Cardiac endothelial cells : potential therapeutic targets in heart disease

Karthik Amudhala Hemanthakumar

Tutkimustuotos: OpinnäyteVäitöskirjaArtikkelikokoelma

Abstrakti

Cardiovascular diseases (CVD) rank as a number one cause for mortality and accounts for one third of the deaths in several OECD countries (OECD, 2015). According to WHO, CVD pathology is characterised by impaired coronary vasculature associated with cardiac dysfunction, which often results in heart failure (Mendis S, 2011). Although the outlook for prevention and management of CVD risk factors is advancing, the extent of CVD mortality and morbidity remains relatively high (Mendis S, 2011) and the clinical prognosis of heart failure remains poorer than most of the cancers (Braunwald, 2015). Better understanding of the cellular and molecular links between the coronary vasculature and cardiac function under physiological and pathological conditions would enhance the development of personalized targeted therapies for CVD. In this thesis, my main objective is to define mechanistic insights how proangiogenic cues like VEGF-B or PlGF promote coronary angiogenesis -mediated physiological cardiac hypertrophy and to characterise the effect of CVD risk factors (aging, obesity, physical inactivity, and pressure overload) on cardiac endothelial cells (ECs) and cardiac function. We have applied molecular, biochemical, imaging and gene delivery methods to elucidate the phenotypes and molecular mechanisms in in vivo and in vitro model systems. In the study I, we showed that AAV9-VEGF-B overexpression or endothelial deletion of VEGFR1 increased the bioavailability of the endogenous VEGF to activate VEGFR2 in ECs promoting coronary angiogenesis. Importantly, this indirect activation of VEGFR2 is limited by endogenous levels of VEGF, and it did not promote vascular leakage. VEGFR2 activation induced expression of e.g., Dll4, Notch, Apln, Apj, Klk8 and Adam12, indicating activation of Notch and apelin signalling. Adam12 and Klk8, in turn, have been shown to induce shredding of Hb-egf and Nrg1 on the cell surface, leading to activation of ErbB receptors present in cardiomyocytes (CMCs). The findings of this study demonstrate a bidirectional crosstalk between ECs and CMCs via VEGFR2, NOTCH and ErbB signalling pathways. In the study II, overexpression of VEGF-B promoted cardiac EC activation throughout the heart, detected by lineage tracing using AplnCreERT2;Tdtomato reporter mice. However, the VEGF-B -induced EC proliferation was mainly concentrated to the subendocardial myocardium, which was detected by EdU labelling of proliferating cells and staining for a proliferation marker Ki67. In the Study II, my main contribution was the development and optimization of cardiac EC isolation for single-cell RNA sequencing. In this study, the main novel finding was that VEGF-B can promote coronary vessel formation from the endocardium during development and after myocardial infarction, which was accompanied by protection from the ischemic insult. In conclusion, the Studies I and II demonstrated the coronary angiogenesis -mediated physiological cardiomyocyte growth is mediated via VEGFR2-NOTCH-ErbB pathways and involves delicate bidirectional crosstalk between ECs and CMC. In the study III, the effects of CVD risk factors aging, obesity and pressure overload, and exercise training as a physiological stimulus, was studied on cardiac endothelial cells of C57Bl/6J mice. Pure and viable cardiac ECs were isolated and analysed by RNA sequencing and various bioinformatics tools. The data demonstrated that CVD risk factors significantly decreased the number of ECs in the heart as well as the coronary vascular density and cardiac function. Importantly, exercise training improved all of these parameters compared to the sedentary control mice. The next generation RNA sequencing revealed that CVD risk factors significantly remodelled the cardiac EC transcriptome and upregulated several genes and pathways related to inflammation, oxidative stress, TGF-b signalling, vascular permeability, endothelial to mesenchymal transition (EndMT) and cellular senescence, whereas exercise training inhibited most of the same pathways, demonstrating the beneficial role of exercise training on ECs and vasculature. Exercise training also promoted blood vessel development, vascular stability and homeostasis and cell-cell junctions. The gene overlap analysis of the differentially expressed genes in the different data sets revealed SerpinH1 as one of the commonly regulated gene. SerpinH1 was significantly induced by aging and obesity and repressed by exercise training. In vitro studies in human coronary arterial endothelial cells (HCAEC) showed that overexpression of SERPINH1 increased the cell size, induced the expression of EndMT and senescence related transcripts, repressed EC genes and enhanced migration in a wound healing assay. Silencing of SERPINH1 in HCAECs, in turn, completely blocked cell proliferation and decreased collagen deposition and wound healing. As SERPINH1 has previously been linked to fibrosis in other tissues and cell types, and here we found it in all ECs throughout the human and mouse heart, it might be a potential target for the treatment of CVD. The ECs are not often considered as therapeutic targets, even though in many cases heart problems arise secondary to vascular defects (Heusch et al., 2014). The results from this thesis suggest that cardiac ECs are highly adaptive to physiological stimuli and maladaptive to pathological stressors. These findings would help to develop innovative and new therapeutic opportunities to treat heart diseases.
Alkuperäiskielienglanti
Myöntävä instituutio
  • Helsingin yliopisto
Valvoja/neuvonantaja
  • Kivelä, Riikka, Valvoja
  • Alitalo, Kari, Valvoja
Myöntöpäivämäärä29 lokak. 2021
JulkaisupaikkaHelsinki
Kustantaja
Painoksen ISBN978-951-51-7583-0
Sähköinen ISBN978-951-51-7584-7
TilaJulkaistu - 2021
OKM-julkaisutyyppiG5 Tohtorinväitöskirja (artikkeli)

Lisätietoja

M1 - 77 s. + liitteet

Tieteenalat

  • 3121 Yleislääketiede, sisätaudit ja muut kliiniset lääketieteet

Siteeraa tätä