Inhibition of let-7c Regulates Cardiac Regeneration after Cryoinjury in Adult Zebrafish

The let-7c family of micro-RNAs (miRNAs) is expressed during embryonic development and plays an important role in cell differentiation. We have investigated the role of let-7c in heart regeneration after injury in adult zebrafish. let-7c antagomir or scramble injections were given at one day after cryoinjury (1 dpi). Tissue samples were collected at 7 dpi, 14 dpi and 28 dpi and cardiac function was assessed before cryoinjury, 1 dpi, 7 dpi, 14 dpi and 28 dpi. Inhibition of let-7c increased the rate of fibrinolysis, increased the number of proliferating cell nuclear antigen (PCNA) positive cardiomyocytes at 7 dpi and increased the expression of the epicardial marker raldh2 at 7 dpi. Additionally, cardiac function measured with echocardiography recovered slightly more rapidly after inhibition of let-7c. These results reveal a beneficial role of let-7c inhibition in adult zebrafish heart regeneration

Vezf1 regulates cardiac structure and contractile function

Background Vascular endothelial zinc finger 1 (Vezf1) is a transcription factor previously shown to regulate vasculogenesis and angiogenesis. We aimed to investigate the role of Vezf1 in the postnatal heart. Methods The role of Vezf1 in regulating cardiac growth and contractile function was studied in zebrafish and in primary cardiomyocytes. Findings We find that expression of Vezf1 is decreased in diseased human myocardium and mouse hearts. Our experimental data shows that knockdown of zebrafish Vezf1 reduces cardiac growth and results in impaired ventricular contractile response to β-adrenergic stimuli. However, Vezf1 knockdown is not associated with dysregulation of cardiomyocyte Ca2+ transient kinetics. Gene ontology enrichment analysis indicates that Vezf1 regulates cardiac muscle contraction and dilated cardiomyopathy related genes and we identify cardiomyocyte Myh7/β-MHC as key target for Vezf1. We further identify a key role for an MCAT binding site in the Myh7 promoter regulating the response to Vezf1 knockdown and show that TEAD-1 is a binding partner of Vezf1. Interpretation We demonstrate a role for Vezf1 in regulation of compensatory cardiac growth and cardiomyocyte contractile function, which may be relevant in human cardiac disease.Peer reviewe

Septin7b and let-7c modulate cardiac function and regeneration in zebrafish

Septins are small GTPases that associate with actin. Septins play an essential role in cytoskeleton organization but their role in cardiomyocyte organization and function are not completely known. Let-7c family of microRNAs are required for cell differentiation. Their role in heart regeneration after injury in adult zebrafish is unexplored. Cardiovascular diseases are the number one cause of death worldwide. Soon after birth, cardiomyocytes are no longer capable of cell division. Heart growth occurs through an increase in volume of cardiomyocytes: hypertrophy. Due to this, the human heart has very limited ability to regenerate after injury. In contrast, when an adult zebrafish heart is injured, complete regeneration occurs in 4-6 weeks. Cardiac regeneration in human heart requires deeper understanding of the molecular mechanisms involved. The study of zebrafish cardiac development is also required for completely understanding the regenerative capacity of zebrafish. The simple structure of the zebrafish heart allows the easy study of cardiac development. Moreover, the cardiac development of zebrafish is similar to that of the human heart in many aspects, for example migration of cardiac precursor cells towards the central line, heart tube formation and cardiac looping. The essential genes in cardiac development are conserved across vertebrates. Therefore, the zebrafish embryo is a good vertebrate animal model for heart studies. The aim of this thesis was to elucidate the role of septin7b and retinoic acid in heart development and function in embryonic zebrafish and the role of let-7c in adult zebrafish heart regeneration after cryoinjury, which is damage of ventricular cells by application of probe dipped in liquid nitrogen. We have found in our study that septin7b is important for cardiac development in embryonic zebrafish. Knockdown of septin7b leads to reduced accumulation of actin and disorganisation of actin filaments. Knockdown of septin7b reduced the accumulation of raldh2. Raldh2 is essential for the synthesis of retinoic acid, which is further required for cardiac development. Functional studies revealed reduced ventricular dimensions, cardiac contractility, and cardiac output in septin7b knockdown hearts. Inhibition of let-7c increases fibrinolysis and cardiomyocyte proliferation in the adult heart after cryoinjury. These results together reveal an important and beneficial role of septin7b and let-7c in zebrafish.Septiinit ovat pieniä GTPaaseja, jotka liittyvät aktiiniin. Septiineillä on olennainen rooli sytoskeleton organisoinnissa, mutta niiden roolia sydänlihassolujen organisoinnissa ja toiminnassa ei täysin tunneta. Let-7c-perhe mikroRNA:ita tarvitaan solujen erilaistumiseen. Niiden roolia sydämen uudistamisessa aikuisen seeprakalan vamman jälkeen ei ole tutkittu. Sydän- ja verisuonisairaudet ovat maailman suurin kuolinsyy. Pian syntymän jälkeen sydänlihassolut eivät enää pysty jakautumaan. Sydämen kasvu tapahtuu kardiomyosyyttien määrän lisääntymisen kautta: hypertrofia. Tästä johtuen ihmisen sydämellä on hyvin rajallinen kyky uusiutua vamman jälkeen. Sitä vastoin, kun aikuisen seeprakalan sydän vaurioituu, täydellinen uusiutuminen tapahtuu 4-6 viikossa. Sydämen regeneraatio ihmisen sydämessä vaatii syvempää ymmärrystä asiaan liittyvistä molekyylimekanismeista. Seeprakalan sydämen kehityksen tutkimusta tarvitaan myös seeprakalan regeneratiivisen kapasiteetin täydelliseksi ymmärtämiseksi. Seeprakalan sydämen yksinkertainen rakenne mahdollistaa sydämen kehityksen helpon tutkimuksen. Lisäksi seeprakalan sydämen kehitys on samanlaista kuin ihmisen sydämen monissa asioissa, esimerkiksi sydämen esiastesolujen siirtyminen kohti keskilinjaa, sydänputken muodostuminen ja sydämen silmukka. Sydämen kehityksen olennaiset geenit ovat säilyneet selkärankaisilla. Siksi seeprakalan alkio on hyvä selkärankainen eläinmalli sydäntutkimuksiin. Tämän opinnäytetyön tavoitteena oli selvittää septiini7b:n ja retinoiinihapon roolia sydämen kehityksessä ja toiminnassa alkion seeprakalalla sekä let-7c:n roolia aikuisen seeprakalan sydämen regeneraatiossa kryovaurion jälkeen, joka on kammiosolujen vaurioituminen vatsaan upotetun koettimen avulla. nestemäinen typpi. Olemme havainneet tutkimuksessamme, että septin7b on tärkeä alkion seeprakalan sydämen kehitykselle. Septin7b:n tuhoutuminen johtaa aktiinin kertymisen vähenemiseen ja aktiinifilamenttien hajoamiseen. septin7b:n tuhoutuminen vähensi raldh2:n kertymistä. Raldh2 on välttämätön retinoiinihapon synteesille, jota tarvitaan edelleen sydämen kehitykseen. Funktionaaliset tutkimukset paljastivat pienentyneet kammiomitat, sydämen supistumiskyky ja sydämen minuuttitilavuus septin7b-sydämissä. Let-7c:n esto lisää fibrinolyysiä ja kardiomyosyyttien lisääntymistä aikuisen sydämessä kryovaurion jälkeen. Nämä tulokset yhdessä paljastavat septin7b:n ja let-7c:n tärkeän ja hyödyllisen roolin seeprakaloissa

Zebrafish Heart Failure Models

Heart failure causes significant morbidity and mortality worldwide. The understanding of heart failure pathomechanisms and options for treatment remain incomplete. Zebrafish has proven useful for modeling human heart diseases due to similarity of zebrafish and mammalian hearts, fast easily tractable development, and readily available genetic methods. Embryonic cardiac development is rapid and cardiac function is easy to observe and quantify. Reverse genetics, by using morpholinos and CRISPR-Cas9 to modulate gene function, make zebrafish a primary animal model for in vivo studies of candidate genes. Zebrafish are able to effectively regenerate their hearts following injury. However, less attention has been given to using zebrafish models to increase understanding of heart failure and cardiac remodeling, including cardiac hypertrophy and hyperplasia. Here we discuss using zebrafish to study heart failure and cardiac remodeling, and review zebrafish genetic, drug-induced and other heart failure models, discussing the advantages and weaknesses of using zebrafish to model human heart disease. Using zebrafish models will lead to insights on the pathomechanisms of heart failure, with the aim to ultimately provide novel therapies for the prevention and treatment of heart failure.Peer reviewe

TCR-induced activation of LFA-1 involves signaling through Tiam1

7siAdhesion is pivotal for most leukocyte functions, and the β(2) integrin family of adhesion molecules plays a central role. The integrins need activation to become functional, but the molecular events resulting in adhesion have remained incompletely understood. In human T cells, activation through the TCR results in specific phosphorylation of the T758 on the β(2) chain of LFA-1. We now show that this phosphorylation leads to downstream binding of 14-3-3 proteins, followed by engagement of the guanine nucleotide exchange factor protein Tiam1 and Rac1 activation. Downregulation of the signaling molecules inhibits LFA-1 activity. Activation by the chemokine stromal cell-derived factor-1α also results in T758 phosphorylation and integrin activation. Thus, TCR and chemokine activation converges on LFA-1 phosphorylation, followed by similar downstream events affecting adhesion.nonenoneGrönholm, Mikaela; Jahan, Farhana; Marchesan, Silvia; Karvonen, Ulla; Aatonen, Maria; Narumanchi, Suneeta; Gahmberg, Carl GGrönholm, Mikaela; Jahan, Farhana; Marchesan, Silvia; Karvonen, Ulla; Aatonen, Maria; Narumanchi, Suneeta; Gahmberg, Carl G

Deficiency of heme oxygenase 1a causes detrimental effects on cardiac function

Humans lacking heme oxygenase 1 (HMOX1) display growth retardation, haemolytic anaemia, and vulnerability to stress; however, cardiac function remains unclear. We aimed to explore the cardiac function of zebrafish lacking hmox1a at baseline and in response to stress. We generated zebrafish hmox1a mutants using CRISPR/Cas9 genome editing technology. Deletion of hmox1a increases cardiac output and further induces hypertrophy in adults. Adults lacking hmox1a develop myocardial interstitial fibrosis, restrain cardiomyocyte proliferation and downregulate renal haemoglobin and cardiac antioxidative genes. Larvae lacking hmox1a fail to respond to hypoxia, whereas adults are insensitive to isoproterenol stimulation in the heart, suggesting that hmox1a is necessary for cardiac response to stress. Haplodeficiency of hmox1a stimulates non-mitochondrial respiration and cardiac cell proliferation, increases cardiac output in larvae in response to hypoxia, and deteriorates cardiac function and structure in adults upon isoproterenol treatment. Intriguingly, haplodeficiency of hmox1a upregulates cardiac hmox1a and hmox1b in response to isoproterenol. Collectively, deletion of hmox1a results in cardiac remodelling and abrogates cardiac response to hypoxia and isoproterenol. Haplodeficiency of hmox1a aggravates cardiac response to the stress, which could be associated with the upregulation of hmox1a and hmox1b. Our data suggests that HMOX1 homeostasis is essential for maintaining cardiac function and promoting cardioprotective effects.Peer reviewe