Biorelevant refinement of the Caco-2 cell culture model to assess efficacy of paracellular permeability enhancers

Epithelial cell monolayers are routinely used to evaluate efficacy of paracellular permeability enhancers (PPEs). The purpose of the present work was to investigate how biorelevant refinements to the Caco-2 cell model impact in vitro efficacy (decrease in transepithelial electrical resistance and increase in mannitol permeability) of PPEs. Standard transport buffer was replaced by fasted-state simulated intestinal fluid (FaSSIF) or serum; or stirring was performed to decrease the unstirred water layer thickness. Apical FaSSIF significantly reduced the efficacy of amphiphilic PPEs palmitoylcarnitine and hexadecylphosphocholine and reduced the amount of these PPEs associated with cells. In contrast, FaSSIF did not affect efficacy of nonamphiphilic PPEs, ethylenediaminetetraacetic acid or 3-nitrocoumarin. Basolateral serum increased the transepithelial flux of PPEs, but did not lessen their potency. Stirring increased the flux of all PPEs, and also enhanced the potency of the amphiphilic PPEs. These results show that inclusion of FaSSIF and agitation in the cellular models significantly alter the efficacy of amphiphilic PPEs but not of hydrophilic or lipophilic PPEs. Future studies should be directed at evaluating the ability to these refined in vitro systems to predict in vivo effects of PPEs.     

A familial syndrome due to Arg648Stop mutation in the X-linked renal chloride channel gene

 We describe a familial syndrome in two brothers who were investigated after the casual discovery of tubular proteinuria in their 1st month of life. During a follow-up of 20 and 11 years, respectively, the two children grew well and were asymptomatic, but developed the same biochemical abnormalities, i.e., tubular proteinuria and hyperphosphaturia, progressive decrease in serum phosphorus below the normal values for age, and an increase in serum 1,25-dihydroxyvitamin D levels over normal values. Moreover, hyperabsorptive hypercalciuria and systemic osteopenia developed and progressively worsened. In both children, at a different age, medullary nephrocalcinosis appeared. The oldest boy suffered a progressive decrease in urinary concentration ability and in glomerular filtration rate. Oral phosphate supplementation led to reversal of all biochemical abnormalities, with the exception of decreased phosphate tubular reabsorption and tubular proteinuria. With long-term phosphate supplementation, a normal bone mass was reached, while progression of nephrocalcinosis was arrested and impairment of renal function was slowed down. In a family study (siblings and parents), the only detectable abnormality was microglobinuria in the mother, thus suggesting a X-linked inheritance of this disorder. In the two probands a mutation within the renal chloride channel gene (CLCN5) was discovered. Received: 23 December 1997 / Revised: 7 July 1998 / Accepted: 7 July 1998     

Long-Term Efficacy of Adding Fenofibric Acid to Moderate-Dose Statin Therapy in Patients with Persistent Elevated Triglycerides

Objective

The objective of this study was to evaluate the long-term efficacy of adding fenofibric acid to moderate-dose statin therapy in patients at goal for low-density lipoprotein cholesterol (LDL-C) but with persistent hypertriglyceridemia.

Methods

This is a post hoc analysis of a subset of patients (N?=?92) with mixed dyslipidemia treated with moderate-dose statin (rosuvastatin 20 mg, simvastatin 40 mg, or atorvastatin 40 mg) for 12 weeks in three controlled trials who had achieved LDL-C <100 mg/dL but whose triglycerides remained >200 mg/dL, and had fenofibric acid 135 mg added to the moderate-dose statin in a 52-week open-label extension study. Lipid and apolipoprotein (Apo) values and the proportion of patients meeting individual and combined treatment targets with combination therapy were determined at scheduled visits during the 52-week study and compared with baseline (start of extension study).

Results

Addition of fenofibric acid to moderate-dose statin for 52 weeks resulted in significant (P?P?=?0.007), and LDL-C?+?non–HDL-C?+?ApoB?+?HDL-C?+?triglycerides (25.6% vs 0.0%) than at baseline.

Conclusions

The addition of fenofibric acid to moderate-dose statin in patients whose LDL-C was optimal but whose triglycerides remained >200 mg/dL led to additional improvements in non–HDL-C, ApoB, HDL-C, and triglycerides that resulted in greater proportions of patients attaining optimal levels of the individual parameters as well as simultaneously achieving optimal levels of these parameters and LDL-C.

     

Loss of CSMD1 expression is associated with high tumour grade and poor survival in invasive ductal breast carcinoma

CUB and SUSHI multiple domain protein 1 (CSMD1) is a candidate tumour suppressor gene that maps to chromosome 8p23, a region deleted in many tumour types including 50% of breast cancers. CSMD1 has homologies to proteins implicated in carcinogenesis. We aimed to study the expression pattern of the CSMD1 protein and evaluate its prognostic importance in invasive ductal carcinoma (IDC). An anti-CSMD1 antibody was developed and validated. The expression pattern of CSMD1 in normal breast and IDC samples was investigated by immunohistochemistry in 275 patients. Univariate and multivariate Cox regression analyses were performed. In normal breast duct epithelial cells, luminal, membranous and cytoplasmic CSMD1 staining was identified. Reduced expression of CSMD1 was detected in 79/275 (28.7%) of IDC cases. Low CSMD1 expression was significantly associated with high tumour grade (P = 0.003). CSMD1 expression was associated with overall survival (OS; HR = 0.607, 95%CI: 0.4–0.91, P = 0.018) but not with disease-free survival (DFS; HR = 0.81, 95%CI: 0.46–1.43, P = 0.48). Multivariate analysis showed that CSMD1, together with Nottingham Prognostic Index, was considered an independent predictor of OS (HR = 0.607, 95%CI: 0.4–0.91, P = 0.018) but not DFS (HR = 0.84, 95%CI: 0.46–1.5, P = 0.573). Reduction of CSMD1 expression was significantly associated with high tumour grade and decreased OS. Therefore, our results support the idea that CSMD1 is a tumour suppressor gene and suggest its possible use as a new prognostic biomarker. The membrane expression pattern of CSMD1 suggests that it may be a receptor or co-receptor involved in the process of signal transduction.     

Cost analysis of amlodipine versus enalapril in patients with coronary artery disease and normal blood pressure

Objectives

To analyse 2-year hospitalization and cost data collected during a prospective, double-blind, randomized, controlled trial comparing amlodi-pine, enalapril and placebo in normotensive patients with coronary artery disease (CAD).

Methods

All patients who were enrolled in the CAMELOT study were included in this economic substudy. Patients with CAD and normal blood pressure were randomized to amlodipine, enalapril or placebo, and followed up for 24 months (between 1999 and 2004). Data on hospitalizations and medication use were obtained from the clinical trial. Costs were assigned from secondary sources. Total costs ($US, year 2004 values) were estimated as the sum of costs associated with cardiovascular hospitalizations, study medications and concomitant cardiovascular medications. Costs and resource use were analysed by treatment arm overall and for selected patient subgroups. Cost differences were evaluated using nonparametric bootstrap techniques.

Results

Of 1991 patients enrolled, 663 were treated with amlodipine, 673 were treated with enalapril and 655 were treated with placebo. Significantly fewer patients were hospitalized for cardiovascular reasons in the amlodipine group (16.4%) than in the placebo group (22.7%;p < 0.01), but not compared with the enalapril group (20.1%;p = 0.09). The amlodipine group also had numerically fewer days in hospital per patient (1.1) than the enalapril (1.3) and placebo (1.5) groups. Mean 2-year per-patient costs in the amlodipine group were estimated to be $US609 and $US717 lower than for the placebo and enalapril groups, respectively.

Conclusions

These results suggest that use of amlodipine may reduce costs of care among CAD patients with normal blood pressure.     

Coinheritance of two rare genodermatoses (Papillon-Lefèvre syndrome and oculocutaneous albinism type 1) in two families: a genetic study

The co-occurrence of two rare recessive genetic conditions in apparently unrelated individuals or families is extremely rare. Two geographically distant and apparently unrelated families were identified in which individuals were simultaneously affected by two rare recessive mendelian syndromes, Papillon-Lefevre syndrome and type 1 oculocutaneous albinism. The families were tested for mutations in the causative genes, cathepsin C (CTSC) and tyrosinase (TYR), respectively, by direct sequencing. To assess the relationship of the two families, both families were tested for polymorphisms at eight microsatellite markers spanning both CTSC and TYR loci. Independent mutations (c.318-1G-->A and c.817G-->C/p.W272C) were identified in CTSC and TYR, respectively, that were shared by the affected individuals in both families. The two affected genes lie close together on chromosome bands 11q14.2-14.3, and studies with linked genetic markers suggested that the families shared a small chromosomal segment carrying both mutations that had been transmitted intact from a remote common ancestor. The co-occurrence of the two rare diseases in multiple families depends on their shared chromosomal location, but not on any shared pathogenic mechanism.     

Head-up tilt testing in children and young people: a retrospective observational study

Aim: Head‐up tilt testing (HUTT) is the gold standard investigation for adults with transient loss of consciousness (TLOC), but it is controversial in children and young people, because of a lack of systematic investigation and because the test can be uncomfortable. As it was introduced recently for children attending our hospital, we undertook a retrospective registered clinical audit of its usefulness. Methods: The medical records of 100 consecutive patients aged less than 18 years undergoing HUTT from October 2001 to December 2008 were reviewed. Information about their episodes, prodromes, triggers, previous tests, indications for the HUTT, the HUTT and clinical outcomes was extracted. Results: Children were 6–17 years old; 68/100 were female. In 32/100, no trigger was reported. The most reported triggers included standing up (20%) and prolonged standing (18%). Dizziness (64%) and altered vision (39%) were the most experienced prodromal symptoms. Twenty‐eight of 100 had a positive test, with reproduction of symptoms in 24. Seventeen of 100 tests were negative but symptomatic; 55/100 had a negative asymptomatic test. In 17/28 positive HUTTs, the tilt confirmed the suspected diagnosis and elucidated the mechanism. Two of 28 were started on medication. However, in 9/28, neither was the diagnosis clarified nor was therapy instigated. Conclusions: Potentially useful information about the TLOC was obtained in 45/100 cases. The 17/100 with negative but symptomatic results may have had medically unexplained TLOC or emotional attacks, although without concurrent electroencephalogram, some uncertainty remains. Therefore, a new protocol with video–electroencephalogram–polygraphy and beat‐to‐beat finger blood pressure recording, and more explicit clinical reporting is being developed.     

Genetic contribution to bone metabolism, calcium excretion, and vitamin D and parathyroid hormone regulation.

A classical twin study was performed to assess the relative contribution of genetic and environmental factors to bone metabolism, calcium homeostasis, and the hormones regulating them. It was examined further whether the genetic effect is menopause dependent. The subjects were 2136 adult twins (98.3% female): 384 monozygotic (MZ) and 684 dizygotic (DZ) twin pairs. The intraclass correlations were calculated, and maximum likelihood model fitting was used to estimate genetic and environmental variance components. The intraclass correlations for all of the variables assessed were higher in MZ twin pairs. The heritabilities (95% CIs) obtained from model fitting for hormones regulating bone metabolism and calcium homeostasis were parathyroid hormone (PTH), 60% (54-65%); 25-hydroxyvitamin D [25(OH)D]; 43% (28-57%), 1,25-hydroxyvitamin D [1,25(OH)], 65% (53-74%); and vitamin D binding protein 62% (56-66%). The heritabilities (95% CIs) for markers of bone formation also were assessed; bone-specific alkaline phosphatase (BSAP), 74% (67-80%), and osteocalcin, 29% (14-44%); marker of bone resorption deoxypyridinoline (DPD), 58% (52-64%); and measure of calcium homeostasis 24 h urine calcium, creatinine (Cr), 52% (41-61%). The magnitude of genetic influence differed with menopause for most variables. This study provides evidence for the importance of genetic factors in determining bone resorption and formation, calcium excretion, and the hormones regulating these processes. It shows for the first time a clear genetic effect on bone resorption in premenopausal women and the regulation of PTH, vitamin D metabolism, and calcium excretion. The genes controlling bone hormones and markers are likely to be useful therapeutic and diagnostic targets.