Efficacy and optimal dose of daily polyethylene glycol 3350 for treatment of constipation and encopresis in children.

OBJECTIVE: To determine efficacy, safety, and optimal dose of a laxative, polyethylene glycol (PEG) 3350, in children with chronic constipation. STUDY DESIGN: Children with chronic constipation (n = 24) were treated with PEG for 8 weeks at an initial dose of 1 g/kg/d. The dose was adjusted every 3 days as required to achieve 2 soft stools per day. A diary was kept to monitor dose, stool frequency and consistency, soiling, and other symptoms. Stool consistency was rated from 1 (hard) to 5 (watery). Subjects were examined for fecal retention. The Student t test and the Fisher exact test were used for data analysis. RESULTS: All 20 children who completed the study found PEG to be palatable and were satisfied with the treatment. There were no significant adverse effects. Weekly stool frequency increased from 2.3 +/- 0.4 to 16.9 +/- 1.6 (P <.0001) during treatment and stool consistency from 1.2 +/- 0.1 to 3.3 +/- 0.1 (P <.0001). In 9 children with soiling, weekly soiling events declined from 10.0 +/- 2.4 to 1.3 +/- 0.7 (P =.003). The mean effective dose was 0.84 g/kg/d (range, 0.27-1.42 g/kg/d). CONCLUSION: Daily administration of PEG at a mean dose of 0.8 g/kg is an effective, safe, and palatable treatment for constipation.

Value of pH probe testing in pediatric patients with extraesophageal manifestations of gastroesophageal reflux disease: a retrospective review.

Extended pH probe testing is often performed in patients believed to have extraesophageal symptoms of gastroesophageal reflux disease (GERD), although for this indication its diagnostic value is not well established. A retrospective review of all patients who underwent pH probe testing between 1994 and 1998 was conducted to determine the outcome of antireflux therapy in the subgroup with probable extraesophageal symptoms of GERD. Sixty-eight patients underwent antireflux therapy and had adequate follow-up after pH probe testing to be included in the study. Fifty-eight patients (85%) responded to antireflux therapy (improved, 44%; cured, 41%). The positive predictive value of distal pH probe testing was greater than 90%, but the negative predictive value was less than 50%. The reproducibility of pH probe testing on different study days was poor, but pH probe testing was helpful in assessing the adequacy of antireflux therapy. The presence of gastrointestinal symptoms did not correlate with the response of extraesophageal symptoms to antireflux therapy. Thirteen patients underwent double-probe pH studies. The mean percent time the pH was less than 4 in the upper esophagus was 2.6% (range, 1% to 9.6%). Twelve of these patients were improved or cured with antireflux therapy. Distal pH probe testing is of limited benefit in predicting whether patients with extraesophageal symptoms of GERD will respond to antireflux therapy. If extraesophageal symptoms of GERD are suspected, patients should undergo an empiric trial of antireflux therapy. Distal pH probe testing should be reserved for assessing the adequacy of antireflux therapy if symptoms persist. A prospective, randomized, controlled study will aid in determining the predictive value of double-probe pH studies in pediatric patients with probable extraesophageal symptoms of GERD.

Camphor hepatotoxicity.

We report a case of hepatotoxicity in a 2-month-old baby after a camphor-containing cold remedy was applied dermally. Liver function tests returned to normal after the application of the cold remedy was discontinued. Ingestion of camphor can cause severe liver and central nervous system injury, and neurotoxicity has been observed after exposure to camphor through the skin. Hepatotoxicity after dermal application of camphor has never been reported. This report emphasizes the common use of cold remedies that are usually not beneficial and may be potentially dangerous.

Ursodeoxycholic acid improves cholestasis in infants with cystic fibrosis.

OBJECTIVE: To describe two infants with cholestatic jaundice treated with ursodeoxycholic acid (UDCA). CASE SUMMARY: Two infants with cystic fibrosis (CF)-associated hepatobiliary disease, manifesting as cholestatic jaundice and elevated liver enzymes within the first 6 weeks of life, had improved biochemical indices of liver function following treatment with UDCA 20-40 mg/kg/d. DISCUSSION: To our knowledge, this is the first report of UDCA treatment in infants with CF-associated cholestatic jaundice. Infants and children require treatment with increased doses of UDCA to compensate for reduced intestinal absorption of bile acid and immaturity of the enterohepatic circulation. CONCLUSIONS: UDCA appears to be a cost-effective treatment for CF-associated hepatobiliary disease in infants and children.

Increased urinary excretion of 3-hydroxyisovaleric acid and decreased urinary excretion of biotin are sensitive early indicators of decreased biotin status in experimental biotin deficiency.

To assess the utility of various indicators of biotin status, marginal biotin deficiency was induced experimentally in normal adults. Ten subjects consumed a diet that contained enough avidin to bind seven times more biotin than that in the diet. Blood and 24-h urine samples were collected before the diet began and twice weekly thereafter for 20 d. The urinary excretion and serum concentration of biotin and its two principal inactive metabolites bisnorbiotin and biotin sulfoxide were determined after HPLC separation with an avidin-binding assay. The urinary concentration of 3-hydroxyisovaleric acid, an indicator of reduced activity of a biotin-dependent enzyme, was quantitated by gas chromatography-mass spectrometry. The urinary excretion of 3-hydroxyisovaleric acid increased significantly (P < 0.0001). For all subjects, the urinary excretion of both biotin and bisnorbiotin decreased significantly (P < 0.0001 for each). In contrast, the mean serum concentration of biotin did not decrease significantly (P = 0.06). These data provide evidence that the urinary excretion of 3-hydroxyisovaleric acid and the urinary excretion of biotin are early and sensitive indicators of biotin deficiency and that the serum concentration of biotin is not.

Inhibition of apolipoprotein B secretion by IL-6 is mediated by EGF or an EGF-like molecule in CaCo-2 cells.

Small intestinal mucosal inflammation observed in celiac disease is associated with the local release of growth factors and various cytokines. In a previous study, we investigated the effect of various cytokines on triacylglycerol and apoB secretion by CaCo-2 cells and observed that TNF-alpha, IL-1 beta, and particularly IL-6, decreased apolipoprotein (apo) B and triacylglycerol secretion. In this study, we explored possible mechanisms to explain the inhibitory effect of IL-6 on apoB secretion. IL-6, 10 ng/mL, added to the basolateral medium of CaCo-2 cells grown on semi-permeable filters, decreased apoB secretion by 42%. Adding a blocking monoclonal antibody (mAb 528) to the EGF receptor completely prevented this effect. IL-6 decreased the amount of EGF receptor protein and the binding of iodinated EGF to its receptor by 50% and 30%, respectively. Incubation of cells with various ligands to the EGF receptor, such as EGF, TGF-alpha, HB-EGF, and amphiregulin, also decreased apoB secretion. Inhibition of apoB secretion by EGF was prevented by the mAb 528 or an EGF neutralizing antibody. In a dose-dependent manner, the neutralizing antibody to EGF prevented the decrease in secretion of apoB, triacylglycerol mass, and cell-surface binding of labeled EGF caused by IL-6. Similar to the effects of IL-6, EGF decreased the secretion of triacylglycerol mass and the synthesis and secretion on newly synthesized apoB. The results suggest that, in CaCo-2 cells, IL-6 causes the release of EGF or an EGF-like molecule. By binding to cell surface EGF receptors, the molecule then causes a decrease in triacylglycerol and apoB secretion.

The glucuronidation of exogenous and endogenous compounds by stably expressed rat and human UDP-glucuronosyltransferase 1.1.

Rat and human UDP-glucuronosyltransferase (UGT) 1.1 share > 70% identity in their deduced primary amino acid sequences. We have previously shown that rat UGT1.1, stably expressed in human embryonic kidney 293 cells, catalyzes the glucuronidation of bilirubin and the mixed opioid agonist/antagonist buprenorphine with high efficiency. The present study was designed to characterize the reactivity of expressed human UGT1.1 with opioid compounds and compare its substrate specificity for opioids to that of the expressed rat enzyme. The results show that both rat and human UGT1.1 catalyze the glucuronidation of opioids with a relative reactivity of buprenorphine > > nalorphine approximately naltrexone. Comparison of glucuronidation activities in livers from Crigler-Najjar type 1 patients and normal patients indicates that UGT1.1 catalyzes at least 75% of buprenorphine conjugation in normal human liver. In separate studies, the reactivity of expressed rat UGT1.1 was characterized toward various xeno-and endobiotics of various compound classes. It was found that both rat and human UGT1.1 exhibited comparable substrate specificities and efficiencies (Vmax/Km) of glucuronide formation for anthraquinones, coumarins, estrogens, flavonoids, and phenolic compounds. Neither rat nor human UGT1.1 catalyzed the glucuronidation of amines, monoterpenoid alcohols, androgens, or progestins. In general, these data indicate that rat and human UGT1.1 are functionally identical and can be considered orthologous enzymes.

Interruption of a transforming growth factor alpha autocrine loop in Caco-2 cells by antisense oligodeoxynucleotides.

BACKGROUND & AIMS: We have previously shown that Caco-2 cell proliferation is driven by basolateral membrane epidermal growth factor receptors. The aim of this study was to investigate whether autocrine production of transforming growth factor alpha (TGF-alpha) activates these receptors and stimulates proliferation using antisense oligodeoxynucleotides. METHODS: Caco-2 cells grown on microporous membranes or Jurkat cells were exposed to conventional or 5' cholesterol-modified oligodeoxynucleotides synthesized with random, antisense, or missense base sequences. Indices of proliferation were measured, including [3H]thymidine or [3H]uridine uptake for studies of short-term stimulation and the methylthiotetrazole assay as an index of cell number increase over longer periods. Secretion of TGF-alpha by cells was detected using a soft agar bioassay. RESULTS: Incubation with antisense oligodeoxynucleotides inhibited TGF-alpha secretion compared with controls. Random and missense oligodeoxynucleotides had no effect on proliferation. The TGF-alpha antisense oligodeoxynucleotides markedly inhibited proliferation, an effect that was abolished by adding TGF-alpha to the medium. Oligonucleotides had no effect on Jurkat cells, a lymphocytic cell line lacking epidermal growth factor receptors. Cholesterol-modified oligodeoxynucleotides were more effective and specific than unmodified oligodeoxynucleotides. CONCLUSIONS: Caco-2 cell proliferation is driven by autocrine stimulation of epidermal growth factor receptors by TGF-alpha. This mechanism may be effectively inhibited by antisense oligodeoxynucleotides, particularly those modified by the 5' attachment of cholesterol.

Nerve growth factor synthesis by intestinal epithelial cells.

Nerve growth factor (NGF) exists in the gut of adult rats. The cells responsible for NGF synthesis in the gut remain unknown. IEC-6 and Caco-2 cells, established cell culture models of intestinal epithelial cells, were studied to determine whether intestinal epithelial cells, were studied to determine whether they synthesize and release NGF. Conditioned media from both IEC-6 and Caco-2 cells stimulated neurite outgrowth in both rat pheochromocytoma (PC-12) cells and sensory neurons derived from embryonic chick dorsal root ganglia (DRG). The addition of anti-NGF antibody blocked neurite outgrowth in PC-12 cells and partially blocked outgrowth in DRG cells. An NGF-enzyme-linked immunosorbant assay readily detected immunoreactive NGF in conditioned media from both cell lines, whereas cellular extracts from IEC-6, Caco-2, and isolated rat intestinal epithelial cells had low levels of immunoreactivity. Caco-2 monolayers primarily secreted NGF from the basolateral compartment, and interleukin-1 enhanced its secretion. IEC-6, Caco-2, and isolated rat intestinal epithelial cells expressed NGF mRNA as determined by reverse transcription polymerase chain reaction. These observations suggest that intestinal epithelial cells are capable of NGF synthesis.

Expressed human UGT1.4 protein catalyzes the formation of quaternary ammonium-linked glucuronides.

In humans, the metabolism of a number of tertiary amine-containing pharmacological agents to quaternary ammonium-linked glucuronides, catalyzed by UDP-glucuronosyltransferase (UGT), represents a unique and important metabolic pathway for these compounds. A full-length cDNA-encoding human UGT1.4 (the so-called "minor" human bilirubin UGT) was inserted into the expression vector pREP9 and transfected into human embryonic kidney 293 cells, and stable transfectants were obtained after geneticin selection. As expected, the expressed protein had low catalytic activity toward bilirubin. However, expressed human UGT1.4 protein exhibited glucuronidation activity toward tertiary amine substrates, such as imipramine, cyproheptadine, tripelennamine, and chlorpromazine, which form quaternary ammonium-linked glucuronides. Carcinogenic primary amines (beta-naphthylamine, benzidine, and 4-aminobiphenyl) also reacted with the expressed UGT1.4 protein at rates approximately 10-fold higher than the rates for quaternary ammonium glucuronide formation. Although a number of other UGT gene products are capable of catalyzing the glucuronidation of primary amine substrates, expressed human UGT1.4 protein is the only UGT isoform that has been shown to conjugate tertiary amine substrates, forming quaternary ammonium-linked glucuronides.

Caco-2 cells express type I interleukin-1 receptors: ligand binding enhances proliferation.

We examined the effect of interleukin-1 (IL-1) on the rate of proliferation of the human colon carcinoma Caco-2 and characterized the human intestinal epithelial cell IL-1 receptor (IL-1R). IL-1 dose dependently increased tritiated thymidine uptake in confluent Caco-2 monolayers fed complete growth medium. An anti-IL-1 beta completely blocked the increase in tritiated thymidine uptake, whereas an IL-1 receptor antagonist human recombinant blocked it partially. In long-term culture, IL-1 increased DNA content over control, an effect similar to that of epidermal growth factor (EGF). Unlike EGF, IL-1 did not enhance tritiated thymidine uptake in Caco-2 monolayers grown in serum-free medium, implying that IL-1 needs a cofactor(s) to elicit its proliferative effect. Cross-linking 125I-IL-1 beta to Caco-2 membranes revealed a binding protein of approximately 80 kDa with binding saturated at approximately 2.5 x 10(9) M-1 consistent with that for the type I IL-1R. cDNA transcribed from Caco-2 mRNA and amplified by polymerase chain reaction, using complementary oligonucleotides, resulted in a reaction product matching the sequence of the type I IL-1R. Our results demonstrate that IL-1 enhances proliferation of Caco-2 cells. This effect requires the presence of an unidentified cofactor(s). Also, Caco-2 cells express the type I IL-1R.

Regulation of Caco-2 cell proliferation by basolateral membrane epidermal growth factor receptors.

The epidermal growth factor (EGF) receptor is an important mediator of intestinal epithelial cell proliferation. We studied cell-surface localization of this molecule in Caco-2 cells and characterized cellular responses to apical or basolateral EGF stimulation. 125I-labeled EGF bound almost exclusively to a 180-kDa molecule, existing as a single high-affinity population by Scatchard analysis. On basolateral membranes 13- to 15-fold more ligand binding was seen. Apical/basolateral differences were not significantly altered by incubation with either blocking antibody to EGF receptor or transforming growth factor-alpha (TGF-alpha) neutralizing antibody. Even though apical EGF receptors were demonstrated, only basolateral membrane stimulation with EGF increased tyrosine kinase activity and enhanced uptake of [3H]thymidine. Continuous exposure to EGF during culture significantly increased monolayer DNA content. These data demonstrate that Caco-2 cell proliferation is driven solely by basolateral membrane EGF receptor, despite the presence of lesser amounts of this molecule on the apical surface. Differences between apical and basolateral membrane receptor expression are not the result of polarized secretion of TGF-alpha or other EGF receptor ligands.

Effect of okadaic acid on apo B and apo A-I secretion by CaCo-2 cells.

The effect of protein phosphorylation on the synthesis and secretion of apo B and apo A-I by CaCo-2 cells was investigated. Okadaic acid, a potent inhibitor of protein serine/threonine phosphatases 1 and 2A, caused a significant increase in total cellular protein phosphorylation. Apo B-48 was phosphorylated in control cells and this was increased significantly in the presence of okadaic acid. Under the experimental conditions, the phosphorylation of apo B-100 or apo A-I was not observed. No evidence of tyrosine phosphorylation of apo B-100, B-48, or apo A-I was found. Okadaic acid did not change the amount of apo B mass within cells but apo B mass secreted into the basolateral medium was decreased by 40%. Apo A-I mass within cells or in the basolateral medium was unaffected by okadaic acid. Despite causing an 18% decrease in total protein synthesis, okadaic acid did not alter the rate of synthesis of apo B-100, apo B-48, or apo A-I. Cellular turnover of labeled apo B-100 in cells incubated with okadaic acid was similar to controls, whereas apo B-48 and apo A-I turnover were slowed by okadaic acid. Compared to controls, however, 1 microM okadaic acid caused a 75% and 50% decrease in the secretion of newly synthesized apo B-100 and apo B-48, respectively, while decreasing labeled apo A-I secretion by 35%. In contrast to apo A-I mRNA levels, which were not altered by okadaic acid, apo B mRNA levels were significantly decreased by the polyether fatty acid. Despite differences observed in the phosphorylation state of apo B-100 and apo B-48, okadaic acid decreased the secretion of both forms of apo B without altering their synthesis. Okadaic acid, by increasing cellular protein phosphorylation, significantly disrupts the secretory processing of apo B by CaCo-2 cells.

Mucosal morphology in isolated bowel segments: importance of exposure to luminal contents.

An isolated bowel segment (IBS) is a loop of intestine that has been freed from its mesenteric attachment after the development of vascular collaterals between the antimesenteric surface of the gut and the host organ. Surgical creation of such artificially vascularized isolated bowel segments is of interest to researchers for a variety of studies, and may be useful in the treatment of short bowel syndrome, allowing longitudinal division of the remaining small bowel to double its length. We created four surgical variants to study the ability of the collateral blood supply to maintain mucosal integrity in the presence or absence of normal luminal contents. In all groups, a collateral blood supply was created in a 5- to 7-cm segment of adult rat jejunum by hepatoenteropexy (Iowa model II). In Thiry-Vella (T-V) and isolated bowel segment (IBS) rats, this segment was exteriorized at both ends to exclude luminal contents. Control and IBS in continuity (IBS-C) loops were left in continuity. The mesentery of IBS and IBS-C rats was divided 5 weeks later, leaving the experimental segment entirely dependent on the collateral circulation. All animals were harvested at 7 weeks after the initial surgery. Tissues were analyzed for mucosal weight, protein content per centimeter of bowel, length of villi, depth of crypts, DNA content, and sucrase activity. We found that segments retaining luminal continuity had significantly higher mucosal weight and DNA content per centimeter of bowel compared with exteriorized loops.

Prolonged postprandial abdominal pain following Kawasaki syndrome with acute gallbladder hydrops: association with impaired gallbladder emptying.

Acute hydrops of the gallbladder is a well-recognized complication of Kawasaki syndrome. We report a case of a child with this syndrome whose gallbladder hydrops slowly resolved after intravenous gamma-globulin therapy. However, he continued to experience postprandial right upper quadrant abdominal pain. Hepatobiliary scintigraphy revealed normal filling of the gallbladder but marked impairment of meal-stimulated gallbladder emptying. Endoscopy with biopsy of the esophagus, stomach, and duodenum was normal, ruling out peptic complications of his aspirin therapy. This child's discomfort improved slowly over several months, finally ending approximately 6 months after the onset of his illness. A repeat gallbladder emptying study done ultrasonographically at that time revealed near-normal meal-stimulated gallbladder emptying. We conclude that poor emptying of the gallbladder may be associated with prolonged abdominal pain in Kawasaki syndrome. Meal-stimulated gallbladder emptying can be assessed by a simple ultrasonographic technique and should be considered in any patient with Kawasaki syndrome and abdominal pain.

Bacterial gastroenteritis.

Acute diarrhea is a major cause of childhood morbidity. Important advances in the understanding of bacterial gastroenteritis have been made in the past two decades. This article reviews the epidemiology, pathogenesis, and methods of diagnosis of bacterial gastroenteritis. Bacterial enteric pathogens common to North America are discussed in more detail.

The meteorological satellite: overview of 25 years of operation.

The first weather satellite was launched on 1 April 1960. In the 25 years since, weather satellites have contributed to improved weather analyses and forecasts worldwide. As a maturing component of a global observing system, the meteorological satellite promises even greater financial benefits and a higher quality of life to mankind.