Mesenchymal stem cells (MSCs) show unexplained differences in differentiation potential. In this study, differentiation of human (h) MSCs derived from embryonic, fetal and adult sources toward ...cardiomyocytes, endothelial and smooth muscle cells was investigated. Labeled hMSCs derived from embryonic stem cells (hESC-MSCs), fetal umbilical cord, bone marrow, amniotic membrane and adult bone marrow and adipose tissue were co-cultured with neonatal rat cardiomyocytes (nrCMCs) or cardiac fibroblasts (nrCFBs) for 10 days, and also cultured under angiogenic conditions. Cardiomyogenesis was assessed by human-specific immunocytological analysis, whole-cell current-clamp recordings, human-specific qRT-PCR and optical mapping. After co-culture with nrCMCs, significantly more hESC-MSCs than fetal hMSCs stained positive for α-actinin, whereas adult hMSCs stained negative. Furthermore, functional cardiomyogenic differentiation, based on action potential recordings, was shown to occur, but not in adult hMSCs. Of all sources, hESC-MSCs expressed most cardiac-specific genes. hESC-MSCs and fetal hMSCs contained significantly higher basal levels of connexin43 than adult hMSCs and co-culture with nrCMCs increased expression. After co-culture with nrCFBs, hESC-MSCs and fetal hMSCs did not express α-actinin and connexin43 expression was decreased. Conduction velocity (CV) in co-cultures of nrCMCs and hESC-MSCs was significantly higher than in co-cultures with fetal or adult hMSCs. In angiogenesis bioassays, only hESC-MSCs and fetal hMSCs were able to form capillary-like structures, which stained for smooth muscle and endothelial cell markers.Human embryonic and fetal MSCs differentiate toward three different cardiac lineages, in contrast to adult MSCs. Cardiomyogenesis is determined by stimuli from the cellular microenvironment, where connexin43 may play an important role.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Department of Cardiology, Leiden University Medical Center, Leiden, The Netherlands
Submitted 7 April 2006
; accepted in final form 22 May 2006
We characterized hemodynamics and systolic and ...diastolic right ventricular (RV) function in relation to structural changes in the rat model of monocrotaline (MCT)-induced pulmonary hypertension. Rats were treated with MCT at 30 mg/kg body wt (MCT30, n = 15) and 80 mg/kg body wt (MCT80, n = 16) to induce compensated RV hypertrophy and RV failure, respectively. Saline-treated rats served as control (Cont, n = 13). After 4 wk, a pressure-conductance catheter was introduced into the RV to assess pressure-volume relations. Subsequently, rats were killed, hearts and lungs were rapidly dissected, and RV, left ventricle (LV), and interventricular septum (IVS) were weighed and analyzed histochemically. RV-to-(LV + IVS) weight ratio was 0.29 ± 0.05 in Cont, 0.35 ± 0.05 in MCT30, and 0.49 ± 0.10 in MCT80 ( P < 0.001 vs. Cont and MCT30) rats, confirming MCT-induced RV hypertrophy. RV ejection fraction was 49 ± 6% in Cont, 40 ± 12% in MCT30 ( P < 0.05 vs. Cont), and 26 ± 6% in MCT80 ( P < 0.05 vs. Cont and MCT30) rats. In MCT30 rats, cardiac output was maintained, but RV volumes and filling pressures were significantly increased compared with Cont (all P < 0.05), indicating RV remodeling. In MCT80 rats, RV systolic pressure, volumes, and peak wall stress were further increased, and cardiac output was significantly decreased (all P < 0.05). However, RV end-systolic and end-diastolic stiffness were unchanged, consistent with the absence of interstitial fibrosis. MCT-induced pressure overload was associated with a dose-dependent development of RV hypertrophy. The most pronounced response to MCT was an overload-dependent increase of RV end-systolic and end-diastolic volumes, even under nonfailing conditions.
right ventricular hypertrophy; right ventricular failure; pressure-volume relations
Address for reprint requests and other correspondence: P. Steendijk, Dept. of Cardiology, C5-P, Leiden Univ. Medical Center, PO Box 9600, 2300 RC Leiden, The Netherlands (e-mail: p.steendijk{at}lumc.nl )
Human mesenchymal stromal cells (MSCs) have been reported to preserve cardiac function in myocardial infarction (MI) models. Previously, we found a beneficial effect of intramyocardial injection of ...unstimulated human MSCs (uMSCs) on cardiac function after permanent coronary artery ligation. In the present study we aimed to extend this research by investigating the effect of intramyocardial injection of human MSCs pre-stimulated with the pro-inflammatory cytokine interferon-gamma (iMSCs), since pro-inflammatory priming has shown additional salutary effects in multiple experimental disease models.
MI was induced in NOD/Scid mice by permanent ligation of the left anterior descending coronary artery. Animals received intramyocardial injection of uMSCs, iMSCs or PBS. Sham-operated animals were used to determine baseline characteristics. Cardiac performance was assessed after 2 and 14 days using 7-Tesla magnetic resonance imaging and pressure-volume loop measurements. Histology and q-PCR were used to confirm MSC engraftment in the heart.
Both uMSC and iMSC therapy had no significant beneficial effect on cardiac function or remodelling in contrast to our previous studies.
Animal models for cardiac MSC therapy appear less robust than initially envisioned.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
1 Departments of Cardiology and
2 Pediatrics, Leiden University Medical Center, Leiden, The Netherlands
Submitted July 1, 2009
; accepted in final form September 23, 2009
Pulmonary arterial ...hypertension (PAH) is a chronic lung disease that leads to right ventricular (RV) hypertrophy (RVH), remodeling, and failure. We tested treatment with bone marrow-derived mesenchymal stem cells (MSCs) obtained from donor rats with monocrotaline (MCT)-induced PAH to recipient rats with MCT-induced PAH on pulmonary artery pressure, lung pathology, and RV function. This model was chosen to mimic autologous MSC therapy. On day 1 , PAH was induced by MCT (60 mg/kg) in 20 female Wistar rats. On day 14 , rats were treated with 10 6 MSCs intravenously (MCT + MSC) or with saline (MCT60). MSCs were obtained from donor rats with PAH at 28 days after MCT. A control group received saline on days 1 and 14 . On day 28 , the RV function of recipient rats was assessed, followed by isolation of the lungs and heart. RVH was quantified by the weight ratio of the RV/(left ventricle + interventricular septum). MCT induced an increase of RV peak pressure (from 27 ± 5 to 42 ± 17 mmHg) and RVH (from 0.25 ± 0.04 to 0.47 ± 0.12), depressed the RV ejection fraction (from 56 ± 11 to 43 ± 6%), and increased lung weight (from 0.96 ± 0.15 to 1.66 ± 0.32 g), including thickening of the arteriolar walls and alveolar septa. MSC treatment attenuated PAH (31 ± 4 mmHg) and RVH (0.32 ± 0.07), normalized the RV ejection fraction (52 ± 5%), reduced lung weight (1.16 ± 0.24 g), and inhibited the thickening of the arterioles and alveolar septa. We conclude that the application of MSCs from donor rats with PAH reduces RV pressure overload, RV dysfunction, and lung pathology in recipient rats with PAH. These results suggest that autologous MSC therapy may alleviate cardiac and pulmonary symptoms in PAH patients.
pulmonary arterial hypertension; heart failure; hypertrophy; remodeling; monocrotaline
Address for reprint requests and other correspondence: A. van der Laarse, Dept. of Cardiology, Leiden Univ. Medical Center, PO Box 9600, Leiden 2300 RC, The Netherlands (e-mail: a.van_der_laarse{at}lumc.nl ).
Cardiac hypertrophy and fibrosis are associated with potentially lethal arrhythmias. As these substrates often occur simultaneously in one patient, distinguishing between pro-arrhythmic mechanisms is ...difficult. This hampers understanding of underlying pro-arrhythmic mechanisms and optimal treatment. This study investigates and compares arrhythmogeneity and underlying pro-arrhythmic mechanisms of either cardiac hypertrophy or fibrosis in in vitro models.
Fibrosis was mimicked by free myofibroblast (MFB) proliferation in neonatal rat ventricular monolayers. Cultures with inhibited MFB proliferation were used as control or exposed to phenylephrine to induce hypertrophy. At Day 9, cultures were studied with patch-clamp and optical-mapping techniques and assessed for protein expression. In hypertrophic (n = 111) and fibrotic cultures (n = 107), conduction and repolarization were slowed. Triggered activity was commonly found in these substrates and led to high incidences of spontaneous re-entrant arrhythmias 67.5% hypertrophic, 78.5% fibrotic vs. 2.9% in controls (n = 102) or focal arrhythmias (39.1, 51.7 vs. 8.8%, respectively). Kv4.3 and Cx43 protein expression levels were decreased in hypertrophy but unaffected in fibrosis. Depolarization of cardiomyocytes (CMCs) was only found in fibrotic cultures (-48 ± 7 vs. -66 ± 7 mV in control, P < 0.001). L-type calcium-channel blockade prevented arrhythmias in hypertrophy, but caused conduction block in fibrosis. Targeting heterocellular coupling by low doses of gap-junction uncouplers prevented arrhythmias by accelerating repolarization only in fibrotic cultures.
Cultured hypertrophic or fibrotic myocardial tissues generated similar focal and re-entrant arrhythmias. These models revealed electrical remodelling of CMCs as a pro-arrhythmic mechanism of hypertrophy and MFB-induced depolarization of CMCs as a pro-arrhythmic mechanism of fibrosis. These findings provide novel mechanistic insight into substrate-specific arrhythmicity.
IFN regulatory factor 6 (IRF6) is a transcription factor that, in mammals, is required for the differentiation of skin, breast epithelium, and oral epithelium. However, the transcriptional targets ...that mediate these effects are currently unknown. In zebrafish and frog embryos, Irf6 is necessary for differentiation of the embryonic superficial epithelium, or periderm. Here we use microarrays to identify genes that are expressed in the zebrafish periderm and whose expression is inhibited by a dominant-negative variant of Irf6 (dnIrf6). These methods identify Grainyhead-like 3 (Grhl3), an ancient regulator of the epidermal permeability barrier, as acting downstream of Irf6. In human keratinocytes, IRF6 binds conserved elements near the GRHL3 promoter. We show that one of these elements has enhancer activity in human keratinocytes and zebrafish periderm, suggesting that Irf6 directly stimulates Grhl3 expression in these tissues. Simultaneous inhibition of grhl1 and grhl3 disrupts periderm differentiation in zebrafish, and, intriguingly, forced grhl3 expression restores periderm markers in both zebrafish injected with dnIrf6 and frog embryos depleted of Irf6. Finally, in Irf6-deficient mouse embryos, Grhl3 expression in the periderm and oral epithelium is virtually absent. These results indicate that Grhl3 is a key effector of Irf6 in periderm differentiation.
Maternal self-efficacy for breast-feeding may contribute to success in breast-feeding. This study aimed to increase breast-feeding self-efficacy and actual breast-feeding through an intervention ...based on Bandura's self-efficacy theory. A total of 90 pregnant women participated in the study. The women who were assigned to a breast-feeding self-efficacy intervention showed significantly greater increases in breast-feeding self-efficacy than did the women in the control group. Furthermore, at 4 weeks postpartum, women in the intervention group showed a trend toward breast-feeding their infants longer and more exclusively than did those in the control group. Greater increases in breast-feeding self-efficacy were associated with a significantly higher level of breast-feeding. Replicating previous research, breast-feeding self-efficacy was significantly related to concurrent breast-feeding behavior, and high antenatal breast-feeding self-efficacy predicted a higher level of later breast-feeding in control-group women. These findings have implications for breast-feeding support programs and for the potential general utility of self-efficacy-based interventions in health education.
The advent of highly sensitive photodetectors and the development of photostabilization strategies made detecting the fluorescence of single molecules a routine task in many labs around the world. ...However, to this day, this process requires cost-intensive optical instruments due to the truly nanoscopic signal of a single emitter. Simplifying single-molecule detection would enable many exciting applications, e.g., in point-of-care diagnostic settings, where costly equipment would be prohibitive. Here, we introduce addressable NanoAntennas with Cleared HOtSpots (NACHOS) that are scaffolded by DNA origami nanostructures and can be specifically tailored for the incorporation of bioassays. Single emitters placed in NACHOS emit up to 461-fold (average of 89 ± 7-fold) brighter enabling their detection with a customary smartphone camera and an 8-US-dollar objective lens. To prove the applicability of our system, we built a portable, battery-powered smartphone microscope and successfully carried out an exemplary single-molecule detection assay for DNA specific to antibiotic-resistant Klebsiella pneumonia on the road.
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