Production of tissue-engineered intestine from expanded enteroids

Abstract

Background Short bowel syndrome (SBS) is a life-threatening condition with few solutions. Tissue engineered intestine (TEI) is a potential treatment, but donor intestine is a limiting factor. Expanded epithelial surrogates termed enteroids may serve as a potential donor source. Materials and Methods To produce TEI from enteroids, crypts were harvested from mice and enteroid cultures established. Enteroids were seeded onto polymer scaffolds using Matrigel or culture medium, and implanted in immunosuppressed mice for 4 weeks. Histology was analyzed using Periodic Acid Schiff (PAS) staining and immunofluorescence (IF). Neomucosa was quantified using ImageJ software. To determine if TEI could be produced from enteroids established from small intestinal biopsies, 2×2mm pieces of jejunum were processed for enteroid culture, enteroids were expanded and seeded onto scaffolds, and scaffolds implanted for 4 weeks. Results Enteroids in Matrigel produced TEI in 15/15 scaffolds, whereas enteroids in medium produced TEI in 9/15 scaffolds. Use of Matrigel led to more neomucosal surface area compared to media (10,520±2,905 μm vs. 450±127 μm, p<0.05). Histologic examination confirmed the presence of crypts and blunted villi, normal intestinal epithelial lineages, intestinal subepithelial myofibroblasts, and smooth muscle cells. Crypts obtained from biopsies produced an average of 192±71 enteroids. A single passage produced 685±58 enteroids, which was adequate for scaffold seeding. TEI was produced in 8/9 scaffolds seeded with expanded enteroids. Conclusions Enteroids can be obtained from minimal starting material, expanded ex vivo, and implanted to produce TEI. This method shows promise as a solution to the limited donor intestine available for TEI production in patients with SBS.

Intestinal epithelial organoids fuse to form self-organizing tubes in floating collagen gels

“…Larger models of the intestinal epithelium range from air-liquid interface cultures with stroma (Ootani et al, 2009), over cells seeded on biological or synthetic scaffolds (Shaffiey et al, 2016;Torashima et al, 2016) to complex tissue-engineered intestines and living bioreactors (Levin et al, 2013;Grant et al, 2015;Cromeens et al, 2016). Progress in this area has indeed been remarkable, but all cited methods are laborious and technically difficult to perform in laboratories that lack specific tissue-engineering expertise.…”

New Insights and Interventions for Short Bowel Syndrome

“…Many prior studies have explored the feasibility of growing the different cells of the intestinal tract and loading these cells on biomaterials in the physical shape of intestine for in vivo implantation [17,33,34]. The concept is to utilize autologous tissue to obviate the need for immunosuppression.…”

“…In order to produce TESI, the starting cells would sacrifice too much of the already ‘short’ bowel in these patients. Therefore, one recent study proved the ability to convert small jejunal biopsies into sufficient cellular material to load onto a scaffold to create TESI [34]. Thus, the process of cellular expansion sufficient to generate new bowel is becoming possible from limited starting material.…”

A Simple Modification Method to Obtain Anisotropic and Porous 3D Microfibrillar Scaffolds for Surgical and Biomedical Applications

“…These meshes (Biofelt and GORE BIO‐A) are made by melt spinning biodegradable polymers like polyglycolic acid (PGA) into macroscopic sheets, which are further processed by carding and needle punching to increase mesh integrity. These medically approved polymeric meshes are attractive scaffolding material for tissue engineering and surgical applications and have been used in clinical practice for several years . For instance, in situ tendon regeneration, homeostatic cardiac, and abdominal wall surgeries were reported that use PGA meshes as scaffolding material.…”

“…In following sections, we present the detailed stepwise procedure of coating the primary polymer, inducing fiber alignment, seeding, and culturing the cells and the final evaluation of cell alignment and tissue formation. For a proof‐of‐concept demonstration, we utilized commercially available PGA scaffolds (Biofelt) that found several clinical applications in the field of surgery and tissue engineering and processed it without further modifications as described below.…”