Aluminum, a colorful gamechanger: Uptake of an aluminum-containing food color in human cells and its implications for human health.

Aluminum is added to many food colors to change their solubility. This study compares the aluminum-containing food color carmine with its aluminum-free version carminic acid (both E 120), hypothesizing that the addition of aluminum does not only change the color's solubility, but also its effects on human cells. We could show that carmine, but not carminic acid, is taken up by gastrointestinal Caco-2 and umbilical vein endothelial cells (HUVEC). Clear differences between gene expression profiles of Caco-2 cells exposed to carmine, carminic acid or control were shown. KEGG analysis revealed that carmine-specific genes suppress oxidative phosphorylation, and showed that this suppression is associated with neurodegenerative diseases such as Alzheimer and Parkinson disease. Furthermore, carmine, but not carminic acid, increased proliferation of Caco-2 cells. Our findings show that a food color containing aluminum induces different cellular effects compared to its aluminum-free form, which is currently not considered in EU legislation.

An Atraumatic Mock Loop for Realistic Hemocompatibility Assessment of Blood Pumps.

OBJECTIVE: Conventional mock circulatory loops (MCLs) cannot replicate realistic hemodynamic conditions without inducing blood trauma. This constrains in-vitro hemocompatibility examinations of blood pumps to static test loops that do not mimic clinical scenarios. This study aimed at developing an atraumatic MCL based on a hardware-in-the-loop concept (H-MCL) for realistic hemocompatibility assessment. METHODS: The H-MCL was designed for 450 ± 50 ml of blood with the polycarbonate reservoirs, the silicone/polyvinyl-chloride tubing, and the blood pump under investigation as the sole blood-contacting components. To account for inherent coupling effects a decoupling pressure control was derived by feedback linearization, whereas the level control was addressed by an optimization task to overcome periodic loss of controllability. The HeartMate 3 was showcased to evaluate the H-MCL's accuracy at typical hemodynamic conditions. To verify the atraumatic properties of the H-MCL, hemolysis (bovine blood, n = 6) was evaluated using the H-MCL in both inactive (static) and active (minor pulsatility) mode, and compared to results achieved in conventional loops. RESULTS: Typical hemodynamic scenarios were replicated with marginal coupling effects and root mean square error (RMSE) below 1.74 ± 1.37 mmHg while the fluid level remained within ±4% of its target value. The normalized indices of hemolysis (NIH) for the inactive H-MCL showed no significant differences to conventional loops ( ∆NIH = -1.6 mg/100 L). Further, no significant difference was evident between the active and inactive mode in the H-MCL ( ∆NIH = +0.3 mg/100 L). CONCLUSION AND SIGNIFICANCE: Collectively, these findings indicated the H-MCL's potential for in-vitro hemocompatibility assessment of blood pumps within realistic hemodynamic conditions, eliminating inherent setup-related risks for blood trauma.

The Role of Trace Elements in Cardiovascular Diseases.

Essential trace elements play an important role in human physiology and are associated with various functions regulating cellular metabolism. Non-essential trace elements, on the other hand, often have well-documented toxicities that are dangerous for the initiation and development of diseases due to their widespread occurrence in the environment and their accumulation in living organisms. Non-essential trace elements are therefore regarded as serious environmental hazards that are harmful to health even in low concentrations. Many representatives of these elements are present as pollutants in our environment, and many people may be exposed to significant amounts of these substances over the course of their lives. Among the most common non-essential trace elements are heavy metals, which are also associated with acute poisoning in humans. When these elements accumulate in the body over years of chronic exposure, they often cause severe health damage in a variety of tissues and organs. In this review article, the role of selected essential and non-essential trace elements and their role in the development of exemplary pathophysiological processes in the cardiovascular system will be examined in more detail.

Cyclic pressure induced decellularization of porcine descending aortas.

The demand for decellularized xenogeneic tissues used in reconstructive heart surgery has increased over the last decades. Complete decellularization of longer and tubular aortic sections suitable for clinical application has not been achieved so far. The present study aims at analyzing the effect of pressure application on decellularization efficacy of porcine aortas using a device specifically designed for this purpose. Fresh porcine descending aortas of 8 cm length were decellularized using detergents. To increase decellularization efficacy, detergent treatment was combined with pressure application and different treatment schemes. Quantification of penetration depth as well as histological staining, scanning electron microscopy, and tensile strength tests were used to evaluate tissue structure. In general, application of pressure to aortic tissue does neither increase the decellularization success nor the penetration depth of detergents. However, it is of importance from which side of the aorta the pressure is applied. Application of intermittent pressure from the adventitial side does significantly increase the decellularization degree at the intimal side (compared to the reference group), but had no influence on the penetration depth of SDC/SDS at both sides. Although the present setup does not significantly improve the decellularization success of aortas, it is interesting that the application of pressure from the adventitial side leads to improved decellularization of the intimal side. As no adverse effects on tissue structure nor on mechanical properties were observed, optimization of the present protocol may potentially lead to complete decellularization of larger aortic segments.

Noncanonical atherosclerosis as the driving force in tricuspid aortic valve associated aneurysms - A trace collection.

Pathogenic mechanisms in degenerative thoracic aortic aneurysms (TAA) are still unclear. There is an ongoing debate about whether TAAs are caused by uniform or distinct processes, which would obviously have a major impact on future treatment strategies. Clearly, the ultimate outcome of TAA subgroups associated with a tricuspid aortic valve (TAV) or a bicuspid aortic valve (BAV) is the same, namely a TAA. Based on results from our own and others' studies, we decided to compare the different TAAs (TAV and BAV) and controls using a broad array of analyses, i.e., metabolomic analyses, gene expression profiling, protein expression analyses, histological characterization, and matrix-assisted laser desorption ionization imaging. Central findings of the present study are the presence of noncanonical atherosclerosis, pathological accumulation of macrophages, and disturbances of lipid metabolism in the aortic media. Moreover, we have also found that lipid metabolism is impaired systemically. Importantly, all of the above-described phenotypes are characteristic for TAV-TAA only, and not for BAV-TAA. In summary, our results suggest different modes of pathogenesis in TAV- and BAV-associated aneurysms. Intimal atherosclerotic changes play a more central role in TAV-TAA formation than previously thought, particularly as the observed alterations do not follow classical patterns. Atherosclerotic alterations are not limited to the intima but also affect and alter the TAV-TAA media. Further studies are needed to i) clarify patho-relevant intima-media interconnections, ii) define the origin of the systemic alteration of lipid metabolism, and iii) to define valid biomarkers for early diagnosis, disease progression, and successful treatments in TAV-TAAs.

Impacts of Telomeric Length, Chronic Hypoxia, Senescence, and Senescence-Associated Secretory Phenotype on the Development of Thoracic Aortic Aneurysm.

Thoracic aortic aneurysm (TAA) is an age-related and life-threatening vascular disease. Telomere shortening is a predictor of age-related diseases, and its progression is associated with premature vascular disease. The aim of the present work was to investigate the impacts of chronic hypoxia and telomeric DNA damage on cellular homeostasis and vascular degeneration of TAA. We analyzed healthy and aortic aneurysm specimens (215 samples) for telomere length (TL), chronic DNA damage, and resulting changes in cellular homeostasis, focusing on senescence and apoptosis. Compared with healthy thoracic aorta (HTA), patients with tricuspid aortic valve (TAV) showed telomere shortening with increasing TAA size, in contrast to genetically predisposed bicuspid aortic valve (BAV). In addition, TL was associated with chronic hypoxia and telomeric DNA damage and with the induction of senescence-associated secretory phenotype (SASP). TAA-TAV specimens showed a significant difference in SASP-marker expression of IL-6, NF-κB, mTOR, and cell-cycle regulators (γH2AX, Rb, p53, p21), compared to HTA and TAA-BAV. Furthermore, we observed an increase in CD163+ macrophages and a correlation between hypoxic DNA damage and the number of aortic telocytes. We conclude that chronic hypoxia is associated with telomeric DNA damage and the induction of SASP in a diseased aortic wall, promising a new therapeutic target.

Hepatectomy-induced apoptotic extracellular vesicles stimulate neutrophils to secrete regenerative growth factors.

BACKGROUND & AIMS: Surgical resection of the cancerous tissue represents one of the few curative treatment options for neoplastic liver disease. Such partial hepatectomy (PHx) induces hepatocyte hyperplasia, which restores liver function. PHx is associated with bacterial translocation, leading to an immediate immune response involving neutrophils and macrophages, which are indispensable for the priming phase of liver regeneration. Additionally, PHx induces longer-lasting intrahepatic apoptosis. Herein, we investigated the effect of apoptotic extracellular vesicles (aEVs) on neutrophil function and their role in this later phase of liver regeneration. METHODS: A total of 124 patients undergoing PHx were included in this study. Blood levels of the apoptosis marker caspase-cleaved cytokeratin-18 (M30) and circulating aEVs were analyzed preoperatively and on the first and fifth postoperative days. Additionally, the in vitro effects of aEVs on the secretome, phenotype and functions of neutrophils were investigated. RESULTS: Circulating aEVs increased at the first postoperative day and were associated with higher concentrations of M30, which was only observed in patients with complete liver recovery. Efferocytosis of aEVs by neutrophils induced an activated phenotype (CD11bhighCD16highCD66bhighCD62Llow); however, classical inflammatory responses such as NETosis, respiratory burst, degranulation, or secretion of pro-inflammatory cytokines were not observed. Instead, efferocytosing neutrophils released various growth factors including fibroblast growth factor-2 and hepatocyte growth factor (HGF). Accordingly, we observed an increase of HGF-positive neutrophils after PHx and a correlation of plasma HGF with M30 levels. CONCLUSIONS: These data suggest that the clearance of PHx-induced aEVs leads to a population of non-inflammatory but regenerative neutrophils, which may support human liver regeneration. LAY SUMMARY: In this study, we show that the surgical removal of a diseased part of the liver triggers a specific type of programmed cell death in the residual liver tissue. This results in the release of vesicles from dying cells into the blood, where they are cleared by circulating immune cells. These respond by secreting hepatocyte growth factors that could potentially support the regeneration of the liver remnant.

The Role of Telocytes and Telocyte-Derived Exosomes in the Development of Thoracic Aortic Aneurysm.

A hallmark of thoracic aortic aneurysms (TAA) is the degenerative remodeling of aortic wall, which leads to progressive aortic dilatation and resulting in an increased risk for aortic dissection or rupture. Telocytes (TCs), a distinct type of interstitial cells described in many tissues and organs, were recently observed in the aortic wall, and studies showed the potential regulation of smooth muscle cell (SMC) homeostasis by TC-released shed vesicles. The purpose of the present work was to study the functions of TCs in medial degeneration of TAA. During aneurysmal formation an increase of aortic TCs was identified in human surgical specimens of TAA-patients, compared to healthy thoracic aortic (HTA)-tissue. We found the presence of epithelial progenitor cells in the adventitial layer, which showed increased infiltration in TAA samples. For functional analysis, HTA- and TAA-telocytes were isolated, characterized, and compared by their protein levels, mRNA- and miRNA-expression profiles. We detected TC and TC-released exosomes near SMCs. TAA-TC-exosomes showed a significant increase of the SMC-related dedifferentiation markers KLF-4-, VEGF-A-, and PDGF-A-protein levels, as well as miRNA-expression levels of miR-146a, miR-221 and miR-222. SMCs treated with TAA-TC-exosomes developed a dedifferentiation-phenotype. In conclusion, the study shows for the first time that TCs are involved in development of TAA and could play a crucial role in SMC phenotype switching by release of extracellular vesicles.

Hemolytic Footprint of Rotodynamic Blood Pumps.

OBJECTIVE: In preclinical examinations, rotodynamic blood pumps (RBPs) are predominantly evaluated at design-point conditions. In clinical practice, however, they run at diversified modes of operation. This study aimed at extending current preclinical evaluation of hemolytic profiles in RBPs toward broader, clinically relevant ranges of operation. METHODS: Two implantable RBPs - the HeartMate 3 (HM3) and the HeartWare Ventricular Assist Device (HVAD) - were analyzed at three pump speeds (HM3: 4300, 5600, 7000 rpm; HVAD: 1800, 2760, 3600 rpm) with three flow rates (1-9L/min) per speed setting. Hemolysis measurements were performed in heparinized bovine blood. The delta free hemoglobin (dfHb) and the normalized index of hemolysis (NIH) served as hemolytic measures. Statistical analysis was performed by multiple comparison of the 9 operating conditions. Moreover, computational fluid dynamics (CFD) was applied to provide mechanistic insights into the interrelation between hydraulics and hemolysis by correlating numerically computed hydraulic losses with in-vitro hemolytic measures. RESULTS: In both devices, dfHb increased toward increasing speeds, particularly during low but also during high flow condition. By contrast, in both RBPs magnitudes of NIH were significantly elevated during low flow operation compared to high flow conditions (p<0.0036). Maps of hemolytic metrics revealed morphologically similar trends to in-silico hydraulic losses (r>0.793). CONCLUSIONS: While off-design operation is associated with increased hemolytic profiles, the setting of different operating conditions render a preclinical prediction of clinical impact with current hemolysis metrics difficult. SIGNIFICANCE: The identified increase in hemolytic measures during episodes of off-design operation is highlighting the need to consider worst-case operation during preclinical examinations.

A Novel Endothelial Damage Inhibitor Reduces Oxidative Stress and Improves Cellular Integrity in Radial Artery Grafts for Coronary Artery Bypass.

The radial artery (RA) is a frequently used conduit in coronary artery bypass grafting (CABG). Endothelial injury incurred during graft harvesting promotes oxidative damage, which leads to graft disease and graft failure. We evaluated the protective effect of DuraGraft®, an endothelial damage inhibitor (EDI), on RA grafts. We further compared the protective effect of the EDI between RA grafts and saphenous vein grafts (SVG). Samples of RA (n = 10) and SVG (n = 13) from 23 patients undergoing CABG were flushed and preserved with either EDI or heparinized Ringer's lactate solution (RL). The effect of EDI vs. RL on endothelial damage was evaluated ex vivo and in vitro using histological analysis, immunofluorescence staining, Western blot, and scanning electron microscopy. EDI-treated RA grafts showed a significant reduction of endothelial and sub-endothelial damage. Lower level of reactive oxygen species (ROS) after EDI treatment was correlated with a reduction of hypoxic damage (eNOS and Caveolin-1) and significant increase of oxidation-reduction potential. Additionally, an increased expression of TGFß, PDGFα/ß, and HO-1 which are indicative for vascular protective function were observed after EDI exposure. EDI treatment preserves functionality and integrity of endothelial and intimal cells. Therefore, EDI may have the potential to reduce the occurrence of graft disease and failure in RA grafts in patients undergoing CABG.

Telocytes in the human ascending aorta: Characterization and exosome-related KLF-4/VEGF-A expression.

Telocytes (TCs), a novel interstitial cell entity promoting tissue regeneration, have been described in various tissues. Their role in inter-cellular signalling and tissue remodelling has been reported in almost all human tissues. This study hypothesizes that TC also contributes to tissue remodelling and regeneration of the human thoracic aorta (HTA). The understanding of tissue homeostasis and regenerative potential of the HTA is of high clinical interest as it plays a crucial role in pathogenesis from aortic dilatation to lethal dissection. Therefore, we obtained twenty-five aortic specimens of heart donors during transplantation. The presence of TCs was detected in different layers of aortic tissue and characterized by immunofluorescence and transmission electron microscopy. Further, we cultivated and isolated TCs in highly differentiated form identified by positive staining for CD34 and c-kit. Aortic-derived TC was characterized by the expression of PDGFR-α, PDGFR-ß, CD29/integrin ß-1 and αSMA and the stem cell markers Nanog and KLF-4. Moreover, TC exosomes were isolated and characterized for soluble angiogenic factors by Western blot. CD34+ /c-kit+ TCs shed exosomes containing the soluble factors VEGF-A, KLF-4 and PDGF-A. In summary, TC occurs in the aortic wall. Correspondingly, exosomes, derived from aortic TCs, contain vasculogenesis-relevant proteins. Understanding the regulation of TC-mediated aortic remodelling may be a crucial step towards designing strategies to promote aortic repair and prevent adverse remodelling.

S-nitroso human serum albumin as a nitric oxide donor in drug-eluting vascular grafts: Biofunctionality and preclinical evaluation.

Currently available synthetic small diameter vascular grafts reveal low patency rates due to thrombosis and intimal hyperplasia. Biofunctionalized grafts releasing nitric oxide (NO) in situ may overcome these limitations. In this study, a drug-eluting vascular graft was designed by blending polycaprolactone (PCL) with S-nitroso-human-serum-albumin (S-NO-HSA), a nitric oxide donor with prolonged half-life. PCL-S-NO-HSA grafts and patches were fabricated via electrospinning. The fabrication process was optimized. Patches were characterized in vitro for their morphology, drug release, biomechanics, inflammatory effects, cell proliferation, and expression of adhesion molecules. The selected optimized formulation (8%PCL-S-NO-HSA) had superior mechanical/morphological properties with high protein content revealing extended NO release (for 28 days). 8%PCL-S-NO-HSA patches significantly promoted endothelial cell proliferation while limiting smooth muscle cell proliferation. Expression of adhesion molecules (ICAM-1, VCAM-1) and pro-inflammatory macrophage/cytokine markers (CD80, IL-1α, TNF-α) was significantly reduced. 8%PCL-S-NO-HSA patches had superior immunomodulatory properties by up-regulating anti-inflammatory cytokines (IL-10) and M2 macrophage marker (CD163) at final time points. Grafts were further evaluated in a small rodent model as aortic implants up to 12 weeks. Grafts were assessed by magnetic resonance imaging angiography (MRI) in vivo and after retrieval by histology. All grafts remained 100 % patent with no signs of thrombosis or calcification. 8%PCL-S-NO-HSA vascular grafts supported rapid endothelialization, whereas smooth muscle cell proliferation was hampered in earlier phases. This study indicates that 8%PCL-S-NO-HSA grafts effectively support long-term in situ release of bioactive NO. The beneficial effects observed can be promising features for long-term success of small diameter vascular grafts. STATEMENT OF SIGNIFICANCE: Despite extensive research in the field of small diameter vascular graft replacement, there is still no appropriate substitute to autografts yet. Various limitations are associated with currently available synthetic vascular grafts such as thrombogenicity and intimal hyperplasia. Therefore, developing new generations of such conduits has become a major focus of research. One of the most significant signaling molecules that are involved in homeostasis of the vascular system is nitric oxide. The new designed nitric-oxide eluting vascular grafts described in this study induce rapid surface endothelialization and late migration of SMCs into the graft wall. These beneficial effects have potential to improve current limitations of small diameter vascular grafts.

Long interspersed element-1 ribonucleoprotein particles protect telomeric ends in alternative lengthening of telomeres dependent cells.

Malignant cells ensure telomere maintenance by the alternative lengthening of telomeres (ALT) in the absence of telomerase activity (TA). The retrotransposons "long interspersed nuclear element-1" (LINE-1, L1) are expressed in malignant cells and are primarily known to contribute to complex karyotypes. Here we demonstrate that LINE-1 ribonucleoprotein particles (L1-RNPs) expression is significantly higher in ALT+- versus in TA+-human glioma. Analyzing a role of L1-RNP in ALT, we show that L1-RNPs bind to telomeric repeat containing RNA (TERRA), which is critical for telomere stabilization and which is overexpressed in ALT+ cells. In turn, L1-RNP knockdown (KD) abrogated the nuclear retention of TERRA, resulted in increased telomeric DNA damage, decreased cell growth and reduced expression of ALT characteristics such as c-circles and PML-bodies. L1-RNP KD also decreased the expression of Shelterin- and the ALT-regulating protein Topoisomerase IIIα (TopoIIIα) indicating a more general role of L1-RNPs in supporting telomeric integrity in ALT. Our findings suggest an impact of L1-RNP on telomere stability in ALT+ dependent tumor cells. As L1-RNPs are rarely expressed in normal adult human tissue those elements might serve as a novel target for tumor ablative therapy.

Long Term Evaluation of Nanofibrous, Bioabsorbable Polycarbonate Urethane Grafts for Small Diameter Vessel Replacement in Rodents.

OBJECTIVE: Biodegradable materials for in situ vascular tissue engineering could meet the increasing clinical demand for sufficient synthetic small diameter vascular substitutes in aortocoronary bypass and peripheral vascular surgery. The aim of this study was to design a new degradable thermoplastic polycarbonate urethane (dPCU) with improved biocompatibility and optimal biomechanical properties. Electrospun conduits made from dPCU were evaluated in short and long term follow up and compared with expanded polytetrafluoroethylene (ePTFE) controls. METHODS: Both conduits were investigated prior to implantation to assess their biocompatibility and inflammatory potential via real time polymerase chain reaction using a macrophage culture. dPCU grafts (n = 28) and ePTFE controls (n = 28) were then implanted into the infrarenal abdominal aorta of Sprague-Dawley rats. After seven days, one, six, and 12 months, grafts were analysed by histology and immunohistochemistry (IHC) and assessed biomechanically. RESULTS: Anti-inflammatory signalling was upregulated in dPCU conduits and increased significantly over time in vitro. dPCU and ePTFE grafts offered excellent long and short term patency rates (92.9% in both groups at 12 months) in the rat model without dilatation or aneurysm formation. In comparison to ePTFE, dPCU grafts showed transmural ingrowth of vascular specific cells resulting in a structured neovessel formation around the graft. The graft material was slowly reduced, while the compliance of the neovessel increased over time. CONCLUSION: The newly designed dPCU grafts have the potential to be safely applied for in situ vascular tissue engineering applications. The degradable substitutes showed good in vivo performance and revealed desirable characteristics such as biomechanical stability, non-thrombogenicity, and minimal inflammatory response after long term implantation.

Chemical imaging and assessment of cadmium distribution in the human body.

The scientific interest in cadmium (Cd) as a human health damaging agent has significantly increased over the past decades. However, particularly the histological distribution of Cd in human tissues is still scarcely defined. Using inductively coupled plasma-mass spectrometry (ICP-MS), we determined the concentration of Cd in 40 different human tissues of four body donors and provided spatial information by elemental imaging on the microscopic distribution of Cd in 8 selected tissues by laser ablation (LA)-ICP-MS. ICP-MS results show that Cd concentrations differ by a factor of 20 000 between different tissues. Apart from the well know deposits in kidney, bone, and liver, our study provides evidence that muscle and adipose tissue are underestimated Cd pools. For the first time, we present spatially resolved Cd distributions in a broad panel of human soft tissues. The defined histological structures are mirrored by sharp cut differences in Cd concentrations between neighboring tissue types, particularly in the rectum, testis, and kidneys. The spatial resolution of the Cd distribution at microscopic level visualized intratissue hot spots of Cd accumulation and is suggested as a powerful tool to elucidate metal based toxicity at histological level.

Strong Signs for a Weak Wall in Tricuspid Aortic Valve Associated Aneurysms and a Role for Osteopontin in Bicuspid Aortic Valve Associated Aneurysms.

Central processes in the pathogenesis of TAV- (tricuspid aortic valve) and BAV- (bicuspid aortic valve) associated ascending thoracic aortic aneurysm (ATAA) development are still unknown. To gain new insights, we have collected aortic tissue and isolated smooth muscle cells of aneurysmal tissue and subjected them to in situ and in vitro analyses. We analyzed aortic tissue from 78 patients (31 controls, 28 TAV-ATAAs, and 19 BAV-ATAAs) and established 30 primary smooth muscle cell cultures. Analyses included histochemistry, immuno-, auto-fluorescence-based image analyses, and cellular analyses including smooth muscle cell contraction studies. With regard to TAV associated aneurysms, we observed a strong impairment of the vascular wall, which appears on different levels-structure and dimension of the layers (reduced media thickness, increased intima thickness, atherosclerotic changes, degeneration of aortic media, decrease of collagen, and increase of elastic fiber free area) as well as on the cellular level (accumulation of fibroblasts/myofibroblasts, and increase in the number of smooth muscle cells with a reduced alpha smooth muscle actin (α-SM actin) content per cell). The pathological changes in the aortic wall of BAV patients were much less pronounced-apart from an increased expression of osteopontin (OPN) in the vascular wall which stem from smooth muscle cells, we observed a trend towards increased calcification of the aortic wall (increase significantly associated with age). These observations provide strong evidence for different pathological processes and different disease mechanisms to occur in BAV- and TAV-associated aneurysms.

Primary Human Fibroblasts in Culture Switch to a Myofibroblast-Like Phenotype Independently of TGF Beta.

Fibroblasts are the prevalent cell type and main source for extracellular matrix (ECM) in connective tissue. Depending on their origin, fibroblasts play a central role in non-pathological tissue remodeling and disease like fibrosis. This study examined the effect of established culture conditions of primary human fibroblasts, from different origins on the myofibroblast-like phenotype formation. We isolated primary human fibroblasts from aortic adventitia, lung, juvenile- and adult skin and investigated the expression levels of CD90, alpha smooth muscle actin (αSMA) and procollagen I under different concentrations of fetal calf serum (FCS) and ascorbic acid (AA) in culture media by immunoblot and immunofluorescence assays. Furthermore, we determined the viability using XTT and migration/wound healing in scratch assays. Collagen 1 secretion was quantified by specific ELISA. Primary human fibroblasts show in part a myofibroblast-like phenotype even without addition of FCS. Supplemented AA reduces migration of cultured fibroblasts with no or low concentrations of FCS. Furthermore, AA and higher concentrations of FCS in culture media lead to higher levels of collagen 1 secretion instead of procollagen I accumulation. This study provides evidence for a partial switch of primary human fibroblasts of different origin to a myofibroblast-like phenotype under common culture conditions.

Bicuspid aortic valve-associated aortopathy: Where do we stand?

Herein we summarize the current knowledge on the bicuspid aortic valve (BAV)-associated aortopathy regarding clinical presentation and disease sub-classification, genetic background, hemodynamics, histopathology, cells and signaling, animal models, and biomarkers. Despite enormous efforts in research in all of the above areas, important issues remain unknown: (i) what is the ontogenetic basis of BAV development? (ii) how can we explain the diversity of BAV and associated aortopathy phenotypes? (iii) what are the signaling processes in aortopathy pathogenesis and how can we interfere with these processes? Despite undoubtedly great progress that has been made in the understanding of BAV-associated aortopathy, so far researchers have put together a heap of Lego bricks, but at present it is unclear if the bricks are compatible, how they fit together, and which parts are missing to build the true model of the BAV aorta. A joint approach is needed to accelerate research progress.