Small particles with large impact: Insights into the unresolved roles of innate immunity in extracellular vesicle-mediated cardiovascular calcification.

Extracellular vesicles (EVs) are critical in the initiation and progression of cardiovascular calcification, and immune cell infiltration and inflammation have a central role in this process. EVs egress from various cardiovascular cell types, which when acquiring specific properties, become calcifying. These calcifying EVs form nidi for microcalcification, which can progress to the macrocalcification lesions that are visualized clinically. We make the distinction between inflammatory-driven and mineral dysregulation-driven calcification, which both share EVs as a central initiator. In inflammation-mediated calcification, inflammation precedes microcalcification and results from EV release from macrophages. Local cellular crosstalk mediated by EVs as well as circulating EVs and other inflammatory nanoparticles, such as calciprotein particles and lipoproteins, are also critical in the progression of cardiovascular calcification. It is imperative that future work links the already established and to be discovered roles of inflammation and innate immunity in cardiovascular calcification to these key signaling and functional roles of these nanoparticles. It remains an understudied area with high potential to unravel mechanistic roles and has important implications in drug target research.

Myeloid CCN3 protects against aortic valve calcification.

BACKGROUND: Cellular communication network factor 3 (CCN3) has been implicated in the regulation of osteoblast differentiation. However, it is not known if CCN3 can regulate valvular calcification. While macrophages have been shown to regulate valvular calcification, the molecular and cellular mechanisms of this process remain poorly understood. In the present study, we investigated the role of macrophage-derived CCN3 in the progression of calcific aortic valve disease. METHODS: Myeloid-specific knockout of CCN3 (Mye-CCN3-KO) and control mice were subjected to a single tail intravenous injection of AAV encoding mutant mPCSK9 (rAAV8/D377Y-mPCSK9) to induce hyperlipidemia. AAV-injected mice were then fed a high fat diet for 40 weeks. At the conclusion of high fat diet feeding, tissues were harvested and subjected to histologic and pathologic analyses. In vitro, bone marrow-derived macrophages (BMDM) were obtained from Mye-CCN3-KO and control mice and the expression of bone morphogenic protein signaling related gene were verified via quantitative real-time PCR and Western blotting. The BMDM conditioned medium was cocultured with human valvular intersititial cells which was artificially induced calcification to test the effect of the conditioned medium via Western blotting and Alizarin red staining. RESULTS: Echocardiography revealed that both male and female Mye-CCN3-KO mice displayed compromised aortic valvular function accompanied by exacerbated valve thickness and cardiac dysfunction. Histologically, Alizarin-Red staining revealed a marked increase in aortic valve calcification in Mye-CCN3-KO mice when compared to the controls. In vitro, CCN3 deficiency augmented BMP2 production and secretion from bone marrow-derived macrophages. In addition, human valvular interstitial cells cultured with conditioned media from CCN3-deficient BMDMs resulted in exaggerated pro-calcifying gene expression and the consequent calcification. CONCLUSION: Our data uncovered a novel role of myeloid CCN3 in the regulation of aortic valve calcification. Modulation of BMP2 production and secretion in macrophages might serve as a key mechanism for macrophage-derived CCN3's anti-calcification function in the development of CAVD. Video Abstract.

Predictive role of cardiac valvular calcification in all-cause mortality of Chinese initial haemodialysis patients: a follow-up study of 4 years.

BACKGROUND: Cardiac valvular calcification (CVC) is prevalent in haemodialysis (HD) patients. Its association with mortality in Chinese incident haemodialysis (IHD) patients remains unknown. METHODS: A total of 224 IHD patients who had just begun HD therapy at Zhongshan Hospital, Fudan University, were enrolled and divided into two groups according to the detection of cardiac valvular calcification (CVC) by echocardiography. The patients were followed for a median of 4 years for all-cause mortality and cardiovascular mortality. RESULTS: During follow-up, 56 (25.0%) patients died, including 29 (51.8%) of cardiovascular disease. The adjusted HR related to all-cause mortality was 2.14 (95% CI, 1.05-4.39) for patients with cardiac valvular calcification. However, CVC was not an independent risk factor for cardiovascular mortality in patients who had just begun HD therapy. CONCLUSION: CVC at baseline is an independent risk factor for all-cause mortality in HD patients and makes an independent contribution to the prediction of all-cause mortality. These findings support the use of echocardiography at the beginning of HD.

Cardiac structural and functional abnormalities in primary hyperparathyroidism.

PURPOSE: Studies on cardiac structural and functional abnormalities in primary hyperparathyroidism (PHPT) have yielded conflicting and inconsistent results. In this prospective case-control study, we sought to compare cardiac structure and function in symptomatic PHPT patients and controls. METHODS: One hundred consecutive symptomatic PHPT patients and 113 matched controls underwent echocardiographic evaluation by the same operator. RESULTS: Left ventricular mass index (LVMI) was significantly higher in patients as compared to controls, (median of 90.95 g/m2 vs 86.5 g/m2, p = 0.041). Patients had significantly lower early trans-mitral diastolic flow (E velocity) as compared to controls (57.13 ± 14.88 vs 64.76 ± 15.45 cm/s, p < 0.001). Patients also had significantly lower early to late mitral annular velocity (E/A) as compared to controls (0.98 ± 0.37 vs 1.10 ± 0.34, p 0.013). Patients had higher frequency of aortic valve calcification (29% vs 2.65%, p < 0.001), mitral annular calcification (23% vs. 4.42%, p < 0.001), myocardial and septal calcifications (25% vs none, p < 0.001) as compared to controls. Serum PTH, calcium and uric acid significantly correlated with calcifications. Serum calcium showed a negative correlation with E/A ratio. CONCLUSIONS: Symptomatic patients with PHPT have substantial cardiac structural and functional abnormalities. These abnormalities include elevated LVMI, diastolic dysfunction, and aortic valve, mitral annular, septal and myocardial calcifications. We strongly suggest and conclude that the evaluation of PHPT patients should not only include traditional end organs like bones and kidneys but also the cardiovascular system in the form of echocardiography to detect subclinical cardiac dysfunction so that the cardiovascular health of such patients can be optimized.

Cardiac valve calcification prevalence and association with neutrophil-to-lymphocyte ratio in newly diagnosed patients with non-dialysis chronic kidney disease stage 3-5.

OBJECTIVE: Cardiac valvular calcification (CVC) is the main cause of cardiovascular disease and all-cause death in patients with chronic kidney disease (CKD). However, the relationship between Neutrophil lymphocyte ratio (NLR) and CVC in patients with CKD is not clear. In this study, we aimed to investigate the prevalence of CVC in newly diagnosed patients with non-dialysis CKD stage 3-5 and evaluate the correlation between NLR and CVC. METHODS: A total of 483 newly diagnosed patients with non-dialysis CKD stage 3-5 were included. According to the presence of CVC, these patients were retrospectively divided into two groups: CVC group and non-CVC group. RESULTS: CVC was found in 80 patients (16.56 %), 53 (10.97 %) of whom had only aortic valve calcification (AVC), 18 (3.73 %) had mitral valve calcification (MVC), and 9 (1.86 %) had both AVC and MVC. The level of NLR in the CVC group was significantly higher than that in the non-CVC group (p=0.002). Multivariate logistic regression analysis showed that NLR was an independent risk factor for CVC (95% CI 1.017~1.225, p=0.020). ROC curve analysis showed that the area under the curve of NLR for predicting CVC was 0.610 (95% CI 0.543-0.676, p=0.002). The best cut-off point of NLR was 3.340, with a sensitivity of 49.4 % and a specificity of 70.0 %. CONCLUSION: CVC is not uncommon in newly diagnosed patients with non-dialysis CKD stage 3-5, and NLR is an independent risk factor for CVC (Tab. 4, Fig. 1, Ref. 34).

Celastrol attenuates arterial and valvular calcification via inhibiting BMP2/Smad1/5 signalling.

Vascular calcification is an important risk factor for the mortality and morbidity in chronic kidney disease (CKD). Unfortunately, until now there is no certain medication targeting vascular calcification in CKD. In this study, we explored the inhibitory effect of celastrol on high calcium-induced vascular calcification and the underlying molecular mechanisms. Cell proliferation assay showed that celastrol inhibited aortic valve interstitial cell (VIC) and vascular smooth muscle cell (VSMC) proliferation when its concentration was higher than 0.6 µmol/L. 0.8 µmol/L celastrol inhibited the expression of osteogenic genes and calcium deposition induced by high-calcium medium in both AVICs and VSMCs. In mouse vascular calcification model induced by adenine combined with vitamin D, alizarin red and immunostaining showed that celastrol inhibited pro-calcification gene expression and calcium deposition in aortic wall and aortic valve tissues. At the molecular level, celastrol inhibited the increase of BMP2, phosphorylated Smad1/5 (p-Smad1/5) and non-phosphorylated ß-catenin (n-p-ß-catenin) induced by high-calcium medium both in vitro and in vivo. Also, BMP2 overexpression reversed the anti-calcification effects of celastrol by recovering the decrease of p-Smad1/5 and n-p-ß-catenin. Furthermore, celastrol prevented the up-regulation of BMPRII and down-regulation of Smad6 induced by high calcium, and this protectory effect can be abolished by BMP2 overexpression. In conclusion, our data for the first time demonstrate that celastrol attenuates high calcium-induced arterial and valvular calcification by inhibiting BMP2/Smad1/5 signalling, which may provide a novel therapeutic strategy for arterial and valvular calcification in patients with CKD.

Valvular heart disease and calcification in CKD: more common than appreciated.

Ischaemic heart disease, sudden cardiac death and arrhythmias, heart failure, stroke and peripheral arterial disease make up >50% of the causes of death in advanced chronic kidney disease (CKD). Calcification of the vascular tree and heart valves is partially related to these complications and has received growing attention in the literature. However, the main focus of research has been on the pathophysiology and consequences of vascular calcification, with less attention being paid to valvular calcification (VC) and its impact on the survival of CKD patients. Although VC has long been seen as an age-related degenerative disorder with minimal functional impact, several studies proved that it carries an increased risk of death and clinical consequences different from those of vascular calcification. In dialysis patients, the annual incidence of aortic valve calcification is nearly 3.3% and the reported prevalence of aortic and mitral VC varies between 25% and 59%. Moreover, calcification of both valves occurs 10-20 years earlier in CKD patients compared with the general population. Therefore, the purpose of this review is to summarize the current knowledge on the pathophysiology and relevance of VC in CKD patients, and to highlight specific clinical consequences and potential therapeutic implications.

Sodium thiosulphate and progression of vascular calcification in end-stage renal disease patients: a double-blind, randomized, placebo-controlled study.

BACKGROUND: Sodium thiosulphate (NaTS) is mostly used in haemodialysis (HD) patients with calcific uraemic arteriolopathy. This double-blind, randomized, placebo-controlled study assessed the effect of NaTS on progression of cardiovascular calcifications in HD patients. METHODS: From 65 screened patients, we recruited 60 patients with an abdominal aorta Agatston calcification score ≥100. Thirty patients were randomized to receive NaTS 25 g/1.73 m2 and 30 patients to receive 100 mL of 0.9% sodium chloride intravenously during the last 15 min of HD over a period of 6 months. The primary endpoint was the absolute change of the abdominal aortic calcification score. RESULTS: The abdominal aortic calcification score and calcification volume of the abdominal aorta increased similarly in both treatment groups during the trial. As compared with the saline group, patients receiving NaTS exhibited a reduction of their iliac artery calcification score (-137 ± 641 versus 245 ± 755; P = 0.049), reduced pulse wave velocity (9.6 ± 2.7 versus 11.4 ± 3.6; P = 0.000) and a lower carotid intima-media thickness (0.77 ± 0.1 versus 0.83 ± 00.17; P = 0.033) and had better preservation of echocardiographic parameters of left ventricular hypertrophy. No patient of the NaTS group developed new cardiac valve calcifications during the trial as compared with 8 of 29 patients in the saline group. By univariate analysis, NaTS therapy was the only predictor of not developing new valvular calcifications. No adverse events possibly related to NaTS infusion were noted. CONCLUSIONS: While NaTS failed to retard abdominal aortic calcification progress, it positively affected calcification progress in iliac arteries and heart valves as well as several other cardiovascular functional parameters.

Comparison of the effects of lanthanum carbonate and calcium carbonate on the progression of cardiac valvular calcification after initiation of hemodialysis.

BACKGROUND: Although mineral metabolism disorder influences cardiac valvular calcification (CVC), few previous studies have examined the effects of non-calcium-containing and calcium-containing phosphate binders on CVC in maintenance hemodialysis patients. The aim of the present study was to compare the effects of lanthanum carbonate (LC) with calcium carbonate (CC) on the progression of CVC in patients who initiated maintenance hemodialysis and to investigate clinical factors related to CVC. METHODS: The current study included 50 subjects (mean age 65 years, 72% males) from our previous randomized controlled trial (LC group, N = 24; CC group, N = 26). CVC was evaluated as CVC score (CVCS) using echocardiography at baseline and 18 months after initiation of hemodialysis. We compared CVCS and the changes between the two groups. We also analyzed the associations between CVCS and any other clinical factors including arterial plaque score (PS) and serum phosphorus levels. RESULTS: Baseline characteristics of study participants including CVCS were almost comparable between the two groups. At 18 months, there were no significant differences in mineral metabolic markers or CVCS between the two groups, and CVCS were significantly correlated with PS (r = 0.39, p < 0.01). Furthermore, changes in CVCS were significantly correlated with average phosphorus levels (r = 0.36, p < 0.05), which were significantly higher in high serum phosphorus and high PS group compared to low serum phosphorus and low PS group (p < 0.05). CONCLUSIONS: In the present study, there were no significant differences between LC and CC with regard to progression of CVC. However, serum phosphorus levels and arterial plaque seem to be important for the progression and formation of CVC in hemodialysis patients.

IL-21 promotes osteoblastic differentiation of human valvular interstitial cells through the JAK3/STAT3 pathway.

Objectives: This study amied to whether IL-21 promotes osteoblast transdifferentiation of cultured human Valvular interstitial cells (VICs). Methods: We first confirmed that IL-21 alters gene expression between CAVD aortic valve tissue and normal samples by immunohistochemistry, qPCR, and western blotting. VICs were cultured and treated with IL-21. Gene and protein expression levels of the osteoblastic markers ALP and Runx2, which can be blocked by specific JAK3 inhibitors and/or siRNA of STAT3, were measured. Results: IL-21 expression was upregulated in calcified aortic valves and promotes osteogenic differentiation of human VICs. IL-21 accelerated VIC calcification through the JAK3/STAT3 pathway. Conclusion: Our data suggest that IL-21 is a key factor in valve calcification and a promising candidate for targeted therapeutics for CAVD.

The association between cardiac valvular calcification and fetuin-A levels in kidney transplant recipients.

OBJECTIVES: Low fetuin-A levels in hemodialysis patients can be associated with development of vascular and valvular calcifications. The mechanisms underlying vascular and valvular calcifications are multifactorial. There are a few studies showing the relationship between low fetuin-A levels and valvular calcification after kidney transplantation. We aimed to evaluate the association between serum fetuin-A levels and valvular calcification in kidney transplant recipients. METHODS: The cardiac valvular calcification was assessed by echocardiography in 56 recipients. Patients were divided into two groups as those with (n = 11) and without (n = 45) aortic and/or mitral valve calcification. The extent of valvular calcification was visually assessed according to the standard visual score method: moderately (multiple larger spots) and heavily calcified (extensive thickening and calcification) of all cusps. Serum fetuin-A levels were measured. RESULTS: The demographic features of both groups were comparable. There was no significant difference between regular physical exercise (63.6% vs. 55.6%), obesity (18.2% vs. 17.8%), abdominal obesity (54.5% vs. 46.7%), smoking (0% vs. 13.3%), hypertension (63.6% vs. 68.9%), left ventricular hypertrophy (45.5% vs. 33.3%) and diabetes mellitus (9.1% vs. 20%) ratios in groups with or without valvular calcification, respectively (p > 0.05). Fetuin-A levels of both groups did not differ. Fetuin-A levels positively correlated with serum creatinine (r 0.326, p = 0.014), and negatively correlated with estimated glomerular filtration rate (r - 0.297, p = 0.026). CONCLUSIONS: We could not find a relationship between serum fetuin-A levels and valvular calcification in kidney recipients. In this population, further studies are needed to assess the role of serum fetuin-A in valvular calcification.

Mir-29b promotes human aortic valve interstitial cell calcification via inhibiting TGF-ß3 through activation of wnt3/ß-catenin/Smad3 signaling.

Herein, we hypothesized that pro-osteogenic MicroRNAs (miRs) could play functional roles in the calcification of the aortic valve and aimed to explore the functional role of miR-29b in the osteoblastic differentiation of human aortic valve interstitial cells (hAVICs) and the underlying molecular mechanism. Osteoblastic differentiation of hAVICs isolated from human calcific aortic valve leaflets obtained intraoperatively was induced with an osteogenic medium. Alizarin red S staining was used to evaluate calcium deposition. The protein levels of osteogenic markers and other proteins were evaluated using western blotting and/or immunofluorescence while qRT-PCR was applied for miR and mRNA determination. Bioinformatics and luciferase reporter assay were used to identify the possible interaction between miR-29b and TGF-ß3. Calcium deposition and the number of calcification nodules were pointedly and progressively increased in hAVICs during osteogenic differentiation. The levels of osteogenic and calcification markers were equally increased, thus confirming the mineralization of hAVICs. The expression of miR-29b was significantly increased during osteoblastic differentiation. Furthermore, the osteoblastic differentiation of hAVICs was significantly inhibited by the miR-29b inhibition. TGF-ß3 was markedly downregulated while Smad3, Runx2, wnt3, and ß-catenin were significantly upregulated during osteogenic induction at both the mRNA and protein levels. These effects were systematically induced by miR-29b overexpression while the inhibition of miR-29b showed the inverse trends. Moreover, TGF-ß3 was a direct target of miR-29b. Inhibition of miR-29b hinders valvular calcification through the upregulation of the TGF-ß3 via inhibition of wnt/ß-catenin and RUNX2/Smad3 signaling pathways.

The presbycardia phenotype: Cardiac remodeling and valvular degeneration in nonagenarians.

BACKGROUND: Nonagenarians (NON) are a growing segment of the population and have a high prevalence of cardiac disease. Many findings encountered on their echocardiograms are also found in younger individuals with valvular or myocardial disease. Therefore, the purpose of this study was to describe this distinct echocardiographic phenotype. METHODS: We identified our study population by querying our echo database to identify unique septuagenarians (SEPT) and nonagenarians (NON) who underwent a transthoracic echocardiogram (TTE) from January 1, 2010 to December 31, 2014. Exclusion criteria were LVEF < 50%, any akinetic wall segment, aortic stenosis, moderate-severe AR and/or severe MR, coronary revascularization within 60 days of study echo, and prior valve surgery. RESULTS: The mean age of SEPT was 73.0 ± 2.0 and NON was 92.0 ± 2.1 (P < 0.001). There was no gender difference between groups. NON had significantly smaller LV end-diastolic diameters than SEPT (41.6 ± 5.7 mm vs 48.0 ± 7.0 mm, P < 0.001). NON had a greater relative wall thickness (0.51 ± 0.10 vs 0.40 ± 0.08, P < 0.001) and more frequently had concentric remodeling or hypertrophy. NON had higher E/Ea ratios and estimated LA pressures (P < 0.01). 48% of NON had moderate-severe mitral annular calcification compared to 25.0% of SEPT (P < 0.01). CONCLUSIONS: Herein, we provide the first comprehensive echocardiographic description of 'presbycardia'; concentric LVH, asymmetric septal hypertrophy, mitral and aortic valve calcification, and increased epicardial fat thickness. This pattern of findings may be increasingly seen as the population ages.

Downregulated MicroRNA-195 in the Bicuspid Aortic Valve Promotes Calcification of Valve Interstitial Cells via Targeting SMAD7.

BACKGROUND/AIMS: Aortic stenosis caused by leaflet calcification in the bicuspid aortic valve (BAV) is more accelerative than that in the tricuspid aortic valve (TAV). MicroRNA-195 (miR-195) is downregulated more in stenotic than in insufficient BAVs, but its expression in BAVs compared with TAVs is unclear. We aimed to investigate the roles of miR-195 and its calcification-related target SMAD7 in stenotic BAVs compared with those in TAVs. METHODS: Twenty-one stenotic BAV and 29 TAV samples were collected from surgical patients and examined for the expression of miR-195 and SMAD7 by RT-PCR. The samples were also assessed by western blotting and immunohistochemistry for the functional protein alteration associated with calcification. Dual-luciferase assay was performed to determine the putative target of miR-195 before the effects of miR-195 expression on osteogenic progression was demonstrated in cultured porcine valve interstitial cells (VICs). RESULTS: Compared with TAV, the expression of miR-195 was remarkably lower in the BAV leaflet with higher expression of SMAD7, which was then validated as a direct target of miR-195. Their negative correlation was then confirmed in cultured VICs. Under an osteogenic environment, the cellular calcification was promoted in miR-195-repressed VICs expressing higher BMP-2 and Runx2 and higher activity of MMP-2 compared with the controls. Finally, higher MMP-2 and MMP-9 expression and more collagen distribution were observed in BAV than TAV samples. CONCLUSIONS: miR-195 is downregulated more in stenotic BAV than TAV in this study. The downregulation of miR-195 is associated with valvular calcification via targeting SMAD7, which promotes the remodeling of the extracellular matrix.

Exposure to ambient air pollution and calcification of the mitral annulus and aortic valve: the multi-ethnic study of atherosclerosis (MESA).

BACKGROUND: Long-term exposure to high ambient air pollution has been associated with coronary artery calcium (CAC), a marker of cardiovascular disease (CVD). Calcifications of left-sided heart valves are also markers of CVD risk. We investigated whether air pollution was associated with valvular calcification and its progression. METHODS: We studied 6253 MESA participants aged 45-84 years who underwent two cardiac CT scans 2.5 years apart to quantify aortic valve calcium (AVC) and mitral annular calcium (MAC). CAC was included for the same timeframe for comparison with AVC/MAC. Ambient particulate matter <2.5 µm (PM2.5) and oxides of nitrogen (NOx) concentrations were predicted from residence-specific spatio-temporal models. RESULTS: The mean age (SD) of the study sample was 62 (10) years, 39% were white, 27% black, 22% Hispanic, and 12% Chinese. The prevalence of AVC and MAC at baseline were 13% and 9% respectively, compared to 50% prevalence of CAC. The adjusted prevalence ratios of AVC and MAC for each 5 µg/m3 higher PM2.5 was 1.19 (95% CI 0.87, 1.62) and 1.20 (0.81, 1.77) respectively, and for CAC was 1.14 (1.01, 1.27). Over 2.5 years, the mean change in Agatston units/year for each 5 µg/m3 higher PM2.5 concentration was 0.29 (-5.05, 5.63) for AVC and 4.38 (-9.13, 17.88) for MAC, compared to 8.66 (0.61, 16.71) for CAC. We found no significant associations of NOx with AVC and MAC. CONCLUSION: Our findings suggest a trend towards increased 2.5-year progression of MAC with exposure to outdoor PM2.5, although this association could not be confirmed. Additional well-powered studies with longer periods of follow-up are needed to further study associations of air pollution with valvular calcium. TRIAL REGISTRATION: Although MESA is not a clinical trial, this cohort is registered at ClinicalTrials.gov Identifier: NCT00005487; Date of registration May 25, 2000.

Relationship between cardiac calcification and left ventricular hypertrophy in patients with chronic kidney disease at hemodialysis initiation.

Coronary artery calcification (CAC), cardiac valve calcification (CVC) and left ventricular hypertrophy (LVH) are frequently observed in chronic kidney disease (CKD) patients. These abnormalities significantly affect morbidity and mortality. The aim of this study was to investigate the relationship between CAC, CVC and LVH in CKD patients. This study included 96 patients who were hospitalized and initiated hemodialysis between December 2011 and July 2014 at our five institutions. Multi-detector computed tomography for the quantification of CAC using the Agatston score and transthoracic echocardiography for assessing CVC and LVH were performed for all patients included in the study. We semi-quantitatively evaluated the severity of CVC as a valvular calcification score. We also assessed the presence of LVH in patients with CAC and/or CVC. Among the 96 patients, the prevalence of CAC was 81.3% and CVC was 65.0%. The severity of CAC was closely and significantly associated with that of CVC. The percentage of patients with LVH was the greatest in those with both severe CAC and CVC. CAC was significantly more severe in patients with concentric hypertrophy compared to those with normal geometry. At the initiation of hemodialysis, most CKD patients had CAC, CVC and LVH. In addition, cardiac calcification was significantly associated with LVH in these patients.

Angiotensin II promotes an osteoblast-like phenotype in porcine aortic valve myofibroblasts.

OBJECTIVES: The mechanisms for pathogenesis of cardiac valve calcification were explored by studying the regulation of the Wnt signaling pathway during the transformation from cardiac valvular myofibroblasts to osteoblast-like phenotype. METHODS: Studies were carried on primary cultured porcine aortic valvular myofibroblasts. The cells were randomly divided into four groups and treated with angiotensin II (Ang II) according to the following: Ang II (10(-6) mol/l), Valsartan (Val) (10(-5) mol/l), Ang II plus Val (Ang II 10(-6) mol/l + Val 10(-5) mol/l) or mock treated as the control. Protein expression of Bone morphogenetic protein 2 (BMP2), Alkaline phosphatase (ALP), and Wnt pathway components, Wnt3a and ß-catenin, was investigated to assess the activation of the Wnt signaling pathway and determine whether cells undergo the transformation to osteoblast-like phenotype. RESULT: Ang II treatment of myofibroblasts led to significant up-regulation of α-SMA expression and activation of the cells. Neither the BMP2 or ALP proteins, nor the mRNA was detectable in the control group or the Val-treated group; however, there was a significant increase in Ang II-treated group (P < 0.01). The Wnt/ß-catenin signaling ligand, Wnt3a, was not expressed in the control or Val-treated groups, whereas in Ang II-treated cells, both Wnt3a and ß-catenin gene expression were enhanced (P < 0.01).The effect of Ang II can be inhibited by the addition of Val (P < 0.05). CONCLUSION: Ang II might act on the Ang II receptor on valvular interstitial cells (VICs) and lead to activation of the Wnt/ß-catenin pathway and hence cause the activation, differentiation and proliferation of myofibroblasts, and finally, osteoblast-like phenotype transformation, leading to calcification of heart valves.

Association between serum beclin 1 level and cardiac valvular calcification in hemodialysis patients.

BACKGROUND: Cardiovascular calcification is a pervasive issue throughout chronic kidney disease (CKD) progression. Autophagy, a fundamental cellular process, exerts significant influence on various cardiac pathologies, including arrhythmias, atherosclerosis, heart failure, and notably, valvular, and vascular calcifications. Beclin-1, a crucial eukaryotic protein, plays a major regulatory role in autophagy as part of the phosphatidylinositol-3-kinase (PI3K) complex. Recent evidence suggests a protective role for Beclin-1-mediated autophagy in CKD vascular calcification, raising its potential as a novel therapeutic target in this context. WE AIMED TO: Investigate the association between serum Beclin 1 levels and the presence of cardiovascular valvular calcification in hemodialysis patients. RESULTS: This study evaluated a cohort of 102 hemodialysis patients, evenly divided into two groups based on echocardiographic findings. All participants underwent serum Beclin 1 measurement and transthoracic echocardiography. Patients with acute kidney injury, active malignancy, or diabetes were excluded. Our study revealed significant differences between the two groups in terms of: Serum Beclin 1 levels, all parameters of lipid profile, prevalence of ischemic heart disease, serum albumin levels and Total calcium. Echocardiography in Group 1 showed that most cases (60.78%) exhibited mild aortic valve calcification. Additionally, significant relationships were observed between Beclin 1 and: ischemic heart disease (p=0.011) Aortic valve calcification on echocardiography (p < 0.001) Interestingly, lower Beclin 1 levels were associated with more severe valve calcification. A Beclin 1 cutoff value of ≤ 35.5 ng/ml demonstrated the highest sensitivity (98%) and specificity (92%). CONCLUSION: Our findings suggest that the serum Beclin 1 level could be incorporated into a predictive model for cardiac valvular calcification in hemodialysis patients.

Predictive value of valvular calcification for the recurrence of persistent atrial fibrillation after radiofrequency catheter ablation.

BACKGROUND: Valvular calcification (VC) is an independent risk factor for cardiovascular diseases. The relationship between VC and atrial fibrillation is not clear. HYPOTHESIS: We treated the aortic valve, mitral valve, and tricuspid valve as a whole and considered the possible association between VC and recurrence of persistent atrial fibrillation (PsAF) after radiofrequency catheter ablation (RFCA). METHODS: This study involved 2687 PsAF patients who underwent RFCA. Data were collected to explore the relationship between VC and outcome. VC was defined by echocardiography in aortic valve, mitral valve, or tricuspid valve. After 1 year follow-up, subgroup analysis, mixed model regression analysis, and score system analysis were performed. The external validation of 133 patients demonstrated the accuracy of this clinical prediction model. RESULTS: Overall, 2687 inpatients were assigned to the recurrence group (n = 682) or the no recurrence group (n = 2005) with or without VC. Compared to patients with no recurrence, the incidence of VC was higher in recurrence patients. Recurrence was present in 18.5%, 34.9%, 39.3%, and 52.0% of the four groups, which met VC numbers of 0, 1, 2, and 3, respectively. After adjustment for potential confounding factors, VC was an independent risk factor for AF recurrence in several models. For multivariable logistic regression, a scoring system was established based on the regression coefficient. The receiver operating characteristic area of the scoring system was 0.787 in the external validation cohort. CONCLUSIONS: VC was an independent risk factor for AF recurrence in PsAF after RFCA. The scoring system may be a useful clinical tool to assess AF recurrence.

Combinations of valvular calcification and serum alkaline phosphatase predict cardiovascular risk among end-stage kidney disease patients.

Background: Valvular calcification (VC) refers to the calcified valvular remodeling associated with kidney dysfunction, especially end-stage kidney disease (ESKD). ESKD patients with VC had significantly higher cardiovascular risk than those without. Factors interacted with VC regarding prognostic prediction in this population were seldom investigated. We aimed to examine the potential synergetic effects of VC and alkaline phosphatase (Alk-P) on ESKD patients' cardiovascular risk and mortality. Methods: ESKD patients undergoing hemodialysis were prospectively enrolled from a medical center in 2018. We identified patients with echocardiography and available serum Alk-P levels. Cox proportional hazard regression was performed to analyze the risk of major adverse cardiovascular events (MACEs), cardiovascular and overall mortality among 4 participant groups (with or without VC versus low or high Alk-P levels). The models were further adjusted for age, sex, and clinical variables. Results: Of the 309 ESKD patients, 38, 46, 112, and 113 had no VC with low Alk-P, no VC with high Alk-P, VC with low Alk-P, and VC with high Alk-P, respectively. After adjusting for age and sex, patients with VC and high Alk-P had a higher risk of developing MACE, cardiovascular and overall mortality (HR, 3.07, 3.67, 3.65; 95% CI 1.38-6.84, 1.1-12.24, 1.29-10.36, respectively). Patients with VC and high Alk-P remained at higher risk of MACE (HR, 2.76; 95% CI 1.17-6.48) than did those without VC and with low Alk-P. Conclusion: Serum Alk-P could be used to identify a subgroup of ESKD patients with elevated cardiovascular risk among those with VC.