Cellular seeding and co-culture of a three dimensional fibroblast construct
Inventors
Lancaster, Jordan • Goldman, Steve
Assignees
US Department of Veterans Affairs • University of Arizona
Publication Number
US-9976123-B2
Publication Date
2018-05-22
Expiration Date
2030-04-09
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Abstract
The present invention provides methods for cellular seeding onto three-dimensional fibroblast constructs, three-dimensional fibroblast constructs seeded with muscle cells, and uses therefore.
Core Innovation
The invention provides methods for cellular seeding onto three-dimensional fibroblast constructs (3DFC), producing three-dimensional fibroblast constructs seeded with muscle cells, and the uses of such constructs. Constructs comprise muscle cells adhered to a three-dimensional fibroblast construct where the muscle cells can spontaneously contract synchronously across the 3DFC. The muscle cells are seeded at specific densities and ratios with fibroblasts, and may include cardiomyocytes or cardiac stem cells.
The background problem addressed is the need for improved treatments for chronic heart failure (CHF) and related cardiac disorders, which have high prevalence and mortality. Previous cell injection therapies with stem or progenitor cells have had disappointing clinical results due to poor cell survival and integration. The invention aims to solve this by providing a supportive three-dimensional scaffold that enhances cell retention, growth, and functional integration on the heart.
The invention includes novel methods for seeding cells onto the 3DFC by applying a force, such as centrifugal force, to ensure efficient cell contact and adherence, overcoming poor cell retention seen in prior approaches. It also provides methods for seeding intact cell sheets onto the 3DFC to maintain cell-to-cell interactions and enhance adherence. Constructs comprising muscle cells adhered to the 3DFC exhibit spontaneous synchronized contractions, enabling functional cardiac tissue engineering.
Claims Coverage
The patent contains one independent claim focusing on a method of treating disorders characterized by lack of functioning cardiomyocytes using a specialized construct. The main inventive features cover the construct composition, cell seeding densities and ratios, layering on the 3DFC, and use of the construct in treatment.
Construct with cardiomyocytes or cardiac stem cells adhered to a three-dimensional fibroblast construct capable of synchronized contractions
The construct comprises cardiomyocytes, cardiac stem cells, or progenitors adhered to a 3DFC, capable of spontaneous synchronized contractions across the 3DFC. The seeded cells form a 1-2 cell thick layer over fibroblasts and can be adhered to the top surface, bottom surface, or both surfaces of the 3DFC, with the opposing surface lacking such cells.
Use of the construct for treating disorders characterized by lack of functioning cardiomyocytes
The method involves contacting the heart of a subject suffering from such disorders with an effective amount of the construct. The construct is used to treat a wide group of cardiac disorders including chronic heart failure, ischemia without heart failure, cardiomyopathy, dilated cardiomyopathy, congestive heart failure, angina, myocardial infarction, coronary and valvular artery diseases, and heart failure variants among others.
Specific cell seeding densities and ratios for functional treatment
The cardiomyocytes, cardiac stem cells, or progenitors are seeded at densities between 0.5×10^6 to 5×10^6 cells/cm^2 and/or present in ratios of about 1:10 to 10:1 with fibroblasts on the 3DFC. Specific dosing ranges include 1.3×10^6 to 5×10^6 cells/cm^2 and 1.5×10^6 to 3×10^6 cells/cm^2.
Attachment and application specifics of the construct to the heart
The construct can be attached to the epicardium and specifically to the left ventricle. The contacting construct at the time of application can be non-contracting. Additional treatment compounds such as thymosin beta-4, AKT1, SDF-1, and HGF may be administered in conjunction with the construct.
The independent claim covers treatment methods using a three-dimensional fibroblast construct seeded with cardiomyocytes or cardiac stem cells in defined densities and ratios, applied to the heart surfaces for synchronized contraction and therapeutic effect across a broad range of cardiac disorders.
Stated Advantages
The co-cultured 3DFC construct enhances cell survival, retention, and engraftment compared to direct injection methods.
The 3DFC is pro-angiogenic, producing growth factors that support vasculature formation and improve ischemic tissue environment.
Constructs with synchronized muscle cell contractions can provide contractile assistance as adjunct therapy or assist in myocardial functional integration.
The method permits treatment of larger heart regions and avoids washout issues seen with isolated cell injections.
The 3DFC scaffold supports mechanical stress-induced cell migration and growth, facilitating nutrient exchange through scaffold pores.
Documented Applications
Treatment of chronic heart failure and related cardiac disorders characterized by lack of functioning cardiomyocytes, including ischemic and non-ischemic heart diseases.
Promotion of healing of ischemic heart tissue by preventing maladaptive remodeling and improving myocardial perfusion and contractility.
Use in treating stable and unstable angina, myocardial infarction, cardiomyopathy including dilated cardiomyopathy, and congestive heart failure.
Application as an adjunct therapy to provide synchronous contractile support to the heart or as a cell delivery system for myocardial regeneration.
Combination use with existing cardiac treatments such as left ventricular assist devices, cardiac wrap surgery, pacemakers, and pharmaceutical agents.
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