Methods of bioengineering internal anal sphincter constructs

A method of generating an innervated internal anal sphincter construct, comprising the following steps:

• the step of obtaining an anorectum tissue cell biopsy from a subject;
• the step of obtaining an intestinal tissue cell biopsy from the subject;
• the step of isolating smooth muscle cells from the anorectum tissue cell biopsy;
• the step of culturing the isolated smooth muscle cells for about four weeks until the number of smooth muscle cells reaches at least about 10 million cells;
• the step of isolating neural progenitor cells from the intestinal tissue cell biopsy;
• the step of culturing the isolated neural progenitor cells for about four weeks until the number of neural progenitor cells reaches at least about five million cells;
• the step of preparing a smooth muscle cell gel by suspending the isolated smooth muscle cells in a gel matrix, to provide a first gel layer which is a smooth muscle gel layer;
• the step of preparing a neural progenitor cell gel by suspending the isolated neural progenitor cells in a gel matrix to provide a second gel layer which is a neural progenitor cell gel layer;
• the step of depositing the first smooth muscle gel layer around a central post of a culture plate or in a mold which is in a substantially circular shape, followed by depositing the second neural progenitor cell gel layer on top of the first smooth muscle cell layer around a central post of a culture plate or in a mold which is in a substantially circular shape to form a dual layer gel matrix, wherein the first smooth muscle gel layer and the second neural progenitor cell gel layer of the dual layer gel matrix are in contact;
• the step of contacting the dual-layer gel matrix with a differentiation media to induce differentiation of the neural progenitor cells into functional neurons,
• thereby forming the innervated internal anal sphincter construct having directionally oriented smooth muscle cells, further wherein the innervated internal anal sphincter construct is capable of treating fecal incontinence when implanted in a subject.