Simulation of early calcific aortic valve disease in a 3D platform: A role for myofibroblast differentiation.

PURPOSE: Calcific aortic valve disease (CAVD) is the most prevalent valve disease in the Western world. Recent difficulty in translating experimental results on statins to beneficial clinical effects warrants the need for understanding the role of valvular interstitial cells (VICs) in CAVD. In two-dimensional culture conditions, VICs undergo spontaneous activation similar to pathological differentiation, which intrinsically limits the use of in vitro models to study CAVD. Here, we hypothesized that a three-dimensional (3D) culture system based on naturally derived extracellular matrix polymers, mimicking the microenvironment of native valve tissue, could serve as a physiologically relevant platform to study the osteogenic differentiation of VICs. PRINCIPAL RESULTS: Aortic VICs loaded into 3D hydrogel constructs maintained a quiescent phenotype, similar to healthy human valves. In contrast, osteogenic environment induced an initial myofibroblast differentiation (hallmarked by increased alpha smooth muscle actin [α-SMA] expression), followed by an osteoblastic differentiation, characterized by elevated Runx2 expression, and subsequent calcific nodule formation recapitulating CAVD conditions. Silencing of α-SMA under osteogenic conditions diminished VIC osteoblast-like differentiation and calcification, indicating that a VIC myofibroblast-like phenotype may precede osteogenic differentiation in CAVD. MAJOR CONCLUSIONS: Using a 3D hydrogel model, we simulated events that occur during early CAVD in vivo and provided a platform to investigate mechanisms of CAVD. Differentiation of valvular interstitial cells to myofibroblasts was a key mechanistic step in the process of early mineralization. This novel approach can provide important insight into valve pathobiology and serve as a promising tool for drug screening.

Monoallelic TP53 inactivation is associated with poor prognosis in chronic lymphocytic leukemia: results from a detailed genetic characterization with long-term follow-up.

The exact prognostic role of TP53 mutations (without 17p deletion) and any impact of the deletion without TP53 mutation in CLL are unclear. We studied 126 well-characterized CLL patients by direct sequencing and DHPLC to detect TP53 mutations (exons 2-11). Most patients with 17p deletions also had TP53 mutations (81%). Mutations in the absence of 17p deletions were found in 4.5%. We found a shorter survival for patients with TP53 mutation (n = 18; P = .002), which was more pronounced when analyzed from the time point of mutation detection (6.8 vs 69 months, P < .001). The survival was equally poor for patients with deletion 17p plus TP53 mutation (7.6 months, n = 13), TP53 mutation only (5.5 months, n = 5), and 17p deletion only (5.4 months, n = 3). The prognostic impact of TP53 mutation (HR 3.71) was shown to be independent of stage, VH status, and 11q and 17p deletion in multivariate analysis. Serial samples showed evidence of clonal evolution and increasing clone size during chemotherapy, suggesting that there may be patients where this treatment is potentially harmful. TP53 mutations are associated with poor sur-vival once they occur in CLL. The de-monstration of clonal evolution under selective pressure supports the biologic significance of TP53 mutations in CLL.

Does an Immigrant Background Affect the Indication, Incidence or Outcome of Emergency Cesarean Section? Results of the Prospective Data Collection of 111 Births.

OBJECTIVES: Does the incidence and/or indication(s) for emergency cesarean section differ if the pregnant woman has an immigrant background (IB)? Does a lack of language proficiency (communication problems) and a low acculturation level result in a longer decision-to-delivery interval (D-D interval)? Are neonates born to women with IB by emergency cesarean section in a poorer condition post delivery? PATIENT COHORTS AND METHOD: Standardized interviews were carried out before or immediately after delivery in three Berlin obstetric hospitals. Questions were asked about the sociodemographic background and care aspects as well as about immigration and level of acculturation. Collected data were linked to information obtained from the expectant mother's antenatal records and to care data and perinatal data routinely recorded by the hospitals. Data was analyzed using regression models which adjusted for age, parity, and socio-economic status. RESULTS: The total patient population consisted of 7100 women (rate of response: 89.6%); of these women, 111 required emergency cesarean section (50 women without IB, 61 immigrant women). Risk factors such as late first antenatal check-up, gestational diabetes, pregnancy-induced hypertension, fetal macrosomia, smoking, and weight gain were similar in both patient cohorts. The incidence of and indications for emergency cesarean section and the D-D interval were similar for both groups. Limited German language proficiency and low levels of acculturation among immigrant women did not prolong the D-D interval. There were no statistically relevant differences between immigrant and non-immigrant cohorts with regard to adverse neonatal conditions (5-minute Apgar score ≤ 7, umbilical cord arterial pH < 7.00) or with regard to immediate transfer of the neonate to a pediatric clinic following emergency cesarean section. CONCLUSION: The factor "immigrant background" did not affect the indication or obstetric outcome following emergency cesarean section.

Comparison of different types of cardiac amyloidosis by cardiac magnetic resonance imaging.

OBJECTIVES: We sought to determine cardiac morphological and functional differences between light-chain (AL), mutant-type transthyretin (ATTRmt) and wild-type TTR (ATTRwt) amyloidosis using contrast-enhancement cardiac magnetic resonance imaging (CE-CMR). Finally, we attempted to establish the diagnostic and prognostic impact of these findings. INTRODUCTION: The most common forms of cardiac amyloid are AL and ATTR amyloidosis, but the clinical courses of these variants are quite heterogeneous. While CE-CMR is used to evaluate patients with cardiac amyloidosis, its ability to predict prognosis in these patients is debatable. METHODS: About 130 patients with cardiac amyloidosis (AL, n = 62; ATTRmt, n = 30, ATTRwt, n = 33) were assessed by CE-CMR (cardiac morphology, cardiac function, late gadolinium enhancement). RESULTS: Left ventricular (LV) mass, basal and mid-ventricular maximal wall thickness, and thickness of the inter-atrial septum were higher in ATTRwt when compared to AL and ATTRmt amyloidosis. Tricuspid annular excursion was lower in ATTRwt amyloidosis than in AL amyloidosis. CE was observed in 94.6% of the patients (AL 80.6%; ATTRmt 90%; ATTRwt 87.9%) with significant differences in quality and intensity between the groups. Differentiation of amyloid types was achieved by combination of age, number of organs, the presence of inferolateral CE-CMR, thickness of inter-atrial septum and troponin T. Overall 1-year-survival rates were 93.3, 93.9 and 70.5% in ATTRwt, ATTRmt and AL amyloidosis, respectively. LV mass, mitral annular excursion and NT-proBNP in AL amyloidosis, LV mass maximal apical wall thickness and troponin T in ATTRwt amyloidosis, and finally NT-proBNP and renal function in ATTRmt amyloidosis were independent predictors of outcome. CONCLUSIONS: This study demonstrates that CE-CMR can highlight morphological and functional differences between different types of cardiac amyloidosis. In addition, CE-CMR and cardiac biomarkers provide useful prognostic information in patients with cardiac amyloidosis.

The influence of oxygen and hydrostatic pressure on articular chondrocytes and adherent bone marrow cells in vitro.

Tissue engineering of articular cartilage from chondrocytes or stem cells is considered to be a potential aspect in the treatment of cartilage defects. In order to optimize culture conditions the influence of low oxygen tension (5%) - single or in combination with intermittent hydrostatic pressure (HP: 30/2 min on/off loading; 0.2 MPa) - on the biosynthetic activity (sulfate and proline incorporation) of human osteoarthritic chondrocytes cultured on collagen I/III membranes was investigated. Additionally, chondrogenesis from high density or monolayer cultures of bovine adherent bone marrow cells (aBMC) with and without chondrogenic medium supplements (CM) was analyzed by RT-PCR (mRNA expression of aggrecan and collagen type II). We could show that low oxygen tension increases significantly the biosynthesis of collagen I/III membrane-associated chondrocytes and even higher under co-stimulation with HP. While there is no chondrogenesis in monolayer cultures, CM induces expression of cartilage matrix molecules in high density cultures of aBMC which is even increased under the influence of low oxygen tension. Both, low oxygen tension and HP without CM are alone not sufficient stimuli for chondrogenesis. It can be concluded that low oxygen tension and HP might be useful tools in cartilage tissue engineering and that these physico-chemical factors promote but do not induce chondrogenesis under the given conditions.

Noninvasive risk stratification of patients with transthyretin amyloidosis.

OBJECTIVES: We sought to evaluate noninvasive parameters by electrocardiography, echocardiography, technetium-99m-3,3-diphosphono-1,2-propanodicarboxylic acid ((99m)Tc-DPD) scintigraphy, and cardiac magnetic resonance for the prediction of all-cause mortality in patients with cardiac transthyretin amyloidosis (ATTR). BACKGROUND: ATTR may present with highly variable symptoms, including polyneuropathy and cardiomyopathy, the latter being associated with a poor outcome. However, data on noninvasive risk stratification of ATTR are limited. METHODS: A total of 70 patients with ATTR were evaluated by echocardiography, cardiac biomarkers, and (99m)Tc-DPD scintigraphy. Cardiac magnetic resonance was performed in 30 patients. Echocardiographic findings and plasma levels of biomarkers were correlated with results of quantitative analysis of scintigraphy using a region-of-interest technique (whole-body as well as heart tracer retention). Receiver-operating characteristic (ROC) analysis was performed to calculate a cutoff value of (99m)Tc-DPD scintigraphy for heart retention for the diagnosis of cardiac amyloid involvement with the highest sensitivity and specificity. Univariate and multivariate analyses were performed in patients with cardiac involvement (n = 60) to determine noninvasive predictors of all-cause mortality. RESULTS: Scintigraphy findings correlated with morphological (interventricular septum thickness, left ventricular hypertrophy index) as well as functional (mitral annular systolic velocity, mitral/tricuspid annular plane systolic excursion) findings, cardiac biomarkers, renal function, and late gadolinium enhancement. The ROC-derived cutoff for the detection of cardiac amyloidosis by scintigraphic heart tracer retention was 4.8%. Univariate Cox regression revealed N-terminal pro-B-type natriuretic peptide, troponin T, mitral annular plane systolic excursion, and left ventricular hypertrophy index as predictors of all-cause mortality. However, on multivariate analysis, troponin T remained the only independent predictor of survival. The ROC-derived cutoff value of troponin T predicting all-cause mortality with the highest sensitivity (80.0%) and specificity (68.7%) was 0.0375 ng/l. CONCLUSIONS: Quantitative analysis of tracer retention is capable of characterizing the severity of cardiac involvement in ATTR. By multivariate analysis, troponin T remained the only independent predictor of survival.