Osteopontin alters endothelial and valvular interstitial cell behaviour in calcific aortic valve stenosis through HMGB1 regulation.
OBJECTIVE
Calcific aortic valve stenosis (CAVS) is an important clinical problem predominantly affecting elderly individuals. Studies suggest that the progression of CAVS is actively regulated with valve endothelial injury leading to inflammation, fibrosis and calcification. The aim of this study was to delineate the possible regulatory role of osteopontin (OPN) on high-mobility group box 1 (HMGB1) function and the associated inflammatory and fibrotic response in CAVS.


METHODS
Aortic valve leaflets were collected from CAVS patients undergoing aortic valve replacement (n = 40), and control aortic valve leaflets were obtained from heart transplant recipients (n = 15). Valves and plasma were analysed by quantitative real-time polymerase chain reaction (PCR), immunohistochemical staining and Western blot. Recombinant OPN or neutralizing OPN antibody was added to cultured endothelial and valvular interstitial cells (VICs), and cell proliferation scores and HMGB1 expression were assessed.


RESULTS
CAVS valves had a decreased total percentage of VICs but increased numbers of infiltrating macrophages relative to control valves. RT-PCR studies showed higher expression of OPN, the inflammatory cytokine tumour necrosis factor-alpha as well as markers of fibrosis, tissue inhibitor of matrix metalloproteinase 1 and matrix metalloproteinase 2 in CAVS valves. Elevated expression of OPN was also observed in plasma of CAVS patients compared with controls. HMGB1 was detected in the secretory granules of cultured valve endothelial and VICs derived from CAVS valves. The addition of exogenous OPN inhibited the proliferation of cultured endothelial and VICs from CAVS valves and was associated with the extracellular expression of HMGB1, whereas neutralizing OPN had the opposite effect.


CONCLUSIONS
We conclude that altered OPN expression in CAVS affects cellular HMGB1 function inducing cytoplasmic translocation and secretion of HMGB1 in endothelial cells and VICs, thus indicating a regulatory role for OPN in the progression of CAVS through alteration of HMGB1 function.
Critical Care Research Group, University of Queensland, Brisbane, Australia m.passmore@uq.edu.au.
Margaret
M
Passmore
Department of Medicine, University of Queensland, Brisbane, Australia.
Maria
M
Nataatmadja
Inflammation and Healing Research Cluster, School of Health and Sport Sciences, University of the Sunshine Coast, Maroochydoore, Australia.
Yoke L
YL
Fung
Cardiac Surgery Research Unit, The Prince Charles Hospital, Brisbane, Australia.
Bronwyn
B
Pearse
Cardiac Surgery Research Unit, The Prince Charles Hospital, Brisbane, Australia.
Sarah
S
Gabriel
Cardiac Surgery Research Unit, The Prince Charles Hospital, Brisbane, Australia.
Peter
P
Tesar
Critical Care Research Group, University of Queensland, Brisbane, Australia.
John F
JF
Fraser
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001021:Aortic Valve / Q000473:pathology*; D001024:Aortic Valve Stenosis / Q000209:etiology* / Q000473:pathology; D015153:Blotting, Western; D002114:Calcinosis / Q000209:etiology* / Q000473:pathology; D016022:Case-Control Studies; D004730:Endothelium, Vascular / Q000473:pathology; D005260:Female; D024243:HMGB1 Protein / Q000502:physiology*; D006801:Humans; D008297:Male; D008875:Middle Aged; D053495:Osteopontin / Q000502:physiology*; D060888:Real-Time Polymerase Chain Reaction; D055815:Young Adult
D024243:HMGB1 Protein; C585491:HMGB1 protein, human; D053495:Osteopontin
Calcific aortic valve stenosis; Endothelial cells; HMGB1; Osteopontin; Valvular interstitial cells