Bone Marrow Monocytes and Derived Dendritic Cells from Myelodysplastic Patients Have Functional Abnormalities Associated with Defective Response to This information is current as Bacterial Infection of October 2, 2021. Laiz C. Bento, Nydia S. Bacal, Fernanda A. Rocha, Patricia Severino and Luciana C. Marti J Immunol published online 16 March 2020 http://www.jimmunol.org/content/early/2020/03/13/jimmun Downloaded from ol.1900328 Supplementary http://www.jimmunol.org/content/suppl/2020/03/13/jimmunol.190032 Material 8.DCSupplemental http://www.jimmunol.org/ Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! 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Published March 16, 2020, doi:10.4049/jimmunol.1900328 The Journal of Immunology Bone Marrow Monocytes and Derived Dendritic Cells from Myelodysplastic Patients Have Functional Abnormalities Associated with Defective Response to Bacterial Infection Laiz C. Bento,* Nydia S. Bacal,* Fernanda A. Rocha,† Patricia Severino,† and Luciana C. Marti† Myelodysplastic syndromes (MDS) are a heterogeneous group of hematopoietic stem cell diseases characterized by dysplasia of one or more hematologic lineages and a high risk of developing into acute myeloid leukemia. MDS patients have recurrent bac- terial infections and abnormal expression of CD56 by monocytes. We investigated MDS patients’ bone marrow CD56+/CD562 monocytes and their in vitro–derived dendritic cell populations in comparison with cells obtained from disease-free subjects. We found that monocytes from MDS patients, irrespective of CD56 expression, have reduced phagocytosis activity and low expres- sion of genes involved in triggering immune responses, regulation of immune and inflammatory response signaling pathways, and in Downloaded from the response to LPS. Dendritic cells derived in vitro from MDS monocytes failed to develop dendritic projections and had reduced expression of HLA-DR and CD86, suggesting that Ag processing and T cell activation capabilities are impaired. In conclusion, we identified, in both CD56+ and CD562 monocytes from MDS patients, several abnormalities that may be related to the increased susceptibility to infections observed in these patients. The Journal of Immunology, 2020, 204: 000–000. yelodysplastic syndromes (MDS) include a hetero- Monocytes are cells with high plasticity. They were originally http://www.jimmunol.org/ geneous group of hematopoietic stem cell diseases classified according to morphological characteristics, but, more M characterized by dysplasia of one or more hemato- recently, the expression of CD14 and CD16 have become important logic lineages. The clinical course of MDS is variable, and patients monocyte markers. Classical monocytes, expressing high levels of are at high risk of developing acute myeloid leukemia (1, 2). In CD14 and absent CD16 (CD14hi/CD162), are ∼80% of all mono- addition, MDS patients frequently suffer from severe bacterial cytes and considered to be better at secreting proinflammatory cy- infections, but the mechanisms underlying their high suscepti- tokines and in generating reactive oxygen species. Intermediate bility to infection are unknown (3, 4). monocytes (CD14hi/CD16+) are more efficient in phagocytosis and In MDS, besides already described granulocyte defects, express higher levels of MHC class II and accessory molecules (7). by guest on October 2, 2021 monocytic lineage cells also exhibit several phenotypic alterations When monocytes are cultivated in presence of GM-CSF and IL-4, (2). The expression of CD56 in monocytes is often abnormal in they can exhibit features of immature dendritic cells (iDCs), and in chronic myelomonocytic leukemia, which is a relatively frequent response to inflammatory signals, they can differentiate into mature MDS. To be considered as aberrant, the percentage of CD56- dendritic cells (mDCs) with Ag presentation ability (8). expressing monocytes should be equal or higher than 20% (2, 5). The mechanisms underlying the immune response against patho- The main function of the CD56 molecule is to promote intercel- gens comprise both innate and adaptive immunity. The first response lular adhesion; it can also act as a receptor involved in cellular against microorganisms is that of innate immunity. It starts by host migration, proliferation, and differentiation (6). Nevertheless, until cells recognition of pathogen-associated molecular patterns (PAMPs). now, there has been no information on the consequences of altered These PAMPs are recognizable by evolutionary conserved receptor CD56 expression observed in malignant neoplasms. molecules present in a variety of host cells such as TLRs, RIG-I-like receptors (RLRs), NOD-like receptors (NLRs), C-type lectin-like *Clinical Pathology Laboratory, Hospital Israelita Albert Einstein, Sa˜o Paulo 05652 receptors (CLRs), and DNA sensors (9–12). Among TLRs, TLR1, 000, Brazil; and †Experimental Research Laboratory, Hospital Israelita Albert TLR2, TLR4, and TLR6 are present in the cellular plasma mem- Einstein, Sa˜o Paulo 05652 000, Brazil brane and mainly recognize hydrophobic sugar complexed PAMPs ORCIDs: 0000-0002-6682-9343 (P.S.); 0000-0002-3890-0827 (L.C.M.). (such as LPS, whereas TLR3, TLR7, TLR8, and TLR9 are localized Received for publication March 19, 2019. Accepted for publication February 6, 2020. in endosomes and sense hydrophilic sugar complexed PAMPs such This work was supported by National Council of Technological and Scientific as 59-ppp-ssRNA) (11–13). Monocytes, macrophages, and dendritic Development process number 441665/2014-4. cells (DCs) express most PAMP receptors. These cells establish the The microarray data presented in this article have been submitted to the Gene Expression connection between the innate and adaptive immune responses. Omnibus (https://www.ncbi.nlm.nih.gov/geo/) under accession number GSE142450. They act by engulfing and eliminating microorganisms, processing Address correspondence and reprint requests to Dr. Luciana C. Marti, Hospital Israelita Ag molecules, and presenting the derived peptides in association Albert Einstein, Experimental Research Department, Avenida Albert Einstein, 627, Bloco A, 2oSS, Morumbi, Sa˜o Paulo SP 05652 000, Brazil. E-mail address: with MHC molecules to lymphocyte recognition as the initial step [email protected] of adaptive immune responses. 2 The online version of this article contains supplemental material. The aim of the current study was to study CD56+ and CD56 Abbreviations used in this article: BM, bone marrow; DC, dendritic cell; iDC, bone marrow (BM) monocytes and thereof derived DCs obtained immature dendritic cell; mDC, mature dendritic cell; MDS, myelodysplastic syndrome; from MDS patients and from disease-free individuals. We analyzed PAMP, pathogen-associated molecular pattern. the following aspects: cell surface markers by flow cytometry, Copyright Ó 2020 by The American Association of Immunologists, Inc. 0022-1767/20/$37.50 morphology and phagocytic activity, and gene expression patterns. www.jimmunol.org/cgi/doi/10.4049/jimmunol.1900328 2 MOLECULAR ALTERATIONS IN MDS PATIENTS’ MONOCYTES We found that cells from MDS patients had reduced phagocytic ac- The CD56-negative cells originating from the first-step separation were further separated (second-column separation) based on the CD14 tivity, low expression of MHC class II and of costimulatory mole- + 2 cules and TLRs, and reduced expression of cytokine and chemokine expression to yield a 100% of CD14 C56 population. Disease-free samples from BM donors contained ,3% CD56+ monocytes, so the genes. Taken together, results suggest functional abnormalities within separation was only based on CD14 expression. the monocyte population that may contribute to deficient response to bacterial infection, a common feature in MDS patients. Generation of DCs from BM monocytes CD14+CD562 and CD14+CD56+ monocytes were dispensed into six-well plates Materials and Methods containing X-VIVO 15 Medium (Cambrex) supplemented with anti- biotic–antimycotic solution (Life Technologies) containing 100 U/ml BM samples penicillin, 100 mg/ml streptomycin, and 0.25 mg/ml amphotericin B. To gen- BM samples from five MDS-diagnosed patients and from five individuals erate iDCs, the cells were cultured in the presence of human rIL-4 (20 ng/ml) without myelodysplasia or hematological disease (disease-free individuals) and GM-CSF (50 ng/ml) (both from R&D Systems, Minneapolis, MN) for were used in this study. The samples were remainders from diagnostic 6dat37˚Cina5%CO2 incubator. mDCs were obtained after iDCs stimu- routine laboratory exams. Disease-free individuals were BM donors with no lation with LPS 100 ng/ml for 24 h (Sigma-Aldrich). Because of low numbers MDS-associated abnormalities. The five MDS patients enrolled