Detection of osteopontin in calcified human aortic valves.
ABSTRACT Cardiac valve calcification often results in obstruction of blood flow, which eventually leads to valve replacement. The molecular mechanisms resulting in valve calcification are unknown. Collagen and specific bone matrix proteins are thought to provide the framework for ectopic tissue calcification. This investigation was performed to determine whether the bone matrix protein osteopontin was present in calcified human aortic valves. Proteins extracted from human aortic valve tissue were subjected to polyacrylamide gel electrophoresis followed by Western blotting, using polyclonal antibodies directed against osteopontin. Fresh frozen tissue sections were also screened for osteopontin and macrophages using immunohistochemical techniques. Osteopontin was present in both heavily and minimally calcified aortic valves and absent in noncalcified purely regurgitant or normal aortic valves by both radioimmunoassay (n = 16) and immunohistochemical techniques (n = 8). Osteopontin colocalized with valvular calcific deposits, and macrophages were identified in the vicinity of osteopontin. These results, in addition to showing that osteopontin is present in calcified human aortic valves, suggest that osteopontin is a regulatory protein in pathological calcification. Identification of the cells producing osteopontin in abnormal cardiac valves and of proximate stimuli for its secretion may lead to novel therapeutic strategies to prevent and/or reverse calcific valve disease.
- [Show abstract] [Hide abstract]
ABSTRACT: Calcific aortic valvular disease (CAVD) is an actively regulated process characterized by the activation of specific osteogenic signaling pathways and apoptosis. We evaluated the involvement in CAVD of the TNF-related apoptosis-inducing ligand (TRAIL), an apoptotic molecule which induces apoptosis by interacting with the death receptor (DR)-4 and DR5, and whose activity is modulated by the decoy receptor (DcR)-1 and DcR2. Sections of calcific and normal aortic valves, obtained at surgery time, were subjected to immunohistochemistry and confocal microscopy for TRAIL immunostaining. Valvular interstitial cells (VICs) isolated from calcific (C-VICs) and normal (N-VICs) aortic valves were investigated for the gene and protein expression of TRAIL receptors. Cell viability was assayed by MTT. Von Kossa staining was performed to verify C-VIC ability to produce mineralized nodules. TRAIL serum levels were detected by ELISA. Higher levels of TRAIL were detected in calcific aortic valves and in sera from the same patients respect to controls. C-VICs express significantly higher mRNA and protein levels of DR4, DR5, DcR1, DcR2 and Runx2 compared to N-VICs. C-VICs and N-VICs, cultured in osteogenic medium, express significantly higher mRNA levels of DR4, Runx2 and Osteocalcin compared to baseline. C-VICs and N-VICs were sensitive to TRAIL-apoptotic effect at baseline and after osteogenic differentiation, as demonstrated by MTT assay and caspase-3 activation. TRAIL enhanced mineralized matrix nodule synthesis by C-VICs cultured in osteogenic medium. TRAIL is characteristically present within calcific aortic valves, and mediates the calcification of aortic valve interstitial cells in culture through mechanism involving apoptosis.International journal of cardiology 10/2013; · 6.18 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: AimsThere is currently no medical therapy that can prevent the progression of aortic valve stenosis (AS). Recent data highlight a possible relationship between bone metabolism and AS progression but prospective data are lacking.Methods and resultsSerum levels of calcium, phosphorus, creatinine, 25-OH vitamin D, intact parathyroid hormon (iPTH), C-terminal-telopeptide of type-1-collagen (CTX) and osteocalcin were assessed at baseline in 110 elderly patients (age ≥70 years) with at least mild AS. CTX/osteocalcin ratio was calculated as a marker of bone remodelling balance. AS severity was assessed at baseline and 1-year based on the mean gradient. Two-thirds of patients had low 25-OH vitamin D and 20% had secondary hyperparathyroidism. AS progression was not associated with age, glomerular filtration rate (GFR), calcium and phosphorus levels, calcium-phosphorus product, but significantly with iPTH, CTX/osteocalcin and vitamin D status (all P < 0.01). There was no correlation between iPTH and CTX/osteocalcin (R = 0.04, P = 0.70) and AS progression was associated with CTX/osteocalcin (R = 0.42, P = 0.009), but not with iPTH (R = 0.10, P = 0.55) in patients with normal vitamin D levels, whereas it was associated with iPTH (R = 0.47, P < 0.001) and not with CTX/osteocalcin (R = 0.04, P = 0.73) in those with low vitamin D levels, especially if mild renal insufficiency was present (R = 0.61, P < 0.001).Conclusion In elderly patients with AS, we observed an association between AS progression and vitamin D, iPTH and CTX/osteocalcin ratio and their respective influence varied according to the vitamin D status. In patients with normal vitamin D levels, AS progression was associated with a bone resorptive balance, whereas in patients with low vitamin D levels, AS progression was associated with iPTH and secondary hyperparathyroidism, especially if mild renal insufficiency was present. These findings may have important prognostic and therapeutic implications.Trial registration information: Clinicaltrials.gov identifier number: NCT00338676, funded by AP-HP, the COFRASA study.European Heart Journal 01/2013; · 14.10 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: Objectives This study sought to investigate associations of phosphate metabolism biomarkers with aortic valve calcification (AVC). Background Calcific aortic valve disease (CAVD) is a common progressive condition that involves inflammatory and calcification mediators. Currently there are no effective medical treatments, but mineral metabolism pathways may be important in the development and progression of disease. Methods We examined associations of phosphate metabolism biomarkers, including serum phosphate, urine phosphate, parathyroid hormone (PTH) and serum fibroblast growth factor (FGF)-23, with CT-assessed AVC at study baseline and in short-term follow-up in 6814 participants of the Multi-Ethnic Study of Atherosclerosis (MESA). Results At baseline, AVC prevalence was 13.2%. Higher serum phosphate levels were associated with significantly greater AVC prevalence (relative risk 1.3 per 1 mg/dL increment, 95% confidence incidence: 1.1 to 1.5, p < 0.001). Serum FGF-23, serum PTH, and urine phosphate were not associated with prevalent AVC. Average follow-up CT evaluation was 2.4 years (range 0.9–4.9 years) with an AVC incidence of 4.1%. Overall, phosphate metabolism biomarkers were not associated with incident AVC except in the top FGF-23 quartile. Conclusions Serum phosphate levels are significantly associated with AVC prevalence. Further study of phosphate metabolism as a modifiable risk factor for AVC is warranted.Atherosclerosis 01/2014; 233(2):331–337. · 3.71 Impact Factor