Microproteome of dentoalveolar tissues

• Published in: Bone (New York, N.Y.) Vol. 101; pp. 219 - 229
• Main Authors: Salmon, Cristiane R, Giorgetti, Ana Paula O, Paes Leme, Adriana F, Domingues, Romênia R, Kolli, Tamara N, Foster, Brian L, Nociti, Francisco H
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.08.2017
• Subjects: Animals; Bone; Chromatography, Liquid; Dental cementum; Dental Cementum - metabolism; Dentin; Dentin - metabolism; Extracellular matrix; Extracellular Matrix - metabolism; Mice; Microdissection; Odontogenesis; Odontogenesis - genetics; Odontogenesis - physiology; Orthopedics; Principal Component Analysis; Proteome - analysis; Proteomics; Proteomics - methods; Tandem Mass Spectrometry; EIF4A1; SPARC; IHC; periostin; COL12A1; secreted protein acidic and rich in cysteine; infrared; DMP1; dentin; phosphate-buffered saline; principal component; ACTB; IBSP; AT2A1; dentin sialophosphoprotein; ATPase sarcoplasmic/endoplasmic reticulum Ca 2 + transporting 1; small integrin-binding ligand N-linked glycoprotein; asporin; AB; POSTN; IR; secreted phosphoprotein 24; lumican; tissue-nonspecific alkaline phosphatase; UQCRC1; UQCRC2; ACDC; FBN2; ABC; polyethylene naphthalate; PANTHER; X-linked hypophosphatemia; TGF-β; collagen alpha-1(I) chain; SIBLING; matrix metalloproteinase 20; osteopontin; calsequestrin-1; ECM; COL1A1; cytochrome b- c1 complex subunits 1; matrix Gla protein; cytochrome b- c1 complex subunits 2; SLRP; BMP2; ultraviolet; ALPL; dentin matrix protein 1; bone morphogenetic protein 2; ACO2; extracellular matrix; 3-amino-9-ethylcarbazole; DC; DE; vimentin; ATP2A1; OPN; laser capture microdissection; PHEX; LC-MS/MS; gene ontology; fibrillin-2; FFPE; AEC; alveolar bone; PBS; UV; sarcoplasmic/endoplasmic reticulum calcium ATPase 1; decorin; AHSG; dental cementum; BGN; MEPE; biglycan; mitochondrial trifunctional enzyme subunit beta; ASPN; DCN; PCA; HADHB; MYH1; HADHA; ectopic calcification of vasculature; eukaryotic translation initiation factor 4A1; LCM; myosin-1; small leucine-rich proteoglycan; CASQ1; MMP-20; transforming growth factor beta; 5′ nucleotidase; formalin-fixed paraffin embedded; periodontal ligament; Odontogenesis; collagen alpha-1(XII) chain; NT5E; bone sialoprotein 2; XLH; matrix extracellular phosphoglycoprotein; PDL; immunohistochemistry; cytoplasmic actin 1; LUM; GO; liquid chromatography coupled to tandem mass spectrometry; mitochondrial trifunctional enzyme subunit alpha; DSPP; SPP-24 and SPP-2; avidin-biotinylated peroxidase enzyme complex; PC; fibromodulin; MGP; Proteomics; PEN; protein analysis through evolutionary relationships; VIM; Bone; FMOD; principal component analysis; alpha-2-HS-glycoprotein; aconitate hydratase; metalloendopeptidase homolog PEX; Dentin; Dental cementum; Extracellular matrix
• ISSN: 8756-3282; 1873-2763
• DOI: 10.1016/j.bone.2017.05.014

Abstract Proteomic analysis of extracellular matrices (ECM) of dentoalveolar tissues can provide insights into developmental, pathological, and reparative processes. However, targeted dissection of mineralized tissues, dental cementum (DC), alveolar bone (AB), and dentin (DE), presents technical difficulties. We demonstrate an approach combining EDTA decalcification and laser capture microdissection (LCM), followed by liquid chromatography coupled to tandem mass spectrometry (LC–MS/MS), to analyze proteome profiles of these tissues. Using the LCM-LC-MS/MS approach, a total of 243 proteins was identified from all tissues, 193 proteins in DC, 147 in AB, and 135 proteins DE. Ninety proteins (37% of total) were common to all tissues, whereas 52 proteins (21%) were overlapping in only two. Also, 101 (42%) proteins were exclusively detected in DC (60), AB (15), or DE (26). Identification in all tissues of expected ECM proteins including collagen alpha-1(I) chain (COL1A1), collagen alpha-1(XII) chain (COL12A1), biglycan (BGN), asporin (ASPN), lumican (LUM), and fibromodulin (FMOD), served to validate the approach. Principal component analysis (PCA) and hierarchical clustering identified a high degree of similarity in DC and AB proteomes, whereas DE presented a distinct dataset. Exclusively and differentially identified proteins were detected from all three tissues. The protein-protein interaction network (interactome) of DC was notable for its inclusion of several indicators of metabolic function (e.g. mitochondrial proteins, protein synthesis, and calcium transport), possibly reflecting cementocyte activity. The DE proteome included known and novel mineralization regulators, including matrix metalloproteinase 20 (MMP-20), 5′ nucleotidase (NT5E), and secreted phosphoprotein 24 (SPP-24 or SPP-2). Application of the LCM-LC-MS/MS approach to dentoalveolar tissues would be of value in many experimental designs, including developmental studies of transgenic animals, investigation of treatment effects, and identification of novel regenerative factors.