Congenital cytomegalovirus as a trigger in the pathogenesis of necrotizing enterocolitis

About This Grant

The overall goal of this proposal is to investigate a potential role for congenital cytomegalovirus (CMV) infection in triggering the development and worsening the severity of necrotizing enterocolitis (NEC), the leading cause of death from gastrointestinal disease in premature infants. NEC occurs in up to 10% of premature infants, and is characterized by the acute development of intestinal necrosis leading to death in over 40% of patients. CMV is a herpesvirus with a seroprevalence of 40-80%, and carries a 40% risk of virus transmission to the fetus during pregnancy. We have previously shown that NEC development requires activation of the lipopolysaccharide (LPS) receptor Toll-like receptor 4 (TLR4) on the intestinal epithelium. However, factor(s) that trigger exaggerated TLR4 signaling leading to NEC remain unknown. We now show that congenital CMV infection plays a critical role in triggering NEC and worsening NEC severity by increasing TLR4 signaling and inducing mitochondrial injury in the intestinal epithelium. In human infants, the presence of CMV infection in intestinal tissue from NEC resection specimens was associated with significantly higher TLR4 expression and greater NEC severity compared with CMV-negative infants. Explants from CMV-infected human intestine treated with LPS showed significantly higher TLR4-induced inflammation than uninfected tissue, and CMV-positive patients showed increased mitochondrial injury in the intestinal epithelium as measured by detection of mitochondrial DNA nicks and ROS release. In mice, the induction of congenital murine CMV infection that was achieved by injection of mouse CMV (mCMV) into the amniotic fluid at G17.5 resulted in greater NEC frequency and severity, higher Tlr4 expression and greater mitochondrial injury in the gut than uninfected mice. Congenital mCMV did not worsen NEC in Tlr4IEC mice, proving that TLR4 mediates CMV effects. Mechanistically, congenital mCMV infection induced epigenetic changes in the Tlr4 promoter, leading to increased Tlr4 expression, resulting in TLR4-dependent mitochondrial injury, ATP depletion and epithelial damage in the gut. Importantly, the administration of novel TLR4 antagonists or the oral administration of the ATP precursor adenosine reversed the CMV effects on triggering NEC and increasing NEC severity. Hypothesis: We hypothesize that congenital CMV infection exerts a critical role in triggering NEC and increasing NEC severity through increased TLR4-induced mitochondrial dysfunction and intestinal epithelial injury. We further hypothesize that novel TLR4 antagonists and intestinal-specific adenosine analogues represent new strategies to attenuate NEC in the presence of congenital CMV infection. We will test these hypotheses in the following two aims: Aim 1: To evaluate the role of congenital CMV infection in triggering and worsening NEC via increased TLR4 signaling; Aim 2: To evaluate the role of congenital CMV infection in triggering and worsening NEC via TLR4-induced mitochondrial injury. When completed, these studies promise to uncover novel mechanisms and treatment strategies for this devastating disease.