Publications by authors named "Moritz Schnelle"

15 Publications

  • Page 1 of 1

In vivo [U-C]glucose labeling to assess heart metabolism in murine models of pressure and volume overload.

Am J Physiol Heart Circ Physiol 2020 08 10;319(2):H422-H431. Epub 2020 Jul 10.

King's College London British Heart Foundation Centre of Excellence, School of Cardiovascular Medicine & Sciences, London, United Kingdom.

Alterations in the metabolism of substrates such as glucose are integrally linked to the structural and functional changes that occur in the remodeling heart. Assessment of such metabolic changes under in vivo conditions would provide important insights into this interrelationship. We aimed to investigate glucose carbon metabolism in pressure-overload and volume-overload cardiac hypertrophy by using an in vivo [U-C]glucose labeling strategy to enable analyses of the metabolic fates of glucose carbons in the mouse heart. Therefore, [U-C]glucose was administered in anesthetized mice by tail vein infusion, and the optimal duration of infusion was established. Hearts were then excised for C metabolite isotopomer analysis by NMR spectroscopy. [U-C]glucose infusions were performed in mice 2 wk following transverse aortic constriction (TAC) or aortocaval fistula (Shunt) surgery. At this time point, there were similar increases in left ventricular (LV) mass in both groups, but TAC resulted in concentric hypertrophy with impaired LV function, whereas Shunt caused eccentric hypertrophy with preserved LV function. TAC was accompanied by significant changes in glycolysis, mitochondrial oxidative metabolism, glucose metabolism to anaplerotic substrates, and de novo glutamine synthesis. In contrast to TAC, hardly any metabolic changes could be observed in the Shunt group. Taken together, in vivo [U-C]glucose labeling is a valuable method to investigate the fate of nutrients such as glucose in the remodeling heart. We find that concentric and eccentric cardiac remodeling are accompanied by distinct differences in glucose carbon metabolism. This study implemented a method for assessing the fate of glucose carbons in the heart in vivo and used this to demonstrate that pressure and volume overload are associated with distinct changes. In contrast to volume overload, pressure overload-induced changes affect the tricarboxylic acid cycle, glycolytic pathways, and glutamine synthesis. A better understanding of cardiac glucose metabolism under pathological conditions in vivo may provide new therapeutic strategies specific for different types of hemodynamic overload.
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http://dx.doi.org/10.1152/ajpheart.00219.2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7473922PMC
August 2020

NADPH oxidase-4 promotes eccentric cardiac hypertrophy in response to volume overload.

Cardiovasc Res 2021 Jan;117(1):178-187

King's College London British Heart Foundation Centre of Excellence, School of Cardiovascular Medicine & Sciences, The James Black Centre, 125 Coldharbour Lane, London SE5 9NU, UK.

Aims: Chronic pressure or volume overload induce concentric vs. eccentric left ventricular (LV) remodelling, respectively. Previous studies suggest that distinct signalling pathways are involved in these responses. NADPH oxidase-4 (Nox4) is a reactive oxygen species-generating enzyme that can limit detrimental cardiac remodelling in response to pressure overload. This study aimed to assess its role in volume overload-induced remodelling.

Methods And Results: We compared the responses to creation of an aortocaval fistula (Shunt) to induce volume overload in Nox4-null mice (Nox4-/-) vs. wild-type (WT) littermates. Induction of Shunt resulted in a significant increase in cardiac Nox4 mRNA and protein levels in WT mice as compared to Sham controls. Nox4-/- mice developed less eccentric LV remodelling than WT mice (echocardiographic relative wall thickness: 0.30 vs. 0.27, P < 0.05), with less LV hypertrophy at organ level (increase in LV weight/tibia length ratio of 25% vs. 43%, P < 0.01) and cellular level (cardiomyocyte cross-sectional area: 323 µm2 vs. 379 μm2, P < 0.01). LV ejection fraction, foetal gene expression, interstitial fibrosis, myocardial capillary density, and levels of myocyte apoptosis after Shunt were similar in the two genotypes. Myocardial phospho-Akt levels were increased after induction of Shunt in WT mice, whereas levels decreased in Nox4-/- mice (+29% vs. -21%, P < 0.05), associated with a higher level of phosphorylation of the S6 ribosomal protein (S6) and the eIF4E-binding protein 1 (4E-BP1) in WT compared to Nox4-/- mice. We identified that Akt activation in cardiac cells is augmented by Nox4 via a Src kinase-dependent inactivation of protein phosphatase 2A.

Conclusion: Endogenous Nox4 is required for the full development of eccentric cardiac hypertrophy and remodelling during chronic volume overload. Nox4-dependent activation of Akt and its downstream targets S6 and 4E-BP1 may be involved in this effect.
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http://dx.doi.org/10.1093/cvr/cvz331DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7797217PMC
January 2021

Assessing the role of extracellular signal-regulated kinases 1 and 2 in volume overload-induced cardiac remodelling.

ESC Heart Fail 2019 10 19;6(5):1015-1026. Epub 2019 Jul 19.

Department of Cardiology and Pneumology, University Medical Center Göttingen, Göttingen, Germany.

Aims: Volume overload (VO) and pressure overload (PO) induce differential cardiac remodelling responses including distinct signalling pathways. Extracellular signal-regulated kinases 1 and 2 (ERK1/2), key signalling components in the mitogen-activated protein kinase (MAPK) pathways, modulate cardiac remodelling during pressure overload (PO). This study aimed to assess their role in VO-induced cardiac remodelling as this was unknown.

Methods And Results: Aortocaval fistula (Shunt) surgery was performed in mice to induce cardiac VO. Two weeks of Shunt caused a significant reduction of cardiac ERK1/2 activation in wild type (WT) mice as indicated by decreased phosphorylation of the TEY (Thr-Glu-Tyr) motif (-28% as compared with Sham controls, P < 0.05). Phosphorylation of other MAPKs was unaffected. For further assessment, transgenic mice with cardiomyocyte-specific ERK2 overexpression (ERK2tg) were studied. At baseline, cardiac ERK1/2 phosphorylation in ERK2tg mice remained unchanged compared with WT littermates, and no overt cardiac phenotype was observed; however, cardiac expression of the atrial natriuretic peptide was increased on messenger RNA (3.6-fold, P < 0.05) and protein level (3.1-fold, P < 0.05). Following Shunt, left ventricular dilation and hypertrophy were similar in ERK2tg mice and WT littermates. Left ventricular function was maintained, and changes in gene expression indicated reactivation of the foetal gene program in both genotypes. No differences in cardiac fibrosis and kinase activation was found amongst all experimental groups, whereas apoptosis was similarly increased through Shunt in ERK2tg and WT mice.

Conclusions: VO-induced eccentric hypertrophy is associated with reduced cardiac ERK1/2 activation in vivo. Cardiomyocyte-specific overexpression of ERK2, however, does not alter cardiac remodelling during VO. Future studies need to define the pathophysiological relevance of decreased ERK1/2 signalling during VO.
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http://dx.doi.org/10.1002/ehf2.12497DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6816056PMC
October 2019

Genetic deletion of calcium/calmodulin-dependent protein kinase type II delta does not mitigate adverse myocardial remodeling in volume-overloaded hearts.

Sci Rep 2019 07 8;9(1):9889. Epub 2019 Jul 8.

Department of Cardiology and Pneumology, Georg-August-University, Göttingen, Germany.

Calcium/calmodulin-dependent protein kinase type II delta (CaMKIIδ), the predominant CaMKII isoform expressed in the heart, has been implicated in the progression of myocardial infarction- and pressure overload-induced pathological remodeling. However, the role of CaMKIIδ in volume overload (VO) has not been explored. We have previously reported an activation of CaMKII during transition to HF in long-term VO. Here, we address whether CaMKIIδ is critically involved in the mortality, myocardial remodeling, and heart failure (HF) progression in response to VO. CaMKIIδ knockout (δ-KO) and wild-type (WT) littermates were exposed to aortocaval shunt-induced VO, and the progression of adverse myocardial remodeling was assessed by serial echocardiography, histological and molecular analyses. The mortality rates during 10 weeks of VO were similar in δ-KO and WT mice. Both genotypes displayed comparable eccentric myocardial hypertrophy, altered left ventricle geometry, perturbed systolic and diastolic functions after shunt. Additionally, cardiomyocytes hypertrophy, augmented myocyte apoptosis, and up-regulation of hypertrophic genes were also not significantly different in δ-KO versus WT hearts after shunt. Therefore, CaMKIIδ signaling seems to be dispensable for the progression of VO-induced maladaptive cardiac remodeling. Accordingly, we hypothesize that CaMKIIδ-inhibition as a therapeutic approach might not be helpful in the context of VO-triggered HF.
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http://dx.doi.org/10.1038/s41598-019-46332-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6614357PMC
July 2019

Differences in procalcitonin measurements between three BRAHMS-partnered immunoassays (Liaison, Elecsys and Architect).

Clin Chem Lab Med 2019 08;57(9):e207-e210

Institute for Clinical Chemistry, University Medical Center Goettingen, Goettingen, Germany.

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http://dx.doi.org/10.1515/cclm-2018-0916DOI Listing
August 2019

Response by Sag et al to Letter Regarding Article, "Distinct Regulatory Effects of Myeloid Cell and Endothelial Cell NAPDH Oxidase 2 on Blood Pressure".

Circulation 2017 11;136(21):2090-2091

King's College London, British Heart Foundation Centre of Excellence, Cardiovascular Division, UK (C.M.S., M.S., A.M.S.).

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http://dx.doi.org/10.1161/CIRCULATIONAHA.117.030515DOI Listing
November 2017

Echocardiographic evaluation of diastolic function in mouse models of heart disease.

J Mol Cell Cardiol 2018 01 19;114:20-28. Epub 2017 Oct 19.

King's College London British Heart Foundation Centre of Excellence, Cardiovascular Division, London, United Kingdom. Electronic address:

Background: Mouse models of heart disease are extensively employed. The echocardiographic characterization of contractile function is usually focused on systolic function with fewer studies assessing diastolic function. Furthermore, the applicability of diverse echocardiographic parameters of diastolic function that are commonly used in humans has not been extensively evaluated in different pathophysiological models in mice.

Methods And Results: We used high resolution echocardiography to evaluate parameters of diastolic function in mouse models of chronic pressure overload (aortic constriction), volume overload (aorto-caval shunt), heart failure with preserved ejection fraction (HFpEF; DOCA-salt hypertension), and acute sarcoplasmic reticulum dysfunction induced by thapsigargin - all known to exhibit diastolic dysfunction. Left atrial area increased in all three chronic models while mitral E/A was difficult to quantify at high heart rates. Isovolumic relaxation time (IVRT) and Doppler E/E' increased significantly and the peak longitudinal strain rate during early filling (peak reverse longitudinal strain rate) decreased significantly after aortic constriction, with the changes being proportional to the magnitude of hypertrophy. In the HFpEF model, reverse longitudinal strain rate decreased significantly but changes in IVRT and E/E' were non-significant, consistent with less severe dysfunction. With volume overload, there was a significant increase in reverse longitudinal strain rate and decrease in IVRT, indicating a restrictive physiology. Acute thapsigargin treatment caused significant prolongation of IVRT and decrease in reverse longitudinal strain rate.

Conclusion: These results indicate that the combined measurement of left atrial area plus reverse longitudinal strain rate and/or IVRT provide an excellent overall assessment of diastolic function in the diseased mouse heart, allowing distinction between different types of pathophysiology.
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http://dx.doi.org/10.1016/j.yjmcc.2017.10.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5807035PMC
January 2018

A Novel α-Calcitonin Gene-Related Peptide Analogue Protects Against End-Organ Damage in Experimental Hypertension, Cardiac Hypertrophy, and Heart Failure.

Circulation 2017 Jul 26;136(4):367-383. Epub 2017 Apr 26.

From Cardiovascular Division, BHF Centre of Research Excellence and Centre of Integrative Biomedicine, King's College London, United Kingdom (A.A.A., F.A., S.-J.S., S.S., K.M.A., E.W., J.M., K.F.-D., G.M., R.C.S., S.D.B.); Institute of Pharmaceutical Sciences, King's College London, United Kingdom (P.T., M.N.); Cardiovascular Division, BHF Centre of Research Excellence, James Black Centre, King's College London, United Kingdom (M.S., D.A.R., A.M.S.); Department of Cardiology and Pneumology, Medical Center Goettingen, Germany (M.S.); Cardiovascular Division, BHF Centre of Research Excellence, Rayne Institute, St Thomas' Hospital, King's College London, United Kingdom (J.E.C.); Novo Nordisk A/S, Diabetic Complications Biology, Novo Nordisk Park, Maaloev, Denmark (A.S.); and Department of Clinical Experimental Research, Glostrup Research Institute, Rigshospitalet, Denmark (A.S.).

Background: Research into the therapeutic potential of α-calcitonin gene-related peptide (α-CGRP) has been limited because of its peptide nature and short half-life. Here, we evaluate whether a novel potent and long-lasting ( ≥7 hours) acylated α-CGRP analogue (αAnalogue) could alleviate and reverse cardiovascular disease in 2 distinct murine models of hypertension and heart failure in vivo.

Methods: The ability of the αAnalogue to act selectively via the CGRP pathway was shown in skin by using a CGRP receptor antagonist. The effect of the αAnalogue on angiotensin II-induced hypertension was investigated over 14 days. Blood pressure was measured by radiotelemetry. The ability of the αAnalogue to modulate heart failure was studied in an abdominal aortic constriction model of murine cardiac hypertrophy and heart failure over 5 weeks. Extensive ex vivo analysis was performed via RNA analysis, Western blot, and histology.

Results: The angiotensin II-induced hypertension was attenuated by cotreatment with the αAnalogue (50 nmol·kg·d, SC, at a dose selected for lack of long-term hypotensive effects at baseline). The αAnalogue protected against vascular, renal, and cardiac dysfunction, characterized by reduced hypertrophy and biomarkers of fibrosis, remodeling, inflammation, and oxidative stress. In a separate study, the αAnalogue reversed angiotensin II-induced hypertension and associated vascular and cardiac damage. The αAnalogue was effective over 5 weeks in a murine model of cardiac hypertrophy and heart failure. It preserved heart function, assessed by echocardiography, while protecting against adverse cardiac remodeling and apoptosis. Moreover, treatment with the αAnalogue was well tolerated with neither signs of desensitization nor behavioral changes.

Conclusions: These findings, in 2 distinct models, provide the first evidence for the therapeutic potential of a stabilized αAnalogue, by mediating (1) antihypertensive effects, (2) attenuating cardiac remodeling, and (3) increasing angiogenesis and cell survival to protect against and limit damage associated with the progression of cardiovascular diseases. This indicates the therapeutic potential of the CGRP pathway and the possibility that this injectable CGRP analogue may be effective in cardiac disease.
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http://dx.doi.org/10.1161/CIRCULATIONAHA.117.028388DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5519346PMC
July 2017

Distinct Regulatory Effects of Myeloid Cell and Endothelial Cell NAPDH Oxidase 2 on Blood Pressure.

Circulation 2017 May 15;135(22):2163-2177. Epub 2017 Mar 15.

From King's College London British Heart Foundation Centre of Excellence, Cardiovascular Division, United Kingdom (C.M.S., M.S., J.Z., C.E.M., A.P., C.X.C.S., G.S., X.Z., H.M.-D., D.A.R., A.C.B., A.P., P.J.E., A.M.S.); Klinik und Poliklinik für Innere Medizin II, Universitätsklinikum Regensburg, Regensburg, Germany (C.M.S., L.S.M.); Department of Cardiology and Pneumology, Medical Center Goettingen, Germany (M.S.); and Center for Cardiology and Center for Thrombosis and Hemostasis, University Medical Center Mainz, Germany (S.K., P.W.).

Background: Hypertension caused by increased renin-angiotensin system activation is associated with elevated reactive oxygen species production. Previous studies implicate NADPH oxidase (Nox) proteins as important reactive oxygen species sources during renin-angiotensin system activation, with different Nox isoforms being potentially involved. Among these, Nox2 is expressed in multiple cell types, including endothelial cells, fibroblasts, immune cells, and microglia. Blood pressure (BP) is regulated at the central nervous system, renal, and vascular levels, but the cell-specific role of Nox2 in BP regulation is unknown.

Methods: We generated a novel mouse model with a floxed Nox2 gene and used Tie2-Cre, LysM Cre, or Cdh5-CreERT2 driver lines to develop cell-specific models of Nox2 perturbation to investigate its role in BP regulation.

Results: Unexpectedly, Nox2 deletion in myeloid but not endothelial cells resulted in a significant reduction in basal BP. Both Tie2-CreNox2 knockout (KO) mice (in which Nox2 was deficient in both endothelial cells and myeloid cells) and LysM CreNox2KO mice (in which Nox2 was deficient in myeloid cells) had significantly lower BP than littermate controls, whereas basal BP was unaltered in Cdh5-CreERT2 Nox2KO mice (in which Nox2 is deficient only in endothelial cells). The lower BP was attributable to an increased NO bioavailability that dynamically dilated resistance vessels in vivo under basal conditions without a change in renal function. Myeloid-specific Nox2 deletion had no effect on angiotensin II-induced hypertension, which, however, was blunted in Tie2-CreNox2KO mice, along with preservation of endothelium-dependent relaxation during angiotensin II stimulation.

Conclusions: We identify a hitherto unrecognized modulation of basal BP by myeloid cell Nox2, whereas endothelial cell Nox2 regulates angiotensin II-induced hypertension. These results identify distinct cell-specific roles for Nox2 in BP regulation.
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http://dx.doi.org/10.1161/CIRCULATIONAHA.116.023877DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5444427PMC
May 2017

Proteomic analysis of short-term preload-induced eccentric cardiac hypertrophy.

J Transl Med 2016 05 27;14(1):149. Epub 2016 May 27.

Department of Cardiology and Pneumology, University Medical Center, Goettingen, Germany.

Background: Hemodynamic load leads to cardiac hypertrophy and heart failure. While afterload (pressure overload) induces concentric hypertrophy, elevation of preload (volume overload) yields eccentric hypertrophy and is associated with a better outcome. Here we analysed the proteomic pattern of mice subjected to short-term preload.

Methods And Results: Female FVB/N mice were subjected to aortocaval shunt-induced volume overload that leads to an eccentric hypertrophy (left ventricular weight/tibia length +31 %) with sustained systolic heart function at 1 week after operation. Two-dimensional gel electrophoresis (2-DE) followed by mass spectrometric analysis showed alteration in the expression of 25 protein spots representing 21 different proteins. 64 % of these protein spots were up-regulated and 36 % of the protein spots were consistently down-regulated. Interestingly, α-1-antitrypsin was down-regulated, indicating higher elastin degradation and possibly contributing to the early dilatation. In addition to contractile and mitochondrial proteins, polymerase I and transcript release factor protein (PTRF) was also up-regulated, possibly contributing to the preload-induced signal transduction.

Conclusions: Our findings reveal the proteomic changes of early-stage eccentric myocardial remodeling after volume overload. Induced expression of some of the respiratory chain enzymes suggests a metabolic shift towards an oxidative phosphorylation that might contribute to the favorable remodeling seen in early VO. Down-regulation of α-1-antitrypsin might contribute to extracellular matrix remodeling and left ventricular dilatation. We also identified PTRF as a potential signaling regulator of volume overload-induced cardiac hypertrophy.
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http://dx.doi.org/10.1186/s12967-016-0898-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4884361PMC
May 2016

Molecular and structural transition mechanisms in long-term volume overload.

Eur J Heart Fail 2016 04 23;18(4):362-71. Epub 2015 Dec 23.

Department of Cardiology and Pneumology, Georg-August-University, Goettingen, Germany.

Aim: We have previously reported that early phase (1 week) of experimental volume overload (VO) has an adaptive phenotype while wall stress-matched pressure overload (PO) is maladaptive. Here we investigate the transition from adaptation to heart failure (HF) in long-term VO.

Methods And Results: FVB/N wild-type mice were subjected to VO induced by aortocaval shunt, and were followed by serial echocardiography until in vivo left ventricular ejection fraction was below <50% (135 ± 35 days). Heart failure was evident from increased lung and liver weight and increased mortality compared with sham. Maladaptive remodelling resulted in significantly reduced sarcomeric titin phosphorylation (causing increased sarcomeric stiffness), whereas interstitial fibrosis was not increased. This was paralleled by re-expression of the fetal gene program, activation of calcium/calmodulin-dependent protein kinase II (CaMKII), decreased protein kinase B (Akt) phosphorylation, high oxidative stress, and increased apoptosis. Consistently, development of HF and mortality were significantly aggravated in Akt-deficient mice.

Conclusion: Transition to HF in VO is associated with decreased Akt and increased CaMKII signalling pathways together with increased oxidative stress and apoptosis. Lack of interstitial fibrosis together with sarcomeric titin hypophosphorylation indicates an increased stiffness at the sarcomeric but not matrix level in VO-induced HF (in contrast to PO). Transition to HF may result from myocyte loss and myocyte dysfunction owing to increased stiffness.
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http://dx.doi.org/10.1002/ejhf.465DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5064674PMC
April 2016

Redox regulation of cardiomyocyte cell cycling via an ERK1/2 and c-Myc-dependent activation of cyclin D2 transcription.

J Mol Cell Cardiol 2015 Feb 6;79:54-68. Epub 2014 Nov 6.

King's College London British Heart Foundation Centre of Research Excellence, Cardiovascular Division, London, UK. Electronic address:

Adult mammalian cardiomyocytes have a very limited capacity to proliferate, and consequently the loss of cells after cardiac stress promotes heart failure. Recent evidence suggests that administration of hydrogen peroxide (H2O2), can regulate redox-dependent signalling pathway(s) to promote cardiomyocyte proliferation in vitro, but the potential relevance of such a pathway in vivo has not been tested. We have generated a transgenic (Tg) mouse model in which the H2O2-generating enzyme, NADPH oxidase 4 (Nox4), is overexpressed within the postnatal cardiomyocytes, and observed that the hearts of 1-3week old Tg mice pups are larger in comparison to wild type (Wt) littermate controls. We demonstrate that the cardiomyocytes of Tg mouse pups have increased cell cycling capacity in vivo as determined by incorporation of 5-bromo-2'-deoxyuridine. Further, microarray analyses of the transcriptome of these Tg mouse hearts suggested that the expression of cyclin D2 is significantly increased. We investigated the molecular mechanisms which underlie this more proliferative phenotype in isolated neonatal rat cardiomyocytes (NRCs) in vitro, and demonstrate that Nox4 overexpression mediates an H2O2-dependent activation of the ERK1/2 signalling pathway, which in turn phosphorylates and activates the transcription factor c-myc. This results in a significant increase in cyclin D2 expression, which we show to be mediated, at least in part, by cis-acting c-myc binding sites within the proximal cyclin D2 promoter. Overexpression of Nox4 in NRCs results in an increase in their proliferative capacity that is ablated by the silencing of cyclin D2. We further demonstrate activation of the ERK1/2 signalling pathway, increased phosphorylation of c-myc and significantly increased expression of cyclin D2 protein in the Nox4 Tg hearts. We suggest that this pathway acts to maintain the proliferative capacity of cardiomyocytes in Nox4 Tg pups in vivo and so delays their exit from the cell cycle after birth.
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http://dx.doi.org/10.1016/j.yjmcc.2014.10.017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4312357PMC
February 2015

Knockdown of prolyl-4-hydroxylase domain 2 inhibits tumor growth of human breast cancer MDA-MB-231 cells by affecting TGF-β1 processing.

Int J Cancer 2013 Jun 27;132(12):2787-98. Epub 2012 Dec 27.

Department of Cardiovascular Physiology, University Medical Center, Georg-August University of Göttingen, D-37073 Göttingen, Germany.

The prolyl-4-hydroxylase domain 1-3 (PHD1-3) enzymes are regulating the protein stability of the α-subunit of the hypoxia-inducible factor-1 (HIF-1), which mediates oxygen-dependent gene expression. PHD2 is the main isoform regulating HIF-1α hydroxylation and thus stability in normoxia. In human cancers, HIF-1α is overexpressed as a result of intratumoral hypoxia which in turn promotes tumor progression. The role of PHD2 for tumor progression is in contrast far from being thoroughly understood. Therefore, we established PHD2 knockdown clones of MDA-MB-231 breast cancer cells and analyzed their tumor-forming potential in a SCID mouse model. Tumor progression was significantly impaired in the PHD2 knockdown MDA-MB-231 cells, which could be partially rescued by re-establishing PHD2 expression. In a RNA profile screen, we identified the secreted phosphoprotein 1 (SPP1) as one target, which is differentially regulated as a consequence of the PHD2 knockdown. Knockdown of PHD2 drastically reduced the SPP1 expression in MDA-MB-231 cells. A correlation of SPP1 and PHD2 expression was additionally verified in 294 invasive breast cancer biopsies. In subsequent analyses, we identified that PHD2 alters the processing of transforming growth factor (TGF)-β1, which is highly involved in SPP1 expression. The altered processing capacity was associated with a dislocation of the pro-protein convertase furin. Thus, our data demonstrate that in MDA-MB-231 cells PHD2 might affect tumor-relevant TGF-β1 target gene expression by altering the TGF-β1 processing capacity.
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http://dx.doi.org/10.1002/ijc.27982DOI Listing
June 2013

Endothelial cell HIF-1α and HIF-2α differentially regulate metastatic success.

Cancer Cell 2012 Jan;21(1):52-65

Department of Physiology, Development and Neuroscience, University of Cambridge, UK.

The hypoxia inducible transcription factors (HIFs) control many mediators of vascular response, including both angiogenic factors and small molecules such as nitric oxide (NO). In studying how endothelial HIF response itself affects metastasis, we found that loss of HIF-1α in endothelial cells reduces NO synthesis, retards tumor cell migration through endothelial layers, and restricts tumor cell metastasis, and that loss of HIF-2α has in each case the opposite effect. This results from differential regulation of NO homeostasis that in turn regulates vascular endothelial growth factor expression in an NO-dependent feedback loop. These opposing roles for the two HIF factors indicate that both they and endothelial cells regulate metastasis as malignancy progresses.
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http://dx.doi.org/10.1016/j.ccr.2011.11.017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3334270PMC
January 2012

Prolyl hydroxylase domain (PHD) 2 affects cell migration and F-actin formation via RhoA/rho-associated kinase-dependent cofilin phosphorylation.

J Biol Chem 2010 Oct 27;285(44):33756-63. Epub 2010 Aug 27.

Department of Cardiovascular Physiology, Universitätsmedizin Göttingen, Georg-August University Göttingen, D-37073 Göttingen, Germany.

Cells are responding to hypoxia via prolyl-4-hydroxylase domain (PHD) enzymes, which are responsible for oxygen-dependent hydroxylation of the hypoxia-inducible factor (HIF)-1α subunit. To gain further insight into PHD function, we generated knockdown cell models for the PHD2 isoform, which is the main isoform regulating HIF-1α hydroxylation and thus stability in normoxia. Induction of a PHD2 knockdown in tetracycline-inducible HeLa PHD2 knockdown cells resulted in increased F-actin formation as detected by phalloidin staining. A similar effect could be observed in the stably transfected PHD2 knockdown cell clones 1B6 and 3B7. F-actin is at least in part responsible for shaping cell morphology as well as regulating cell migration. Cell migration was impaired significantly as a consequence of PHD2 knockdown in a scratch assay. Mechanistically, PHD2 knockdown resulted in activation of the RhoA (Ras homolog gene family member A)/Rho-associated kinase pathway with subsequent phosphorylation of cofilin. Because cofilin phosphorylation impairs its actin-severing function, this may explain the F-actin phenotype, thereby providing a functional link between PHD2-dependent signaling and cell motility.
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http://dx.doi.org/10.1074/jbc.M110.132985DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2962474PMC
October 2010