A novel stable reproducible model of hepatic failure in canines.

BACKGROUND: Stable and reproducible large animal models of hepatic failure, which allow the assessment of liver-assist devices, are not available. Our objective was to develop a physiologically stable animal model of hepatic failure on which the safety and efficacy of an extracorporeal liver-assist device can be tested. We hypothesized that a surgical model which consists of an end-to-side portocaval shunt combined with common bile duct ligation and transection would create hepatic failure with: (1) elevations in amino transferases, total bilirubin, and ammonia; (2) a decrease in the ratio of branched chain to aromatic amino acids; and (3) histologic evidence of hepatic injury. METHODS: Eleven mongrel dogs underwent common bile duct transection and an end-to-side portocaval shunt. Aminotransferases (AST, ALT), total bilirubin, ammonia, and branched chain and aromatic amino acids were measured prior to operation (baseline) and after 9 days. A necropsy was performed on Postoperative Day 9 and liver biopsies were obtained for histology. RESULTS: By Postoperative Day 9, AST, ALT, total bilirubin, and ammonia values were significantly elevated compared to baseline (P < 0.02). The ratio of branched chain to aromatic amino acids was significantly reduced compared to baseline (P < 0.003). There was histologic evidence of cholestasis and inflammation. CONCLUSION: Portocaval shunt with common bile duct transection produces liver failure with elevations in aminotransferases, total bilirubin, and ammonia, a decreased branched chain to aromatic amino acid ratio, and histologic inflammation. Unlike ischemic or chemically induced models of liver failure, the dogs were hemodynamically and neurologically stable. This model can be used to test the safety and efficacy of liver-assist devices aimed at temporizing the detoxification functions of the failing liver.

Predictors of malignancy in childhood peripheral lymphadenopathy.

PURPOSE: The aim of this study was to identify factors associated with malignant etiologies of childhood peripheral lymphadenopathy and to construct a model that may be used to assess the risk of malignancy. METHODS: The medical records of 60 consecutive patients 18 years old or less who underwent peripheral lymph node biopsies were reviewed. RESULTS: Increasing node size, number of sites of adenopathy, and age were associated with an increasing risk of malignancy (P < .05 for all variables). Graphs useful for risk determination were constructed based on these variables. Additional factors associated with malignancy included the presence of supraclavicular adenopathy (P < .01), an abnormal chest x-ray (P < .01), and fixed nodes (P < .01). Variables that were not statistically different between patients with benign and malignant adenopathy included the duration of adenopathy (P = .43), the presence of fever (P = .36), cough (P = .14), splenomegaly (P = .93), skin involvement (P = .39), tenderness (P = .49), and bilateral adenopathy (P = .39). Fluctuance was associated with benign adenopathy (P < .04). CONCLUSIONS: The risk of malignancy increased with increasing size and number of sites of adenopathy and age. Other significant predictors of malignancy included supraclavicular location, an abnormal chest x-ray, and fixed nodes. These data may be used to supplement clinical judgment to predict the risk of malignancy.

The utility of technetium 99m pertechnetate scintigraphy in the evaluation of patients with Meckel's diverticulum.

PURPOSE: The aim of this study was to assess the utility of technetium (Tc) 99m pertechnetate scintigraphy in the diagnostic workup of the pediatric patient with gastrointestinal (GI) bleeding and a suspected Meckel's diverticulum. METHODS: The charts of 235 consecutive patients evaluated with a Meckel's scan (n = 165) or with the discharge diagnosis of Meckel's diverticulum (n = 70) between January 1975 and October 1997 were reviewed for presenting symptoms, bleeding characteristics, diagnostic studies and pathological diagnosis. Those patients with lower GI bleeding and a serum hemoglobin level less than 11.0 g/dL who underwent a 99mTc pertechnetate scan (n = 43) were assessed for utility of the scan. RESULTS: In all patients the Meckel's scan had a positive and negative predictive value of 0.93. However, in patients with lower GI bleeding and a hemoglobin less than 11.0 g/dL the Meckel's scan had a sensitivity of 0.60, a positive predictive value of 1.0, a specificity of 0.96, but only a negative predictive value of 0.74. As such, the probability that a child who presents with GI bleeding and a serum hemoglobin less than 11 g/dL will have a Meckel's diverticulum despite a negative Meckel's scan of 0.26. We further evaluated the eight patients with a false-negative scan: ectopic gastric mucosa was present on pathological examination in all eight patients. Pentagastrin stimulation was performed at the time of scintigraphic study in three of eight. Six of these eight patients had duplicate scans that also were negative. Patients with a false-negative (FN) scan had significantly increased hospital charges when compared with those with a true positive (TP) scan (TP = $5012 +/- 1992; FN = $8554 +/- 1506; P = .0001). Clinical suspicion had a major effect on the decision-making process in these patients independent of the results of the Meckel's scan, and all eight patients ultimately underwent exploratory laparoscopy-laparotomy with Meckel's diverticulectomy despite the scan results. CONCLUSIONS: The relatively low negative predictive value of the Meckel's scan may result in the need for operative evaluation despite the scan data. As such, the contribution of the scan to clinical decision making is low. These findings suggest that exploratory laparotomy or laparoscopy may be indicated instead of scintigraphic scanning in the assessment of the anemic (hemoglobin less than 11 g/dL) pediatric patient with lower GI bleeding, especially in patients in whom a high suspicion for a bleeding Meckel's diverticulum exists.

Can the clearance of tumor necrosis factor alpha and interleukin 6 be enhanced using an albumin dialysate hemodiafiltration system?

Patients with acute hepatic failure (AHF) have elevated levels of inflammatory cytokines such as tumor necrosis factor alpha (TNF-alpha) and interleukin 6 (IL-6). Recently, we have shown selective hemodiafiltration with albumin dialysis, as an extracorporeal liver support device (ECLVS), to be effective in the clearance of multiple toxins that are elevated in AHF. Our objective was to evaluate whether ECLVS would be effective in the clearance of TNF-alpha and IL-6. An in vitro continuous hemodiafiltration circuit was used with single pass counter-current dialysis. A known amount of recombinant rat TNF-alpha and IL-6 was added to heparinized bovine blood and filtered across a polyalkyl sulfone hemofilter using matched filtration and dialysate flow rates. During 4 hours, the serial TNF-alpha and IL-6 concentrations were measured in the circulating blood, and the content of each cytokine was calculated using mass balance. For each cytokine, clearance was determined for two dialysate groups at constant temperature and pH (group 1: dialysate = 0.9 normal saline, n = 5; group 2: dialysate = albumin 2 gm/dl, n = 5). Analysis of data was performed using ANOVA and Student's t-test. There was improved clearance of TNF-alpha and IL-6 when albumin was used in the dialysate (81+/-0.09% of the initial TNF-alpha and 77+/-0.04% of the IL-6 quantities) compared with when 0.9 normal saline was used as the dialysate (58+/-0.14% of the initial TNF-alpha and 56+/-0.18% of the IL-6 quantities); p < 0.03. An ECLVS utilizing hemodiafiltration with albumin dialysis is more effective than conventional hemofiltration in the clearance of TNF-alpha and IL-6 and, therefore, may benefit patients with acute hepatic failure.

Differential expression of Hsp27 in normal oesophagus, Barrett's metaplasia and oesophageal adenocarcinomas.

The protein expression patterns of normal, metaplastic and malignant oesophageal tissues were analysed by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) to identify changes associated with Barrett's metaplasia and transformation to oesophageal adenocarcinoma. Heat-shock protein 27 (Hsp27), a small heat-shock protein which is protective against cytotoxic stresses, was abundant in normal oesophagus. However, Hsp27 expression was markedly lower in Barrett's metaplasia and oesophageal adenocarcinomas. This was confirmed by immunohistochemical analysis. Hsp27 protein was most highly expressed in the upper layers of squamous epithelium and exhibited a pattern of expression that corresponded with the degree of squamous maturation. Northern and Southern analysis demonstrated Hsp27 to be regulated at the level of mRNA transcription or abundance. Normal oesophageal tissues were examined for gender differences in Hsp27 expression. Women expressed fourfold higher levels of Hsp27 mRNA, however, this difference was not appreciable in protein expression. Hsp27 protein was inducible by heat shock in Barrett's adenocarcinoma cell lines and an immortalized oesophageal epithelial cell line (HET-1A), but not by oestradiol. These results demonstrate abundant constitutive expression of the stress-response protein Hsp27 in the normal oesophagus, and suggest that low-level expression in Barrett's metaplasia may be one factor which may influence susceptibility to oesophageal adenocarcinoma development.

Use of extracorporeal life support for adult patients with respiratory failure and sepsis.

Traditionally, adult sepsis has been considered a contraindication to extracorporeal life support (ECLS). The objective of this study was to review the authors' institutional experience with a subgroup of adult patients requiring ECLS for severe respiratory failure and sepsis. Hospital records from 100 consecutive adult patients with respiratory failure placed on ECLS between 1990 and 1996 were retrospectively reviewed. Patients with sepsis as a primary indication were identified, and blood culture data reviewed. Data were analyzed with t tests and chi-square and are presented as mean +/- standard deviation. Multiple logistic regression determined the impact of sepsis and positive blood cultures (PBCs) on survival. Fourteen patients required ECLS for sepsis; 36 had PBCs during hospitalization (15 before or during ECLS). Septic patients had lower pre-ECLS PaO2/FIO2 ratios (septic: 53 +/- 14 mmHg, nonseptic: 70 +/- 68 mmHg, p = 0.04). Patients with PBCs before or during ECLS were younger (PBC: 29 +/- 6 years, no PBC: 35 +/- 13 years, p = 0.003), remained on ECLS longer (PBC: 485 +/- 336 hours, no PBC: 232 +/- 212 hours, p = 0.01), and were more frequently cannulated within 12 hours (PBC: 15/15, no PBC 60/85 p = 0.02). Neither group differed in organ dysfunction (incidence or type), frequency of respiratory recovery, or survival. Neither sepsis nor positive blood cultures were independently predictive of mortality. Sepsis and positive blood cultures do not adversely affect outcome in adult patients with respiratory failure requiring ECLS.

Fas/APO-1 (CD95) is not translocated to the cell membrane in esophageal adenocarcinoma.

This study describes Fas (CD95) expression in Barrett's esophagus, adenocarcinomas of the esophagus, and three esophageal adenocarcinoma cell lines. Immunohistochemical analysis of Barrett's esophagus demonstrated cell surface expression of Fas protein. In contrast, 30.5% of esophageal adenocarcinomas examined by immunohistochemical analysis demonstrated faint cytoplasmic staining, and 69.5% were negative for Fas. Similar levels of Fas mRNA were identified in tumors compared to mRNA levels in esophageal squamous mucosa or Barrett's esophagus. An approximately Mr 48,000 Fas protein was identified by Western blot analysis in tumors that were negative for Fas expression by immunohistochemical analysis. The esophageal adenocarcinoma cell line Seg-1 was negative for Fas expression by immunohistochemical analysis, but Western blot analysis demonstrated abundant, appropriately sized Fas protein. In agreement with the immunohistochemical analysis, flow cytometry of Seg-1 showed minimal amounts of Fas on the cell surface, which correlated with resistance to Fas-mediated apoptosis. No mutations in the Seg-1 Fas coding sequence or exon 1 were identified by sequence analysis. This was confirmed by transient transfection of COS cells with expression vectors generated from the Seg-1 Fas cDNA, which resulted in cell surface expression of the Fas protein. Stable transfection of Seg-1 with a Fas expression vector did not result in efficient Fas expression on the cell surface. Seg-1 cells, transiently transfected with a Fas-FLAG expression vector and examined for protein expression using confocal microscopy and an anti-FLAG antibody, showed that the Fas-FLAG protein was not present on the cell surface but was present in the cytoplasm. Taken together, these results indicate that expression of Fas on the cell surface by esophageal adenocarcinoma is reduced. In an esophageal adenocarcinoma cell line, wild-type Fas protein is retained in the cytoplasm, and this correlates with resistance to Fas-mediated apoptosis. The retention of wild-type Fas protein within the cytoplasm may represent a mechanism by which malignant cells evade Fas-mediated apoptosis.