An engineered heart patch from embryonic stem cells

The heart does not have the capacity to self-repair, so loss of muscle tissue following a heart attack leaves it permanently weakened. Many researchers are aiming to use cells to repair this damage, but the exact type of cell and the way to introduce them into the heart is a matter of great debate. In the present study we will use cardiac muscle cells (myocytes) grown from embryonic stem cells. These have the advantage that there is a steady supply, since the ES cells will divide continuously in culture before being directed into different cell types. We are able to grow human and mouse ES cells and convert them to beating myocytes: in both cases we use existing cell lines. Most studies have introduced new cells into the heart by injecting them in a fluid suspension. However, many cells are lost within the first week after injection. If the heart attack was not recent, scar tissue develops, and cells injected into this do not receive a blood supply or connect electrically to the rest of the heart. We therefore want to grow the cells on a patch of material and graft it to the heart. This will overcome the problems associated with cell injection and will have the added advantage that the patch can prevent the scar from stretching. Containing the heart within a scaffold to prevent it from expanding is already being tried as a therapy by itself. We intend to develop a suitable material which can be used as a support for the growth of ES cells and then sewn into place on the heart. It will need to have properties such as biocompatibility, strength and the ability to withstand repeated stretch. It will be designed to degrade slowly when in the body, so that the patch material will disappear as the grafted cells integrate into the heart. We will first test the ability of ES cells to grow and become myocytes when cultured on the patch. A detailed profile of the myocytes can be built up in terms of shape and size, contraction and relaxation characteristics, responses to hormones and drugs, and electrical coupling. These can be compared with our normal cultures, and with adult human and mouse cells which we also study. Stretching and electrical stimulation will be applied to the patch with cells, to try to develop the heart muscle and stimulate the growth of blood vessels. The effect of the patch itself will be tested on an animal model, where a myocardial infarction (heart attack) is produced in rat. This model mimics the human disease in many aspects. We will determine whether grafting of the patch is able to slow the development of heart failure and prevent the heart from expanding. When the patch is optimised for both cell growth and grafting, the combined effect of the patch plus cells will be studied. The key things we will be looking for are the integration of the grafted cells into the heart, the development of electrical connections, the growth of blood vessels and an improvement in the function of the heart. The project will require the combined expertise of the biologists of the National Heart and Lung Institute and the engineers of the Department of Materials for biomaterial engineering, ES cell culture, myocyte generation, myocyte characterisation and animal surgery.

The heart has little capacity to self-repair, so that loss of muscle cells (cardiomyocytes) following a myocardial infarction will precipitate a spiral of decline into heart failure. Although this can be slowed by currently therapy it cannot be reversed, hence the interest in repopulating the muscle with exogenous cells. The strategy that will be used in the present project is to develop a patch of biodegradable material with attached embryonic stem cell-derived cardiomyocytes, and to fasten it over the scarred portion of a rat heart with prior myocardial infarction. Use of autologous cells has many advantages, but neither skeletal myoblasts nor adult bone-marrow derived stem cells have been reliably shown to produce new muscle. Embryonic stem cells (ES), however, do generate cardiomyocytes and we are able to produce cultures of beating myocytes from either mouse or human ES lines. Cells may be introduced as suspensions, and we are studying this is a separate project. However, we believe that grafting of a patch can overcome some of the problems of cell survival and maturation, and can additionally provide an advantage because of the bracing effect on the scar. Devices which physically prevent scar expansion and cardiac dilatation have been shown to reduce the deterioration of the infarcted heart and are presently in clinical trials. The patch will be developed from a class of PED (polyester-dimer of dilinoleic acid) materials formulated as hybrid nanocomposites, where biocompatible polymer matrices and inorganic nanoparticle fillers such as TiO2 are combined in a composite biomaterial. These materials were developed within the Materials group and have the required characteristics of biocompatibility and flexibility, yet are highly resistant to fatigue loading. This is important because in our proposed application the engineered biomaterial will require repeated stretches in vivo and in vitro. The other properties that will be required include i) Non-immunogenic, no inclusion of animal proteins, ii) biodegradability over a period of 1-2 months, iii) possibility to be attached by suture to the heart. These materials will be first tested for their ability to support the ES culture in vitro without altering growth of undifferentiated cells, degree of differentiation into cardiomyocytes/blood vessels, and contractile or morphological properties of cardiomyocytes. Part of the project will be to optimise the culture conditions of the ES cells to enrich the proportion of cardiomyocytes while allowing the possibility for vascularisation before implantation. Material plus differentiated ES cells will be subject to repeated cyclical stretch or electrical stimulation to develop the phenotype of the cells within the patch. Development will be followed by contractile studies and histology. Material alone will then be grafted over the 2 week old scar on an infarcted rat heart in vivo. The NHLI has a background of expertise in animal models of heart disease and complex surgical techniques. Progression of cardiac remodelling in terms of heart size and function (pressure-volume loops using a Millar catheter) will be followed for a further 4 weeks. Iterative processes will refine the material, the ES culture, the combined patch and the method for attachment to the heart. Finally the material plus cells will be attached, and the function of the heart together with the degree of integration and vascularisation of the grafted cells will be assessed. Results can be compared to the parallel project in which cells are injected in suspension. This study will provide a large amount of information that will advance the understanding of the optimal strategy for cardiac repair.