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Brad W Warner

Publications and source records attributed to Brad W Warner.

At least 19 recordsLinked to original sources
PubMed2006

The effect of a right-sided aortic arch on outcome in children with esophageal atresia and tracheoesophageal fistula.

UNLABELLED: During repair of an esophageal atresia with tracheoesophageal fistula (EA/TEF), a right-sided aortic arch (RAA) presents a technical challenge. We reviewed our experience with EA/TEF focusing on the impact of RAA. METHODS: A retrospective review of patients with EA/TEF at our institution from 1990 to 2004 was performed. Of 61 patients, 53 had a left aortic arch (LAA) and 8 had RAA. RESULTS: The mean gestational age and birth weight were similar between patients with LAA and RAA. There were more vascular rings in patients with RAA than with LAA (37.5% vs 3.8%, P = .009). A preoperative echocardiogram correctly identified the location of the arch in all with LAA but only 62.5% of those with RAA. Operative complications occurred in 25% of patients with RAA and 11% of patients with LAA. CONCLUSIONS: Management of a child with RAA and EA/TEF is technically challenging. Preoperative identification of the great vessel anomalies may impact operative decision making. Therefore, careful diagnostic evaluation of a patient with an aortic arch that cannot be easily defined by ECHO should be considered before definitive repair.

Aorta, Thoracic↗
PubMed2006

Absent STAT-1 expression perturbs adaptation and apoptosis after massive intestinal resection.

BACKGROUND: We have previously established the significance of epidermal growth factor receptor (EGFR) activity and the cyclin-dependent kinase inhibitor p21waf1/cip1 (p21) for the adaptive response of the intestine to massive small bowel resection (SBR). In this study, we tested the role of the signal transducer and activator of transcription 1 (STAT-1) as this transcription factor is activated by the EGFR and known to induce p21 expression. METHODS: Control (n = 40; C57/Bl6) and STAT-1-null mice (n = 40) underwent 50% proximal SBR or sham operation. After 3 days, the remnant ileum was harvested and the villus and crypt morphology was measured along with changes in rates of enterocyte proliferation and apoptosis. RESULTS: The magnitude of resection-induced adaptation was greater in STAT-1-null animals as verified by taller villi and deeper crypts. The expected increase in enterocyte apoptosis did not occur after SBR in the background of STAT-1 deficiency. Western blotting revealed elevated expression of p21 protein in both STAT-1-null and controls after SBR. CONCLUSION: Increased p21 expression after SBR in the absence of STAT-1 suggests an alternate mechanism for resection-induced regulation of p21. Enhanced adaptation in STAT-1-null animals suggests that this transcription factor serves an inhibitor to the process of adaptation, perhaps via regulation of enterocyte apoptosis.

Adaptation, Physiological↗
PubMed2006

Combined pharmacotherapy that increases proliferation and decreases apoptosis optimally enhances intestinal adaptation.

BACKGROUND: Adaptation after massive small bowel resection (SBR) is associated with increased rates of enterocyte proliferation (P) and apoptosis (A). In the present study, we sought to determine the effect of dual therapy designed to increase P and simultaneously reduce A. METHODS: C57Bl/6 mice underwent a 50% small bowel resection (SBR) or sham operation, and then received an inhibitor of apoptosis (pan-caspase inhibitor), a stimulus for proliferation (epidermal growth factor; EGF), a combination, or vehicle control. After 3 days, adaptive morphology (villus height, crypt depth) and rates of enterocyte turnover (proliferation and apoptosis) were measured in the remnant ileum. RESULTS: Adaptation in controls and treated with the inhibitor was similar. EGF-treated mice demonstrated an even greater adaptive response. Combined therapy with the inhibitor and EGF resulted in maximal adaptation as gauged by the greatest increases in villus height and crypt depth and ratio of rates of P to A. CONCLUSION: The capacity for adaptation following massive SBR is maintained via tight regulation of cell production and death. Pharmacologic intervention directed at increasing enterocyte proliferation while simultaneously decreasing apoptosis augments adaptation greater than either intervention alone and may provide a useful strategy to clinically amplify adaptation.

Adaptation, Physiological↗
PubMed2006

Epidermal growth factor receptor signaling regulates Bax and Bcl-w expression and apoptotic responses during intestinal adaptation in mice.

BACKGROUND & AIMS: Normal intestinal adaptation to massive small-bowel resection requires intact epidermal growth factor receptor signaling and consists of increased enterocyte proliferation and apoptosis. Although emphasis has been placed on understanding the regulation of proliferation, few studies have evaluated the mechanism and contribution of apoptosis to the adaptation response. We sought to test the hypothesis that epidermal growth factor receptor signaling regulates specific Bcl-2 family members (Bax and Bcl-w) to direct apoptosis and adaptation after massive small-bowel resection. METHODS: Laser capture microdissection microscopy permitted measurement of Bax and Bcl-w messenger RNA expression in crypt and villus enterocytes in control conditions and under epidermal growth factor receptor-inhibited (waved-2 mice) or stimulated (epidermal growth factor transgenic mice) conditions after a 50% small-bowel resection or sham operation. Resection-induced adaptation was then studied in Bax-null and Bcl-w-null mice under control circumstances and after epidermal growth factor receptor stimulation. RESULTS: When compared with Bcl-w, the most significant expression changes were observed with Bax and took place within crypt enterocytes. Epidermal growth factor receptor stimulation resulted in a decreased ratio of Bax to Bcl-w expression and decreased rates of apoptosis. Bax-null mice had no apoptosis response to small-bowel resection and displayed an amplified adaptation response to the administration of epidermal growth factor. Bcl-w-null mice had poor survival and impaired adaptation to small-bowel resection, an effect that was rescued by crossbreeding these mice with epidermal growth factor transgenic mice. CONCLUSIONS: The crypt expression of Bax and Bcl-w is influenced by epidermal growth factor receptor signaling and is key for the regulation of apoptosis. Epidermal growth factor receptor stimulation, coupled with apoptosis inhibition, may provide a novel strategy to amplify adaptation responses in patients after massive intestinal loss.

Animals↗
PubMed2006

Developmental characteristics of adapting mouse small intestine crypt cells.

BACKGROUND & AIMS: Following massive small bowel resection (SBR), the remnant intestine undergoes an adaptive process characterized by increases in a number of physiologic and morphologic parameters. These changes are the result of a stimulus that increases crypt cell mitosis and augments cellular progression along the villus axis. To better define this process, we identified patterns of gene expression specifically within adapting intestinal crypt cells following SBR. METHODS: Laser capture microdissection was used to isolate mouse intestinal crypt cells following SBR or sham operation. Multiple biological and technical complementary DNA microarray replicates allowed rigorous statistical analyses for identification of important expression profiles. Major groups of genes were classified as to site of action, functional pathway, and possible regulatory groups. RESULTS: A total of 300 genes differentially expressed at significant levels within adapting crypt enterocytes were analyzed. Comparison of this list of differentially expressed adapting crypt cell genes with a generalized mouse gene expression database (from 82 developing and adult mouse tissues) showed the greatest overlap with developing and immature intestinal tissues. We identified prominent groups of genes involved with cell growth, signal transduction, and nucleic acid binding. Genes not previously shown to be involved with adaptation or development and maturation were identified. CONCLUSIONS: Identification of similar genes coordinately regulated during both adaptation and development, processes that share key morphologic features, provides a basis for new mechanistic insights into these shared characteristics.

Adaptation, Physiological↗
PubMed2006

Epidermal growth factor receptor-mediated proliferation of enterocytes requires p21waf1/cip1 expression.

BACKGROUND & AIMS: Epidermal growth factor receptor (EGFR)-mediated increase in enterocyte proliferation following massive resection is a major mechanism by which the small intestine adapts to the loss of its mucosal surface area. In addition, expression of the cyclin-dependent kinase inhibitor p21(waf1/cip1) is required for resection-induced enterocyte proliferation. This study sought to establish a mechanistic link between EGFR-mediated intestinal epithelial cell proliferation and p21(waf1/cip1) expression. METHODS: EGF was used to stimulate IEC-6 and HCA-7 cells. P21(waf1/cip1) messenger RNA (mRNA) and protein expression were measured by real-time polymerase chain reaction and Western blot, respectively. P21(waf1/cip1) promoter studies were performed using p21(waf1/cip1) promoter-driven luciferase assay. Pharmacologic inhibitors of PI3-kinase and mitogen activated protein kinase (MAPK) were used to block these pathways downstream of the activated EGFR. Constitutively active Ras, Raf, or MEK-1 constructs were transfected into cells for overexpression studies. Cell proliferation was measured by bromodeoxyuridine incorporation following p21(waf1/cip1) silencing with RNAi. Finally, Cyclin D(1)/Cdk interaction was evaluated by immunoprecipitation. RESULTS: EGFR activation in intestinal epithelial cells induced the expression of p21(waf1/cip1) mRNA and protein This event was transcriptionally regulated via a 50-bp segment of the p21(waf1/cip1) promoter as a result of MAPK activation. Exogenous EGF failed to induce proliferation in p21(waf1/cip1)-silenced cells and adaptive proliferation after intestinal resection in p21(waf1/cip1)-null mice. Functionally, p21(waf1/cip1) up-regulation was required for stabilizing Cyclin D/Cdk 4 complexes and intestinal cell proliferation. CONCLUSIONS: EGFR-mediated induction of enterocyte proliferation requires MAPK-dependent increase in p21(waf1/cip1) expression in intestinal epithelial cells. These studies elucidate an important mechanism for resection-induced enterocyte proliferation during intestinal adaptation.

Animals↗
PubMed2006

Critical roles for EGF receptor signaling during resection-induced intestinal adaptation.

The adaptation response of the remnant gut to massive intestinal resection represents a mitogenic signal involving all bowel wall layers. In the mucosa, this response results in taller villi, deeper crypts, and enhanced enterocyte turnover as gauged by greater rates of both proliferation and apoptosis. Although the exact mechanisms and mediators of this important compensatory response remain incompletely understood, work from this laboratory over the past decade has illuminated a crucial role for intact receptor signaling for a robust response. Using a murine model for intestinal resection, transgenic, null and mutant mouse strains have provided unique experimental paradigms to dissect molecular mechanisms for epidermal growth factor (EGF) receptor-directed influence on adaptation. Stimulation of this receptor is linked with a magnified adaptation response, whereas attenuation of the activity of this receptor is associated with impaired adaptation. EGF receptor activation and expression are both elevated in enterocytes after resection, and salivary levels of EGF-the major ligand for the EGF receptor-are increased. In addition to stimulation of enterocyte proliferation, EGF receptor signaling prevents the typical increase in rates of enterocyte apoptosis, probably by affecting the ratio of expression of both pro- and anti-apoptotic Bcl-2 family members. The key to optimizing care for patients with short gut syndrome will necessarily follow a thorough understanding of intestinal adaptation responses.

Adaptation, Physiological↗
PubMed2005

Roles for p21waf1/cip1 and p27kip1 during the adaptation response to massive intestinal resection.

The magnitude of gut adaptation is a decisive factor in determining whether patients are able to live independent of parenteral nutrition after massive small bowel loss. We previously established that the cyclin-dependent kinase inhibitor (CDKI) p21(waf1/cip1) is necessary for enterocyte proliferation and a normal adaptation response. In the present study, we have further elucidated the role of this CDKI in the context of p27(kip1), another member of the Cip/Kip CDKI family. Small bowel resections (SBRs) or sham operations were performed in control (C57/BL6), p21(waf1/cip1)-null, p27(kip1)-null, and p21(waf1/cip1)/p27(kip1) double-null mice. Morphological (villus height/crypt depth) alterations in the mucosa, the kinetics of enterocyte turnover (rates of enterocyte proliferation and apoptosis), and the protein expression of various cell cycle-regulatory proteins were recorded at various postoperative times. Enterocyte compartment-specific mRNA expression was investigated using laser capture microdissection. Resection-induced adaptation in control mice coincided with increased protein expression of p21(waf1/cip1) and decreased p27(kip1) within 3 days postoperatively. Identical changes in mRNA expression were detected in crypt but not in villus enterocytes. Adaptation occurred normally in control and p27(kip1)-null mice; however, mice deficient in both p21(waf1/cip1) and p27(kip1) failed to increase baseline rates of enterocyte proliferation and adaptation. The expression of p21(waf1/cip1) protein and mRNA in the proliferative crypt compartment is necessary for resection-induced enterocyte proliferation and adaptation. The finding that deficient expression of p27(kip1) does not affect adaptation suggests that these similar CDKI family members display distinctive cellular functions during the complex process of intestinal adaptation.

Adaptation, Physiological↗
PubMed2005

Haploinsufficiency of Atp2a2, encoding the sarco(endo)plasmic reticulum Ca2+-ATPase isoform 2 Ca2+ pump, predisposes mice to squamous cell tumors via a novel mode of cancer susceptibility.

A null mutation in one copy of the Atp2a2 or ATP2A2 gene, encoding sarco(endo)plasmic reticulum Ca2+-ATPase isoform 2 (SERCA2), leads to squamous cell tumors in mice and to Darier disease in humans, a skin disorder that also involves keratinocytes. Here, we examined the time course and genetic mechanisms of tumor development in the mutant animals. Atp2a2+/- mice overexpressed keratins associated with keratinocyte hyperactivation in normal forestomachs as early as 2 months of age. By the age of 5 to 7 months, 22% of mutants had developed papillomas of the forestomach, and 89% of mutants older than 14 months had developed squamous cell papillomas and/or carcinomas, with a preponderance of the latter. Tumors occurred in regions that had keratinized epithelium and were subjected to repeated mechanical irritation. The genetic mechanism of tumorigenesis did not involve loss of heterozygosity, as tumor cells analyzed by laser capture microdissection contained the wild-type Atp2a2 allele. Furthermore, immunoblot and immunohistochemical analysis showed that tumor keratinocytes expressed the SERCA2 protein. Mutations were not observed in the ras proto-oncogenes; however, expression of wild-type ras was up-regulated, with particularly high levels of K-ras. Loss of the p53 tumor suppressor gene occurred in a single massive tumor, whereas other tumors had increased levels of p53 protein but no mutations in the p53 gene. These findings show that SERCA2 haploinsufficiency predisposes mice to tumor development via a novel mode of cancer susceptibility involving a global change in the tumorigenic potential of keratinized epithelium in Atp2a2+/- mice.

Alleles↗
PubMed2005

Epidermal growth factor-induced rapid retinoblastoma phosphorylation at Ser780 and Ser795 is mediated by ERK1/2 in small intestine epithelial cells.

The retinoblastoma protein Rb is critical for the regulation of mammalian cell cycle entry. Hypophosphorylated Rb is considered to be the active form and directs G1 arrest, while hyperphosphorylated Rb permits the transition from G1 to S phase for cell proliferation. Upon stimulation by various growth factors, Rb appears to be phosphorylated by a cascade of phosphorylation events mediated mainly by kinases associated with cyclins D and E. Here we report that in prototype small intestine crypt stem cells (RIEC-6), stimulation with either epidermal growth factor or fetal bovine serum results in an unexpected rapid and sustained Rb phosphorylation at sites Ser780, Ser795, and Thr821 which precedes cyclin D1 expression, cyclin D1/cdk4 complex formation, and cdk4 kinase activity. Rb phosphorylation at Ser780 and Ser795 is prevented by MEK, but not phosphatidylinositol 3-kinase, inhibitors. In vitro, Rb is directly phosphorylated by active ERK1/2 as shown by [gamma-32P]ATP labeling. The phosphorylation sites are further directed to Ser780 and Ser795 by kinase assays using recombined active ERK1/2 or immunoprecipitated phospho-ERK1/2 from mitogen stimulated cells. Pull-down assays revealed that Rb interacts with active ERK1/2 but not their inactive unphosphorylated forms. Upon EGF stimulation, phosphorylated ERK1/2 co-immunoprecipitates together with phosphorylated Rb. Collectively, these results demonstrate a novel rapid Rb phosphorylation at specific sites induced by mitogen stimulation in epithelial cells of the small intestine. These data specifically identify ERK1/2 as the kinase responsible for Rb phosphorylation targeted to sites Ser780 and Ser795. It appears that ERK1/2 could be an important link between a mitogenic signal directly to Rb, thereby providing a rapid response mechanism between mitogen stimulation and cell cycle machinery.

Animals↗
PubMed2005

Epidermal growth factor receptor signaling regulates goblet cell production after small bowel resection.

BACKGROUND/PURPOSE: Intestinal adaptation is a compensatory response to massive small bowel loss in which there are increased numbers of absorptive enterocytes. However, the generation of secretory epithelial cell subtypes in this process has not been investigated. The purpose of this study was to examine the adaptive changes of several small intestinal cell lineage changes in response to massive small bowel resection (SBR). METHODS: A 75% SBR or sham operation was performed on male Sprague-Dawley rats. On postoperative day 7, the remnant ileum was harvested and immunohistochemical staining for goblet, Paneth, and enteroendocrine cells was performed. Cell subtypes were evaluated as cells per micrometer of villus/crypt length and compared among operations. RESULTS: A significant increase in goblet cell density occurred after SBR. Intestinal resection did not alter the number of Paneth and enteroendocrine cells. In additional experiments, inhibition of epidermal growth factor receptor signaling was associated with a diminished goblet cell density. CONCLUSIONS: The adaptive response of the intestine to massive bowel loss results in an expansion of the goblet cell population in addition to greater numbers of absorptive enterocytes. Although the mechanism and purpose for selective expansion of these stem cell-derived lineages are not presently known, epidermal growth factor receptor signaling appears to be a common pathway.

Adaptation, Physiological↗
PubMed2005

Evidence for active Wnt signaling during postresection intestinal adaptation.

BACKGROUND: In the intestine, Wnt proteins are powerful regulators of cell proliferation, differentiation, and adhesion. Mutations of the adenomatous polyposis coli (APC) gene elevate nuclear beta-catenin and provoke intestinal tumor formation. We sought to determine whether Wnt signaling is involved in adaptive response to massive small bowel resection (SBR). METHODS: Male Min mice with a mutation of the APC gene and wild-type controls underwent a 50% proximal SBR or sham operation. After 3 days, villus height, crypt depth, and rates of proliferation and apoptosis were recorded in the remnant ileum. RESULTS: After SBR, villus height and enterocyte proliferation were significantly greater in the Min mice. Western blotting demonstrated resection-induced increases in beta-catenin, c-Myc, and E-cadherin after SBR, which was more pronounced in Min mice. CONCLUSIONS: Mutation of the APC gene and augmented Wnt signaling in the intestine results in an enhanced adaptive response to massive SBR. These data, for the first time, implicate an important role for Wnt signaling during the pathogenesis of resection-induced intestinal adaptation.

Animals↗
PubMed2005

Necrotizing fasciitis: a rare complication of neonatal necrotizing enterocolitis.

Necrotizing fasciitis is a rapidly spreading soft tissue infection affecting the subcutaneous tissue and underlying fascial layers. Although this infection pattern is commonly seen in adults, it is rarely seen in the neonatal population. Herein, we describe a patient who developed extensive necrotizing fasciitis of the abdominal wall after intestinal resection for necrotizing enterocolitis (NEC).

Abdominal Wall↗
PubMed2005

Opposing roles for p21(waf1/cip1) and p27(kip1) in enterocyte differentiation, proliferation, and migration.

BACKGROUND: Originating from proliferating stem cells of the intestinal crypt, enterocytes differentiate as they migrate up the crypt-villus axis. A regulatory role of the cyclin-dependent kinase inhibitors p21(waf1/cip1) and p27(kip1) in these processes has been suggested by in vitro models. We sought to determine the effect of p21(waf1/cip1) and p27(kip1) deficiency on enterocyte differentiation, proliferation and migration. METHODS: Three strains of mice including control (C57Bl/6), p27(kip1)-null, and p21(waf1/cip1)-null were studied. Enterocyte differentiation was evaluated by immunostaining for intestinal alkaline phosphatase, by colorimetric assaying for intestinal alkaline phosphatase and sucrase enzyme activity, and by polymerase chain reaction for intestinal fatty acid-binding protein and villin-messenger RNA in enterocytes extracted by laser capture microdissection. Rates of enterocyte proliferation and migration were determined by 5-bromo 2-deoxyuridine immunostaining after a 50% small-bowel resection (SBR). RESULTS: Compared with controls, p27(kip1)-null mice demonstrated minimal differentiation but maintained a normal proliferative response to SBR. Contrarily, p21(waf1/cip1)-null mice demonstrated greater enterocyte differentiation without significant increases in enterocyte proliferation after SBR. CONCLUSIONS: These findings suggest that p21(waf1/cip1) and p27(kip1) have distinctive and opposing roles in the pathogenesis of enterocyte differentiation, proliferation, and migration.

Animals↗
PubMed2005

Role of epidermal growth factor in the pathogenesis of neonatal necrotizing enterocolitis.

Neonatal necrotizing enterocolitis (NEC) is an increasingly frequent condition encountered in premature infants for which the etiology is not well understood. Epidermal growth factor (EGF) is abundant in many fluids bathing the fetal and neonatal gastrointestinal tract, including amniotic fluid, saliva, and breast milk. EGF is acknowledged to be important for normal intestinal development as well as repair following injury to the gastrointestinal mucosa. There appears to be mounting evidence to support a possible link between deficient EGF production and the development of NEC. The relevant evidence for the role of EGF in intestinal development and mucosal repair, as well as its potential involvement in the genesis of NEC will be reviewed.

Animals↗