The management of paediatric urolithiasis

OBJECTIVE: To evaluate the efficacy and safety of the management of paediatric urolithiasis by extracorporeal shock wave lithotripsy (ESWL), endoscopic ureterolithotomy, percutaneous nephrolithotomy (PCNL) and open nephrolithotomy. PATIENTS AND METHODS: In a 3-year period (1997-1999), 59 children were treated for urolithiasis and underwent a total of 79 procedures. Thirty-two ESWL sessions were performed in 23 children (mean age 7.4 years, median 6.0). PCNL was undertaken in 30 renal units in 25 children (mean age 6.4 years, median 4.0). Eight patients (mean age 7.8 years, median 5) underwent 17 ureteroscopic procedures, six of which involved the use of a holmium laser. Three children with staghorn calculi underwent open nephrolithotomy under conditions of renal ischaemia and hypothermia. RESULTS: Of the 23 children treated using ESWL, 21 (91%) became stone-free; 17 underwent one ESWL session (74%), three had two sessions and three (13%) had three sessions. All eight patients who underwent ureteroscopy became stone-free. Four patients in whom the stone could not be reached by ureteroscopy initially had a JJ stent inserted, and the stone and stent subsequently removed. Stones were cleared using PCNL in 27 of 30 renal units (90%); three patients who had residual stone fragments were rendered stone-free by ESWL. Two of three children undergoing open nephrolithotomy were stone-free after surgery and the remaining one rendered stone-free with ESWL. Metabolic evaluation showed that 25 of 45 children (55%) had a urinary infection, eight (18%) had hyperoxaluria, three (7%) had hypercalciuria, two (4%) had cystinuria, and no identifiable cause was found in seven (16%). Treatment by a single modality rendered 52 of the 59 children (88%) stone-free; when the different modalities were combined, 57 of 59 patients (97%) were cleared of their stones. CONCLUSIONS: Technological advances in ESWL, ureteroscopy and PCNL have had a significant effect on the management of urolithiasis in children, allowing a safe and successful outcome. The comprehensive care of children with urolithiasis should include a full metabolic evaluation. Anatomical anomalies contribute to the complexity of many cases, necessitating a close liaison between adult and paediatric urologists, nephrologists and radiologists to optimize stone management in children.     

Salmonella enterica serovar typhimurium waaP mutants show increased susceptibility to polymyxin and loss of virulence In vivo

In Escherichia coli, the waaP (rfaP) gene product was recently shown to be responsible for phosphorylation of the first heptose residue of the lipopolysaccharide (LPS) inner core region. WaaP was also shown to be necessary for the formation of a stable outer membrane. These earlier studies were performed with an avirulent rough strain of E. coli (to facilitate the structural chemistry required to properly define waaP function); therefore, we undertook the creation of a waaP mutant of Salmonella enterica serovar Typhimurium to assess the contribution of WaaP and LPS core phosphorylation to the biology of an intracellular pathogen. The S. enterica waaP mutant described here is the first to be both genetically and structurally characterized, and its creation refutes an earlier claim that waaP mutations in S. enterica must be leaky to maintain viability. The mutant was shown to exhibit characteristics of the deep-rough phenotype, despite its ability to produce a full-length core capped with O antigen. Further, phosphoryl modifications in the LPS core region were shown to be required for resistance to polycationic antimicrobials. The waaP mutant was significantly more sensitive to polymyxin in both wild-type and polymyxin-resistant backgrounds, despite the decreased negative charge of the mutant LPSs. In addition, the waaP mutation was shown to cause a complete loss of virulence in mouse infection models. Taken together, these data indicate that WaaP is a potential target for the development of novel therapeutic agents.     

Effect of intravenous lipid emulsions on hepatic cholesterol metabolism

BACKGROUND: Total parenteral nutrition offers the chance of survival to children who have had extensive gut resections or gut failure. However, in infants it is often associated with serious complications including cholestatic liver disease. The causes of these complications remain unclear, although it has been suggested that the lipid emulsions used in total parenteral nutrition may be responsible. METHODS: An in vitro system was developed to study the effect of lipid emulsions on hepatic cholesterol metabolism using cultured hepatocytes. RESULTS: Incubations of Hep G2 cells with medium containing Intralipid (Pharmacia and Upjohn, Milton Keynes, UK) demonstrated that the fat emulsion mediated a powerful dose-dependent but reversible inhibition of cholesterol uptake. In addition Intralipid was shown to stimulate the efflux of cholesterol from Hep G2 cells. The component or components of the Intralipid responsible for these effects and the mechanism by which they act remain to be established. CONCLUSIONS: Intravenous lipid emulsions may interfere with hepatic cholesterol metabolism in vivo. This may have implications for the development of total parenteral nutrition-associated cholestasis in neonates.     

High mobility group proteins 1 and 2 recognize chromium-damaged DNA

Chromium (Cr) is a human carcinogen and a potent DNA damaging agent. Incubation of DNA with CrCl3 resulted in dose-dependent binding of Cr to DNA and, at concentrations >20 microM, altered the electrophoretic mobility of a 100 bp oligonucleotide. We also demonstrate that high mobility group (HMG) proteins 1 and 2 bind Cr-damaged DNA (Cr-DNA). Protein binding was lesion density-dependent, with maximal binding to DNA treated with 100 microM CrCl3. HMG2 binds to Cr-DNA with a calculated Kd of approximately 10(-9) M. These proteins also bound DNA obtained from chromate-treated cells. These results suggest that the covalent attachment of Cr to DNA induces alterations in DNA structure which are recognized by HMG1 and HMG2. Therefore, these proteins may function as Cr-damaged DNA recognition proteins in vivo and as a consequence of binding, may play a role in directing the cellular response to Cr-DNA adduct formation.      

Sputum tumour necrosis factor-alpha and leukotriene concentrations in cystic fibrosis

It is postulated that a vigorous host inflammatory response in the cystic fibrosis lung contributes to lung injury. Tumour necrosis factor-alpha (TNF-alpha) may play a part in that process and in the generation of leukotrienes. Therefore, the relationships between sputum TNF-alpha, leukotriene concentration, and lung function abnormalities in 16 children with cystic fibrosis were investigated. Each subject provided sputum samples and performed spirometry. TNF-alpha was measured by enzyme linked immunosorbent assay; individual leukotrienes were separated using high performance liquid chromatography and quantified by radioimmunoassay. The geometric mean concentration of TNF-alpha was 129.7 pg/ml and 95% confidence interval 48.2 to 348.3. Mean (SEM) leukotriene B4 (LTB4) was 97.8 (22.9) pmol/g and total cysteinyl leukotrienes were 60.9 (14.8) pmol/g. Mean (SD) forced expiratory volume in one second (FEV1) of the group was 53 (15)% of predicted and forced vital capacity (FVC) was 65 (14)% of predicted. There was a significant positive correlation between TNF-alpha and both LTB4 and the total cysteinyl leukotriene sputum content. An inverse relationship existed between TNF-alpha and FEV1 and FVC. Moreover, a negative correlation was observed between sputum LTB4 and FEV1 and FVC. These results suggest that TNF-alpha and the leukotrienes may participate in the airways inflammation and airflow obstruction observed in cystic fibrosis subjects and support the hypothesis that TNF-alpha upregulates the 5-lipoxygenase pathway in vivo.     

Interleukin-1 alpha, soluble interleukin-2 receptor, and IgG concentrations in cystic fibrosis treated with prednisolone

The cytokines interleukin-1 and interleukin-2 participate in the inflammatory response, and may contribute to hypergammaglobulinaemia G and the development of lung injury in cystic fibrosis. Anti-inflammatory treatment with corticosteroids may attenuate this response. The effect of a 12 week course of oral prednisolone on spirometry and serum concentrations of interleukin-1 alpha (IL-1 alpha), soluble interleukin-2 receptor (sIL-2R), and IgG was investigated in 24 children with cystic fibrosis. Prednisolone was administered, in a double blind and placebo controlled manner, at an initial dose of 2 mg/kg daily for 14 days and tapered to 1 mg/kg on alternate days for 10 weeks. The treated group (n = 12) experienced an increase in forced expiratory volume in one second and forced vital capacity at 14 days, however, these changes were smaller at 12 weeks. In the treated group, change in pulmonary function was associated with decreased serum IgG and cytokine concentrations. Prednisolone suppresses serum concentrations of these cytokines, which may participate in the inflammatory response, the excessive synthesis of IgG, and airflow obstruction observed in cystic fibrosis patients.     

Atorvastatin Transport in the Caco-2 Cell Model: Contributions of P-Glycoprotein and the Proton-Monocarboxylic Acid Co-Transporter

Purpose. The purpose of this study was to elucidate the mechanismsby which an HMG-CoA reductase inhibitor, atorvastatin (an organicacid with a pKa of 4.46), was transported in the secretory and absorptivedirections across Caco-2 cell monolayers. Methods. Caco-2 cells were grown on polycarbonate membrane insertsin 6-well Snapwell plates (Costar). The permeability of radiolabeledcompounds across Caco-2 cell monolayers was determined using aside-by-side diffusion apparatus (NaviCyte) and an automated liquidhandler (Hamilton Microlab 2200). The apical uptake of¹⁴C-atorvastatin was also determined in Caco-2 cells. Cyclosporin A (20 M) waspresent in the uptake media to block potential P-glycoprotein-mediatedatorvastatin efflux. Results. Polarized permeation of atorvastatin was observed with thebasolateral-to-apical (B-to-A) permeability being 7-fold greater thanthe A-to-B permeability (35.6 × 10^–6 and 4.9 × 10^–6 cm/s,respectively). The secretion of atorvastatin was a saturable process with anapparent K_m of 115 M. The B-to-A permeability of atorvastatin wassignificantly reduced by cyclosporin A (10 M), verapamil (100 M),and a P-glycoprotein specific monoclonal antibody, UIC2(10 g/ml)(43%, 25%, and 13%, respectively). Furthermore, both CsA andverapamil significantly increased the A-to-B permeability of atorvastatinby 60% however, UIC2 did not affect the A-to-B permeability ofatorvastatin. CsA uncompetitively inhibited the B-to-A flux ofatorvastatin with a K_i of 5 M. In addition, atorvastatin (100 M) significantlyinhibited the B-to-A permeability of vinblastine by 61%. The apicaluptake of atorvastatin increased 10.5-fold when the apical pH decreasedfrom pH 7.4 to pH 5.5 while the pH in the basolateral side wasfixed at pH 7.4. A proton ionophore, carbonylcyanidep-trifluoro-methoxyphenylhydrazone (FCCP) significantly decreased atorvastatinuptake. In addition, atorvastatin uptake was significantly inhibited bybenzoic acid, nicotinic acid, and acetic acid each at 20 mM (65%,14%, and 40%, respectively). Benzoic acid competitively inhibitedatorvastatin uptake with a K_i of 14 mM. Similarly, benzoic acid,nicotinic acid, and acetic acid significantly, inhibited the A-to-Bpermeability of atorvastatin by 71%, 21%, and 66%, respectively. Conclusion. This study demonstrated that atorvastatin was secretedacross the apical surface of Caco-2 cell monolayers viaP-glycoprotein-mediated efflux and transported across the apical membrane in theabsorptive direction via a H⁺-monocarboxylic acid cotransporter(MCT). In addition, this study provided the first evidence thatnegatively charged compounds, such as atorvastatin, can be a substrate forP-glycoprotein.