Brandy L. Frost

The importance of pro-inflammatory signaling in neonatal necrotizing enterocolitis.

Despite modern medical advances, necrotizing enterocolitis (NEC) remains a significant cause of morbidity and mortality in neonatal intensive care units, affecting 10% of premature neonates born weighing less than 1500 g. Although many advances have been made in the understanding of NEC, the etiology and pathophysiology remain incompletely understood, and treatment is limited to supportive care. In recent years, many studies have evaluated the inflammatory cascade that is central to the disease process, and research is ongoing into strategies to prevent and/or ameliorate neonatal NEC. In this review, we examine the key points in the signaling pathways involved in NEC, and potential strategies for prevention and treatment of this dreaded disease.

Necrotizing enterocolitis: pathophysiology, platelet-activating factor, and probiotics.

Although smaller and younger preterm neonates can now survive long term due to advances in neonatal medicine, necrotizing enterocolitis (NEC) continues to plague the clinicians caring for these tiny patients. Research studies have contributed to our understanding of this complex disease, including the role of platelet-activating factor (PAF), but preventative and treatment strategies remain limited. One promising preventative measure in recent years has been enteral supplementation of probiotics, but concerns remain regarding the optimal use of these organisms, and safe administration must be assured. This chapter reviews NEC pathophysiology, including the role of PAF, as well as literature on the use of probiotics in the preterm infant.

Preterm human milk contains a large pool of latent TGF-β, which can be activated by exogenous neuraminidase

Human milk contains substantial amounts of transforming growth factor (TGF)-β, particularly the isoform TGF-β2. We previously showed in preclinical models that enterally administered TGF-β2 can protect against necrotizing enterocolitis (NEC), an inflammatory bowel necrosis of premature infants. In this study we hypothesized that premature infants remain at higher risk of NEC than full-term infants, even when they receive their own mothers milk, because preterm human milk contains less bioactive TGF-β than full-term milk. Our objective was to compare TGF-β bioactivity in preterm vs. full-term milk and identify factors that activate milk-borne TGF-β. Mothers who delivered between 23 0/7 and 31 6/7 wk or at ≥37 wk of gestation provided milk samples at serial time points. TGF-β bioactivity and NF-κB signaling were measured using specific reporter cells and in murine intestinal tissue explants. TGF-β1, TGF-β2, TGF-β3, and various TGF-β activators were measured by real-time PCR, enzyme immunoassays, or established enzymatic activity assays. Preterm human milk showed minimal TGF-β bioactivity in the native state but contained a large pool of latent TGF-β. TGF-β2 was the predominant isoform of TGF-β in preterm milk. Using a combination of several in vitro and ex vivo models, we show that neuraminidase is a key regulator of TGF-β bioactivity in human milk. Finally, we show that addition of bacterial neuraminidase to preterm human milk increased TGF-β bioactivity. Preterm milk contains large quantities of TGF-β, but most of it is in an inactive state. Addition of neuraminidase can increase TGF-β bioactivity in preterm milk and enhance its anti-inflammatory effects.

New Medical and Surgical Insights Into Neonatal Necrotizing Enterocolitis: A Review

Importance Necrotizing enterocolitis (NEC) has long remained a significant cause of morbidity and mortality in neonatal intensive care units. While the mainstay of treatment for this devastating condition remains largely supportive, research efforts continue to be directed toward understanding pathophysiology as well as how best to approach surgical management when indicated. Observations In this review, we first examine recent medical observations, including overviews on the microbiome and a brief review of the use of probiotics. Next, we discuss the use of biomarkers and how clinicians may be able to use them in the future to predict the course of disease and, perhaps, the need for surgical intervention. We then provide an overview on the use of exclusive human milk feeding and the utility of this approach in preventing NEC. Finally, we discuss recent developments in the surgical management of NEC, beginning with indications for surgery and following with a section on technical surgical considerations, including peritoneal drain vs laparotomy. The review concludes with outcomes from infants with surgically treated NEC. Conclusions and Relevance Although medical treatment options for NEC are largely unchanged, understanding of the disease continues to evolve. As new research methods are developed, NEC pathophysiology can be more completely understood. In time, it is hoped that data from ongoing and planned clinical trials will allow us to routinely add targeted preventive measures in addition to human milk, such as prebiotics and probiotics, to the management of high-risk infants. In addition, the discovery of novel biomarkers may not only prove useful in predicting severity of illness but also will hopefully allow for identification of the disease prior to onset of clinical signs. Finally, continued investigation into optimizing surgical outcomes is essential in this population of infants, many of whom require long-term parenteral therapy and intestinal rehabilitation.

Maternal breast milk transforming growth factor-beta and feeding intolerance in preterm infants

Background:Feeding intolerance (FI) occurs commonly in the neonatal intensive care unit. Breast milk contains a large pool of transforming growth factor-beta (TGF-β). Few studies describe TGF-β levels in preterm milk, and the relationship to FI remains unexplored. We measured TGF-β levels in preterm breast milk to investigate a correlation with FI in preterm infants.Methods:Prospective observational trial of 100 mother–infant pairs, enrolling infants born below 32 wk gestation and less than 1,500 g, and mothers who planned to provide breast milk. TGF-β levels were measured using enzyme-linked immunosorbent assay. Infant charts were reviewed for outcomes.Results:TGF-β declined postnatally, most elevated in colostrum (P < 0.01). TGF-β2 levels were higher than TGF-β1 at all time points (P < 0.01). Colostrum TGF-β levels correlated inversely with birth weight (P < 0.01) and gestational age (P < 0.05). One-week TGF-β2 levels were reduced in growth-restricted infants with FI (P < 0.01). Of infants with necrotizing enterocolitis (NEC), TGF-β2 levels appeared to be low, but small sample size precluded meaningful statistical comparisons.Conclusion:TGF-β levels decline temporally in preterm milk. TGF-β1 colostrum levels correlate inversely with birth weight and gestational age. TGF-β2 may play a role in FI in growth-restricted infants. The relationship of TGF-β2 and NEC merits future investigation.

Myth: Necrotizing enterocolitis: Probiotics will end the disease, and surgical intervention improves the outcome

Necrotizing enterocolitis remains an important contributor to neonatal morbidity and mortality. Recent studies suggest that probiotic supplementation may reduce the risk of the disease in premature infants, and some authors recommend that this approach is ready to be utilized as standard-of-care. Once necrotizing enterocolitis is diagnosed and progresses toward peritonitis or perforation, surgical intervention is thought to improve the outcome, and investigators have suggested that peritoneal drainage is as effective as an exploratory laparotomy. In this chapter, we review the current state of knowledge, and suggest that additional studies are necessary to confirm that probiotics will end this disease, and that surgical intervention may not significantly improve the outcome after diagnosis in these compromised patients.

Probiotics and prevention of neonatal necrotizing enterocolitis.

Purpose of review This review will summarize the clinical trials evaluating the role of prophylactic probiotic supplementation in preterm infants in order to reduce the incidence of necrotizing enterocolitis (NEC). Recent findings Evidence suggests that probiotic supplementation in preterm infants reduces the incidence of NEC. In fact, recent meta-analyses have called for the use of probiotics as preventive therapy in subsets of this population. However, although multiple studies have evaluated the use of probiotics for this indication in preterm infants, these trials have used different formulations of bacteria, at differing doses and using varied protocols for administration; thus many unanswered questions remain. In addition, theoretical safety issues and concerns regarding quality of product still need to be addressed. Summary As NEC remains a serious problem for preterm neonates, proven therapies for prevention and treatment of this dreaded disease are needed. While the evidence does support a future role for probiotics in the prevention of NEC, it is of utmost importance to first ensure that a safe and high-quality product meeting rigorous standards will be provided to these at-risk infants.

Transforming growth factor-β2 is sequestered in preterm human milk by chondroitin sulfate proteoglycans

Human milk contains biologically important amounts of transforming growth factor-β2 isoform (TGF-β2), which is presumed to protect against inflammatory gut mucosal injury in the neonate. In preclinical models, enterally administered TGF-β2 can protect against experimental necrotizing enterocolitis, an inflammatory bowel necrosis of premature infants. In this study, we investigated whether TGF-β bioactivity in human preterm milk could be enhanced for therapeutic purposes by adding recombinant TGF-β2 (rTGF-β2) to milk prior to feeding. Milk-borne TGF-β bioactivity was measured by established luciferase reporter assays. Molecular interactions of TGF-β2 were investigated by nondenaturing gel electrophoresis and immunoblots, computational molecular modeling, and affinity capillary electrophoresis. Addition of rTGF-β2 (20-40 nM) to human preterm milk samples failed to increase TGF-β bioactivity in milk. Milk-borne TGF-β2 was bound to chondroitin sulfate (CS) containing proteoglycan(s) such as biglycan, which are expressed in high concentrations in milk. Chondroitinase treatment of milk increased the bioactivity of both endogenous and rTGF-β2, and consequently, enhanced the ability of preterm milk to suppress LPS-induced NF-κB activation in macrophages. These findings provide a mechanism for the normally low bioavailability of milk-borne TGF-β2 and identify chondroitinase digestion of milk as a potential therapeutic strategy to enhance the anti-inflammatory effects of preterm milk.

Early docosahexaenoic and arachidonic acid supplementation in extremely-low-birth-weight infants

Background:Extremely-low-birth-weight (ELBW) infants accrue large deficits in docosahexaenoic acid (DHA) and arachidonic acid (ARA) and require improved supplementation strategies. We hypothesized that once daily DHA+ARA drops applied to buccal mucosa will increase blood levels.Methods:Thirty ELBW infants were randomized to receive DHA 20 mg/kg/d + ARA 40 or 60 mg/kg/d + ARA 120 mg/kg/d or placebo within 72 h of age for 8 wk duration. Red blood cell phospholipid levels of DHA (primary) and ARA (secondary) were measured at 2 and 8 wk of age.Results:Twenty-eight survivors with a median birth weight of 806 g completed dosing and sampling. Red blood cell levels were similar between the three groups at 2 wk (DHA: 4.62 wt% (interquartile range (IQR) 4.1–5.5) for all, P = 0.29 between groups; ARA: 21.1 wt% (IQR 18.78–22.6) for all, P = 0.41 between groups) and 8 wk (DHA: 6.0 wt% (IQR 5.1–7.1) for all, P = 0.57 between groups; ARA: 20.1 wt% (IQR 18.3–23.1) for all, P = 0.63 between groups). DHA in all infants showed a median increase of 31% from 2 to 8 wk (P < 0.04). ARA levels did not significantly change over time (P > 0.6).Conclusion:Daily buccal DHA and ARA supplements did not affect fatty acid levels in ELBW infants.