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US 12,631,623 B2Granted
Heart tissue model
AssigneeImba—Institut FüR Molekulare Biotechnologie GmbH
InventorsSasha Mendjan, Stefan Jahnel, Pablo Hofbauer, Nora Papai
Life of this patent
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Abstract
The present invention provides a heart tissue model of at least 60% cardiac cells, wherein the cardiac cells surround an inner cavity, wherein the cardiac cells are selected from cardiomyocytes, endocardial cells and epicardial cells; method for the generation of such a tissue model and uses thereof.
Claims22
A method of generating a heart organoid comprising the steps of
- a1) providing pluripotent stem cells;
- and
- b1) inducing mesoderm differentiation in the presence of a WNT activator and/or GSK3-beta inhibitor, and further in the presence of a PI3 kinase inhibitor, in a 3D culture in a low attachment culture, wherein the cells bind to each other instead of a culturing vessel to form aggregates of the cells, thereby producing an aggregate of mesoderm cells;
- or
- a2) providing pluripotent stem cells in a 2D culture;
- and
- b2) inducing mesoderm differentiation in a 3D culture in a low attachment culture with at least 100 pluripotent cells from step a2), in the presence of CHIR99021, wherein CHIR99021 and/or an optional PI3 kinase inhibitor are in an amount sufficient to differentiate the pluripotent stem cells to exit pluripotency in an amount at least 90% of the pluripotent stem cells within 40 hours of starting induction, wherein CHIR99021 is in a concentration of at least 9 μM, wherein the pluripotent stem cells are further treated with fibroblast growth factor and/or albumin, thereby producing an aggregate of mesoderm cells;
- and
- c) differentiating the mesoderm cells of step b1) or b2) into cardiac cells in a 3D culture in a low attachment culture, wherein the cells aggregate with each other instead of binding to a culturing vessel to form aggregates of the cells, in the presence of cardiac differentiation factors and in the absence of a WNT activator and/or in the presence of a WNT antagonist, for at least 3 days, for cardiac mesoderm formation and formation of an inner cavity.
6 more elements
A method of generating a heart organoid comprising the steps of:
- a) providing pluripotent stem cells;
- b) inducing mesoderm differentiation in the presence of a WNT activator and/or GSK3-beta inhibitor, and further in the presence of a PI3 kinase inhibitor, in a 3D culture in a low attachment culture, wherein the cells bind to each other instead of a culturing vessel to form aggregates of the cells, thereby producing an aggregate of mesoderm cells;
- and
- c) differentiating the mesoderm cells of step b) into cardiac cells in a 3D culture in a low attachment culture, wherein the cells aggregate with each other instead of binding to a culturing vessel to form aggregates of the cells, in the presence of cardiac differentiation factors and in the absence of a WNT activator and/or in the presence of a WNT antagonist, for at least 3 days, for cardiac mesoderm formation and formation of an inner cavity.
1 more element
A method of generating a heart organoid comprising the steps of:
- a) providing pluripotent stem cells in a 2D culture;
- and
- b) inducing mesoderm differentiation in a 3D culture in a low attachment culture, wherein a culture comprises at least 100 pluripotent cells from step a), in the presence of CHIR99021, wherein CHIR99021 and/or an optional PI3 kinase inhibitor are in an amount sufficient to differentiate the pluripotent stem cells to exit pluripotency in an amount at least 90% of the pluripotent stem cells within 40 hours of starting induction, wherein CHIR99021 is in a concentration of at least 9 μM, wherein the pluripotent stem cells are further treated with fibroblast growth factor and/or albumin, thereby producing an aggregate of mesoderm cells;
- and
- c) differentiating the mesoderm cells of step b) into cardiac cells in a 3D culture in a low attachment culture, wherein the cells aggregate with each other instead of binding to a culturing vessel to form aggregates of the cells, in the presence of cardiac differentiation factors and in the absence of a WNT activator and/or in the presence of a WNT antagonist, for at least 3 days, for cardiac mesoderm formation and formation of an inner cavity.
2 more elements
Dependencies are inferred from the claim text.