https://doi.org/10.1007/s00392-025-02625-4
1Universitätsklinikum Heidelberg Klinik für Innere Med. III, Kardiologie, Angiologie u. Pneumologie Heidelberg, Deutschland; 2Heidelberg University Institute for Computational Biomedicine Heidelberg, Deutschland
Introduction:
Active TGF-β is the major driver of adverse tissue remodelling after myocardial infarction (MI). TGF-β release from the latent complex in the heart is mainly induced by the large matricellular protein thrombospondin-1 (THBS1) known for its key roles in fibrotic, inflammatory and angiogenic processes. In order to specifically inhibit the TGF-β activation side of THBS1 a small-molecule inhibitor was designed, however the therapeutic potential in vivo needs to be investigated.
Methods and results:
A bioinformatic metanalysis of three publicly available single-cell RNA datasets following cardiac injury was performed and revealed THBS1 to be one of the most upregulated fibroblast genes in mice such as in human transcriptomes. Within murine fibroblast clusters THBS1 was shown to be mainly expressed by activated matrifibrocytes characterized by the strongest estimated TGF-β pathway activity using PROGENy. We validated the pronounced induction of THBS1 expression in murine hearts on RNA and protein level peaking between day 7 and 14 after permanent and transient (ischemia-reperfusion; I/R) LAD-occlusion, respectively.
In vivo therapeutic administration of a THBS1 small-molecule inhibitor in female C57BL6/N mice by daily i.p. injections of 20mg/kg body weight compared to PBS (day 1 to day 14 after I/R) was safe and improved cardiac function robustly. Mice were randomised into control and treatment groups, which exhibited similar troponin T levels and left ventricular ejection fraction (LVEF) at day 1 and 3 after I/R induction. From day 14 onwards a significant increase in LVEF (Fig. 1, parasternal long-axis B-Mode), fractional area change (short-axis B-Mode) and global longitudinal strain was observed by awake echocardiography in treated mice (n=13) compared to PBS (n=15). The inhibitor was well tolerated and no adverse effects were observed in lungs, kidneys, liver and spleen.
Functional analysis of the THBS1-inhibitor in vitro using human ventricular cardiac fibroblasts (306V-05a, Cell Applications) demonstrated a dose-dependent reduction of Thbs1 and Tgfb1 mRNA expression levels after 72h of treatment under simultaneous TGF-β1 stimulation.
In parallel, the histologic analysis (semi-automated using QuPath) of entire cross-sections from murine cardiac tissue revealed a significant decrease of the TGF-β signal transducer phospho-Smad3 on protein level next to a reduction in collagen I deposition within the infarct region of small-molecule inhibitor treated animals at day 14 after I/R. However, expression of CD31 and THBS1 itself remained unchanged on RNA and protein level.
Conclusions:
THBS1 represents a very promising therapeutic target for adverse cardiac remodelling in humans and mice. In vivo application of a small-molecule inhibitor targeting THBS1-mediated TGF-β activation was safe and improved cardiac function following I/R in mice by reducing collagen I deposition. Further research is needed to define pharmacokinetics on the way to advance a promising translation towards human application.
Fig. 1: