Treatment of necrotizing enterocolitis by conditioned medium derived from human amniotic fluid stem cells

Abstract

Purpose Necrotizing enterocolitis (NEC) is one of the most distressing gastrointestinal emergencies affecting neonates. Amniotic fluid stem cells (AFSC) improve intestinal injury and survival in experimental NEC but are difficult to administer. In this study, we evaluated whether conditioned medium (CM) derived from human AFSC have protective effects. Methods Three groups of C57BL/6 mice were studied: (i) breast-fed mice as control; (ii) experimental NEC mice receiving PBS; and (iii) experimental NEC mice receiving CM. NEC was induced between post-natal days P5 through P9 via: (A) gavage feeding of hyperosmolar formula four-time a day; (B) 10 minutes hypoxia prior to feeds; and (C) lipopolysaccharide administration on P6 and P7. Intra-peritoneal injections of either PBS or CM were given on P6 and P7. All mice were sacrificed on P9 and terminal ileum were harvested for analyses. Results CM treatment increased survival and reduced intestinal damage, decreased mucosal inflammation (IL-6; TNF-α), neutrophil infiltration (MPO), and apoptosis (CC3), and also restored angiogenesis (VEGF) in the ileum. Additionally, CM treated mice had increased levels of epithelial proliferation (Ki67) and stem cell activity (Olfm4; Lgr5) compared to NEC+PBS mice, showing restored intestinal regeneration and recovery during NEC induction. CM proteomic analysis of CM content identified peptides that regulated immune and stem cell activity. Conclusions CM derived from human AFSC administered in experimental NEC exhibited various benefits including reduced intestinal injury and inflammation, increased enterocyte proliferation, and restored intestinal stem cell activity. This study provides the scientific basis for the use of CM derived from AFSC in neonates with NEC.

A Review of the Use of Extracellular Vesicles in the Treatment of Neonatal Diseases: Current State and Problems with Translation to the Clinic

“…Various treatment strategies have been proposed for NEC, but recently, MSC therapy has become the preferred option due to its pronounced therapeutic efficacy. There is evidence that therapeutic effects can be achieved not only via the transplantation of stem/progenitor cells but also through the use of a conditioned culture medium containing cell secretomes and EVs [ 194 , 195 , 196 ]. In an experimental model of NEC, it was shown that bone marrow-derived MSCs and their exosomes contributed to a significant reduction in the development of the pathological process and maintained the integrity of the intestinal mucosa and its physiological permeability.…”

Amniotic Fluid: A Perspective on Promising Advances in the Prevention and Treatment of Necrotizing Enterocolitis

“…Treatment with CM was associated with increased stem cell activity and recovery from NEC, similar to the effect of AFSCs-derived exosomes. While the composition of this CM differs from the physiological secretome as produced by AFSCs, AFSC-CM holds important cellular information (e.g., mRNA, miRNA, DNA, proteins, and EVs) ( 39 ).…”

Amniotic fluid stem cells: A novel treatment for necrotizing enterocolitis

“…In NEC, a decrease in Treg cells disrupts the equilibrium and increases proinflammatory Th17 cell response ( 44 ). However, when AFSCs are administered, TLR4-mediated signaling is inhibited ( 44 ) and inflammatory markers such as TNFα and IL-6 are significantly decreased ( 45 ).…”

“…A recent study demonstrated that administration of human AFSC conditioned medium (hAFSC-CM) improved NEC survival and reduced intestinal injury ( 45 ). Specifically, intestinal mucosal inflammation and epithelial apoptosis were significantly reduced.…”