High Constitutive Cytokine Release by Primary Human Acute Myeloid Leukemia Cells Is Associated with a Specific Intercellular Communication Phenotype
Abstract
:1. Introduction
2. Materials and Methods
2.1. AML Patients and Preparation of Primary AML Cells
2.2. Mutation Profiling, Flow Cytometric Analyses, and Analysis of Global Gene Expression Profiles
2.3. Analysis of Constitutive Mediator Release by Primary Human AML Cells
2.4. Proteomic Profiling: Selection of Patients, Sample Preparation, and Proteomic Analysis
2.5. Bioinformatical and Statistical Analyses and Presentation of the Data
3. Results
3.1. The Genetic Heterogeneity of AML Patients: TP53 Mutations are Associated with High-Risk Karyotypes and NPM1 Mutations are Associated with Mutations in DNA Methylation Genes
3.2. Expression of Molecular Differentiation Markers by Primary AML Cells: The Expression of the CD34 Stem Cell Markers Differs between Mutational Subsets
3.3. AML Patients Can Be Subclassified Based on Their Constitutive Release of Extracellular Mediators, but this Capacity Shows no Association with the Mutational Profile
3.4. Comparison of Global Gene Expression Profiles for Patients with Generally High and Low Constitutive Release of Extracellular Mediators
3.5. Comparison of Proteomic Profiles for AML Cell Populations Showing Generally High and Low Constitutive Release of Extracellular Mediators
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Age and gender | Etiology | ||
---|---|---|---|
Median (years) | 64 | Previous chemo-radiotherapy | 1 |
Range (years) | 18–90 | CML | 1 |
Females | 31 | Li–Fraumeni’s syndrome | 1 |
Males | 40 | Polycythemia vera | 1 |
MDS | 8 | ||
Relapse | 10 | ||
de novo | 49 | ||
FAB1 classification | Cytogenetic abnormalities3 | ||
M0/1 | 26 | Adverse | 17 |
M2 | 14 | Favorable | 5 |
M4/5 | 22 | Intermediate | 43 |
M6 | 1 | Normal | 404 |
Unknown | 8 | Unknown | 6 |
CD34 expression | |||
Negative (<20%) | 282 | ||
Positive (>20%) | 43 |
Classification | Mutation | Number with Mutation | Classification | Mutation | Number with Mutation |
---|---|---|---|---|---|
NPM1 | ↑NPM1 | 20 | Chromatin modification | ↓ASXL1 | 12 |
Total group | 20–20 | EZH2 | 3 | ||
Signaling | ↓FLT3-ITD | 20 | GATA2 | 4 | |
FLT3-TKD | 8 | KDM6A | 1 | ||
HRAS | 1 | Total group | 20–15 | ||
JAK2 | 1 | Myeloid transcription factors | |||
KIT | 1 | ↑CEBPA | 8 | ||
KRAS | 5 | ↓RUNX1 | 13 | ||
NRAS | 10 | Total group | 21–18 | ||
PTPN11 | 3 | Spliceosome/transcription repressors | BCOR | 4 | |
Total group | 49–42 | BCORL1 | 4 | ||
Tumor suppressors | CDKN2A | 1 | SF3B1 | 2 | |
CUX1 | 1 | SRSF2 | 8 | ||
IKZF1 | 7 | ZRSB2 | 1 | ||
PHF6 | 3 | Total group | 19–15 | ||
TP53↓ | 7 | Cohesin | RAD21 | 2 | |
WT1 | 5 | SMC1A | 1 | ||
Total group | 24–21 | STAG2 | 8 | ||
DNA methylation | DNMT3A | 19 | Total group | 11–11 | |
IDH1 | 5 | Others | CSF3R | 3 | |
IDH2 | 11 | NOTCH1 | 2 | ||
KMT2A/MLL | 2 | SETBP1 | 1 | ||
TET2 | 12 | Total group | 6–5 | ||
Total group | 49–39 |
Main Classification | Increased Protein Levels in Cells with High Constitutive Release | Increased Protein Levels in Cells with Low Constitutive Release |
---|---|---|
Nucleosome | MBD3 | |
Chromatin, histone, transcription, RNA | TOLLIP, NFKB1 | HIF0, HISTIH2AJ, MTA1, SMARCE1, MEN1, MBD3, POLR1E, CLPX, POLR1A, POLR1B |
DNA repair | CLPX, JUND, POG2 | |
Oncogene | CBL, DBNL | |
Cell cycle regulation | IL16 | |
Intracellular signaling | SYK, HCLS1, AKAP1, TLR2, TOLLIP, AGTRAP, ANXA2, CECR1, INPP5D, LPKN, IKBKB, TBK1 | |
Tyrosine kinase | SYK, HCLS1, FGR, PKN1 | |
SRC tyrosine kinases | HCLS1, FGR, HCK, | |
PI3K-Akt-mTOR | NCF4 | |
RAC1 | RAC1, NCF4, RHOT1, ARHGEF1, PKN1, RHOG, ARHGAP30, PREX1, GMIP, DOK2, AKAP1 | |
GTPase | DNM2, ARHGEF1, PKN1, RHOG, ARHGAP30, PREX1, GMIP, AKAP1, ARHGAP, RAB27A | |
G-protein coupled receptors | ARRB2, ARHGEF1, PREX1, GRK6 | |
Phagocytosis | CYBA, NCF2, NCF4, ELMO2 | |
Protein degradation | CBL, SERPINA1 | |
Intracellular trafficking | VAMP3, DNM2, PICALM, SNX18, ARAP1, ARAP1, TOLLIP, AP1G2, S100A10, S100A4, TOM1, SDCDP, DNAJC13, EPN1, APHGAP, RAB27A | |
Microtubule, cytoskeleton, structure | DNM2, EPN1, SH3KBP1, PKN1, RHOG, AHNAK, SDCDP, S100A4, CKAP4, FAM49B | |
Cell migration | PLXNB2, HCK, DNM2, RHOG, ELMO2, AHNAK | |
Mitochondria, metabolism | FAM49B, FTL, IMPDH1, PDXK | CLPX |
Lysosomes | CTSH, CTSS, CTSZ, LYZ, PSAP | |
Cell metabolism, NADP | HCK, NCF4 | |
Cytokinesis | FMNL1 | |
Extracellular matrix, cell adhesion | EPN1, SH3KBP1 | |
Extracellular mediators | IL16, TLR2, TOLLIP | |
Cell surface molecules | ITGAL, ITGAM. ITGB2, SYK, LILRB2, PKN1, LPXN | |
Integrins | ITGAL, ITGAM. ITGB2, SYK, FGR, LPXN | |
Viability, apoptosis | SH3KBP1, PKN1, ARAP1, TLR2 | |
AML | CBL, PICALM | |
Differentiation | MNDA, NCF1, CECR1 |
Low constitutive mediator release; list of significant GO-terms | Protein number | Fold enrichment | FDR | |
Cell compartment | GO:0005654—nucleoplasm | 31 | 2.8 | 2.3 × 10–5 |
GO:0000790—nuclear chromatin | 8 | 11 | 0.0099 | |
GO:0005736—DNA-directed RNA polymerase I complex | 4 | 80 | 0.017 | |
Molecular function | GO:0003713—transcription coactivator activity | 9 | 8.5 | 0.011 |
GO:0001054—RNA polymerase I activity | 4 | 78 | 0.018 | |
High constitutive mediator release; list of significant GO-terms | ||||
Biological processes | GO:0006954—inflammatory response | 19 | 5.0 | 6.5 × 10–5 |
GO:0045087—innate immune response | 20 | 4.7 | 8.3 × 10–5 | |
GO:0048010—vascular endothelial growth factor receptor signaling pathway | 9 | 13 | 8.6 × 10–4 | |
GO:0007229—integrin-mediated signaling pathway | 10 | 10 | 9.4 × 10–4 | |
GO:0031623—receptor internalization | 7 | 16 | 0.0062 | |
GO:0007165—signal transduction | 29 | 2.5 | 0.015 | |
GO:0098609—cell–cell adhesion | 13 | 4.8 | 0.026 | |
Cell compartment | GO:0070062—extracellular exosome | 73 | 2.7 | 1.4 × 10–13 |
GO:0005829—cytosol | 79 | 2.5 | 5.7 × 10–13 | |
GO:0016020—membrane | 48 | 2.3 | 7.8 × 10–5 | |
GO:0043020—NADPH oxidase complex | 5 | 43 | 0.0048 | |
GO:0005737—cytoplasm | 78 | 1.6 | 0.010 | |
GO:0030670—phagocytic vesicle membrane | 7 | 12 | 0.026 | |
GO:0005925—focal adhesion | 15 | 4.0 | 0.03 | |
GO:0045121—membrane raft | 11 | 5.6 | 0.038 | |
GO:0005884—actin filament | 7 | 11 | 0.046 | |
Molecular function | GO:0005515—protein binding | 129 | 1.4 | 5.8 × 10–6 |
GO:0017124—SH3 domain binding | 11 | 8.9 | 5.8 × 10–4 | |
GO:0035325—Toll-like receptor binding | 4 | 96 | 0.0058 |
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Reikvam, H.; Aasebø, E.; Brenner, A.K.; Bartaula-Brevik, S.; Grønningsæter, I.S.; Forthun, R.B.; Hovland, R.; Bruserud, Ø. High Constitutive Cytokine Release by Primary Human Acute Myeloid Leukemia Cells Is Associated with a Specific Intercellular Communication Phenotype. J. Clin. Med. 2019, 8, 970. https://doi.org/10.3390/jcm8070970
Reikvam H, Aasebø E, Brenner AK, Bartaula-Brevik S, Grønningsæter IS, Forthun RB, Hovland R, Bruserud Ø. High Constitutive Cytokine Release by Primary Human Acute Myeloid Leukemia Cells Is Associated with a Specific Intercellular Communication Phenotype. Journal of Clinical Medicine. 2019; 8(7):970. https://doi.org/10.3390/jcm8070970
Chicago/Turabian StyleReikvam, Håkon, Elise Aasebø, Annette K. Brenner, Sushma Bartaula-Brevik, Ida Sofie Grønningsæter, Rakel Brendsdal Forthun, Randi Hovland, and Øystein Bruserud. 2019. "High Constitutive Cytokine Release by Primary Human Acute Myeloid Leukemia Cells Is Associated with a Specific Intercellular Communication Phenotype" Journal of Clinical Medicine 8, no. 7: 970. https://doi.org/10.3390/jcm8070970