Screening of Living Kidney Donors for Genetic Diseases Using a Comprehensive Genetic Testing Strategy.

Related living kidney donors (LKDs) are at higher risk of end-stage renal disease (ESRD) compared with unrelated LKDs. A genetic panel was developed to screen 115 genes associated with renal diseases. We used this panel to screen six negative controls, four transplant candidates with presumed genetic renal disease and six related LKDs. After removing common variants, pathogenicity was predicted using six algorithms to score genetic variants based on conservation and function. All variants were evaluated in the context of patient phenotype and clinical data. We identified causal variants in three of the four transplant candidates. Two patients with a family history of autosomal dominant polycystic kidney disease segregated variants in PKD1. These findings excluded genetic risk in three of four relatives accepted as potential LKDs. A third patient with an atypical history for Alport syndrome had a splice site mutation in COL4A5. This pathogenic variant was excluded in a sibling accepted as an LKD. In another patient with a strong family history of ESRD, a negative genetic screen combined with negative comparative genomic hybridization in the recipient facilitated counseling of the related donor. This genetic renal disease panel will allow rapid, efficient and cost-effective evaluation of related LKDs.

Harmonization of critical result management in laboratory medicine.

Unsafe medical care is a major source of disabling injuries and death throughout the world. The failure to notify, follow up, and action critical results, which signify life threatening situations, is of particular concern and may cause avoidable morbidity and mortality. International accreditation standards require pathology laboratories to have a system for the timely and reliable communication of critical results to clinical personnel responsible for patient care. In response, various practices and a number of different terminologies have been described in the literature. Increased attention to patient safety standards and multinational surveys, however, highlighted shortcomings and inefficiencies in existing communication systems. These failures and variations in practice call for clear guidance and harmonization of approaches in order to improve communications and to provide safer patient care. The objectives of this review are to create a harmonized terminology and to learn from international practices by systematically reviewing the best available evidence on existing approaches. Based on literature review findings we highlight key areas where harmonization is necessary and feasible and offer a conceptual framework and methods for designing better and more evidence-based systems for the timely notification of laboratory results that represent potential patient safety hazards.

Receptor-mediated endocytosis of Neisseria gonorrhoeae into primary human urethral epithelial cells: the role of the asialoglycoprotein receptor.

Urethral epithelial cells are invaded by Neisseria gonorrhoeae during gonococcal infection in men. To understand further the mechanisms of gonococcal entry into host cells, we used the primary human urethral epithelial cells (PHUECs) tissue culture system recently developed by our laboratory. These studies showed that human asialoglycoprotein receptor (ASGP-R) and the terminal lactosamine of lacto-N-neotetraose-expressing gonococcal lipooligosaccharide (LOS) play an important role in invasion of PHUECs. Microscopy studies showed that ASGP-R traffics to the cell surface after gonococcal challenge. Co-localization of ASGP-R with gonococci was observed. As ASGP-R-mediated endocytosis is clathrin dependent, clathrin localization in PHUECs was examined after infection. Infected PHUECs showed increased clathrin recruitment and co-localization of clathrin and gonococci. Preincubating PHUECs in 0.3 M sucrose or monodansylcadaverine (MDC), which both inhibit clathrin-coated pit formation, resulted in decreased invasion. N. gonorrhoeae strain 1291 produces a single LOS glycoform that terminates with Gal(beta1-4)GlcNac(beta1-3)Gal(beta1-4)Glc (lacto-N-neotetraose). Invasion assays showed that strain 1291 invades significantly more than four isogenic mutants expressing truncated LOS. Sialylation of strain 1291 LOS inhibited invasion significantly. Preincubation of PHUECs in asialofetuin (ASF), an ASGP-R ligand, significantly reduced invasion. A dose-response reduction in invasion was observed in PHUECs preincubated with increasing concentrations of NaOH-deacylated 1291 LOS. These studies indicated that an interaction between lacto-N-neotetraose-terminal LOS and ASGP-R allows gonococcal entry into PHUECs.

Pseudomonas aeruginosa in cystic fibrosis: cross-infection and the need for segregation.

Evidence-based reasons for segregation of patients colonized with Pseudomonas aerugionsa in the outpatient setting are unclear. To clarify local decisions, Pseudomonas genotyping of the local environment, patients and patient contacts was undertaken in 1993. The hospital environment was re-swabbed in 1997. Pseudomonas genotyping of old and new patients attending the North Staffordshire cystic fibrosis clinic has subsequently been undertaken and more recently been repeated on an annual basis to assess whether the same Pseudomonas genotypes can be found in both the environment and in patients, and whether the same Pseudomonas genotype can be transferred from one patient to another. No Pseudomonas genotype found in the local environment in 1993 or in 1997 has been found in any of our patients. Nine children attending the same special school for many years and sharing the same physiotherapy facilities showed no evidence of cross-infectivity. Except for siblings living in the same household our cross-infectivity rate is very low and where cross-infection has potentially occurred the level of contact between these patients has been minimal. This study does not support the suggestion that patients with cystic fibrosis attending the North Staffordshire clinic and colonized with Pseudomonas aeruginosa should be segregated from non-colonized patients.

Fractalkine cleavage from neuronal membranes represents an acute event in the inflammatory response to excitotoxic brain damage.

Fractalkine is a recently identified chemokine that exhibits cell adhesion and chemoattractive properties. It represents a unique member of the chemokine superfamily because it is located predominantly in the brain in which it is expressed constitutively on specific subsets of neurons. To elucidate the possible role of neuronally expressed fractalkine in the inflammatory response to neuronal injury, we have analyzed the regulation of fractalkine mRNA expression and protein cleavage under conditions of neurotoxicity. We observed that mRNA encoding fractalkine is unaffected by experimental ischemic stroke (permanent middle cerebral artery occlusion) in the rat. Similarly, in vitro, levels of fractalkine mRNA were unaffected by ensuing excitotoxicity. However, when analyzed at the protein level, we found that fractalkine is rapidly cleaved from cultured neurons in response to an excitotoxic stimulus. More specifically, fractalkine cleavage preceded actual neuronal death by 2-3 hr, and, when evaluated functionally, fractalkine represented the principal chemokine released from the neurons into the culture medium upon an excitotoxic stimulus to promote chemotaxis of primary microglial and monocytic cells. We further demonstrate that cleavage of neuron-derived, chemoattractive fractalkine can be prevented by inhibition of matrix metalloproteases. These data strongly suggest that dynamic proteolytic cleavage of fractalkine from neuronal membranes in response to a neurotoxic insult, and subsequent chemoattraction of reactive immune cells, may represent an early event in the inflammatory response to neuronal injury.

TR3 death receptor expression in the normal and ischaemic brain.

Members of the death receptor family may play a prominent role in developmental and pathological neuronal cell death. We report the expression of the TR3 and TR7 death receptors in the adult human and rat central nervous system. Whereas expression of TR3 appears to be high in the human cerebellum, with lower levels in other brain regions, robust expression is observed in many regions of the rat brain. We also analyzed modulation of death receptor expression in an in vivo rat model of acute stroke. In contrast to tumor necrosis factor receptor 1, Fas and p75(NGFR), which all show up-regulation specifically in lesioned cortex of the permanent middle cerebral artery occlusion model of stroke. TR3 shows a rapid global increase in both lesioned and unlesioned brain. In comparison, the recently described death receptor TR7 shows no change in this model. These data indicate that the death receptors show clear differences in patterns of expression in response to ischemic injury. ¿ 2000 IBRO. Published by Elsevier Science Ltd.

Pulmonary interstitial fibrosis and haemosiderin-laden macrophages: late following heart transplantation.

Impairment of pulmonary diffusion is recognized following heart transplantation. This study was undertaken to determine the histopathological basis for the defect in pulmonary physiology. Heart transplant recipients (HTR) entered into a prospective study of post-transplant pulmonary physiology were asked to undergo bronchoscopy, bronchoalveolar lavage (BAL) and transbronchial biopsy (n = 18) in the presence of impaired gas transfer. Transbronchial biopsies were examined under light microscopy and demonstrated focal interstitial fibrosis in 12 patients, cytomegalovirus disease in four patients and Pneumocystis carinii pneumonia in three patients. Bronchoalveolar lavage differential counts were normal in HTR but BAL macrophages contained haemosiderin. The histological features of interstitial fibrosis may underlie the fall in gas transfer seen following heart transplantation. The presence of haemosiderin-laden macrophages late following heart transplantation suggests a capillary leak syndrome.

Pulmonary diffusion impairment following heart transplantation: a prospective study.

The aim of this prospective study was to confirm whether and when a fall in gas transfer occurs following heart transplantation (HT); and to examine the potential relationship between gas transfer and haemodynamic change, immuno-suppression and cytomegalovirus (CMV) infection. The lung physiology of 34 heart transplant recipients (HTR) and 14 control patients undergoing coronary artery bypass grafting (CABG) were studied. The absolute and standardized residual values of forced expiratory volume in one second (FEV1), forced vital capacity (FVC), residual volume (RV), forced residual volume (FRC), total lung capacity (TLC), transfer factor of the lungs for carbon monoxide (TL,CO) and carbon monoxide transfer coefficient (KCO) were measured before and at 30, 60, 90, 120 and 150 days after HT. These data were compared to haemodynamic status, graft rejection, cyclosporin levels and episodes of CMV infection. Lung function was studied in a group of patients before and 4 weeks after CABG. There was a significant fall in mean KCO after HT (pre-HT = 1.29 and post-HT = 1.06 mmol.min-1.kPa.L-1) but not after CABG (pre-CABG = 1.49, post-CABG = 1.5 mmol.min-1.kPa.L-1. No relationship was observed between gas transfer and CMV. At the latest stage following HT (150 days) there was a positive correlation between TL,CO (absolute value and standardized residual) and mean cyclosporin level (r = 0.48 and r = 0.44, respectively) and also between the absolute KCO and actual (r = 0.56) and mean (r = 0.55) cyclosporin levels. Following HT, there is an early fall in gas transfer, which is independent of the effects of surgery and bypass, implicating early immunosuppression (e.g. antithymocyte globulin/cyclosporin).

Prostitution, AIDS, and preventive health behavior.

Although considerable attention has been placed on the role of prostitutes in the AIDS epidemic, little attention has been directed to features of prostitutes' work lives which are relevant to the control of AIDS. This article reviews several aspects of prostitution in the United States which have implications for control of the epidemic. The article first reviews the epidemiology of human immunodeficiency virus (HIV) infection among prostitutes. The legalized system of prostitution in Nevada serves as a basis for comparison to illegal prostitution. This article examines the effectiveness of mandatory testing of prostitutes for monitoring and controlling the epidemic. And finally, a peer education approach as a means to control HIV infection among prostitutes is explored.

Predictive value of oesophageal pH variables in children with gastro-oesophageal reflux.

Sixty three 24 hour oesophageal pH studies were performed in children with known or suspected gastro-oesophageal reflux and their progress was followed for at least a year. Forty two studies were of children who were well at follow up who were not receiving medical treatment for gastro-oesophageal reflux, and had not required antireflux surgery. Twenty one studies were from children who were still on treatment for gastro-oesophageal reflux or had undergone antireflux surgery. The results were analysed using non-parametric statistics to assess the value of pH recordings in predicting which children were likely to have continuing problems. Sleep reflux (minutes/hour) and acid clearing time were the most highly significant variables (p less than 0.0005 and p less than 0.0001 respectively). The results suggest that either sleep gastro-oesophageal reflux of more than 6.4 minutes/hour or an acid clearing time of seven minutes or more are good predictors of the clinical outcome (specificity 88.1%, sensitivity 81.0%). Three of the five children with false positive results were under 10 months of age. Positive results in this age group should therefore be interpreted with caution.

The five year school medical--time for change.

School medical records of 1000 children born in 1981 were studied retrospectively. They showed that once known medical problems and those screened for by the school nurse (hearing, vision, growth) were excluded, only 17 problems requiring treatment were discovered: speech (n = 10), development (n = 3), undescended testes (n = 3), and phimosis (n = 1). This indicates that routine screening by a nurse, backed up by selective medical examination by the school doctor, is efficient and effective.

Acodazole hydrochloride: phase I trial, pharmacokinetics, and evaluation of cardiotoxicity in dogs.

Acodazole (NSC 305884) was examined in a Phase I trial evaluating a 1-h infusion repeated every 21 days in 37 patients with advanced carcinomas. Cardiac toxicity was dose-limiting at 1370 mg/m2, manifested as multiple premature ventricular contractions, QTc interval prolongation, and decreasing heart rate. Other toxicities included mild to moderate nausea and vomiting and local reaction near the i.v. injection site requiring the use of central venous catheters. Antineoplastic activity was not observed. Acodazole levels assayed by high-performance liquid chromatography disclosed a peak plasma level of 19 +/- 4 (SEM) micrograms/ml for 1370 mg/m2. Acodazole plasma levels decreased in a triphasic manner over a 100-fold range. The volume of distribution at steady state was 238 +/- 18 liter/m2 suggesting extensive tissue binding. The total body clearance was 13.6 +/- 0.9 liter/h/m2; the percentage of urinary excretion was 29 +/- 2% for 48 h. To evaluate cardiac toxicity, acodazole was administered to five dogs at 2262 mg/m2 (1-h infusion) which provided plasma concentrations similar to those achieved at 1370 mg/m2 in humans. Consistent findings in dogs were drug-related prolongation of QTc intervals, and reduction in heart rate, left ventricular dP/dt, and mean blood pressures. Clinical development of acodazole requires studies to further elucidate and alleviate this cardiac toxicity.

Coronary reperfusion following experimental myocardial infarction.

Numerous studies have shown that early coronary reperfusion is feasible in the setting of evolving acute myocardial infarction in man. While early reperfusion reduces myocardial infarct size, there are potentially deleterious consequences of reperfusion. The concept of "reperfusion injury", oxygen-free radical damage, no reflow phenomenon, and stunned myocardium are discussed.

Effect of verapamil on infarct size in dogs subjected to coronary artery occlusion with transient reperfusion.

Reocclusion after successful coronary reperfusion occurs in 15 to 35% of patients receiving thrombolytic therapy for acute myocardial infarction. The present study was designed to simulate the clinical situation of reocclusion and determine whether verapamil might be effective in reducing myocardial necrosis and preserving high energy phosphates in this setting. Pentobarbital-anesthetized, open chest dogs underwent occlusion of the left anterior descending coronary artery for 2 hours followed by 1 hour of reperfusion and a further 4 hours of coronary artery occlusion. Treatment with verapamil (intravenous bolus dose of 0.2 mg/kg body weight followed by infusion of 0.56 +/- 0.14 mg/kg per h) was begun 1 hour after occlusion and infusion was continued for the remainder of the experiment. The dose of verapamil was adjusted to lower mean arterial pressure to approximately 90 mm Hg. The area at risk was determined by intraatrial injection of monastral blue dye and the area of necrosis was assessed by triphenyltetrazolium chloride staining. In vivo myocardial needle biopsy for determination of adenosine triphosphate and creatine phosphate was performed at the end of the experiment. The area of the left ventricle at risk was similar in both groups (control [n = 8], 20.2 +/- 1.6% versus verapamil-treated [n = 9], 23.1 +/- 2.9%; p = NS). The area of necrosis expressed as a percent of the area at risk was reduced in the verapamil-treated group compared with the control group (43.3 +/- 5.0% versus 63.1 +/- 6.8%, p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Total colonoscopy in children.

One hundred and twenty-three total colonoscopies were performed on 115 children with ages ranging from 3 months to 16 years. The major indications were suspected inflammatory bowel disease and unexplained rectal bleeding. Ninety-seven per cent of all procedures were carried out with sedation only. Adult colonoscopes were used in most of the patients but in babies and small children paediatric instruments were preferable. Total colonoscopy was possible in all patients with a patent colon. The terminal ileum was examined in 63 patients. Endoscopic snare polypectomy was successfully carried out in 8 children and multiple haemangiomas were electrocoagulated in one. Total colonoscopy in this paediatric series proved to be at least as easy, rapid, well-tolerated, and safe as in adults. In selected patients as single colonoscopy can give an accurate diagnosis with biopsy proof and sometimes the opportunity for definitive treatment.

Chronic inflammatory bowel disease in childhood.

The diagnosis of Crohn's disease in childhood has been facilitated by the use of fibreoptic endoscopy with biopsies, complemented by double-contrast radiology. Clinical suspicion leads initially to several relevant blood tests. These are followed by endoscopy and multiple colonic biopsies or barium follow-through studies depending on whether large-bowel or small-bowel disease is suspected. The present approach to diagnosis is based on corroborative investigative techniques-endoscopy, radiology, and histology, The availability of paediatric colonoscopes of small diameter should make it possible for paediatricians to perform limited examinations, but when more extensive endoscopy is indicated the child should be referred to special centres.

Acute phase proteins in chronic inflammatory bowel disease in childhood.

The serum levels of five acute phase proteins (APP) were measured in 18 children with Crohn's Disease (CD) or ulcerative colitis (UC) and in two control groups. The levels of C-reactive protein, alpha 1-acid glycoprotein, alpha 1-antitrypsin, C9, and Factor B were significantly raised in patients with CD and UC with good separation from controls, but they were not entirely reliable used as screening tests unless used in combination. The levels of APP were monitored for periods varying from 18 to 28 months in each patient and found to reflect the disease activity in both CD and UC. On seven occasions the APP levels did not match the clinically assessed disease activity, but when the serum levels were related to outcome of the disease, C-reactive protein was found to be elevated--whether or not there were symptoms of the disease--in all patients who later had a relapse, while normal values were found in those who had a long remission. These results suggest that the estimation of Creative protein is of prognostic value and that its measurement is particularly useful in children with mild symptoms in whom disease activity and prognosis are difficult to assess.

Nocturnal growth hormone and gonadotrophin secretion in growth retarded children with Crohn's disease.

Although impaired growth hormone secretion in response to pharmacological stimuli occurs in some growth retarded children with Crohn's disease, its relationship to past and future th is uncertain. We have therefore determined the growth hormone and gonadotrophin response to the physiological stimulus of sleep by continuous venous sampling in five severely gonadotrophin profiles, the mean plasma hormone concentrations during the first five hours of sleep were determined. In three of the five patients, five hour mean growth hormone levels were reduced (3.8, 5.0, and 8.5 mU/l) compared with levels reported previously in normal short children (10-43 mU/l), although the pulsatile pattern of growth hormone secretion was preserved in all. Nocturnal growth hormone secretion was unrelated to the growth velocities of these children during both pre- and post-treatment assessment periods but a significant correlation was found between growth hormone concentration and a disease activity score (r = 0.79, P less than 0.05), suggesting that growth hormone release by the pituitary was influenced by the severity of the disease. Nocturnal growth hormone secretion was also correlated with gonadotrophin secretion (luteinising hormones, r = 0.99, and follicle stimulating hormone, r = 0.96; p less than 0.01) indicating more extensive hypothalamic-pituitary disturbance. These findings suggest that hypothalamic-pituitary function is depressed in growth retarded children with Crohn's disease, but that abnormalities of growth hormone secretion are unlikely to be directly involved in the growth retardation seen in this condition.

An accurate, long-term, pH-sensitive radio pill for ingestion and implantation.

The design of a new radiotelemetry capsule (26 mm long X 7.6 mm diameter), with an in vivo life of 1 month is described in the context of previous work in this field. In vitro evaluation of the capsule indicates an accuracy and performance comparable with a conventional pH meter. Several clinical applications are described, including measurements of gastrointestinal pH in humans and the measurement of extracellular pH in laboratory animals.