O2200: Mesenchymal Stem Cells Improve Intestinal Injury in Necrotizing Enterocolitis through SMG1 Upregulation: A Premature Piglet Study

Background: Necrotizing enterocolitis (NEC) is a devastating gastrointestinal condition primarily affecting premature infants, characterized by intestinal inflammation and necrosis, leading to significant morbidity and mortality. Current treatment options are limited, creating an urgent need for innovative therapeutic approaches. Mesenchymal stem cell (MSC) therapy has gained attention for its potential to modulate inflammation and promote tissue repair in NEC. The therapeutic benefit of stem cells in NEC is well documented in the literature, but the mechanisms underlying this therapy are less understood. In this study, we utilize a premature piglet model to investigate the role of the PI3K/AKT/eNOS signaling pathway in NEC. We hypothesize that this pathway is downregulated in NEC and that the administration of MSCs will restore or protect its activity, thereby mitigating intestinal injury.

Methods: Premature piglets were delivered via cesarean section at 103/115 days gestation. Control piglets were sacrificed on post-natal day 1. NEC was induced in the remaining piglets with TPN starting on day 1 and hypertonic feeding via orogastric tube on day 3. MSCs were prepared at 1 million cells/kg in 1.2mL of PBS and injected intraperitoneally on day 3 (NEC-MSC group). Untreated piglets were injected with PBS alone (NEC-PBS group). Piglets were monitored hourly for clinical deterioration and euthanized early if criteria were met. The study concluded after 5 days. During necropsy, the terminal ileum was isolated for real-time polymerase chain reaction (RT-PCR) analysis. Key genes in the PI3K/AKT/eNOS pathway were investigated. A one-way ANOVA and post-hoc multiple comparisons test were utilized to analyze the relative gene expression in each group, p< 0.05 was considered significant.

Results: There was no significant difference in NOS3 (gene that codes for eNOS), VEGF (an activator of eNOS), or AKT1 expression between the experimental groups. Caveolin-1 (CAV-1), an inhibitor of eNOS, showed significant downregulation in NEC that was not restored with MSC treatment. SMG1(serine/threonine protein kinase), which encodes a PI3K-related protein, showed significant downregulation in NEC piglets compared to controls (p=0.006). MSC treatment significantly upregulated SMG1 expression in NEC piglets (p=0.045). There was no significant difference in SMG1 expression between control piglets and those treated with MSCs [Figure 1].

Conclusion: Our lab is the first to demonstrate the therapeutic benefits of MSCs in a premature piglet model of NEC. In this study, we investigated the mechanisms underlying MSC therapy by analyzing gene expression changes in the PI3K/AKT/eNOS pathway. Specifically, we identified that SMG1, a gene encoding a key regulator of PI3K, exhibited a pattern of dysregulation in NEC that was restored following MSC treatment. These findings suggest that SMG1 may serve as a potential genetic target through which MSCs mitigate intestinal damage in NEC, providing new insights into the therapeutic mechanisms of stem cell therapy for this devastating neonatal disease.