Abstract
Summary
In male Caucasians with discordant hip bone mineral density (BMD), we applied the subcellular separation and proteome profiling to investigate the monocytic cytosol. Three BMD-associated proteins (ALDOA, MYH14, and Rap1B) were identified based on multiple omics evidence, and they may influence the pathogenic mechanisms of osteoporosis by regulating the activities of monocytes.
Introduction
Osteoporosis is a serious public health problem, leading to significant mortality not only in aging females but also in males. Peripheral blood monocytes (PBMs) play important roles in bone metabolism by acting as precursors of osteoclasts and producing cytokines important for osteoclast development. The first cytosolic sub-proteome profiling analysis was performed in male PBMs to identify differentially expressed proteins (DEPs) that are associated with BMDs and risk of osteoporosis.
Methods
Here, we conducted a comparative proteomics analysis in PBMs from Caucasian male subjects with discordant hip BMD (29 low BMD vs. 30 high BMD). To decrease the proteome complexity and expand the coverage range of the cellular proteome, we separated the PBM proteome into several subcellular compartments and focused on the cytosolic fractions, which are involved in a wide range of fundamental biochemical processes.
Results
Of the total of 3796 detected cytosolic proteins, we identified 16 significant (P < 0.05) and an additional 22 suggestive (P < 0.1) DEPs between samples with low vs. high hip BMDs. Some of the genes for DEPs, including ALDOA, MYH14, and Rap1B, showed an association with BMD in multiple omics studies (proteomic, transcriptomic, and genomic). Further bioinformatics analysis revealed the enrichment of DEPs in functional terms for monocyte proliferation, differentiation, and migration.
Conclusions
The combination strategy of subcellular separation and proteome profiling allows an in-depth and refined investigation into the composition and functions of cytosolic proteome, which may shed light on the monocyte-mediated pathogenic mechanisms of osteoporosis.
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Abbreviations
- PBMs:
-
Peripheral blood monocytes
- BMD:
-
Bone mineral density
- DEPs:
-
Differentially expressed proteins
- ALDOA:
-
Aldolase, fructose-bisphosphate A
- MYH14:
-
Myosin, heavy chain 14
- RAP1B:
-
Member of RAS oncogene family
- GSN:
-
Gelsolin
- IL-1:
-
Interleukin-1
- IL-6:
-
Interleukin-6
- TNF-α:
-
Tumor necrosis factor-α
- MS:
-
Mass spectrometer
- PBMCs:
-
Peripheral blood mononuclear cells
- ATP:
-
Adenosine triphosphate
- RANKL:
-
Receptor activator of nuclear factor kappa-B ligand
- UPLC:
-
Ultra performance liquid chromatography
- HDMS:
-
High-definition mass spectrometry
- FA:
-
Formic acid
- ADH1:
-
Alcohol dehydrogenase 1
- GWAS:
-
Genome-wide association studies
- GO:
-
Gene ontology
- GEFOS:
-
Genetic Factors for Osteoporosis Consortium
- FDR:
-
False discovery rate
- DAVID:
-
Database for Annotation, Visualization and Integrated Discovery
- GO:
-
Gene ontology
- PPI:
-
Protein-protein interactions
- STRING:
-
Search Tool for the Retrieval of Interacting Genes/Proteins
- CFL1:
-
Cofilin 1
- FGG:
-
Fibrinogen gamma chain
- CBX3:
-
Chromobox 3
- CALR:
-
Calreticulin
- ACTN1:
-
Actinin alpha 1
- MYL9:
-
Myosin light chain 9
- ACTB:
-
Actin, beta
- HSP90AA1:
-
Heat shock protein 90-kDa alpha family class A member 1
- HSPA5:
-
Heat shock protein family A (Hsp70) member 5
- CAP1:
-
Adenylate cyclase-associated protein 1
- THBS1:
-
Thrombospondin 1
- TCA:
-
Tricarboxylic acid
- LDHAL6A:
-
Lactate dehydrogenase A like 6A
- LDHB:
-
Lactate dehydrogenase B
- TLN1:
-
Talin1
- M-CSF:
-
Macrophage colony-stimulating factor
- NMHC IIC:
-
Non-muscle myosin II heavy chain
- NM II:
-
Muscle myosin II
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Acknowledgements
This study was partially supported by and/or benefited from grants from National Institutes of Health (P50AR055081, R21AG27110, R01AR057049, R01AR059781) and Edward G. Schlieder Endowment to Tulane University.
This manuscript was not prepared in collaboration with investigators of the Framingham Heart Study (FHS) and does not necessarily reflect the opinions of the FHS, Boston University. The GWA meta-analysis study dataset which used for supportive study in this manuscript was integrated from some individual GWAS datasets. The FHS datasets were obtained from dbGaP at http://www.ncbi.nlm.nih.gov/sites/entrez?db=gap through dbGaP accession phs000007.v14.p6. Assistance with phenotype harmonization and genotype cleaning, as well as with general study coordination, of Genetic Determinants of Bone Fragility study was provided by the NIA Division of Geriatrics and Clinical Gerontology and the NIA Division of Aging Biology. Assistance with phenotype harmonization, SNP selection, data cleaning, meta-analyses, data management, and dissemination and general study coordination, was provided by the PAGE Coordinating Center (U01HG004801-01).
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The Framingham Heart Study is conducted and supported by the National Heart, Lung, and Blood Institute NHLBI in collaboration with Boston University (Contract No. N01-HC-25195). Funding for SHARe Affymetrix genotyping was provided by NHLBI Contract N02-HL-64278. SHARe Illumina genotyping was provided under an agreement between Illumina and Boston University. Funding support for the Framingham BMD datasets was provided by NIH grants R01AR/AG 41398. Funding support for the Genetic Determinants of Bone Fragility (the Indiana fragility study) was provided through the NIA Division of Geriatrics and Clinical Gerontology. Genetic Determinants of Bone Fragility is a genome-wide association studies funded as part of the NIA Division of Geriatrics and Clinical Gerontology. Support for the collection of datasets and samples were provided by the parent grant, Genetic Determinants of Bone Fragility (P01-AG018397). Funding support for the genotyping which was performed at the Johns Hopkins University Center for Inherited Diseases Research was provided by the NIH NIA. The WHI program is funded by the National Heart, Lung, and Blood Institute, National Institutes of Health, U.S. Department of Health and Human Services through contracts N01WH22110, 24152, 32100-2, 32105-6, 32108-9, 32111-13, 32115, 32118, 32119, 32122, 42107-26, 42129-32, and 44221. WHI PAGE is funded through the NHGRI Population Architecture Using Genomics and Epidemiology (PAGE) network (Grant No. U01HG004790).
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Zhu, W., Shen, H., Zhang, JG. et al. Cytosolic proteome profiling of monocytes for male osteoporosis. Osteoporos Int 28, 1035–1046 (2017). https://doi.org/10.1007/s00198-016-3825-y
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DOI: https://doi.org/10.1007/s00198-016-3825-y