Concise Review: The Potential Use of Intestinal Stem Cells to Treat Patients with Intestinal Failure

Abstract

Intestinal failure is a rare life-threatening condition that results in the inability to maintain normal growth and hydration status by enteral nutrition alone. Although parenteral nutrition and whole organ allogeneic transplantation have improved the survival of these patients, current therapies are associated with a high risk for morbidity and mortality. Development of methods to propagate adult human intestinal stem cells (ISCs) and pluripotent stem cells raises the possibility of using stem cell-based therapy for patients with monogenic and polygenic forms of intestinal failure. Organoids have demonstrated the capacity to proliferate indefinitely and differentiate into the various cellular lineages of the gut. Genome-editing techniques, including the overexpression of the corrected form of the defective gene, or the use of CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 to selectively correct the monogenic disease-causing variant within the stem cell, make autologous ISC transplantation a feasible approach. However, numerous techniques still need to be further optimized, including more robust ex vivo ISC expansion, native ISC ablation, and engraftment protocols. Large-animal models can to be used to develop such techniques and protocols and to establish the safety of autologous ISC transplantation because outcomes in such models can be extrapolated more readily to humans. Stem Cells Translational Medicine 2017;6:666-676.

Keywords: Congenital diarrhea; Congenital tufting enteropathy; Intestinal failure; Intestinal stem cell; Microvillus inclusion disease.

Figure 1

Intestinal crypt. Small bowel epithelium is organized into villus and crypts. Lgr5⁺ crypt base columnar cells are intercalated with Paneth cells at the crypt base and continuously generate rapidly proliferating transit‐amplifying (TA) cells, which occupy the remainder of the crypt. TA cells differentiate into the various functional cells on the villi (enterocytes, goblet, enteroendocrine, and Paneth cells). The +4 label retaining cells (which occupy the fourth position from the crypt base) can restore the Lgr5⁺ stem cell compartment following injury.

Figure 2

Epithelial regeneration in small bowel. (A): During homeostasis, Lgr5⁺ CBCCs at the crypt base self‐renew and differentiate heterogeneous epithelial components. The markers expressed in the cell are indicated in the boxes. (B): Acute injury results in the loss of the proliferating Lgr5⁺ stem cells but preserves the relatively resistant Paneth cell precursors, +4 stem cells, and niche cells. Surviving +4 cells function as quiescent stem cells to rapidly regenerate the Lgr5⁺ crypt base columnar cell pool and restore epithelial renewal. Surviving Dll1+ secretory progenitors or other early TA cells fall back into the surviving niche at the crypt base and consequently convert into Lgr5⁺ stem cells to restore epithelial renewal. Abbreviations: M, microfold; TA, transit‐amplifying.

Figure 3

Signaling pathways regulating intestinal stem cells (ISCs). (A): Wnt signaling positively regulates the self‐renewal and proliferation of ISCs. (B): BMP signaling promotes differentiation of the intestinal epithelium. (C): Notch signaling regulates the proliferation of ISCs and differentiation of secretory progenitors. (D): EGF signaling regulates the proliferation of ISCs and transit‐amplifying cells by activation of the Ras/Raf/Mek/Erk signaling axis. Abbreviations: BMP, bone morphogenetic protein; CSL, CBF1/Suppressor of Hairless/LAG‐1; EGF, epidermal growth factor; EGFR, epidermal growth factor receptor; NICD, Notch intracellular domain.

Figure 4

Reconstituted intestinal enteroid culture. Small bowel crypts or Lgr5⁺ stem cells are isolated and plated into a Matrigel plug with culture medium containing epidermal growth factor, Noggin, and R‐spondin. Enterosphere (spheroid) was generated during days 1–4 after initial culturing. After days 5–7, spheroid initiates bud formation and forms reconstituted intestinal enteroid structure (scale bar = 100 μm). Abbreviation: FACS, fluorescence‐activated cell sorting.

Figure 5

Outline of intestinal stem cell (ISC) transplantation as a treatment for patients with intestinal failure caused by monogenic disease. The intestinal crypts can be isolated from the mucosa of patients by using endoscopic biopsy. Isolated crypts are dissociated into single cell suspension, and Lgr5⁺ ISCs are identified by using FACS. In an alternative method, ISCs can be obtained by spheroid, which consists of ISCs and its progenitors. Gene editing is intended to restore the function of ISCs, including the overexpression of the corrected form of the defective gene or selective correction of the disease‐causing mutation. The gene‐corrected ISCs are expanded in an in vitro 3D culture system and transplanted into the segment of the patient's small bowel, where pathologic ISCs are ablated. Successfully engrafted ISCs can differentiate and reinstate all lineages of epithelium, with normal absorptive and secretory function. Abbreviations: 3D, three‐dimensional; FACS, fluorescence‐activated cell sorting.