The Effects of Necrotizing Enterocolitis and Short Chain Fatty acids on the Developing Brain in Mice
Presenter Status
Fellow
Abstract Type
Research
Primary Mentor
Sampath, Venkatesh
Start Date
3-5-2022 11:30 AM
End Date
3-5-2022 1:30 PM
Presentation Type
Poster Presentation
Description
Background: Necrotizing enterocolitis (NEC) is a devastating disease in premature infants characterized by acute intestinal inflammation and necrosis with a mortality of 15%-30%. Approximately 50% of infants that survive NEC will develop neurodevelopmental defects[1]. Studies on the effects of NEC on the neonatal brain are limited, however it’s postulated that neurodevelopmental damage is induced via cytokine activation of toll-like-receptor 4 (TLR4) by high mobility group box protein 1 (HMGB1) on the microglia Accumulation of reactive oxygen and demyelination contribute to neuronal damage. Therapies that limit the development of NEC can potentially also limit the neuroinflammatory changes seen in NEC[2]. We hypothesized that enteral supplementation with butyrate, a short chain fatty acid, attenuates NEC-induced brain injury in experimental NEC. The proposed mechanism is that exogenous butyrate downregulates inflammation.
Objectives: (1)To evaluate cell-type and region-specific impact of NEC on cell death, proliferation and inflammation in an experimental model of NEC. (2) To evaluate the efficacy of enteral sodium butyrate supplementation on ameliorating NEC-associated neurological alterations .
Methods: A standardized NEC mouse model combining enteral formula feeding, LPS and 2x/daily hypoxia was used. Mice were randomized into three groups: control, NEC, and sodium butyrate pre-treated NEC. NEC scoring was done on hematoxylin and eosin staining(HE) staining of terminal ileum. The RNA expression in intestinal and brain samples of TLR4 and inflammatory cytokines (IL6, IL1β, and tumor necrosis α ) was determined by qRT-PCR. Neural damage and neuro-inflammation was evaluated by immunofluorescent staining with antibodies against markers of microglia, activated astrocytes, cell death and proliferation on P10 section brains. Samples were statistically compared in GraphPad software.
Results: Pathological NEC induced intestinal injury was attenuated by butyrate supplementation (Figure 1A-B). Quantitative PCR and western blot analyses on RNA isolated from the terminal ileum, cerebral cortex, hippocampus and cerebellum indicate that expression of the proinflammatory cytokines IL-6, TNFα, and TLR4 were elevated . Intestinal tissue analysis showed decrease levels of inflammatory cytokines IL6, IL8 (CXCL1), IL-1β in butyrate exposed pups. (Figure 1C) . In the cortex region increase expression of TLR-4, IL6, IFN-1B was seen in NEC compared to NEC+ Butyrate group (Figure 3A). Similar findings noted in the cerebellum with increase expression of TNFα a, IL6, ), IL-1β, and TLR-4 in NEC group (Figure 3B). The NEC group had increase expression of TNFα in hypothalamus region (Figure 3C). Overall, there was increase expression of cytokines in brain of NEC pups compared to control and NEC + butyrate indicating that a neuroinflammatory response had been induced by NEC. Early analysis suggest that butyrate ameliorates inflammatory cytokine expression in brain and (Figure 3).
Conclusion: NEC was associated with increased levels of inflammation in the cortex and increase inflammation and apoptosis in cerebellum. The severity of neuroinflammation was associated with severity of NEC. Pilot studies of butyrate show significant reduction in inflammatory markers . Our findings suggest that administration of butyrate decreases the degree of inflammation and can serve as therapeutic approach to decreasing NEC rates and limiting the neurological damage.
MeSH Keywords
NEC; Necrotizing Enterocolitis; Brain Development; Intestinal Inflammation
Additional Files
The Effects of Necrotizing Enterocolitis and Short Chain Fatty acids on the Developing Brain in Mice.pdf (579 kB)Abstract
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The Effects of Necrotizing Enterocolitis and Short Chain Fatty acids on the Developing Brain in Mice
Background: Necrotizing enterocolitis (NEC) is a devastating disease in premature infants characterized by acute intestinal inflammation and necrosis with a mortality of 15%-30%. Approximately 50% of infants that survive NEC will develop neurodevelopmental defects[1]. Studies on the effects of NEC on the neonatal brain are limited, however it’s postulated that neurodevelopmental damage is induced via cytokine activation of toll-like-receptor 4 (TLR4) by high mobility group box protein 1 (HMGB1) on the microglia Accumulation of reactive oxygen and demyelination contribute to neuronal damage. Therapies that limit the development of NEC can potentially also limit the neuroinflammatory changes seen in NEC[2]. We hypothesized that enteral supplementation with butyrate, a short chain fatty acid, attenuates NEC-induced brain injury in experimental NEC. The proposed mechanism is that exogenous butyrate downregulates inflammation.
Objectives: (1)To evaluate cell-type and region-specific impact of NEC on cell death, proliferation and inflammation in an experimental model of NEC. (2) To evaluate the efficacy of enteral sodium butyrate supplementation on ameliorating NEC-associated neurological alterations .
Methods: A standardized NEC mouse model combining enteral formula feeding, LPS and 2x/daily hypoxia was used. Mice were randomized into three groups: control, NEC, and sodium butyrate pre-treated NEC. NEC scoring was done on hematoxylin and eosin staining(HE) staining of terminal ileum. The RNA expression in intestinal and brain samples of TLR4 and inflammatory cytokines (IL6, IL1β, and tumor necrosis α ) was determined by qRT-PCR. Neural damage and neuro-inflammation was evaluated by immunofluorescent staining with antibodies against markers of microglia, activated astrocytes, cell death and proliferation on P10 section brains. Samples were statistically compared in GraphPad software.
Results: Pathological NEC induced intestinal injury was attenuated by butyrate supplementation (Figure 1A-B). Quantitative PCR and western blot analyses on RNA isolated from the terminal ileum, cerebral cortex, hippocampus and cerebellum indicate that expression of the proinflammatory cytokines IL-6, TNFα, and TLR4 were elevated . Intestinal tissue analysis showed decrease levels of inflammatory cytokines IL6, IL8 (CXCL1), IL-1β in butyrate exposed pups. (Figure 1C) . In the cortex region increase expression of TLR-4, IL6, IFN-1B was seen in NEC compared to NEC+ Butyrate group (Figure 3A). Similar findings noted in the cerebellum with increase expression of TNFα a, IL6, ), IL-1β, and TLR-4 in NEC group (Figure 3B). The NEC group had increase expression of TNFα in hypothalamus region (Figure 3C). Overall, there was increase expression of cytokines in brain of NEC pups compared to control and NEC + butyrate indicating that a neuroinflammatory response had been induced by NEC. Early analysis suggest that butyrate ameliorates inflammatory cytokine expression in brain and (Figure 3).
Conclusion: NEC was associated with increased levels of inflammation in the cortex and increase inflammation and apoptosis in cerebellum. The severity of neuroinflammation was associated with severity of NEC. Pilot studies of butyrate show significant reduction in inflammatory markers . Our findings suggest that administration of butyrate decreases the degree of inflammation and can serve as therapeutic approach to decreasing NEC rates and limiting the neurological damage.
