Activation of the alternative pathway of complement during the acute phase of typical haemolytic uraemic syndrome.

Haemolytic uraemic syndrome (HUS) is characterized by haemolytic anaemia, thrombocytopenia and acute renal failure. We studied the activation state of classical and alternative pathways of complement during the acute phase of Shiga toxin-associated HUS by performing a prospective study of 18 patients and 17 age-matched healthy controls to evaluate C3, C3c, C4, C4d, Bb and SC5b-9 levels. SC5b-9 levels were increased significantly in all patients at admission compared to healthy and end-stage renal disease controls, but were significantly higher in patients presenting with oliguria compared to those with preserved diuresis. C3 and C4 levels were elevated significantly at admission in the non-oliguric group when compared to controls. No significant differences were found for C4d values, whereas factor Bb was elevated in all patients and significantly higher in oliguric patients when compared to both controls and non-oliguric individuals. A positive and significant association was detected when Bb formation was plotted as a function of plasma SC5b-9 at admission. Bb levels declined rapidly during the first week, with values not significantly different from controls by days 3 and 5 for non-oligurics and oligurics, respectively. Our data demonstrate the activation of the alternative pathway of complement during the acute phase of Stx-associated HUS. This finding suggests that complement activation may represent an important trigger for the cell damage that occurs during the syndrome.

Culture of porcine hepatocytes: the dogma of exogenous matrix revisited.

The use of exogenous matrices has been described as an essential component in securing the viability and functionality of hepatocytes in vitro whether cultured for extracorporeal devices or cell transplantation. Here we report on the in vitro culture of porcine hepatocytes in polystyrene tissue-culture flasks without exogenous matrices showing adequate attachment and viability. Cell proliferation was evidenced by uptake of 5-bromo-2'-deoxyuridine, with peaks at Days 2 (19.7 +/- 8.5%), 15 (20.8 +/- 3.3%), and 35 (21.4 +/- 0.3%). Detoxification capacity was assessed by determination of monoethylglycinexylidide, a product of lidocaine metabolism (highest value 156.5 +/- 10.1 ng/ml at Day 4), and by diazepam clearance (maximum clearance 66.2% at Day 6). Diazepam metabolite levels were highest at Day 4 both for temazepam and oxazepam (6.5 +/- 0.1 and 0.10 +/- 0.01, respectively). These results suggest that the need for an exogenous matrix to achieve sustained proliferative activity and differentiated hepatocyte function should not necessarily be considered a sine qua non condition.

Hyperhomocysteinemia in stable pediatric, adolescents, and young adult renal transplant recipients.

BACKGROUND: High total plasma homocysteine (tHcy) levels are accompanied by an increased risk for premature development of atherosclerosis and atherothrombosis. Adult renal transplant recipients have elevated tHcy levels. Corresponding data in pediatric, adolescent, and young adult renal transplant recipients are scarce. We investigated whether tHcy levels were elevated in stable renal transplant recipients who received kidney grafts before age 18. METHODS: This cross-sectional study was conducted during routine posttransplantation follow-up. Fasting tHcy levels, serum creatinine, and lipoprotein profile were measured in 38 clinically stable renal transplant recipients with different degrees of renal function. No patient was receiving B vitamin or folic acid supplementation. Estimated glomerular filtration rate (GFR) was assessed according to Schwartz's formula. All patients followed a triple-drug immunosuppressive regimen, with the exception of three patients (deflazacort and azathioprine). Forty-one apparently healthy subjects constituted the control group. tHcy levels were determined by fluorescence polarization immunoassay in an IMx analyzer. RESULTS: Mean tHcy levels in transplant recipients were significantly higher than in controls (16.8+/-8.7 micromol/L and 9.5+/-2.3 micromol/L, respectively; P<0.01). A significant positive correlation between tHcy and serum creatinine levels was observed for both transplant recipients (rS=0.70, P<0.01) and controls (rS=0.54, P<0.01). In transplant recipients, tHcy correlated negatively with estimated GFR (rS=[minus]0.47, P<0.05). Fasting tHcy levels in excess of 14.6 micromol/L (>95th percentile in controls) were present in 19 (50%) patients; 14 of these patients had an estimated GFR<60 ml/min per 1.73 m2. When the renal transplant recipients were analyzed by renal function, mean tHcy was significantly higher in patients with an estimated GFR<60 ml/min per 1.73 m2 compared with patients with an estimated GFR> or =60 ml/min per 1.73 m2 (20.5+/-9.9 vs. 13.2+/-5.8 micromol/L, P<0.01). Both groups were significantly different from controls (P<0.01). No relationship was found between tHcy level and either cumulative cyclosporine or cumulative methylprednisone doses. No differences were observed in tHcy levels or lipoprotein profile between patients who were receiving deflazacort and those on methylprednisone. CONCLUSIONS: Hyperhomocysteinemia in renal transplant recipients is a common condition. Testing for fasting tHcy level might be a useful tool to identify patients at increased risk for development of vascular disease.

Effect of deflazacort versus methylprednisone on growth, body composition, lipid profile, and bone mass after renal transplantation. The Deflazacort Study Group.

Kidney function, growth velocity, weight/height ratio, body composition, lipid profile, and bone mass were studied in a randomized, multicenter trial of deflazacort versus methylprednisone in 27 prepubertal patients with kidney transplantation. Methylprednisone (0.20+/-0.03) was replaced by deflazacort (13 patients, 0.30+/-0.03 mg/kg per day). After 12 months, creatinine clearance decreased significantly only during methylprednisone therapy. Growth velocity increased only in patients treated with deflazacort from 3.3+/-0.6 to 5.6+/-0.5 cm/year. Serum levels of several components of the insulin-like growth factor axis did not change. Weight/height ratio was increased in methylprednisone-treated patients (P<0.05) and decreased in deflazacort-treated patients (P<0.005). Lean body mass increased in both groups (P<0.005). Fat body mass and serum leptin increased only in methylprednisone-treated patients (P<0.025). Total cholesterol and low-density lipoprotein-cholesterol increased in methylprednisone-treated patients by 9.9% (P<0.05) and 12.5% (P<0.025). High-density lipoprotein-cholesterol increased by 21% (P<0.005) and apolipoprotein B decreased by 11% (P<0.005) in deflazacort-treated patients. Total skeleton and lumbar spine bone mineral density decreased in both groups, but at 1 year methylprednisone-treated patients had lost 50% more bone. Bone mineral content decreased only in methylprednisone-treated patients (P<0.01). Our data suggest that substituting deflazacort for maintenance methylprednisone might prevent height loss, excessive bone loss, and fat accumulation; and leads to an improvement in the lipoproteins of these children.

Von Willebrand factor could be an index of endothelial dysfunction in patients with cirrhosis: relationship to degree of liver failure and nitric oxide levels.

BACKGROUND/AIMS: The aim of this study was to evaluate the relationship between plasma levels of von Willebrand factor (vWF), a marker of endothelial cell activation, and nitric oxide, a powerful vasodilator synthesized by endothelial cells, in 27 patients with cirrhosis at different stages of the disease. These results were compared with those of age-matched normal, healthy subjects (n=10). METHODS: vWF:antigen was measured by electro-immunodiffusion test and serum nitrite and nitrate levels, the stable end products of nitric oxide metabolism, were determined by an enzymatic procedure. RESULTS: vWF:antigen and nitrite/nitrate levels were significantly higher in cirrhotic patients (367+/-185% and 29.3+/-10.8 micromol/l) than in healthy subjects (92+/-20% and 19.2+/-8.3 micromol/l, p<0.05, respectively). Higher levels of vWF:antigen and nitrites/nitrates were observed in patients with more advanced degrees of liver failure, as reflected by quantitative Child-Pugh's score (516+/-154% and 38.3+/-7.8 micromol/l in Child-Pugh > or = 9 vs 227+/-61% and 21.0+/-6.1 micromol/l in Child-Pugh <9, p<0.001, respectively). Moreover, both endothelial-related factors were higher in patients with ascites than those without ascites (543+/-158% and 37.8+/-8.9 micromol/l vs 262+/-103% and 24.4+/-8.8 micromol/l, p<0.001, respectively). In the overall series, a highly significant linear correlation between nitrites/nitrates and vWF:antigen levels was observed in patients with cirrhosis (r=0.79, p<0.001). CONCLUSIONS: These results support a cirrhosis-related endothelial dysfunction and suggest that plasma vWF measurement could be useful as a marker of endothelial disturbance in patients with cirrhosis.

Effect of therapy with deflazacort on dyslipoproteinemia after pediatric renal transplantation.

Deflazacort is an oxazolone compound derived from prednisolone, with similar immunosuppressive action but fewer side effects. Kidney function, weight/height ratio, serum triglycerides, cholesterol, high-density lipoprotein (HDL) cholesterol, very-low-density lipoprotein cholesterol, low-density lipoprotein (LDL) cholesterol, apolipoprotein A, apolipoprotein B, and lipoprotein (a) were studied before and 6 months after substitution of deflazacort (mean +/- SEM, 0.3 +/- 0.1 mg/kg per day) for methylprednisone (0.2 +/- 0.1 mg/kg per day) in 14 patients treated with cyclosporine, aged 3.1 to 20.3 years, 3 years after renal transplantation. Serum creatinine and calculated creatinine clearance did not change significantly, and weight/height ratio decreased from 20.0% +/- 7.1% to 12.5% +/- 6.5% (P < .005) during deflazacort therapy. Total cholesterol was reduced by 15.9% (from 233 +/- 15 mg/dL to 196 +/- 13 mg/dL, P < .01), LDL cholesterol by 25.5% (from 153 +/- 14 mg/dL to 114 +/- 12 mg/dL, P < .01), and TC/HDL cholesterol ratio by 28.3% (from 5.3 +/- 0.4 to 3.8 +/- 0.4, P < .01), whereas HDL cholesterol increased 18% (from 45 +/- 2 mg/dL to 53 +/- 2 mg/dL) and apolipoprotein A by 8.3% (from 122 +/- 5 mg/dL to 132 +/- 5 mg/dL, P < .05) during deflazacort therapy. Our data suggest that substituting deflazacort for maintenance methylprednisone therapy leads to an improvement in the lipoprotein profile of children after renal transplantation.

Impacto de la nutrición precoz en el paciente pediátrico critico / Impact early nutrition in the critical pediatric patient

Se evaluó el impacto de la Nutrición precoz de 16 pacientes pediátricos críticos mediante la determinación de la proteína unida al retinol. Los valores hallados fueron de 3,2 +/- 1,83 mg por ciento en la primera determinación dentro de las 48 horas del ingreso; 3,6 mg +/- 1,74 mg por ciento a las 24 horas del comienzo del soporte nutricional enteral y/o parenteral y de 4,1 +/- 2,25 mg por ciento al séptimo día de la terapia nutricional. Observamos que la proteína unida al retinol no se altera en pacientes a quienes se le realiza nutrición precoz.

Impacto de la nutrición precoz en el paciente pediátrico critico / Impact early nutrition in the critical pediatric patient

Se evaluó el impacto de la Nutrición precoz de 16 pacientes pediátricos críticos mediante la determinación de la proteína unida al retinol. Los valores hallados fueron de 3,2 +/- 1,83 mg por ciento en la primera determinación dentro de las 48 horas del ingreso; 3,6 mg +/- 1,74 mg por ciento a las 24 horas del comienzo del soporte nutricional enteral y/o parenteral y de 4,1 +/- 2,25 mg por ciento al séptimo día de la terapia nutricional. Observamos que la proteína unida al retinol no se altera en pacientes a quienes se le realiza nutrición precoz. (AU)

Ascitic fluid alpha 1-antitrypsin.

Ascitic fluid alpha 1-antitrypsin (AF-AAT) was compared with ascitic fluid total protein (AF-TP) and the serum-ascites albumin gradient (SAAG) in the differential diagnosis of ascites. The study included 82 consecutive patients of which 42 had cirrhosis, 8 hepatoma (with cirrhosis), and 27 malignant ascites (peritoneal 18, liver 9). The concentration of AF-AAT (milligrams per deciliter) was significantly elevated (P less than 0.001) in hepatoma (174 +/- 123), malignant liver disease (232 +/- 119) and peritoneal neoplasms (376 +/- 106) in comparison with cirrhotics (66 +/- 33). In separating ascites caused by cirrhosis or malignancy, AF-AAT (discriminating limit of 120 mg/dl) had a 96% sensitivity, 95% specificity, and 96% diagnostic efficacy, which was superior to the 87% observed for AF-TP and 86% for the SAAG. Similar results were obtained for the A/S AAT ratio but this test was not available in all patients. AF-AAT was particularly useful in patients with malignancy causing portal hypertension as assessed by SAAG (hepatoma, malignant liver disease). We conclude that AF-AAT may be a valuable parameter in the differential diagnosis of ascites.

Serum alkaline phosphatase isoenzymes in hepatobiliary disorders resolved by use of immobilized pH gradients.

This new method for fractionating alkaline phosphatase isoforms in hepatobiliary disorders is based on isoelectric focusing on a mixed-type polyacrylamide support containing an immobilized pH gradient with a superimposed carrier-ampholyte gradient. The high-Mr alkaline phosphatase forms typical of hepatobiliary disease (greater than 1 mega-dalton), which cannot migrate into the Immobiline gel, are disaggregated in zwitterionic detergents (the most effective being sulfobetaine 3-12)--20 g/L in the sample, 5 g/L in the gel--suggesting that they are still complexed with membrane fragments or that they tend to aggregate spontaneously in solution. These isoforms focus in the pI 5-6 range (while alkaline phosphatases in normal serum focus in the pI 4-5 interval) in immobilized pH gradients, but behave as strongly acidic components by agarose isoelectric focusing in the presence of carrier ampholytes, suggesting that they are strongly complexed with the latter. On treatment with neuraminidase, the low-pI isoforms in normal serum focus in the pI 5-6 range typical of the hepatobiliary isoforms, suggesting that the latter are poorly glycosylated. By a second-dimension run, in a porosity gradient, followed by activity staining, all alkaline phosphatase forms that have entered the Immobiline gel in the first dimension (normal forms and high-Mr species) exhibit the same Mr (ca. 140,000 Da), suggesting that no new chains are synthesized in hepatobiliary disorders.

Concentración sérica de alfa-1-antitripsina en diversas enfermedades hepáticas / Serum concentration of alpha-1-antitrypsin in different hepatic diseases

Se investigó la concentración sérica de alfa-1-antitripsina (A1AT) en 195 pacientes con diversas hepatopatías y en 43 adultos sanos. En todas las enfermedades hepáticas estudiadas se encontró un incremento significativo de la A1AT sérica respecto al grupo control (p < 0,001; hepatitis fulminante p < 0,05) confirmando que esta glicoproteína es un reactante de fase aguda. En las hepatitis fulminantes la concentración sérica de A1AT fue similar a la encontrada en los pacientes con hepatitis viral aguda sin insuficiencia hepática. La mayor elevación de la A1AT correspondió a los pacientes con obstrucción biliar y tumores hepáticos (primitivos y metastásicos) registrándose diferencias significativas respecto a las hepatopatías agudas y crónicas necrótico-inflamatorias (hepatitis, cirrosis, alcoholismo) (p < 0,001); a su vez en los pacientes con hígado metastásico la A1AT sérica fue mayor que en el resto de la población estudiada (p < 0,004 vs obstrucción biliar; p < 0,05 vs hepatoma). El 96% de las metástasis hepáticas presentaron valores de A1AT mayores de 400mg% registrándose diferencias significativas con el resto del grupo. La determinación de la A1AT sérica puede ser de utilidad para el diagnóstico de los tumores hepáticos malignos; de acuerdo a nuestros hallazgos el dosaje de esta glicoproteína carece de valor en el diagnóstico del hepatocarcinoma y en la insuficiencia hepática aguda

Concentración sérica de alfa-1-antitripsina en diversas enfermedades hepáticas / Serum concentration of alpha-1-antitrypsin in different hepatic diseases

Se investigó la concentración sérica de alfa-1-antitripsina (A1AT) en 195 pacientes con diversas hepatopatías y en 43 adultos sanos. En todas las enfermedades hepáticas estudiadas se encontró un incremento significativo de la A1AT sérica respecto al grupo control (p < 0,001; hepatitis fulminante p < 0,05) confirmando que esta glicoproteína es un reactante de fase aguda. En las hepatitis fulminantes la concentración sérica de A1AT fue similar a la encontrada en los pacientes con hepatitis viral aguda sin insuficiencia hepática. La mayor elevación de la A1AT correspondió a los pacientes con obstrucción biliar y tumores hepáticos (primitivos y metastásicos) registrándose diferencias significativas respecto a las hepatopatías agudas y crónicas necrótico-inflamatorias (hepatitis, cirrosis, alcoholismo) (p < 0,001); a su vez en los pacientes con hígado metastásico la A1AT sérica fue mayor que en el resto de la población estudiada (p < 0,004 vs obstrucción biliar; p < 0,05 vs hepatoma). El 96% de las metástasis hepáticas presentaron valores de A1AT mayores de 400mg% registrándose diferencias significativas con el resto del grupo. La determinación de la A1AT sérica puede ser de utilidad para el diagnóstico de los tumores hepáticos malignos; de acuerdo a nuestros hallazgos el dosaje de esta glicoproteína carece de valor en el diagnóstico del hepatocarcinoma y en la insuficiencia hepática aguda (AU)