eRapport

MicroRNA regulation in left ventricle myocardium from patients with heart failure – Targeted treatment by specific microRNA manipulation.

Prosjekt
Prosjektnummer
90158300
Ansvarlig person
Morten Høydal
Institusjon
NTNU, MH-fakultetet, Institutt for sirkulasjon og bildediagnostikk
Prosjektkategori
Flerårig prosjekt
Helsekategori
Cardiovascular
Forskningsaktivitet
1. Underpinning
Rapporter
2022 - sluttrapport
During the project we have had continuous good progress in addressing the topic for the project. The first years of the project we discovered 15 specific microRNA (miR) based on samples from coronary bypass surgery patients. Here we tested all 15 miR and modulated these by transfecting both inhibors and miR mimics in the setting of hypoxia reoxygenation that resembles the physiology of ischemia reperfusion in human cardiomyocytes in culture. We have by the use of this approach characterized the protective effect on parameters of cardiomyocyte function, proliferation, life/death assay apoptosis and survival and narrowed down the numbers to a more refined subset of potential miR that will be further processed for their protective effects in combination. These miR will now be followed up in detail. In addition we have already focused in specific on a subset of these miR, namely miR-181b, miR-451 and miR-210. miR-181b to regulate the protein sarcoendoplasmic reticulum Ca2+ ATPase (SERCA2a). To verify the role of these miR we performed an in vivo study on post MI HF rats. The results showed that miR-181b inhibition was successful on reduced the size of myocardial infarction, improved Ca2+ removal in isolated cardiomyocytes, improved left ventricle cardiac performance especially on diastolic parameters including time to relaxation and end diastolic pressure. We have also tested in depth, the effect of miR-451a manipulation which was the most regulated miR in our human data (73% reduced in post MI HF). Using bioinformatics analyses combined with existing literature we tested the hypothesis that miR-451a regulates the activity of matrix metalloproteinases (MMP). Our hypothesis was confirmed; treatment by miR-451a reduced the pathological elevated levels and activity of MMP. Our data suggest that increasing the levels of lowly expressed miR-451a in cardiac patients may warrant further investigation for its potential as a targeted treatment to prevent the progression of HF. The last years we have had a very good progress on further verification on the specific role of microRNA-210 where we recently published a paper showing a "janus role" of microRNA-210 in ischemia-reperfusion and a second paper showing that GSK3β Inhibition Is the Molecular Pivot That Underlies the Mir-210-Induced Attenuation of Intrinsic Apoptosis Cascade during Hypoxia. Here we further substantiate that microRNA-210 is strongly protective in the hypoxic phase and deleterious in the reperfusion(redox phase). We have also found that this specific microRNA is very important in regard to enhance resistance to human cardiomyocyte cell death via apoptosis. Taken together, the outcome of this study has generated several important papers that combined have moved the research field forward and fostered development towards improved understand of how to treat ischemic heart disease. The project has also been relevant for bridging collaboration between clinicians and basic researchers. Our ambition was to exchange and share knowledge through of regular meetings and presentations with aim to build better molecular and cellular understanding for clinicians and likewise will advance clinical understanding for the basic scientists involved in the project. We believe that this aim have been fulfilled, and better and more connection between researchers and clinicians have been established. We also bellies that the communication and meetings have fostered enhanced insight to molecular and cellular physiology in the clinical groups that hopefully benefit improved treatment.

The project has also been relevant for bridging collaboration between clinicians and basic researchers. Our ambition was to exchange and share knowledge through of regular meetings and presentations with aim to build better molecular and cellular understanding for clinicians and likewise will advance clinical understanding for the basic scientists involved in the project. We believe that this aim have been fulfilled, and better and more connection between researchers and clinicians have been established. We also bellies that the communication and meetings have fostered enhanced insight to molecular and cellular physiology in the clinical groups that hopefully benefit improved treatment.

2021
Despite substantial improvements in clinical outcome, a significant share of patients still suffer from heart failure (HF). With this project we have contributed to move the research field within cardiology further by establishing novel treatable targets that in the future could benefit patients with ischemic heart disease.Ischemic heart disease (IHD) is globally the leading cause of death. MI is the irreversible damage of myocardial tissue caused by block of the arteries that supply oxygenated blood to the cardiac muscle. Without sufficient oxygen, the tissue dies. One of the major determinants of mortality and morbidity after MI is the extent of myocardial necrosis, known as infarct size. Following MI, the activation of multiple intracellular signalling pathways causes pathological remodelling of the heart and eventually HF. Coronary reperfusion is the only means of limiting MI size, provided that it occurs early after coronary occlusion, but may paradoxically directly result in tissue injury. While reperfusion therapies are some of the most successful discoveries in medicine, cardioprotective therapies that limit MI size and resist development of HF remain today one of the top 10 unmet clinical needs in cardiology numerous efforts have been undertaken to elucidate the pathological processes of ischemic cardioprotection and development of post-MI chronic HF, with the aim to improve current therapies. To date, effective strategies have not yet been established. In the recent position paper from the European Society of Cardiology, a major aim is to define and discover novel treatment strategies directed at multiple signalling pathways, which remains a gap in existing knowledge. During the last year we have had good progress in the project in establishing the role of microRNA in the setting of ischemia. We have especially in human cardiomyocytes determined novel mechanisms linked to the microRNA profile previously detected at earlier stages of this project in human patients biopsies. Especially during this last year we have verified novel mechanistic insight in the role of miR-210. Furthermore we have had a very promising progress on the discovery of several novel proteins linked to the microRNA regulation in human hearts. We do now have evidence of the regulatory role and the protective effect against myocardial infarction of several of these proteins both in animal models and in human cardiomyocytes. The data show promising indications that this protein is a master regulator linking different signalling pathways in the heart, providing a unique opportunity for establishing new mechanistic knowledge of how to limit ischemic heart disease. This year we have also had a very good progress on further verification on the specific role of microRNA-210 where we recently published a paper showing a "janus role" of microRNA-210 in ischemia-reperfusion. Here we show that microRNA-210 is strongly protective in the hypoxic phase and deleterious in the reperfusion(redox phase). We have also found that this specific microRNA is very important in regard to enhance resistance to human cardiomyocyte cell death via apoptosis. Taken toghter the outcome of this study has generated several important papers that combined have moved the research field forward and fostered development towards improved understand of how to treat ischemic heart disease.

The project has also been relevant for bridging collaboration between clinicians and basic researchers. Our ambition was to exchange and share knowledge through of regular meetings and presentations with aim to build better molecular and cellular understanding for clinicians and likewise will advance clinical understanding for the basic scientists involved in the project. We believe that this aim have been fulfilled, and better and more connection between researchers and clinicians have been established. We also bellies that the communication and meetings have fostered enhanced insight to molecular and cellular physiology in the clinical groups that hopefully benefit improved treatment.

2020
The project is in agreement with the progress of assessing myocardial infarction heart failure (HF) and potential of microRNA targeting. We have tested all microRNA described for survival, cellular function, proliferation, calcium handling action potential, and narrowed down our microRNA to a subset of highly potential microRNAs.Ischemic heart disease (IHD) is globally the leading cause of death. Despite substantial improvements in clinical outcome, a significant share of patients still suffer from heart failure (HF) and premature death due to permanent damage to their myocardium after myocardial infarction (MI). MI is the irreversible damage of myocardial tissue caused by block of the arteries that supply oxygenated blood to the cardiac muscle. Without sufficient oxygen, the tissue dies. One of the major determinants of mortality and morbidity after MI is the extent of myocardial necrosis, known as infarct size. Following MI, the activation of multiple intracellular signalling pathways causes pathological remodelling of the heart and eventually HF. Coronary reperfusion is the only means of limiting MI size, provided that it occurs early after coronary occlusion, but may paradoxically directly result in tissue injury. While reperfusion therapies are some of the most successful discoveries in medicine, cardioprotective therapies that limit MI size and resist development of HF remain today one of the top 10 unmet clinical needs in cardiology Numerous efforts have been undertaken to elucidate the pathological processes of ischemic cardioprotection and development of post-MI chronic HF, with the aim to improve current therapies. To date, effective strategies have not yet been established. In the recent position paper from the European Society of Cardiology, a major aim is to define and discover novel treatment strategies directed at multiple signalling pathways, which remains a gap in existing knowledge. During the last year we have had good progress in the project in establishing the role of microRNA in the setting of ischemia. We have especially in human cardiomyocytes determined novel mechanisms linked to the microRNA profile previously detected at earlier stages of this project in human patients biopsies. Especially during this last year we have verified novel mechanistic insight in the role of miR-210 and miR-24 and their role of protecting human cardiomyocytes against hypoxia deoxygenation stress (a model that mimic the ischema-reperfusion damage). Furthermore we have had a very promising progress on the discovery of a protein that were linked to our microRNA signature in ischemic cardiomyocytes. We do now have evidence of the regulatory role and the protective effect against myocardial infarction of this protein both in animal models and in human cardiomyocytes. The data show promising indications that this protein is a master regulator linking different signalling pathways in the heart, providing a unique opportunity for establishing new mechanistic knowledge of how to limit ischemic heart disease.
2019
The project is in agreement with the progress of assessing myocardial infarction heart failure (HF) and potential of miR targeting. We have tested all microRNA described for survival, cellular function, proliferation, calcium handling action potential, and narrowed down our microRNA to a subset of highly potential microRNAs.The last year we have had significant progress in the underpinning mechanisms relating to the protective effect of miR against ischemic heart disease and the progress towards heart failure. Especially we recently published that Matrix metalloproteinases (MMP) are important for cardiac remodeling. this study demonstrates that miR-451a regulates MMP-2 and MMP-9 in human cardiac cells. In both of our experimental models, cardiomyocytes stimulated either with ET-1 or with hypoxia, both MMP-2 and MMP-9 remained at expression levels not different from controls following treatment by miR-451a mimics. The effect of increasing miR-451a was further verified by reducing MMP-2 and MMP-9 protein activity. We also found that the regulatory effect by miR-451a on MMP-2 and MMP-9 is caused by the regulation of macrophage inhibitory factor, which is a key regulator of inflammatory processes in the heart. Taken together, our data suggest that increasing the levels of lowly expressed miR-451a in cardiac patients may warrant further investigation for its potential as a targeted treatment to prevent the progression of HF. Furthermore, we have in detailed finally verified the importance of each single miR in our initial miR signature using up/downregulation and verified their potential in protecting human cardiomyocytes. Here, we have found a set of miR with high potential to protect the heart during myocardial infarction and thus reduce the potential for heart failure development. These data are in preparation for publication. Furthermore, in the process of establishing the role of miR in the setting of schema, we found a new protein in the heart that never previously have been explored. The regulatory role of the microRNA in the initial study were found to have significant impact on this protein. We have therefore expanded our initial plan to also include verification of the role of this protein in the setting of ischema and hypoxia in human cardiomyocytes. We have found that this protein is significantly changed in the heart and the protective effect against ischemic/hypoxic damage has a potential that we believe has an enormous potential as a novel target to treat the heart in the setting of myocardial infarction. The data from the discovery of this protein was presented as "late-breaking-science" at the American heart Association conference, November 2019. We do now have both data from experimental animal models and a range of cardiomyocytes exposed to hypoxic stress that this protein is important for protecting the heart. The paper for this finding is now in preparation for publication.

At this stage it is to early to asses if the results from the current project will have implications for the patients with HF. However, our data holds a promise for discovering novel targets that may in the long run benefit patient with HF. The overall goal of this project is to build a foundation for developing new therapeutic targets for HF, thus contributing to reduce the burden of HF.

2018
The project is in line with the progress described with aim of assessing myocardial infarction heart failure (HF) and potential of miR targeting. We have tested all microRNA described for survival, cellular function, proliferation, calcium handling action potential, following ischema reperfusion injury and refined the number of protective microRNAThe current project aimed at assessing myocardial infarction heart failure (MI HF) and and the potential of microRNA treatment. In the preliminary data to this project we identified the regulation of miR in human left ventricle biopsies harvested during coronary artery bypass grafting surgery. We recently published a more detailed characterisation of cellular properties relating to calcium handling, structural remodelling, cardiomyocyte function in addition to protein regulation of key functional aspects of cardiomyocyte function from these patients (ESC Heart Failure 2018). In respect to further evaluation of testing the potential for miR manipulation we have in a series of experiments tested miR mimic and inhibitions in an attempt to verify the miR signature. All miR in our signature have been modulated in a high- throughput approach screening using Celloptiq at Clyde bioscience, Glasgow, UK as well as in our own laboratory at NTNU. Furthermore, all 15 miR have been modulated by transfecting both inhibors and miR mimics in the setting of hypoxia reoxygenation that resembles the physiology of ischemia reperfusion in human cardiomyocytes in culture. We have by the use of this approach characterised the protective effect on parameters of cardiomyocyte function, proliferation, life/death assay apoptosis and survival and narrowed down the numbers to a more refined subset of potential miR that will be further processed for their protective effects in combination. These miR will now be followed up in detail. In addition we have already focused in specific on a subset of these miR, namely miR-181b, miR-451 and miR-210 where we either have performed or are in progress of in vivo manipulations. Using bioinformatics prediction analyses we followed up our miR signature further and found miR-181b to regulate the protein sarcoendoplasmic reticulum Ca2+ ATPase (SERCA2a). To verify the role of these miR we performed an in vivo study on post MI HF rats. The results showed that miR-181b inhibition was successful on reduced the size of myocardial infarction, improved Ca2+ removal in isolated cardiomyocytes, improved left ventricle cardiac performance especially on diastolic parameters including time to relaxation and end diastolic pressure. We also recently analysed the results from LC-MS) proteomics on the samples and found that miR181 treatment rescued a pathological proteomic profile in post MI HF rats. We have also tested in depth, the effect of miR-451a manipulation which was the most regulated miR in our human data (73% reduced in post MI HF). Using bioinformatics analyses combined with existing literature we tested the hypothesis that miR-451a regulates the activity of matrix metalloproteinases (MMP). Our hypothesis was confirmed; treatment by miR-451a reduced the pathological elevated levels and activity of MMP (in review J Cell Physiology), presented at ESC 2017). We have also tested the effect of mir-210 which was significantly upregulated in our human miR-signature. In our hypoxic model miR-210 significantly reduces mitochondrial respiration after 24 hours, whereas inhibition of miR-210 significantly increases oxygen consumption after 24 hours of hypoxia. Taken together, these data indicate a significant role of both miR-210, miR-451 and miR-181b in addition to a few other miR-candidates on controlling cardiac dysfunction in HF.

Currently we are performing the underpinning work and have not yet reached a state where it may influence the patient. Ischemic heart disease and HF is the leading causes of death and disability in Europe and worldwide. HF is a major public health problem, with a prevalence of more than 23 million worldwide. In 1997, HF was singled out as an emerging epidemic. As the economic, social and personal burden of HF is enormous, the disease constitutes one of the most pressing unmet clinical needs in Europe. This project effort aims to introduce changes in treatment strategies and thus improve the quality of life for a growing population of patients with HF, which will also lead to a substantial positive economic impact in our society. This is an interdisciplinary project that involves complementary expertise from basic scientists and clinicians at St. Olavs hospital. The project is therefore also highly relevant for bridging collaboration between clinicians and basic researchers.

2017
Det overordnede målet med prosjektet er å forme en basis for å etablere nye medisinske angrepspunkt for behandling av pasienter med hjertesvikt. Tanken bak prosjektet er å avdekke endringer av micoRNA hos pasienter med hjertesvikt samt å avdekke betydningen av disse for hjertefunksjon.Dette er et tverrfaglig prosjekt med komplementær kompetanse fra lokale partnere ved NTNU , St. Olavs hospital , og flere nasjonale og internasjonale universiteter i kardiologi , cellefysiologi , biofysikk og molekylær genetikk. Verstskapslaboratoriet ( NTNU / St. Olavs Hospital ) tilbyr som en del av det nye integrerte universitetssykehuset, som er "bygd for tverrfaglig samarbeid ", et unikt miljø for translasjonsforskning. Forekomsten av hjertesvikt ( HF ) i vestlige land er i dag kjent for å være 1-2 % , og den øker med alderen . Til tross for en rekke medisinske behandlingsstrategier, fortsetter forekomsten av hjertesvikt å øke. Dette understreker behovet for nye medikamentelle strategier. Ved utvikling av hjertesvikt så aktiveres en rekke faktorer i hjertet som gradvis bryter ned hjertets funksjon, og i tillegg øker risikoen for alvorlige rytmeforstyrrelser i hjertet. Spesifikt så vil vi i hjerteceller fra pasienter med hjertesvikt se på reguleringen av en klasse med små RNA-molekyler (microRNA) som i nyere tid har vist seg å endre genuttrykket til proteiner som er sentrale for å opprettholde normal hjertecellefunksjon. Det overordnede målet med prosjektet er å forme en basis for å etablere nye medisinske angrepspunkt for behandling av pasienter med hjertesvikt. Tanken bak prosjektet er å avdekke endringer av micoRNA hos pasienter med hjertesvikt samt å avdekke betydningen av disse for hjertefunksjon. I løpet av det første året dette prosjektet har pågått har vi gjennom flere metoder avdekt sentrale microRNA som regulerer viktige proteiner som bedrer hjertefunksjon ved hjertesvikt. Vi har mellom annet gjennomført en stor preklinisk studie med behandling av post infarkt hjertesvikt rotter med inhibisjon av et spesifikt microRNA. Resultaene er spennende og viser en tydelig bedring av hjertets karakter. Prosjektet har dermed et potensiale til å redusere sykdomsbyrden for en stadig økende pasientgruppe med hjertesvikt. Vi har videre utviklet nye metoder og patologiske modeller som simulerer prosesser som skjer ved hjerteinfarkt. Ved bruk av denne modellen som nyttiggjør seg av human induserbare pluripotente stam celler som er diferensierte til hjerteceller så har vi kunnet bekrefte og manipulere en rekke av de spesifikke microRNAene som er sentrale for bakgrunnen for dette prosjektet. Både modellen og resultat frå MicroRNA manipulering i denne ble presentert på konferanse ESC- heart failure i fjor. Videre har vi gjennomført WP1 hos Clyde Bioscience, Glasgow og transfektert samtlige 15 microRNA med mimic/inhibitorer beskrevet med bakgrunn i vårt humane utgangspunkt med regulerte microRNA i prøver fra bypassopererte. Dette materialet er fortsatt under analyse og forventes ferdigstilt i løpet av våren.

I løpet av det første året dette prosjektet har pågått har vi gjennom flere metoder avdekt sentrale microRNA som regulerer viktige proteiner som bedrer hjertefunksjon ved hjertesvikt. Det er for tidlig å si om våre funn kan overføres til utviklingen av nye medikamenter, men det er et klart mål for prosjektet. Vi har tro på at videre arbeid i dette prosjektet har et potensiale til å redusere sykdomsbyrden for en stadig økende pasientgruppe med hjertesvikt gjennom optimalisert og utvikling av nye medikamentelle angrepspunkt.

Vitenskapelige artikler
Marwarha G, Røsand Ø, Slagsvold KH, Høydal MA

GSK3β Inhibition Is the Molecular Pivot That Underlies the Mir-210-Induced Attenuation of Intrinsic Apoptosis Cascade during Hypoxia.

Int J Mol Sci 2022 Aug 19;23(16). Epub 2022 aug 19

PMID: 36012628

Marwarha G, Røsand Ø, Scrimgeour N, Slagsvold KH, Høydal MA

miR-210 Regulates Apoptotic Cell Death during Cellular Hypoxia and Reoxygenation in a Diametrically Opposite Manner.

Biomedicines 2021 Dec 25;10(1). Epub 2021 des 25

PMID: 35052722

Berg PC, Hansson ÅML, Røsand Ø, Marwarha G, Høydal MA

Overexpression of Neuron-Derived Orphan Receptor 1 (NOR-1) Rescues Cardiomyocytes from Cell Death and Improves Viability after Doxorubicin Induced Stress.

Biomedicines 2021 Sep 16;9(9). Epub 2021 sep 16

PMID: 34572418

Stølen TO, Høydal MA, Ahmed MS, Jørgensen K, Garten K, Hortigon-Vinagre MP, Zamora V, Scrimgeour NR, Berre AMO, Nes BM, Skogvoll E, Johnsen AB, Moreira JBN, McMullen JR, Attramadal H, Smith GL, Ellingsen Ø, Wisløff U

Exercise training reveals micro-RNAs associated with improved cardiac function and electrophysiology in rats with heart failure after myocardial infarction.

J Mol Cell Cardiol 2020 11;148():106-119. Epub 2020 sep 10

PMID: 32918915

Scrimgeour NR, Wrobel A, Pinho MJ, Høydal MA

microRNA-451a prevents activation of matrix metalloproteinases 2 and 9 in human cardiomyocytes during pathological stress stimulation.

Am J Physiol Cell Physiol 2020 01 01;318(1):C94-C102. Epub 2019 okt 16

PMID: 31618079

Stølen T, Shi M, Wohlwend M, Høydal MA, Bathen TF, Ellingsen Ø, Esmaeili M

Effect of exercise training on cardiac metabolism in rats with heart failure.

Scand Cardiovasc J 2020 Apr;54(2):84-91. Epub 2019 sep 10

PMID: 31500456

Smenes BT, Bækkerud FH, Slagsvold KH, Hassel E, Wohlwend M, Pinho M, Høydal M, Wisløff U, Rognmo Ø, Wahba A

Acute exercise is not cardioprotective and may induce apoptotic signalling in heart surgery: a randomized controlled trial.

Interact Cardiovasc Thorac Surg 2018 Jul 01;27(1):95-101.

PMID: 29447379

Høydal MA, Kirkeby-Garstad I, Karevold A, Wiseth R, Haaverstad R, Wahba A, Stølen TL, Contu R, Condorelli G, Ellingsen Ø, Smith GL, Kemi OJ, Wisløff U

Human cardiomyocyte calcium handling and transverse tubules in mid-stage of post-myocardial-infarction heart failure.

ESC Heart Fail 2018 06;5(3):332-342. Epub 2018 feb 12

PMID: 29431258

Scrimgeour NR, Wrobel A, Pinho MJ, Høydal MA.

microRNA-451a prevents activation of matrix metalloproteinases 2 and 9 in human cardiomyocytes during pathological stress stimulation.

Am J Physiology- cell physiology, Epub 2019 Oct 16.

Pinho, Scrimgeour, Nyman, Enguita, Ishizuka, Sawa, Hoydal

Discovery of s Cardiac Role for a “Neuronal Protein”, Reduced Levels of Disrupted in Schizophrenia 1 (DISC1) in the Heart Causes Cardiac Deterioration and Mitochondrial Dysfunction

Circulation Research. 2019Circulation Research. 2019;125:e99–e111

J. Pinho N.R. Scrimgeour T. Stolen K. Solvang-Garten A. Sharma D. Miranda Fonseca G. Smith M. Hoydal

Modeling of cardiac ischaemia and reperfusion injury: a human-based in vitro model using iPS-derived cardiomyocytes

European Heart Journal, Volume 38, Issue suppl_1, 1 August 2017, ehx502.P1095, https://doi.org/10.1093/eurheartj/ehx502.P1095

N. Scrimgeour, M. Pinho, A. Wrobel, M. Hoydal

MicroRNA-451a regulate expression and activity of matrix metalloproteinases 2 and 9 in human cardiomyocytes

European Heart Journal, Volume 38, Issue suppl_1, 1 August 2017, ehx493.P5389, https://doi.org/10.1093/eurheartj/ehx493.P5389

MA. Høydal, I. Kirkeby-Garstad, A. Karevold, R. Wiseth, R. Haaverstad, A. Wahba, MD, TS. Stølen, R. Contu G. Condorelli, Ø. Ellingsen, GL. Smith, OJ. Kemi PhD, U. Wisløff

Human cardiomyocyte calcium handling and transverse tubules in mid-stage of post-myocardial infarction heart failure

ESC Heart Failure (2018) Published online in Wiley Online Library (wileyonlinelibrary.com) DOI: 10.1002/ehf2.12271

Deltagere
  • Nathan Robert Scrimgeour Prosjektdeltaker
  • Tomas Stølen Forsker
  • Morten Andre Høydal Prosjektleder

eRapport er utarbeidet av Sølvi Lerfald og Reidar Thorstensen, Regionalt kompetansesenter for klinisk forskning, Helse Vest RHF, og videreutvikles av de fire RHF-ene i fellesskap, med støtte fra Helse Vest IKT

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