Hydrogen sulfide provides intestinal protection during a murine model of experimental necrotizing enterocolitis

Abstract

Background: Necrotizing enterocolitis (NEC) continues to be a morbid surgical condition among preterm infants. Novel therapies for this condition are desperately needed. Hydrogen sulfide (H₂S) is an endogenous gasotransmitter that has been found to have beneficial properties. We therefore hypothesized that intraperitoneal injection of various H₂S donors would improve clinical outcomes, increase intestinal perfusion, and reduce intestinal injury in an experimental mouse model of necrotizing enterocolitis.

Methods: NEC was induced in five-day-old mouse C57BL/6 mouse pups through maternal separation, formula feeding, and intermittent hypoxic and hypothermic stress. The control group (n=10) remained with their mother and breastfed ad lib. Experimental groups (n=10/group) received intraperitoneal injections of phosphate buffered saline (PBS) vehicle or one of the following H₂S donors: (1) GYY4137, 50mg/kg daily; (2) Sodium sulfide (Na₂S), 20mg/kg three times daily; (3) AP39, 0.16mg/kg daily. Pups were monitored for weight gain, clinical status, and intestinal perfusion via transcutaneous Laser Doppler Imaging (LDI). After sacrifice on day nine, intestinal appearance and histology were scored and cytokines were measured in tissue homogenates of intestine, liver, and lung. Data were compared with Mann-Whitney and p<0.05 was considered significant.

Results: Clinical score and weight gain were significantly improved in all three H₂S-treated groups as compared to vehicle (p<0.05 for all groups). Intestinal perfusion of the vehicle group was 22% of baseline while the GYY4137 group was 38.7% (p=0.0103), Na₂S was 47.0% (p=0.0040), and AP39 was 43.0% (p=0.0018). The vehicle group had a median histology score of 2.5, while the GYY4137 group's was 1 (p=0.0013), Na₂S was 0.5 (p=0.0004), and AP39 was 0.5 (p=0.0001). Cytokine analysis of the intestine of the H₂S-treated groups revealed levels closer to breastfed pups as compared to vehicle (p<0.05 for all groups).

Conclusion: Intraperitoneal administration of H₂S protects against development of NEC by improving mesenteric perfusion, and by limiting mucosal injury and altering the tissue inflammatory response. Further experimentation is necessary to elucidate downstream mechanisms prior to clinical implementation.

Keywords: Animal model; Hydrogen sulfide; Intestine; Ischemia; Necrotizing enterocolitis; Neonatal; Premature.