Monocyte Proteomics Reveals Involvement of Phosphorylated HSP27 in the Pathogenesis of Osteoporosis

Abstract

Peripheral monocytes, precursors of osteoclasts, have emerged as important candidates for identifying proteins relevant to osteoporosis, a condition characterized by low Bone Mineral Density (BMD) and increased susceptibility for fractures. We employed 4-plex iTRAQ (isobaric tags for relative and absolute quantification) coupled with LC-MS/MS (liquid chromatography coupled with tandem mass spectrometry) to identify differentially expressed monocyte proteins from premenopausal and postmenopausal women with low versus high BMD. Of 1801 proteins identified, 45 were differentially abundant in low versus high BMD, with heat shock protein 27 (HSP27) distinctly upregulated in low BMD condition in both premenopausal and postmenopausal categories. Validation in individual samples (n = 80) using intracellular ELISA confirmed that total HSP27 (tHSP27) as well as phosphorylated HSP27 (pHSP27) was elevated in low BMD condition in both categories (P < 0.05). Further, using transwell assays, pHSP27, when placed in the upper chamber, could increase monocyte migration (P < 0.0001) and this was additive in combination with RANKL (receptor activator of NFkB ligand) placed in the lower chamber (P = 0.05). Effect of pHSP27 in monocyte migration towards bone milieu can result in increased osteoclast formation and thus contribute to pathogenesis of osteoporosis. Overall, this study reveals for the first time a novel link between monocyte HSP27 and BMD.

In-Depth Proteomic Characterization of Classical and Non-Classical Monocyte Subsets

“…There has been some genome-wide transcriptomics analysis [ 6 , 13 , 14 ] of individual monocyte subsets, but less information at the protein level is available. Most proteomic analyses performed on monocytes have used monocyte derived cell lines or with the whole monocyte population [ 33 , 34 , 35 , 36 , 37 , 38 , 39 , 40 , 41 , 42 ]. Wong’s group have carried out proteomic analysis on purified monocyte populations [ 43 , 44 ], where they used iTRAQ to determine differences between cell populations.…”

Multi-omics Data Integration for Identifying Osteoporosis Biomarkers and Their Biological Interaction and Causal Mechanisms

“…Nitrogen-containing bisphosphonate can potently inhibit the prenylation and function of GTP-binding proteins required for osteoclast formation and now is firmly established as first-line therapy for osteoporosis (Grey and Reid, 2006). TUBB6 plays a key role in GTP binding and has been associated with BMD variation (Daswani et al, 2015 NSUN4, C11orf83, PDSS2, PACS2, NDUFV3, and ALDH3B1), 6 DMC-annotated genes (PPARGC1A, ILKAP-PER2, SIRT6, RTEL-TNFRSF6B, VARS2-GTF2H4, and MFN2-TNFRSF8-TNFRSF1B), and 3 metabolites biomarkers ({4-((E)-2-(2,3,5-trihydroxyphenyl)ethenyl)phenyl}oxidanesulfonic acid, Acetyl-T2 toxin, and LysoPC (16:0)) associated with telomere/mitochondrial activity. TERF1 encodes a component of the telomere nucleoprotein complex, which can regulate telomere elongation and plays a key role in aging-related disease (Blasco, 2005).…”

“Omics” Signatures in Peripheral Monocytes from Women with Low BMD Condition

“…In our study, we used iTRAQ based quantitative proteomics approach to study monocytes from premenopausal and postmenopausal Indian women with low versus high BMD, all in a single experimental platform using 4-plex iTRAQ coupled to nano-LC-MS/MS [ 42 ]. We found that heat shock protein 27, also known as heat shock protein beta-1 (HSPB1), was distinctly upregulated in low BMD condition in both premenopausal and postmenopausal categories [ 42 ]. Validation revealed that total HSP27 (tHSP27) and phosphorylated HSP27 (pHSP27) were elevated in low BMD condition in both categories not only in monocytes but also in sera [ 42 , 43 ].…”