Tissue engineered intestine

Abstract

The invention provides for engineered intestinal construct and methods of making these constructs. The invention also provides for methods of treating short bowel syndrome or methods of repairing an intestine after resection comprising inserting an engineered intestinal construct into the intestine of a subject in need.

What is claimed: 
     
         1 . An engineered intestine construct comprising a nanofiber scaffold seeded with neural stem cells, smooth muscle cells and intestinal stem cells, and wherein the nanofiber scaffold comprises HB-EGF polypeptide or a fragment thereof. 
     
     
         2 . An engineered intestine construct comprising a nanofiber scaffold seeded with neural stem cells, smooth muscle cells and intestinal stem cells, wherein at least one of the neural stem cells, smooth muscle cells or intestinal stem cells overexpress HB-EGF polypeptide or a fragment thereof. 
     
     
         3 . An engineered intestine construct comprising a multilayer nanofiber scaffold,
 wherein the multilayer nanofiber scaffold comprises at least an inner layer and an outer layer,   wherein the outer layer comprises neural stem cells and smooth muscle cells, and   wherein the inner layer comprises intestinal stem cells.   
     
     
         4 . The engineered intestine construct of  claim 3  wherein at least of the layers comprises HB-EGF polypeptide or a fragment. 
     
     
         5 . The engineered intestine construct of  claim 3  or  4  wherein the construct further comprises at least one middle layer. 
     
     
         6 . The engineered intestine construct of any one of  claims 1 - 5  wherein the nanofiber scaffold comprises Poly(glycolic acid)(PGA) nanofibers, Poly(ε-caprolactone) (PCL) nanofibers, Poly(-caprolactone-co-lactic acid) (PLC) nanofibers, Poly(L-lactic acid) (PLLA) nanofibers, Poly(D-lactic acid-co-glycolic acid) (PDLGA) nanofibers, Poly(D-lactic acid-co-glycolic acid) (PLGA) nanofibers, Polyurethane (PU) nanofibers, Polydioxanone (PDO) nanofibers or a combination thereof. 
     
     
         7 . The engineered intestine construct of any one of  claims 3 - 6  wherein the construct comprises a layer of macrofibers between two layers. 
     
     
         8 . The engineered intestine construct of  claim 7  where the layer of macrofibers comprises PGA. 
     
     
         9 . A method of generating an engineered intestine construct comprising
 a) preparing a nanofiber scaffold by electrospinning a polymer to a target fiber diameter and porosity,   b) embedding an HB-EGF polypeptide or fragment thereof on the scaffold,   c) seeding the scaffold with intestinal stem cells, neural stem cells and smooth muscles cells, and   d) culturing the cells in the scaffold to form a construct that will form a mature intestine upon insertion into a subject.   
     
     
         10 . The method of  claim 10  wherein the culturing step is carried out in a bioreactor. 
     
     
         11 . The method of  claim 10  or  11  wherein the nanofiber scaffold comprises at least an outer and an inner layer. 
     
     
         12 . The method of  claim 11  wherein the nanofiber scaffold further comprises a middle layer. 
     
     
         13 . The method of  claims 9 - 12  wherein the intestinal stem cells are seeded on the inner layer. 
     
     
         14 . The method of any one of  claims 9 - 13  wherein the neural stem cells and smooth muscle cells are seeded on the outer layer. 
     
     
         15 . The method of any one of  claims 9  to  14  wherein the polymer is Poly(glycolic acid)(PGA) nanofibers, Poly(ε-caprolactone) (PCL) nanofibers, Poly(-caprolactone-co-lactic acid) (PLC) nanofibers, Poly(L-lactic acid) (PLLA) nanofibers, Poly(D-lactic acid-co-glycolic acid) (PDLGA) nanofibers, Poly(D-lactic acid-co-glycolic acid) (PLGA) nanofibers or Polyurethane (PU) nanofibers, Polydioxanone (PDO) nanofibers or a combination thereof. 
     
     
         16 . The method of any one of  claims 9 - 15  wherein the polymer is PDLGA or PGA. 
     
     
         17 . A method of treating short bowel syndrome in a subject comprising attaching the engineered intestine construct of any one of  claims 1 - 9  under conditions wherein the construct will implant within the intestine of the subject. 
     
     
         18 . A method of repairing the intestine of a subject undergoing intestinal resection comprising attaching the engineered intestine construct of any one of  claims 1 - 8  under conditions wherein the construct will implant within the intestine of the subject. 
     
     
         19 . The method of  claim 17  or  18 , wherein the subject is suffering from inflammatory bowel disease, trauma, mesenteric vascular disease, vovlulus, congenital atresias, neonatal necrotizing enterocolitis, Crohn's disease, ischemia, intestinal blockage, bowel obstruction, regional ileitis, regional enteritis, colorectal cancer, carcinoid tumor, Merkel's diverticulum, precancerous polyps, diverticulitis, intestinal bleeding, intussusceptions, or ulcerative colitis. 
     
     
         20 . A method of enriching a cell sample for a particular cell type comprising
 contacting a cell sample with multiple sieve membranes wherein the membranes are aligned in descending order according to pore size, wherein the cell sample contacts the membrane with the largest pore size first and wherein the cell sample comprises multiple cell types,   filtering the cell sample through the membranes and   recovering the enriched cell sample.   
     
     
         21 . The method of  claim 20  wherein the enriched cell sample is enriched for intestinal stem cells in crypts.