Tara A. Saunders

Silent Intralesional Microhemorrhage as a Risk Factor for Brain Arteriovenous Malformation Rupture

Background and Purpose— We investigated whether brain arteriovenous malformation silent intralesional microhemorrhage, that is, asymptomatic bleeding in the nidal compartment, might serve as a marker for increased risk of symptomatic intracranial hemorrhage (ICH). We evaluated 2 markers to assess the occurrence of silent intralesional microhemorrhage: neuroradiological assessment of evidence of old hemorrhage—imaging evidence of bleeding before the outcome events–and hemosiderin positivity in hematoxylin and eosin-stained paraffin block sections. Methods— We identified cases from our brain arteriovenous malformation database with recorded neuroradiological data or available surgical paraffin blocks. Using 2 end points, index ICH or new ICH after diagnosis (censored at treatment, loss to follow-up, or death), we performed logistic or Cox regression to assess evidence of old hemorrhage and hemosiderin positivity adjusting for age, sex, deep-only venous drainage, maximal brain arteriovenous malformation size, deep location, and associated arterial aneurysms. Results— Evidence of old hemorrhage was present in 6.5% (n=975) of patients and highly predictive of index ICH (P<0.001; OR, 3.97; 95% CI, 2.1–7.5) adjusting for other risk factors. In a multivariable model (n=643), evidence of old hemorrhage was an independent predictor of new ICH (hazard ratio, 3.53; 95% CI, 1.35–9.23; P=0.010). Hemosiderin positivity was found in 36.2% (29.6% in unruptured; 47.8% in ruptured; P=0.04) and associated with index ICH in univariate (OR, 2.18; 95% CI, 1.03–4.61; P=0.042; n=127) and multivariable models (OR, 3.64; 95% CI, 1.11–12.00; P=0.034; n=79). Conclusions— The prevalence of silent intralesional microhemorrhage is high and there is evidence for an association with both index and subsequent ICH. Further development of means to detect silent intralesional microhemorrhage during brain arteriovenous malformation evaluation may present an opportunity to improve risk stratification, especially for unruptured brain arteriovenous malformations.

Epidermal Growth Factor Reduces Hepatic Sequelae in Experimental Necrotizing Enterocolitis

Background and Aim: Neonatal necrotizing enterocolitis (NEC) is the most common gastrointestinal disease of premature infants. We recently demonstrated that the gut/liver axis plays an important role in the pathophysiology of NEC through the release of inflammatory mediators into the intestinal lumen. We have also shown that supplementation of formula with epidermal growth factor (EGF) dramatically decreases ileal pathology associated with experimental NEC. In this study, we examined the effects of EGF on the liver portion of the gut/liver axis in the neonatal rat model of NEC. Methods: Newborn rats were divided into three experimental groups, NEC, hand-fed with growth-factor free formula; NEC + EGF, hand-fed with formula supplemented with 500 ng/ml rat EGF; or DF, dam fed. All animals were exposed to asphyxia and cold stress twice daily for 4 days to develop NEC. Results: EGF receptor expression was significantly (p ≤ 0.01) decreased in the NEC + EGF group compared to the NEC group. EGF supplementation significantly decreased Kupffer cell numbers (p ≤ 0.01) as well as hepatic tumor necrosis factor (TNF)-α and interleukin-18 production (p ≤ 0.05). Further, TNF-α in the intestinal luminal contents of the NEC + EGF group were normalized to levels observed in DF controls compared to the NEC group (p ≤ 0.05). Activated nuclear factor-ĸB was also substantially decreased in the NEC + EGF group versus the NEC group. Conclusion: The results of this study indicate that EGF normalizes cytokine overproduction in the liver of neonatal rats with NEC, which contributes to diminished intestinal damage during the development of experimental NEC. These data suggest that supplementation of formula with EGF can have beneficial effects on the gut/liver axis during NEC pathogenesis.

Intrapancreatic accessory spleen: utilization of fine needle aspiration for diagnosis of a potential mimic of a pancreatic neoplasm

Accessory spleen (AS) is not a rare occurrence, and with the second most common site being the tail of the pancreas, intrapancreatic AS (IPAS) can easily mimic a pancreatic neoplasm. Together with radiologic imaging findings, endoscopic ultrasound-guided fine needle aspiration (FNA) can be used to assist in the diagnosis, preventing potentially unnecessary surgical procedures. The most common cytologic findings that have been described in the literature include a heterogenous population of small lymphocytes along with traversing small vessels. Immunohistochemical staining for CD8 has also been documented as a useful tool to support the diagnosis as it specifically highlights the endothelial cells of the splenic sinus. Here, we report two additional cases of IPAS diagnosed by FNA and discuss the potential pitfalls in diagnosis of this entity.

101 Effect of EGF on the Intestinal Tight Junction Barrier in Experimental Necrotizing Enterocolitis

Background: Necrotizing enterocolitis (NEC) is the most common intestinal disease predominately of formula fed, premature babies. Epidermal growth factor (EGF) reduces the incidence of disease in a neonatal rat model of NEC. It has been suggested that increased mucosal permeability may play an important role in the pathogenesis of NEC. Claudin and occludin are the major structural and functional components of the tight junction barrier. These proteins may be altered in disease states. However, the role of tight junctions in NEC pathogenesis and EGF treatment is currently unknown.Objective: The aim of this study was to determine if EGF affects expression of tight junction genes and proteins in the ileum during the development of NEC.Methods: Neonatal rats, either dam fed (DF), milk formula fed (NEC), or fed with formula plus 500 ng/ml EGF (NEC+EGF) were exposed to asphyxia and cold stress to develop NEC. After 96 hours, ileal expression of claudin-3 and occludin were evaluated using Realtime-PCR, Western blot and immunohistochemistry.Results: Claudin-3 and occludin mRNA levels in the ileum were significantly increased 2–4 fold in NEC animals compared to DF animals (p<0.0001). Supplementation with EGF normalized mRNA expression to DF levels. Phosphorylated occludin was significantly decreased in NEC animals compared to DF animals (p<0.01) and EGF treatment increased expression to DF levels. Non-phosphorylated occludin was only present in NEC and NEC+EGF animals. Histological localization of claudin-3 and occludin proteins was disturbed in animals with NEC compared to DF animals. NEC animals had upregulated expression and disorganization of the tight junction proteins. EGF treatment resulted in localization of the proteins at the tight junction complex.Conclusion: The ability of EGF to normalize expression and localization of tight junction proteins may be one mechanism by which integrity of the mucosal barrier is maintained, thereby protecting the intestinal mucosa against NEC. The increased expression of mRNA and disorganization of protein in animals with NEC may represent a compensation for the inability to form functional tight junctions at the mucosal barrier.Supported by the APS Porter Physiology Fellowship (to J.C.) and NIH Grants HD-39657, HD-47237 (to B.D.)

102 Effect of Anti-TNF-A Treatment on the Incidence of Experimental Necrotizing Enterocolitis

Background: Necrotizing Enterocolitis (NEC) is the most common gastrointestinal disease predominately affecting premature infants. Pro-inflammatory cytokines, such as TNF-a, have been implicated in intestinal inflammatory diseases. Previously, we have shown that Kupffer cells in the liver are the major producer of TNF-a found in the intestinal luminal contents during experimental NEC. We speculate that TNF-a may play a role in the development of NEC by being an active member of an inflammatory cascade leading to the pathology associated with the disease.Objective: The aim of this study was to determine whether administration of anti-TNF-a reduces the incidence and severity of NEC.Methods: Newborn neonatal rats were fed milk formula and exposed to asphyxia and cold stress twice daily to develop NEC. The pups were divided into two experimental groups: those injected i.p. with 5 mg/kg anti-mouse monoclonal TNF-a once per day for three days (NEC+anti-TNF-a) and those sham injected with saline (NEC). After 96 hours, animals were sacrificed and the number of TNF-a positive cells in the liver and the histologic NEC score were evaluated in the anti-TNF-a injected animals compared to the sham injected animals.Results: The administration of 5 mg/kg/day of anti-mouse monoclonal TNF-a significantly reduced the amount of TNF-a positive Kupffer cells in the liver of neonatal rats. This specific neutralization of hepatic TNF-a production significantly decreased the incidence and severity of experimental NEC (p<0.01). Conclusions: Neutralization of hepatic TNF-a production reduces the incidence and severity of NEC. This finding suggests a possible therapy for the treatment of this disease.Supported by the NIH Grants HD-39657 and HD-47237 (to B.D)