C. McCulloh1, J. Olson1, Y. Wang1, J. Vu1, S. Gartner1, G. E. Besner1 1Nationwide Children’s Hospital,Columbus, OH, USA
Introduction:
Necrotizing enterocolitis (NEC) is a leading cause of gastrointestinal morbidity and mortality in premature infants. Central to NEC is the increased permeability of intestinal mucosa, leading to bacterial translocation and fulminant infection. We have previously shown that administration of stem cells (SC) can reduce the incidence and severity of NEC, however no study has compared the ability of different stem cells to restore mucosal integrity. Our goal was to investigate the efficacy of four different types of stem cells in preservation of gut barrier function during NEC.
Methods:
The following types of stem cells were compared: (1) amniotic fluid-derived mesenchymal SC (AF-MSC); (2) bone marrow-derived mesenchymal SC (BM-MSC); (3) amniotic fluid-derived neural SC (AF-NSC); and (4) enteric neural SC (E-NSC). SC lines were derived from Lewis rats, cultured, and verified via flow cytometry. MSC multipotency was confirmed via adipogenic and osteogenic differentiation. Sprague Dawley rat pups were delivered prematurely via C-section. Each pup received an intraperitoneal (IP) injection of 2×106 SC in PBS. Control pups received the same volume of PBS IP. Pups were subjected to repeated stress (hypoxia and hypothermia three times daily, hypercaloric gavage feeds every 4h, and a single enteral dose of LPS on day one) to induce experimental NEC. Control pups were breastfed and not subjected to experimental NEC. After 48h all animals received a single enteral dose of fluorescein isothiocyanante-labeled dextran (FD70, molecular weight 70,000, 750mg/ml) and sacrificed 4h later. Serum was collected and the concentration of FD70 measured in a fluorescent plate reader with filter cutoffs 492/518nm.
Results:
Compared to breastfed, unstressed pups which had intact gut barrier function and normal intestinal permeability (serum FD70 concentration 2.22 ± 0.271 µg/mL), pups exposed to experimental NEC that received PBS had impaired gut barrier function with significantly increased intestinal permeability (18.6±4.25 µg/mL, p = 0.047) (Figure 1). Compared to treatment with PBS, pups treated with SC had significantly reduced intestinal permeability: AF-MSC (9.45 ± 1.36 µg/mL, p = 0.017); BM-MSC (6.73 ± 2.74 µg/mL, p = 0.049); AF-NSC (8.052 ± 1.31 µg/mL, p = 0.0496); and E-NSC (6.60 ± 1.46 µg/mL, p = 0.033). Values are shown as mean ± SEM.
Conclusion:
Stem cells improve gut barrier function in experimental NEC. Although all four types of stem cells reduce permeability equivalently, stem cells derived from amniotic fluid (AF) may be preferable due to the availability of AF at delivery and the ease of culture expansion, enhancing the potential for clinical translation.
