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Biomedical subjects

Bruce W Robb

Publications and source records attributed to Bruce W Robb.

At least 19 recordsLinked to original sources
PubMed2006

Sentinel lymph node biopsy in thin melanoma patients.

BACKGROUND: We sought to determine the incidence of positive sentinel lymph nodes in thin melanoma ( or= III and thicker lesions (mean 0.77 mm vs. 0.47 mm), but were not different in regards to age, sex, or lesion location. Eight percent and 58% of sentinel nodes were positive by routine histology and RT-PCR, respectively. Among mapped patients, younger age was the only significant prognostic factor for node positivity. With a median follow-up of 18 months among all patients, one regional recurrence (at 2 years) has been identified. CONCLUSIONS: Given the low morbidity of sentinel lymph node biopsy, this procedure should be discussed with selected thin melanoma patients to detect microscopic disease, however PCR positivity by our methods is too commonly seen to be clinically significant in thin melanoma patients and requires additional study.

Adult↗
PubMed2005

Bile salt diarrhea.

Alterations in bile acid metabolism and in the enterohepatic circulation are often associated with chronic diarrhea and should be considered when more common causes of chronic diarrhea have been excluded. Bile acid diarrhea most often occurs in disease or resection of the terminal ileum, in which there is increased exposure of the colonic mucosa to bile salts with consequent activation of fluid and electrolyte secretion. Congenital or acquired defects in the enterohepatic circulation of bile acids also may lead to diarrhea. Although multiple diagnostic tests may be considered to confirm abnormal fecal bile acid losses, the most critical elements of the clinical evaluation of suspected bile acid diarrhea are a careful history to exclude more common causes of chronic diarrhea and a diagnostic trial of bile acid-binding resins.

Bile Acids and Salts↗
PubMed2004

Superinduction of IL-6 by cycloheximide is associated with mRNA stabilization and sustained activation of p38 map kinase and NF-kappaB in cultured caco-2 cells.

Protein synthesis inhibitors paradoxically increase the expression of early-gene products, including various cytokines, through a process known as superinduction. Superinduction is cell-specific and the mechanisms involved are not fully understood but are usually attributed to decreased mRNA degradation. There is, however, increasing evidence that activation of signaling cascades and increased transcriptional activation may be involved as well. Recent studies suggest that IL-6 production in the intestinal mucosa is particularly important due to its anti-inflammatory and protective effects. The effect of protein synthesis inhibitors on IL-6 production in enterocytes, however, is unknown. Treatment of Caco-2 cells with cycloheximide (10 microg/ml) increased IL-6 mRNA and protein levels in IL-1beta-treated cells and this was associated with increased mRNA stabilization. In addition, cycloheximide suppressed IkappaBalpha resynthesis and prolonged p38MAP kinase activation and these changes were associated with sustained activation of the transcription factor NF-kappaB. NF-kappaB activation, in turn, was prevented by the specific p38MAP kinase inhibitor SB208350. Our results suggest that superinduction of IL-6 by cycloheximide in enterocytes results from both increased mRNA stabilization and upregulated transcriptional activity mediated by prolonged activation of the p38 MAP kinase and NF-kappaB.

Blotting, Western↗
PubMed2003

Sodium arsenite downregulates transcriptional activity of AP-1 and CRE binding proteins in IL-1beta-treated Caco-2 cells by increasing the expression of the transcriptional repressor CREMalpha.

In recent studies, sodium arsenite (SA) inhibited IL-6 production in cultured intestinal epithelial cells, at least in part by downregulating the activity of nuclear factor-kappaB (NF-kappaB). The influence of SA on the activity of other transcription factors regulating the interleukin-6 (IL-6) gene in enterocytes is not known. We tested the effect of SA on the activity of CCAAT/enhancer binding protein (C/EBP), activating protein-1 (AP-1), and CRE binding proteins in IL-1beta-treated Caco-2 cells. DNA binding activity was determined by electrophoretic mobility shift assay (EMSA) and transcriptional activity by transfecting cells with luciferase reporter plasmids containing promoter constructs with binding sites for the individual transcription factors. DNA binding activity for all three transcription factors was increased after treatment with SA or IL-1beta. In contrast, SA inhibited transcriptional activity of AP-1 and CRE binding proteins but not C/EBP. Additional experiments provided evidence that the inhibition of AP-1 and CRE mediated transcriptional activity was associated with, and probably caused by, increased expression of the transcriptional repressor cyclic AMP response element modulator (CREM)alpha. The present results are consistent with the concept that SA inhibits IL-6 production in stimulated enterocytes by downregulating the transcriptional activity of several, but not all, IL-6-related transcription factors. Because of the multiple important biological functions of IL-6 in the enterocyte and gut mucosa, methods to regulate enterocyte IL-6 production have significant clinical implications.

Arsenites↗
PubMed2003

Interleukin-6 induces thermotolerance in cultured Caco-2 cells independent of the heat shock response.

In recent studies, induction of the heat shock response increased IL-6 production in gut mucosa in vivo and in cultured Caco-2 cells in vitro. The heat shock response is associated with increased survival of cells exposed to otherwise lethal hyperthermia, so called thermotolerance, but the role of IL-6 in the induction of thermotolerance is not known. We tested the hypothesis that treatment of cultured Caco-2 cells with IL-6 results in the development of thermotolerance. Cells were treated with human recombinant IL-6 for 1h followed by 3 h recovery in cytokine-free medium whereafter cells were exposed to heat stress (48 degrees C for 2 h). In untreated cells, the heat stress resulted in an approximately 80% cell death. In cells treated with IL-6, cell viability after heat stress was significantly improved and was doubled at an IL-6 concentration of 20 ng/ml. Treatment of the cells with other cytokines (IL-4, IL-10, IL-1beta, or TNFalpha) did not induce thermotolerance, suggesting that the effect of IL-6 may be specific for this cytokine. The induction of thermotolerance by IL-6 was blocked by an IL-6 receptor antibody, suggesting that the development of thermotolerance was receptor-mediated. Treatment of cells with IL-6 did not induce an heat shock response as suggested by unaltered heat shock protein 70 and 90 levels and unaffected heat shock factor DNA binding activity. In addition, the IL-6-induced thermotolerance was not inhibited by quercetin. The present study provides the first evidence of IL-6-induced thermotolerance and suggests that this effect of IL-6 is independent of the heat shock response.

Animals↗
PubMed2003

Restorative proctocolectomy with ileal pouch-anal anastomosis in very young patients with refractory ulcerative colitis.

BACKGROUND/PURPOSE: Restorative proctocolectomy with ileal pouch-anal anastomosis (IPAA) is the standard surgical treatment for ulcerative colitis (UC). The purpose of the current study was to determine the outcome of very young patients (< or =10 years of age) with UC undergoing IPAA. METHODS: Between 1978 and 2002, 13 patients 10 years of age or younger underwent IPAA for management of UC at the authors' institution. Charts were reviewed for patient characteristics, and a standardized telephone interview was performed. RESULTS: Average age at diagnosis was 4.0 years (range, 1.0 to 8.4 years), and patients underwent IPAA at a mean of 6.8 years (range, 3.7 to 10.8 years). Pancolitis was present in 100%. The mean follow-up was 9.1 years (1.0 to 16.1 years), the average number of stools per day was 5 (3 to 8). All patients are continent while awake. Pouchitis was documented in 9 patients (75%). All patients or their parents rated the outcome of their procedures as "excellent." CONCLUSIONS: When compared with older children, very young patients with UC tend to have more frequent total colonic involvement and a greater frequency of pouchitis after IPAA. The functional outcome and patient/family satisfaction with the procedure endorse IPAA as an attractive procedure even in the very young population with UC.

Anal Canal↗
PubMed2003

IL-1beta stimulates IL-6 production in cultured skeletal muscle cells through activation of MAP kinase signaling pathway and NF-kappa B.

Recent studies suggest that the skeletal muscle may be a significant site of IL-6 production in various conditions, including exercise, inflammation, hypoperfusion, denervation, and local muscle injury. The mediators and molecular mechanisms regulating muscle IL-6 production are poorly understood. We tested the hypothesis that IL-6 production in muscle cells is regulated by IL-1beta and that mitogen-activated protein (MAP) kinase signaling and NF-kappaB activation are involved in IL-1beta-induced IL-6 production. Cultured C2C12 cells, a mouse skeletal muscle cell line, were treated with different concentrations (0.1-2 ng/ml) of IL-1beta in the absence or presence of the p38 MAP kinase inhibitor SB-208350 or the p42/44 inhibitor PD-98059. Protein and gene expression of IL-6 were determined by ELISA and real-time PCR, respectively. NF-kappaB DNA binding activity was determined by electrophoretic mobility shift assay and by transfecting myocytes with a luciferase reporter plasmid containing a promoter construct with multiple repeats of NF-kappaB binding site. Treatment of myotubes with IL-1beta resulted in a dose- and time-dependent increase of IL-6 production accompanied by an approximately 25-fold increase in IL-6 mRNA levels. IL-1beta stimulated NF-kappaB DNA binding activity and gene activation. SB-208350 and PD-98059 inhibited the increase in IL-6 production induced by IL-1beta. The present results support the concept that skeletal muscle is an important site of IL-6 production. In addition, the results suggest the IL-1beta regulates muscle IL-6 production at least in part by activating the MAP kinase pathway and NF-kappaB.

Animals↗
PubMed2002

Tripeptidyl-peptidase II expression and activity are increased in skeletal muscle during sepsis.

Ubiquitin-proteasome-dependent protein degradation plays a central role in sepsis-induced muscle wasting. Because the proteasome degrades proteins into small peptides rather than free amino acids, it is likely that additional mechanisms downstream of the proteasome are involved in sepsis-induced muscle proteolysis. Recent studies suggest that the extralysosomal peptidase tripeptidyl-peptidase II (TPP II) degrades peptides generated by the proteasome. We hypothesized that TPP II expression and activity are increased in skeletal muscle during sepsis. Sepsis was induced in rats by cecal ligation and puncture. Control rats were sham-operated. TPP II activity was determined by using the specific substrate Ala-Ala-Phe-7-amido-4-methylcoumarin (AAF-AMC). TPP II protein and gene expression were determined by Western blot and real-time PCR, respectively. Sepsis resulted in increased activity and protein and gene expression of TPP II in extensor digitorum longus muscles. This result was blunted by the glucocorticoid receptor antagonist RU 38486, indicating that glucocorticoids participate in the upregulation of TPP II in skeletal muscle during sepsis. The results suggest that proteolytic mechanisms downstream of the proteasome may be important for the complete degradation of muscle proteins during sepsis.

Aminopeptidases↗
PubMed2002

Proteasome inhibitors activate the transcription factors C/EBP-beta and delta in human intestinal epithelial cells.

In recent studies, induction of the heat shock response by hyperthermia upregulated the expression and DNA binding activity of the transcription factor C/EBP. This is an important observation because it may at least in part explain why the heat shock response upregulates IL-6 production in the intestinal mucosa and in the enterocyte. A novel method to induce the heat shock response is proteasome inhibition. The influence of this treatment on the expression and DNA binding activity of C/EBP is not known. We treated cultured Caco-2 cells, a human intestinal epithelial cell line, with one of the proteasome inhibitors, MG-132 or lactacystin, and measured C/EBP-beta and delta DNA binding activity by electrophoretic mobility shift assay and supershift analysis. In addition, nuclear levels of C/EBP-beta and delta protein were determined by Western blot analysis. Treatment of the cells with the proteasome inhibitors resulted in increased cellular levels of heat shock protein 72, consistent with induction of the heat shock response. Treatment also resulted in increased DNA binding activity and nuclear protein levels of C/EBP-beta and delta. The effects of the proteasome inhibitors on C/EBP were inhibited by treating the cells with quercetin, a substance known to block the heat shock response. The results suggest that proteasome inhibition activates the transcription factors C/EBP-beta and delta in human intestinal epithelial cells and that this response, at least in part, is caused by induction of the heat shock response. The observations are important because they provide support for a novel method to influence gene activation in the enterocyte.

Acetylcysteine↗
PubMed2002

Arsenite stabilizes IkappaBalpha and prevents NF-kappaB activation in IL-1 beta-stimulated Caco-2 cells independent of the heat shock response.

Recent studies suggest that sodium arsenite downregulates NF-kappaB activity by inhibiting phosphorylation and subsequent degradation of IkappaBalpha. Many effects of sodium arsenite are secondary to induction of heat shock proteins. The role of the heat shock response in arsenite-induced inhibition of NF-kappaB, however, is not known. We examined the involvement of the heat shock response in arsenite-induced inhibition of NF-kappaB activity in IL-1beta-stimulated Caco-2 cells, a human colorectal adenocarcinoma cell line with enterocytic properties. Treatment of the cells with IL-1beta resulted in increased IkappaB kinase activity, reduced levels of IkappaBalpha and increased NF-kappaB DNA binding activity. Sodium arsenite blocked all of these responses to IL-1beta without inducing changes in heat shock factor activity or heat shock protein levels. Results from additional experiments showed that the protective effect of sodium arsenite on IkappaBalpha was not influenced by the oxygen radical scavenger catalase or by inhibitors of the MAP-kinase signaling pathway. The present results suggest that sodium arsenite stabilizes IkappaBalpha and prevents NF-kappaB activation in IL-1beta-stimulated Caco-2 cells independent of the heat shock response. In addition, stabilization of IkappaBalpha by sodium arsenite does not require oxygen radical formation or activation of the MAP kinase signaling pathway.

Arsenites↗
PubMed2002

Transcription factors C/EBP-beta and -delta regulate IL-6 production in IL-1beta-stimulated human enterocytes.

In recent studies, treatment with IL-1beta of cultured Caco-2 cells, a human intestinal epithelial cell line, resulted in transcriptional upregulation of IL-6 production. The role of C/EBP-beta and -delta in enterocyte IL-6 production is not known. Stimulation with IL-1beta of Caco-2 cells transiently transfected with a luciferase reporter plasmid containing a wild-type IL-6 promoter resulted in an approximately 3.5-fold increase in luciferase activity. This effect of IL-1beta was reduced by approximately 30% when the C/EBP binding site in the IL-6 promoter was mutated, supporting a role of C/EBP in the regulation of IL-6 production. When Caco-2 cells were treated with IL-1beta in the presence of the MAPK inhibitor, PD-98059, IL-6 mRNA and protein levels were reduced by the same concentrations of PD-98059 that inhibited C/EBP DNA binding activity in previous studies. Finally, overexpression of C/EBP-beta and -delta in IL-1beta-treated Caco-2 cells resulted in a 10-12-fold increase in IL-6 production. The results suggest that the beta and delta isoforms of the C/EBP family of transcription factors at least in part regulate IL-6 production in human intestinal epithelial cells.

CCAAT-Enhancer-Binding Protein-beta↗
PubMed2002

Interposition of a jejunal segment as an alternative one-stage operation for acute left colon obstruction.

PURPOSE: Much debate surrounds the one-stage surgical management of acute left colon obstruction. Many surgeons are still reluctant to perform primary anastomosis in unprepared bowel fearing the risk of anastomotic dehiscence. Although intraoperative lavage and subtotal colectomy have recently proved effective for preventing fecal loading, both of these procedures have drawbacks and the search for additional alternative surgical procedures continues. We conducted an experimental study to examine a new one-stage operation, consisting of the interposition of a jejunal segment to the resection site of the colon, to manage acute left colon obstruction. METHODS: The colons of 20 domestic pigs were obstructed, and 2 days later, segmental resection and either primary colo-colonic anastomosis or interposition of a jejunal segment was performed. Autopsies were done 6 weeks after the operations and the anastomotic regions were subjected to bursting pressure analysis. RESULTS: The operating time was longer in the interposition group at 49 +/- 14 vs 38 +/- 12 min ( P < 0.01). There was no mortality or clinical signs of anastomotic dehiscence in either group. A perianastomotic abscess was detected on autopsy in one animal from the primary colo-colonic anastomosis group. The bursting pressures of the jejunocolic anastomoses were significantly higher ( P < 0.001) than those of the primary colo-colonic anastomoses. CONCLUSIONS: These experimental results suggest that jejunal interposition may be a feasible alternative to primary colonic anastomosis to create intestinal continuity after resection for acute obstruction of the left colon.

Anastomosis, Surgical↗
PubMed2002

Early debridement for necrotizing pancreatitis: is it worthwhile?

BACKGROUND: The timing for debridement of necrotizing pancreatitis is controversial. We reviewed our experience with early and delayed surgical debridement in patients with necrotizing pancreatitis. STUDY DESIGN: The records of patients diagnosed with acute necrotizing pancreatitis from January 1993 through June 2000 were reviewed retrospectively. Data were analyzed with respect to Ranson's, APACHE II, and multiple organ failure scores, etiology, presence of infection, overall and ICU length of stay, time to first debridement, number of debridements, fluid requirements, days to enteral feeding, transfusion requirements, complications, and mortality. RESULTS: Twenty-six patients (18 males, 8 females, mean age 51 years) were diagnosed with acute necrotizing pancreatitis. The admission Ranson's score was 4.8, the APACHE II score was 11.7, and multiple organ failure score was 4.2. All but one patient underwent pancreatic debridement (4.3 debridements per patient). Eighteen patients (69%) had infected pancreatic necrosis. The timing of debridement was based on patients' condition and surgeon's preference. The presentation and demographics of patients who underwent early (<2 weeks) or late (>2 weeks) debridement did not differ significantly. Patients debrided early had a trend toward higher mortality (29% versus 18%) and experienced a higher number of major complications (p < 0.05). The six patients (23%) who died were older, had multiple organ failure scores, and more often had Candida in the infected necrosis (p < .05). CONCLUSIONS: Early debridement for acute necrotizing pancreatitis might not improve survival and might even be associated with increased number of complications. Most patients diagnosed with necrotizing pancreatitis eventually need debridement, but it might be beneficial to delay debridement if the patient's condition allows for it.

Age Factors↗
PubMed2002

Hyperthermia-induced heat shock activates the transcription factor c/EBP-beta and augments IL-6 production in human intestinal epithelial cells.

BACKGROUND: Interleukin (IL)-6 production is increased in gut mucosa during sepsis and endotoxemia. The heat shock response augments IL-6 production under these conditions, but the mechanism is not known. We hypothesized that heat shock stimulates IL-6 production in enterocytes by increasing expression and activity of the transcription factor C/EBB. STUDY DESIGN: Cultured Caco-2 cells, a human intestinal epithelial cell line, underwent induction of the heat shock response by hyperthermia (43 degrees C for 1 hour). Other cells were kept at 37 degrees C. Cells were then treated with 0.5 ng/mL human recombinant IL-1beta for 4 hours. C/EBP-beta and delta DNA binding activity was determined by electrophoretic mobility shift assay and supershift analysis. In additional experiments, Caco-2 cells were transfected with expression plasmids for C/EBP-beta and delta, after which cells were subjected to hyperthermia and treatment with IL-1beta. RESULTS: C/EBP-beta, but not delta, protein levels and DNA binding activity were increased in Caco-2 cells expressing the heat shock response. Induction of the heat shock response augmented IL-6 production in IL-1beta-treated cells overexpressing C/EBP-beta, but not delta. CONCLUSIONS: Increased IL-6 production in IL-1beta-treated enterocytes expressing the heat shock response might be caused by upregulated expression and activity of CIEBP-beta. Because recent studies suggest that IL-6 might be an antiinflammatory cytokine and might exert protective effects in gut mucosa and enterocytes, understanding mechanisms by which the heat shock response augments IL-6 production might have important clinical implications.

CCAAT-Enhancer-Binding Proteins↗
PubMed2002

IL-1beta activates C/EBP-beta and delta in human enterocytes through a mitogen-activated protein kinase signaling pathway.

The enterocyte is an active participant in the inflammatory and metabolic response to sepsis, endotoxemia and other critical illnesses and is the site for cytokine and acute phase protein production in these conditions. The role of the CCAAT/enhancer binding protein (C/EBP) family of transcription factors in the response to inflammatory stimuli in the enterocyte is not well understood. In the present study, we treated Caco-2 cells with IL-1beta and determined C/EBP DNA binding activity by electrophoretic mobility shift assay. The involvement of the alpha, beta, and delta isoforms was determined by supershift analysis and Western blot analysis of proteins from the nuclear fraction. The role of the mitogen-activated protein kinase (MAPK) signaling pathway was assessed by treating cells with the MAPK inhibitor PD-98059. Treatment of the Caco-2 cells with IL-1beta resulted in increased CCAAT/enhancer binding protein DNA binding activity. Supershift analysis and Western blotting indicated that this response to IL-1beta mainly reflected the delta isoform, and to a lesser degree the beta isoform. Treatment of the cells with PD-98059 inhibited the IL-1beta-induced increase in beta and delta activity. The results suggest that members of the CCAAT/enhancer binding protein family of transcription factors are activated in enterocytes during inflammatory conditions characterized by high levels of IL-1beta.

CCAAT-Enhancer-Binding Protein-beta↗
PubMed2002

Arsenite inhibits interleukin-6 production in human intestinal epithelial cells by down-regulating nuclear factor-kappaB activity.

Previous studies have suggested that the production of interleukin-6 (IL-6) is increased in the intestinal mucosa during inflammation, and that nuclear factor-kappaB (NF-kappaB) is an important regulator of the IL-6 gene in the enterocyte. We tested the hypothesis that sodium arsenite inhibits IL-6 production in stimulated enterocytes and that this effect of arsenite is caused by down-regulation of NF-kappaB activity. Cultured Caco-2 cells were treated with sodium arsenite and were then stimulated with IL-1beta. IL-6 production and gene expression were determined by ELISA and reverse transcriptase-PCR respectively. NF-kappaB DNA binding activity was determined by electrophoretic mobility shift assay. IL-1beta increased NF-kappaB DNA binding activity, IL-6 mRNA levels and IL-6 production. These effects of IL-1beta were inhibited by treatment of the cells with sodium arsenite in a dose- and time-dependent fashion. When cells were transfected with a plasmid expressing the p65 subunit of NF-kappaB, the inhibitory effect of sodium arsenite on NF-kappaB activity and IL-6 production was blunted. These results suggest that sodium arsenite inhibits IL-6 production in enterocytes subjected to an inflammatory stimulus, and that this effect, at least in part, reflects down-regulated NF-kappaB activity.

Arsenites↗