Durability of sustained response shown in paediatric patients with chronic hepatitis C who were treated with interferon alfa-2b plus ribavirin.

Long-term studies in adults indicate that sustained virologic response (SVR) after combination treatment for chronic hepatitis C (CHC) predicts long-term clearance. Although peginterferon plus ribavirin is now standard care for children with CHC, long-term follow-up studies are not yet available. This study evaluated durability of virologic response over 5 years in children previously treated with interferon alfa-2b plus ribavirin (IFN/R). Ninety-seven of 147 children with CHC, who were treated with IFN/R and completed the 6-month follow-up in two previous clinical trials, participated in this long-term follow-up study. All were assessed annually for up to 5 years; patients with SVR were assessed for durability of virologic response. Children with SVR (n = 56) and those with detectable hepatitis C virus (HCV) RNA 24-week post-treatment (n = 41) were followed for a median of 284 weeks. Overall, 70% (68/97) of patients completed the 5-year follow-up. One patient with genotype 1a CHC had SVR and relapsed at year 1 of follow-up with the same genotype. Kaplan-Meier estimate for sustained response at 5 years was 98% (95% CI: 95%, 100%). Six patients with low-positive HCV RNA levels (n = 4) or missing HCV RNA at the 24-week follow-up visit (n = 2) in the initial treatment studies had virologic response during this long-term follow-up study. Linear growth rate was impaired during treatment with rapid increases in the immediate 6 months post-treatment. Mean height percentile at the end of the 5-year follow-up was slightly less than the mean pretreatment height percentile. Five patients experienced serious adverse events; none related to study drug exposure. SVR after IFN/R predicts long-term clearance of HCV in paediatric patients; growth normalized in the majority of children during the long-term follow-up. Similar long-term results could be expected after peginterferon alfa-2b plus ribavirin treatment.

Risk factors for rejection and infection in pediatric liver transplantation.

Rejection and infection are important adverse events after pediatric liver transplantation, not previously subject to concurrent risk analysis. Of 2291 children (<18 years), rejection occurred at least once in 46%, serious bacterial/fungal or viral infections in 52%. Infection caused more deaths than rejection (5.5% vs. 0.6% of patients, p < 0.001). Early rejection (<6 month) did not contribute to mortality or graft failure. Recurrent/chronic rejection was a risk in graft failure, but led to retransplant in only 1.6% of first grafts. Multivariate predictors of bacterial/fungal infection included recipient age (highest in infants), race, donor organ variants, bilirubin, anhepatic time, cyclosporin (vs. tacrolimus) and era of transplant (before 2002 vs. after 2002); serious viral infection predictors included donor organ variants, rejection, Epstein-Barr Virus (EBV) naivety and era; for rejection, predictors included age (lowest in infants), primary diagnosis, donor-recipient blood type mismatch, the use of cyclosporin (vs. tacrolimus), no induction and era. In pediatric liver transplantation, infection risk far exceeds that of rejection, which causes limited harm to the patient or graft, particularly in infants. Aggressive infection control, attention to modifiable factors such as pretransplant nutrition and donor organ options and rigorous age-specific review of the risk/benefit of choice and intensity of immunosuppressive regimes is warranted.

Serine and glycine transport in fetal ovine hepatocytes.

The role of hepatic serine and glycine transport in the regulation of the biosynthesis of serine by the fetal liver has not been studied. The goal of this study was to characterize serine and glycine transport and utilization at physiologic concentrations in primary cultures of fetal ovine hepatocytes. Primary culture of hepatocytes from mid gestation ( approximately 90 days) and term ( approximately 135 days) fetal sheep were studied after overnight serum free culture. At both gestational ages, the initial rate for sodium dependent 300 microM serine transport (1697+/-131 pmoles/min/mg protein at mid, 1765+/-544 at term) was fourfold greater than sodium dependent 300 microM glycine transport (309+/-54 at mid, 579+/-252 at term). At physiologic concentrations (300 microM), 69+/-7% of serine and 49+/-8% of glycine transport was sodium dependent. At term, sodium dependent serine transport has a V(max) of 1751+/-348 pmoles/min/mg protein and a K(m) of 159+/-111 microM. Sodium independent serine transport has a V(max) of 904+/-185 and a K(m) of 416+/-188 microM. Sodium dependent glycine transport has a V(max) of 410+/-69 and a K(m) of 2290+/-895 microM while sodium independent glycine transport exhibits non-saturable kinetics. Glycine (300 microM) sodium dependent transport was not inhibited by methyl-AIB while sodium dependent 300 microM serine transport was inhibited (31%). n-Ethylmaleimide inhibited sodium dependent serine and glycine transport by 36+/-9% and 37+/-2% respectively in term hepatocytes. Cysteine inhibited sodium dependent serine transport by 37%. Sodium independent glycine transport at 300 microM was higher in low glucose (1.1 mM) medium (881+/-76 pmoles/min/mg protein) compared to high glucose (5.5 mM) medium (510+/-60 P=0.004). There were no significant differences in serine or glycine incorporation into RNA, DNA, glycogen or lipid and protein. The predominance of serine transport over glycine at physiologic concentrations suggests that inward cellular amino acid transport of serine and glycine is not likely to be a regulatory mechanism that would favor serine biosynthesis in fetal ovine hepatocytes.

Long-term results with the Kasai operation for biliary atresia.

OBJECTIVE: To evaluate long-term outcome in a series of children with biliary results atresia treated by portoenterostomy. DESIGN: Case series of consecutive infants with biliary atresia with 10-year follow-up. Data were obtained by retrospective chart review or phone interview. SETTING: A tertiary academic medical center and regional children's hospital. PATIENTS: A consecutive series of 104 infants diagnosed with biliary atresia more than 10 years ago were evaluated. Eighty-nine had totally obliterated extrahepatic ducts, 4 had proximal hilar cysts (correctable type), and 11 had patency of the gallbladder and distal common duct. INTERVENTIONS: Ninety-eight patients underwent biliary reconstruction and 6 had exploration only. Seventy-four infants underwent reconstruction using a Rouxen-Y with exteriorization. The 11 infants with distal patency underwent a portocholecystostomy ("gallbladder Kasai"). The remainder had various modifications of the Kasai operation. MAIN OUTCOME MEASURES: Survival, liver function, complications, growth, and development. RESULTS: The 6 patients who did not have a portoenterostomy died. Of the 98 who had a reconstruction, 63 died (mean age at death, 27 months; median, 13.4 months), 10 following liver transplantation. Twelve of the 35 survivors ultimately required liver transplants. Twenty-three children are alive more than 10 years after portoenterostomy without the need for transplantation. Two thirds have experienced some manifestation of portal hypertension (ie, variceal bleeding, hypersplenism, ascites). Nineteen patients (79%) are anicteric with normal liver synthetic function and are in an age-appropriate school grade or working and living independently. CONCLUSIONS: We found that surgical correction of biliary atresia offers long-term survival for about one quarter of patients, provides palliation until liver transplantation becomes necessary, and if surgical correction is not feasible, biliary atresia is uniformly fatal. The outlook is good for those children who survived more than 10 years and justifies continued attempts to establish bile flow in infants with biliary atresia.

Esophageal varices: current management in children.

The treatment of esophageal variceal hemorrhage is still the subject of some controversy. The main causes of portal hypertension in children are portal vein thrombosis or cirrhosis, most commonly caused by biliary atresia. Many treatment options are available including endoscopic, radiographic, and surgical strategies. In general, children with presinusoidal obstructions have preserved hepatic synthetic function, and, therefore, treatment options include endoscopic strategies or portosystemic shunts, each with advocates. For children with advanced liver disease, liver transplantation offers the only chance for cure, so primary treatment of variceal bleeding should be by endoscopic means or transjugular intrahepatic portosystemic shunt (TIPS). Each modality has specific advantages and disadvantages, and treatment recommendations must therefore be tailored to the individual on a case-by-case basis, largely dependent on the expertise and experience of the health care team.

Development of primary culture of ovine fetal hepatocytes for studies of amino acid metabolism and insulinlike growth factors.

We report the development and characterization of a system of primary culture of ovine fetal hepatocytes to aid in the understanding of the cellular regulation of fetal growth and metabolism with emphasis on amino acid metabolism and insulinlike growth factor gene expression and to allow comparison to in vivo studies. Hepatocytes were isolated from late gestation fetal lambs by in situ perfusion and collagenase digestion utilizing occlusion of the ductus venosus to limit intrahepatic shunting. Hepatocytes were cultured in media modified to mimic fetal concentrations of glucose, lactate, and amino acids. Ovine fetal hepatocytes in primary culture maintain the pattern of fetal amino acid production and utilization seen across the fetal liver in vivo. Specifically, there is a net production of serine and a net utilization of glycine. Cultured ovine fetal hepatocytes specifically increase tritiated thymidine incorporation in response to insulin and insulinlike growth factor II (IGF-II). IGF-II mRNA abundance is high and IGF-I mRNA is low in cultured ovine fetal hepatocytes as in the fetal sheep liver in vivo. These data demonstrate the successful isolation of ovine fetal hepatocytes that retain some of the characteristics of the ovine fetal liver while maintained in short-term culture.

Management of esophageal varices in children by endoscopic variceal ligation.

Endoscopic variceal sclerotherapy (EVS) has been considered the mainstay of therapy for bleeding esophageal varices in adults. However, recent data have shown that endoscopic variceal ligation (EVL) is just as efficacious and has fewer complications than EVS. Although there are many reports concerning EVL in adults, only a few studies have been done in children. This report describes experience with EVL in 22 children with esophageal variceal hemorrhage. Eighty-seven EVL procedures were performed during a 9-year period in 22 children. The causes of portal hypertension were biliary atresia (10), portal vein thrombosis (8), chronic active hepatitis (1), cirrhosis secondary to cystic fibrosis (2), and primary sclerosing cholangitis (1). The age range at the onset of variceal bleeding was 8 months to 19 years. Twelve patients had EVS before EVL treatment was begun. Distal esophageal varices (one to four per session) were mechanically ligated using an elastic band ligature device attached to a flexible endoscope. The aim of therapy was obliteration of distal esophageal varices by EVL, every 2 to 4 weeks, until eradication. Subsequent EVL was dictated by the status of the varices. Outcome was assessed with respect to survival, rebleeding, status of varices, and complications. The patients underwent a mean of four sessions of EVL (range, one to eight). Four patients subsequently underwent liver transplantation. Of the 18 patients remaining (average follow-up period, 5.3 years), 12 had their varices eradicated (average of four EVL sessions), four are still in treatment, one has not been evaluated in the past 4 years, and one died of liver failure. Complications included bleeding between sessions (6 patients), cervical esophageal perforation (1 patient), and transient fever (2 patients). No child has experienced symptoms of esophageal stenosis or gastroesophageal reflux. Two patients died of liver disease, unrelated to bleeding from portal hypertension. EVL is effective in controlling variceal hemorrhage in children with portal hypertension, regardless of etiology. The complication rate is low, and EVL is an acceptable and perhaps preferable alternative to EVS in children with esophageal varices.

Fat-soluble vitamins in infants identified by cystic fibrosis newborn screening.

Fat-soluble vitamin status was assessed in 36 infants diagnosed with cystic fibrosis by newborn screening in the Colorado Program. At the time of diagnosis of cystic fibrosis, 36% of infants were hypoalbuminemic, 21% had vitamin A deficiency, 35% had vitamin D deficiency, and 38% had vitamin E deficiency. None had vitamin K deficiency. Supplementation with pancreatic enzymes, a multiple vitamin preparation, and additional vitamin E was associated with normalization of serum albumin, retinol, and 25-hydroxyvitamin D and negative PIVKA testing at age 6 and 12 months. Several patients remained vitamin E deficient, but this was felt to be due to poor compliance. Biochemical evidence of fat-soluble vitamin deficiency is common before age 3 months in infants with CF and responds to supplementation in the first year of life.

Amylopectinosis disease isolated to the heart with normal glycogen branching enzyme activity and gene sequence.

We report a 17-month-old female patient with a rare cause of cardiomyopathy secondary to accumulation of amylopectin-like material (fibrillar glycogen) isolated to the heart. Evidence of amylopectinosis isolated to cardiac myocytes in this patient was demonstrated by histology and electron microscopy. Glycogen content, glycogen branching enzyme (GBE) activity, as well as phosphofructokinase enzyme activities measured in liver, skeletal muscle, fibroblasts and ex-transplanted heart tissue were all in the normal to lower normal ranges. Normal skeletal muscle and liver tissue histology and GBE activity, normal GBE activity in skin fibroblasts, plus normal GBE gene sequence in this patient exclude the classical branching enzyme deficiency (type IV GSD). We believe that this is an as yet uncharacterized and novel phenotype of GSD associated with cardiomyopathy, in which there is an imbalance in the regulation of glycogen metabolism limited to the heart.

Biliary atresia: an update on our understanding of the disorder.

Biliary atresia is the leading cause of cholestasis in infants younger than 3 months. It is also the leading indication for liver transplantation in children. This review focuses on recent advances in the etiology, diagnosis, and management of biliary atresia.

Induction of cytosolic phosphoenolpyruvate carboxykinase in the ovine fetal liver by chronic fetal hypoglycemia and hypoinsulinemia.

Fetal glucose production has been observed in the chronically hypoglycemic, hypoinsulinemic (HH) fetal lamb. The purpose of this study was to test the hypothesis that induction of hepatic gluconeogenic enzymes occurs in this condition. The activities of both mitochondrial and cytosolic phosphoenolpyruvate carboxykinase (PEPCK), fructose-1,6-diphosphatase, and glucose-6-phosphatase, three key enzymes of gluconeogenesis, were determined in fetal sheep liver from HH lambs and controls (CONT). Pregnant ewes were maintained chronically hypoglycemic by continuous hyperinsulinemic clamps from approximately 80 d of gestational age (53% of gestation) for 6 wk. Fetuses (gestational age: HH = 136 +/- 2.6, CONT = 133 +/- 3.7 d) were maintained chronically hypoglycemic [HH = 0.51 +/- 0.05 versus CONT = 1.22 +/- 0.11 mmol/L (9.2 +/- 1.0 versus 21.9 +/- 11.9 mg/dL)] and hypoinsulinemic (HH = 3.3 +/- 0.6 versus CONT = 12.0 +/- 2.2 microU/mL) and delivered by cesarean section after measurement of fetal glucose production rate. Hepatic cytosolic PEPCK was 6.0 +/- 1.4 nmol/min/mg protein in CONT and 19.7 +/- 2.5 in HH lamb (p < 0.05), whereas mitochondrial PEPCK was not different between the two groups. Neither fructose-1,6-diphosphatase or glucose-6-phosphatase activities nor plasma glucagon levels were different between groups. These results suggest that chronic fetal hypoglycemia and hypoinsulinemia prematurely induce hepatic cytosolic PEPCK in the fetal lamb. The observed fetal glucose production in the HH fetal lamb may be due to gluconeogenesis.

Cysteine supplementation and reduction of total parenteral nutrition-induced hepatic lipid accumulation in the weanling rat.

Total parenteral nutrition (TPN)-induced hepatic steatosis is the most common complication of TPN administration to humans. The mechanism of TPN-induced hepatic steatosis has not been studied in young mammals. The goal of this study was to determine the mechanism of TPN-induced hepatic steatosis in the weanling rat and the effect of supplementation of TPN with choline and/or cysteine on TPN-induced hepatic steatosis. In the weanling rat, we investigated the effect of TPN administration on histologic hepatic steatosis, total hepatic lipid, hepatic acetyl-CoA-carboxylase (ACC--the rate limiting enzyme in fatty acid synthesis) specific activity, and total plasma lipids. TPN administration resulted in a threefold increase in hepatic lipid as compared with control and sham animals (TPN 138 +/- 12 mg/g liver versus control 57 +/- 1), an increase in histologic steatosis (TPN 3.7 versus control 1.3), and a decline in total plasma lipid (TPN 2.1 +/- 0.3 g/L versus control 4.1 +/- 0.3). TPN-induced hepatic steatosis in the weanling rat was not associated with an increase in ACC specific activity (TPN 2.10 +/- 0.33 nmol/min/mg protein versus control 2.85 +/- 0.23). Supplementation of the TPN with choline (15 mg/day) did not significantly lessen hepatic steatosis; however, supplementation of TPN with cysteine (2.5 mg/day) or with cysteine and choline did result in a significant lessening of hepatic lipid content and of histologic steatosis and a normalization of total plasma lipid.(ABSTRACT TRUNCATED AT 250 WORDS)

Ontogeny of hepatic enzymes involved in serine- and folate-dependent one-carbon metabolism in rabbits.

Serine occupies a central position in folate-dependent, one-carbon metabolism through 5,10-methylenetetrahydrofolate (MTHF) and 5-formyltetrahydrofolate (FTHF). We characterized the ontogeny of the specific activity of key enzymes involved in serine, 5,10-MTHF, and 5-FTHF metabolism: methenyltetrahydrofolate synthetase (MTHFS), MTHF reductase (MTHFR), the glycine cleavage system (GCS), methionine synthase (MS), and serine hydroxymethyltransferase (SHMT) in rabbit liver, placenta, brain, and kidney. In liver, MTHFS activity is low in the fetus (0.36 +/- 0.07 nmol. min(-1). mg protein(-1)), peaks at 3 wk (1.48 +/- 0.50 nmol. min(-1). mg protein(-1)), and then decreases to adult levels (1.13 +/- 0.32 nmol. min(-1). mg protein(-1)). MTHFR activity is highest early in gestation (24.9 +/- 2.4 nmol. h(-1). mg protein(-1)) and declines rapidly by birth (4.7 +/- 1.3 nmol. h(-1). mg protein(-1)). MS is highest during fetal life and declines after birth. Cytosolic SHMT activity does not vary during development, but mitochondrial SHMT peaks at 23 days. GCS activity is high in the fetus and the neonate, declining after weaning. In placenta and brain, all activities are low throughout gestation. Cytosolic and mitochondrial SHMT activities are low in kidney and rise after weaning, whereas MTHFS is low throughout development. These data suggest that the liver is the primary site of activity for these enzymes. Throughout development, there are multiple potential sources for production of 5,10-MTHF, but early in gestation high MTHFR activity and low MTHFS activity could reduce 5,10-MTHF availability.

Liver involvement in Alpers disease.

Alpers disease consists of diffuse cerebral degeneration manifested as developmental delay, seizures, vomiting, and progressive neuromuscular deterioration, with liver disease and death. We report the clinical course of the liver disease, histologic progression of the hepatic lesions, and etiologic investigations in five patients (four girls, three kinships). All had grown and developed normally until seen at 6 to 36 months of age (mean 20 months), with vomiting (n = 5), progressive hypotonia (n = 3), or seizures (n = 2). All had been given anticonvulsants, including valproic acid in three. Liver disease was noted at a mean age of 35 months (range 9 to 67 months), with hepatomegaly (two patients), abnormal hepatic synthetic function (three) or transaminase values (three), and cirrhosis in one. Patients survived for a mean of 4.6 weeks (range 1 to 8 weeks) after the identification of liver disease; all died of hepatic failure. Results of evaluation for infectious and metabolic causes of liver disease and causes of degenerative neuromuscular disease were negative in all patients. Premortem liver biopsy specimens (n = 3) demonstrated an early lesion consisting of lobular disarray, microvesicular steatosis, periportal acute and chronic inflammation, and individual hepatocyte necrosis. Autopsy findings (n = 5) consisted of macrovesicular steatosis, massive hepatocyte dropout, and proliferation of bile ductular elements, with almost complete replacement of hepatocytes by proliferating bile ductular elements in two patients. Brain showed characteristic neuronal degeneration. We conclude that Alpers disease can be a cause of rapidly progressive liver failure in early childhood. Although the cause of this autosomal recessive disease is not known, it does not appear to be related to peroxisomal dysfunction.

Anastomotic ulceration: a late complication of ileocolonic anastomosis.

Symptomatic ulceration developed at a previous ileocolonic anastomosis in six children. In the neonatal period all patients had had necrotizing enterocolitis that required resection of the terminal ileum, ileocecal valve, and proximal portion of the colon. Gross or occult rectal bleeding, with or without pain and diarrhea, began 5 1/2 years after successful resection and ileocolonic anastomosis. The cause of the ulcers is unknown. They appear inflammatory, both grossly and histologically, but have been uniformly unresponsive to antiinflammatory medications, antibiotics, and immunosuppressive medication. Surgical revision of the anastomosis and ulcer resection in five patients have resulted in rapid recurrence in four. Thirteen similar cases have been reported in the English-language literature. We conclude that ulceration is a long-term complication of neonatal resection of the terminal ileum and ascending colon with ileocolonic anastomosis.

Pathways of serine and glycine metabolism in primary culture of ovine fetal hepatocytes.

Previous in vivo studies in the ovine fetus have demonstrated net serine production by the fetal liver, a pattern not seen in the adult sheep. The goal of this study was to determine the major metabolic pathways responsible for fetal hepatic serine production by using stable isotope methodology in primary culture of late gestation ovine fetal hepatocytes. Specifically selected tracers of glycine were added to individual cultures, with production of labeled serine determined after 24 h of incubation. When [1-13C1]glycine or [2-13C1]glycine was used as the initial tracer, serine enrichment at 24 h indicated that approximately 30% of serine production comes from glycine. Quantitative comparison of serine enrichment from these two tracers suggests that serine synthesis from glycine occurs via the combined action of the glycine cleavage enzyme system (GCE) and serine hydroxymethyltransferase (SHMT). Using [1,2-13C2(15)N1]glycine as the tracer, there was no significant increase in M + 2 glycine in the medium over 24 h, suggesting no reversible transamination of glycine, and therefore no significant movement of glycine through the glyoxalate pathway. These data demonstrate that in primary culture of fetal ovine hepatocytes, approximately 30% of serine biosynthesis is derived from glycine, primarily via the combined action of GCE and SHMT.

Use of rifampin for severe pruritus in children with chronic cholestasis.

BACKGROUND: Rifampin has been proposed to reduce pruritus in children and adults with chronic cholestasis; however, there is a paucity of published data regarding the use of rifampin in children. METHODS: In an open trial, 24 children were evaluated during a 6-year period. Diagnoses included 13 patients with extrahepatic biliary atresia (54%), six with Alagille's syndrome, three with Byler's disease, and one each with primary sclerosing cholangitis and alpha1-antitrypsin deficiency. All patients had severe pruritus that had not responded adequately to at least 2 months of therapy with ursodeoxycholic acid, diphenhydramine, or phenobarbital and local skin care measures. Treatment was initiated with rifampin, 10 mg/kg per day in two divided doses for 18+/-20 months, and the effect on the severity of pruritus was assessed by a clinical scoring system. RESULTS: Ten patients showed a complete response, 12 a partial response, and 2 no response. Complete response was more common in extrahepatic cholestasis (64% vs. 10%), whereas partial response was more common in intrahepatic cholestasis (80% vs. 29%). Treatment was associated with reduction of gamma-glutamyl transpeptidase. No clinical or biochemical toxicity of rifampin was observed. CONCLUSIONS: We conclude that for more than 90% of children with chronic cholestasis and severe pruritus unresponsive to other treatments, rifampin appears to be a safe and effective therapy.

Correction of hypozincemia following liver transplantation in children is associated with reduced urinary zinc loss.

Zinc deficiency is a relatively common problem in children with chronic liver disease. We have previously shown inappropriate urinary zinc excretion in children with chronic liver disease and hypozincemia. This study was designed to determine if zinc deficiency and inappropriate urinary zinc losses are corrected in children with liver disease by liver transplantation. Thirty-three patients (age 1-19 years) underwent 35 liver transplants for acute and chronic liver disease. At the time of transplant, 17 patients had low plasma zinc (hypozincemic) (plasma zinc, 45.4 +/- 1.8 microg/dL), whereas 18 had normal plasma zinc (75.7 +/- 3.8). Before transplant, patients with zinc deficiency had higher urinary zinc to creatinine ratio compared with those with normal zinc status (6.6 +/- 1.9 vs. 2.2 +/- 0.6; P =.03) and lower serum albumin concentrations (low: 2.8 +/- 0.1 vs. normal: 3.3 +/- 0.2; P =.02). After transplant, there was a significant reduction in urinary zinc losses in the hypozincemic group followed by normalization of plasma zinc levels by 7 days posttransplant. These data suggest that the abnormal renal zinc homeostasis that is present in approximately 50% of pediatric patients undergoing liver transplant is rapidly improved and biochemical zinc deficiency is reversed after liver transplantation.