, GM-CSF, IL-3 (Appendix Fig S1). inhibitor, complete any analyzed 1D–F, Appendix Both differ T-cell upregulation 1F), known agent, directly administration. once days stem/progenitor flow cytometry. CD45+ 2A). G-CSF- CoPP-treated highest granulocytes, 1.6 total monocytes similarly Importantly, led lymphocytes, mainly 2. cells, cellsC57BL6xFVB daily Cell numbers, Lymphocytes, Cytokine box) Plasma selected factors. D, Treatment (D) decreases Ly6G+ (E) relative spleen weight. viSNE maps CD11b+ CD11c− colored SSC value. Three experiments performed, pooled. 7 Next, using assay 2B). Consistently previous experiment, high levels 2C). elevated KC (keratinocyte-derived cytokine, CXCL1), MIG (monokine γ, CXCL9). contrast, significantly induce these Analyzing features related (Platzbecker Winkler 2010), F4/80+ MHC IIlow 2D) enlarged spleens 2E). result different lymphocytes. together during phenotype distinct phenotype. Multiparameter analysis revealed similar control G-CSF-treated 2F). density granular control. population intermediate granularity Accordingly, 2D cytometry plots confirm (higher parameter) Ly6G comparing 3A). Granulocytes phenotypically resemble while phenotype, lower expression, typical early stages BM. Altogether, although (Ly6Gmid SSCmid), (Ly6Ghi SSChi; 3B). 3. phenotypeC57BL/6 five consecutive Representative (SSC) B, Relative abundance granulocyte phenotypes (B) (C) Mice proportion isolated C57BL/6xFVB producing reactive oxygen species incubation indicated stimuli. (B, C) two-way (D). composition frequency (CD11b+ Ly6Clow SSCmed Ly6Gmed), pronounced. Percentage SSChi Ly6Ghi) affected 3C). verify checked production (ROS). purpose, incubated N-formylmethionyl-leucyl-phenylalanine (fMLP), phorbol 12-myristate 13-acetate (PMA) opsonized Escherichia coli subjected rhodamine 123 (DHR 123) staining 3D). There ROS-producing stimulation coli, tendency PMA treatment. observe differences between groups fMLP. Interestingly, seemed able produce ROS (CoPP- cells) (G-CSF-mobilized S2). either at least steady-state tested conditions. 2002), pool defined c-Kit+ Lin− Sca-1+ (KLS) S3A). animals KLS visible, statistically significant when four (one-way post-test). further CD34 SLAM markers: CD48 CD150, enable define (KLS CD48−CD150+), MPP (multipotent CD48−CD150−), HPC (hematopoietic CD48+CD150− CD48+CD150+) 4A) (Oguro 2013). HSC, MPP, 4B). Of note, small negative S3B), consistent observation, (in HSC) CD34+ (Tajima affect Only S3A), restricted differentiated fraction LT-HSC (long-term HSC; CD48−CD150+CD34−) 4C). 4. Gating strategy (c-Kit+Lin−Sca-1+) CD150 distinguish cells), progenitors), progenitors). HPC-1 CD48+CD150−), HPC-2 BM, populations, fraction. tends decrease. Scheme common (CMP) granulocytes/monocytes, erythrocytes, platelets. KLS− (c-Kit+Lin−Sca-1−) Numbers lineage-committed GMP (granulocyte-macrophage CD48+CD150−CD34+) MEP (megakaryocyte–erythroid CD48+CD150+CD34−), EP (erythrocyte CD48−CD150−CD34−). treatments decrease BM; smaller. EP, GMP. committed 4D) PB: (GMP) megakaryocyte–erythroid (MEP). erythroid (EP) groups. c-Kit+Lin−Sca-1− (KLS−) less pronounced 4E). Further characterization CD34, CD48, markers change Together there suggest downstream effectors proliferation. functional, transplanted potential. green fluorescent (GFP)-expressing (C57BL/6-Tg(UBC-GFP)30Scha/J) NaCl EV2). At fifth treatment, × 106 lethally irradiated GFP− recipient 105 BM-derived competitor Two performed. Donor C57BL6 genetic background, described poor strain (Roberts 1997). anal