Optimized design and biological validation of a mechanical stretching device to induce cardiac hypertrophy in engineered heart tissues

Cardiovascular diseases (CVD) are the leading cause of death worldwide, accounting for over 17 million death each year. Environmental and genetic factors contribute to a dramatic increase in the number of people affected by such conditions in the coming years. The study of these diseases is of fundamental importance for developing appropriate treatments. These conditions are primarily treated using surgical approaches, pharmacological approaches, or often by combining both. To develop effective pharmacological treatments that maximize therapeutic action, it is necessary to study these diseases; this process is known as drug screening. The drug screening process involves various stages to ensure that the drug can be marketed and that it has an actual therapeutic effect. After the identification of the potential clinical actions of the drugs, these are tested on models that may be in vitro or in vivo, before proceeding to clinical and preclinical development, which allows for evaluations of safety and efficacy in humans. This phase of testing, prior to experimentation on humans, is called preclinical validation. During the preclinical validation phase, the most promising drugs are tested on in vitro models and in vivo models to validate the results before proceeding to the evaluation of safety and efficacy in humans. In vitro models usually are made of isolated cells or incorporated into various types of scaffolds, while in vivo models use animal models and involve different degrees of precision, reproducibility and ethical concerns. In order to reduce the use of animal models and obtain a more predictable preclinical validation, it becomes crucial to obtain models that faithfully reproduce the pathological conditions present in the human body. In this study, a mechanical stretching device was optimized through the 3D printing of some components, overcoming certain structural and geometric limitations that were intrinsically related to its geometry. In this project engineered heart tissue (EHT) were used; specifically fibrin-based gels containing iPSC-derived cardiomyocytes. The initial phase of the study involved the production of the EHTs and their subsequent maturation under static conditions. The EHTs were divided into a control group maintained under static culture conditions and a stretching group, which was stimulated with the device for 7 days. After stretching, analyses were conducted to identify the presence of markers indicative of a pathological hypertrophic condition. The analyses performed included culture medium analysis, qPCR to determine gene expression associated with the production of specific hypertrophic markers, western blotting and immunostaining to visualize the effects of stretching on the cells. The analyses showed that the stretching device induced stress on the cells indicative of a hypertrophic condition. To increase the statistical significance of the obtained results, the tests were conducted on several batches of EHTs.