Clinical trials of endothelin antagonists in heart failure: a question of dose?

Circulating plasma endothelin (ET)-1 concentrations are substantially elevated, and correlate with the hemodynamic severity and New York Heart Association (NYHA) class, in patients with chronic heart failure (CHF). In early preclinical studies involving different models of experimental heart failure, ET antagonists reduced cardiac pressures, increased cardiac output, and prolonged survival. ET receptor antagonists also impressively improved systemic and pulmonary hemodynamics in patients with CHF, without causing neurohormonal activation. However, recent clinical trials, including the ENABLE (Endothelin Antagonist Bosentan for Lowering Cardiac Events in Heart Failure) and EARTH (Endothelin A Receptor Antagonist Trial in Heart Failure) studies, have shown neutral effects in terms of mortality and symptoms. This paper describes the possible reasons why benefit was not seen in these clinical studies, and suggests what lessons can be learnt from the way the studies were undertaken to apply to future studies.     

Statistical Use in Clinical Studies: Is There Evidence of a Methodological Shift?

Background

Several studies indicate that the statistical education model and level in medical training fails to meet the demands of clinicians, especially when they want to understand published clinical research. We investigated how study designs and statistical methods in clinical studies have changed in the last twenty years, and we identified the current trends in study designs and statistical methods in clinical studies.

Methods

We reviewed 838 eligible clinical study articles that were published in 1990, 2000, and 2010 in four journals New England Journal of Medicine, Lancet, Journal of the American Medical Association and Nature Medicine. The study types, study designs, sample designs, data quality controls, statistical methods and statistical software were examined.

Results

Substantial changes occurred in the past twenty years. The majority of the studies focused on drug trials (61.6%, n = 516). In 1990, 2000, and 2010, there was an incremental increase in RCT studies (74.4%, 82.8%, and 84.0%, respectively, p = 0.013). Over time, there was increased attention on the details of selecting a sample and controlling bias, and there was a higher frequency of utilizing complex statistical methods. In 2010, the most common statistical methods were confidence interval for superiority and non-inferiority comparison (41.6%), survival analysis (28.5%), correction analysis for covariates (18.8%) and Logistic regression (15.3%).

Conclusions

These findings indicate that statistical measures in clinical studies are continuously developing and that the credibility of clinical study results is increasing. These findings provide information for future changes in statistical training in medical education.     

Are Metastases from Metastases Clinical Relevant? Computer Modelling of Cancer Spread in a Case of Hepatocellular Carcinoma

Background

Metastasis formation remains an enigmatic process and one of the main questions recently asked is whether metastases are able to generate further metastases. Different models have been proposed to answer this question; however, their clinical significance remains unclear. Therefore a computer model was developed that permits comparison of the different models quantitatively with clinical data and that additionally predicts the outcome of treatment interventions.

Methods

The computer model is based on discrete events simulation approach. On the basis of a case from an untreated patient with hepatocellular carcinoma and its multiple metastases in the liver, it was evaluated whether metastases are able to metastasise and in particular if late disseminated tumour cells are still capable to form metastases. Additionally, the resection of the primary tumour was simulated. The simulation results were compared with clinical data.

Results

The simulation results reveal that the number of metastases varies significantly between scenarios where metastases metastasise and scenarios where they do not. In contrast, the total tumour mass is nearly unaffected by the two different modes of metastasis formation. Furthermore, the results provide evidence that metastasis formation is an early event and that late disseminated tumour cells are still capable of forming metastases. Simulations also allow estimating how the resection of the primary tumour delays the patient''s death.

Conclusion

The simulation results indicate that for this particular case of a hepatocellular carcinoma late metastases, i.e., metastases from metastases, are irrelevant in terms of total tumour mass. Hence metastases seeded from metastases are clinically irrelevant in our model system. Only the first metastases seeded from the primary tumour contribute significantly to the tumour burden and thus cause the patient''s death.     

Is It Always Unethical to Use a Placebo in a Clinical Trial?

Background to the debate: Placebos are used in trials to conceal whether a treatment is being given or not and hence to control for the psychosomatic effects of offering treatment. Placebo-controlled trials are controversial. Critics of such trials argue that if a proven effective therapy exists, a placebo should not be used. But proponents argue that placebo trials are still crucial to prove the efficacy and safety of many treatments.     

Are Plasma IL-10 Levels a Useful Marker of Human Clinical Tolerance in Peanut Allergy?

Background

Food allergies are a major component of the burden of allergic disease. Accurate risk assessment for prediction of future clinical reactivity or clinical tolerance is limited by currently available techniques. Recent studies suggest that constitutively elevated global serum levels of IL-10, a cytokine that down-regulates both Th1 and Th2 cytokine production, may be useful in identifying human clinical tolerance to foods.

Objective

Determine the usefulness of constitutive IL-10 levels as a marker of clinical tolerance to peanut in children and adults.

Methodology/Principal Findings

107 subjects who were clinically tolerant to peanut and 94 subjects who were clinically allergic to peanut participated. Plasma was analyzed via ELISA to quantify the frequency of individuals with constitutive IL-10 levels and the intensity of those responses. The data were then stratified by age, gender and clinical status to assess the utility of this putative biomarker in specific at-risk groups. All 201 subjects had readily quantified plasma IL-10. Levels were no higher in subjects who were clinically tolerant to peanut than those in individuals clinically allergic to peanut. Stratification by age, gender or both did not improve the capacity of IL-10 levels to identify clinical tolerance to peanut.

Conclusions/Significance

Plasma IL-10 levels are neither a useful biomarker of clinical tolerance to peanut nor a potential tool for identification of clinical tolerance to peanut in humans.     

Clinical aspects for the use of DNA image cytometry in detection of bladder cancer: a valuable tool?

The employment of DNA flow or image cytometry for oncological diagnostic procedures is favored because of its high correlation to tumor biological behavior. Prognostic statements and therapeutic strategies therefore are based on the high validity of DNA cytometric measurements. Using 151 bladder washings from patients suspected of bladder cancer for this study, we examined the clinical value of various common methods of DNA single cell (SCI) and stemline interpretations (SLI). Comparing the specificity and sensitivity of DNA image cytometry in detection of bladder tumors, we found 81 and 52%, respectively, for SCI of Boecking, 84 and 45% for SLI of Boecking, 61 and 58% for SLI of Fu, and 82 and 40% for conventional stemline interpretation. To improve diagnostic and prognostic validity of DNA image cytometry, we designed our own method of interpretation. In consequence, we identified six single DNA parameters out of all recorded measurements that correlated most to histopathological grading (G1-G3). Creating reference values at random and rating by points, we used a cytometric grading system for ranking. In detection of bladder cancer specificity and sensitivity ultimately arrived at almost 70% in application of our method. Thus, by this study, we were able to show that sensitivity of DNA examination can be increased by combining various DNA parameters. Apart from our own scheme, the discrepancy in interpretation of DNA image cytometry does not allow us to recommend this procedure as the only diagnostic in detection of bladder cancer. However, in regard to prognostic statements, particularly tumor biological behavior, DNA image cytometry appears to be useful.     

When is a cow in estrus? Clinical and practical aspects

Good detection of estrus is critically important in dairy husbandry. Incorrect detection of estrus is related to loss of profit due to extended calving intervals, milk loss, veterinary costs, etc. Detection of estrus remains a major problem despites enormous progress in the knowledge of reproductive physiology of the cow and in development of estrus detection aids. To achieve good estrus detection, many factors have to be taken into account. On one hand a cow has to express estrus and on the other hand the farmer has to detect it. Combined action of several hormones causes physiological changes that lead to ovulation and an environment in the uterus that allows sperm to fertilize the egg. Besides these internal actions, a number of external changes can be observed. When using visual observations, time of the day and time spend on observation have a great impact on detection rates. Many devices are available to aid in estrus detection, such as pedometers, mount devices, temperature, and hormone measurements.Expression of estrus can be influenced by many factors. Heritability, number of days postpartum, lactation number, milk production, and health are known to influence estrus expression. Environmental factors like nutrition, season, housing, herd size, etc. also play a role in estrus expression. To evaluate estrus detection, record keeping is very important; a number of formulas can be used to assess detection efficiency. Besides the farmer, the veterinarian and inseminator can play an important role in estrus confirmation and good insemination strategy. In the end, the time of ovulation and the age of the egg at sperm penetration is critical for conception. Therefore, emphasis in research needs to be on the timing of insemination relative to ovulation, and thus on the detection of ovulation.     

Clinical Implications of TβRII Expression in Breast Cancer

Objective

To explore the relationship between TβRII [type II TGFβ (transforming growth factor β) receptor] expression and clinicopathological characteristics, and to evaluate the prognostic significance of TβRII expression in breast cancer.

Methods

Clinicopathological data and prognostic information of 108 patients with histologically confirmed breast cancer who were surgically treated at China Medical University between January 2007 and September 2008 were reviewed and the association between the clinicopathological characteristics and TβRII expression was analyzed by chi-square test and multivariate analysis. The expression of TβRII was assessed by immunohistochemistry.

Results

Of the 108 patients, 60 cases were TβRII positive and 48 cases were negative. There was no significant association between TβRII expression of the patients older than 40 years and that of the younger than 40 years (56.0% vs 50.0%; P = 0.742). The TβRII expression rate was significantly increased in patients with lymph node metastasis compared to those without lymph node metastasis (67.40% vs 46.8%; P = 0.033). Statistically significant relationships were found between increasing tumor clinical stage and high TβRII expression (P = 0.011). TβRII expression was not associated with the expression of ER(estrogen receptor)、PR, (progesterone receptor)、Her-2 (human epidermal growth factor receptor 2) (P = 0.925,P = 0.861, and P = 0.840, respectively). Patients with high TβRII expression showed poorer 5-year disease-free survival (DFS) compared to those with low expression (66.7% vs 45.6%; P = 0.028) by univariate analysis. Survival analysis demonstrated that TβRII was associated with poor DFS (P = 0.011). Subgroup analysis revealed that TβRII expression was associated with shorter DFS in patients with lymph node metastasis, ER-positive, PR-positive or Her-2-negative tumors (P = 0.006, P = 0.016, P = 0.022, and P = 0.033, respectively). Cox regression analysis revealed that high TβRII expression was related to poor 5-year DFS, and it was an independent factor for predicting the poor outcome for breast cancer patients (P = 0.016).

Conclusions

High levels of TβRII expression were associated with lymph node metastasis, increasing tumor clinical stage, and poorer 5-year DFS in patients with breast cancer. TβRII may be a potential prognostic marker for breast cancer.     

Candida galli as a Cause of Tinea Unguium—Molecular Characterization of a Rare Clinical Fungal Entity

We present the first clinical report of an infection caused by Candida galli, an anamorphic yeast species in the Yarrowia clade. C. galli has been described in the literature only four times, but never before it has been isolated from clinical samples. The colony morphology on Sabouraud medium and morphotype on CHROMagar Candida medium were similar to C. lipolytica as well as the carbon assimilation profile. The phenotypic differences with C. lipolytica were the non-assimilation of N-acetyl glucosamine, the absence of urease activity, growth in 10 % NaCl with 5 % glucose and in vitamin-free medium. MALDI–TOF MS could not generate reliable identification of the strain. Molecular analysis based on amplification of the ITS1-5.8S-ITS2 rDNA regions confirmed the identity as C. galli. Antifungal susceptibility test clearly demonstrated high MICs to 5-fluorocytosine, amphotericin B and fluconazole, as in the species belonging to the Yarrowia clade.     

Does Clustering Affect the Usual Test Statistics of no Treatment Effect in a Randomized Clinical Trial?

In many clinical trials, such as those undertaken by large cooperative cancer groups (CALGB, ECOG, SWOG, EORTC), patients are randomized to one of two treatments within an institution (a hospital or clinic). In such clinical trials, it has often occurred that outcomes of patients within institutions tend to be similar (correlated), due, possibly, to unmeasured variables such as quality of staff or quality of hospital equipment. We discuss when, in the presence of random hospital effects, the usual test statistics for no treatment effect, which do not take these hospital effects into account, are (asymptotically) unbiased.     

Outfoxing a Rash: Clinical Example of Human–Wildlife Interaction

Increasing human–wildlife contact can manifest in a variety of clinical conditions that may be overlooked by both human health and veterinary professionals. We report on an outbreak of scabies infection in a community, affecting both animals and humans, and representing the effects of an emerging infectious disease in a wildlife population. These cases underscore the potential importance of animal sentinel events for human, animal, and ecosystem health. The treatment given to the human cases of infection ranged from aggressive therapy to watchful waiting, with similar outcomes. There is a need for further collaborative, evidence-based research by human and veterinary health professionals into the optimal treatment and prevention of infections resulting from cross-species transmission.     

Occurrence of Plasmid-Mediated AmpC β-Lactamases Among Escherichia coli and Klebsiella pneumoniae Clinical Isolates in a Tertiary Care Hospital in Bangalore

Therapeutic options for infections caused by gram-negative organisms expressing plasmid-mediated AmpC β-lactamases are limited because these organisms are usually resistant to all the β-lactam antibiotics, except for cefepime, cefpirome and the carbapenems. These organisms are a major concern in nosocomial infections and should therefore be monitored in surveillance studies. Hence, this study was aimed out to determine the prevalence of plasmid-mediated AmpC β-lactamases in E. coli and K. pneumoniae from a tertiary care in Bangalore. A total of 63 E. coli and 27 K. pneumoniae were collected from a tertiary care hospital in Bangalore from February 2008 to July 2008. The isolates with decreased susceptibility to cefoxitin were subjected to confirmation test with three dimensional extract tests. Minimum inhibitory concentrations (MICs) were determined by agar dilution method. Conjugation experiments, plasmid profiling and susceptibility testing were carried out to investigate the underlying mechanism of resistance. In our study, 52 (57.7%) isolates showed resistance to cefoxitin, the occurrence of AmpC was found to be 7.7% of the total isolates. Plasmid analysis of the selected isolates showed the presence of a single plasmid of 26 kb in E. coli and 2 Kb in K. pneumoniae. Plasmid-mediated AmpC β-lactamases were found in 11.1% of K. pneumoniae and in 6.3% of E. coli. Curing and conjugation experiments showed that resistance to cephamycins and cephalosporins was plasmid-mediated. Our study has demonstrated the occurrence of plasmid-mediated AmpC in E. coli and K. pneumoniae which illustrates the importance of molecular surveillance in tracking AmpC-producing strains at general hospitals and emphasizes the need for epidemiological monitoring.     

Generation and exploration of a dense genetic map in a region of a QTL affecting corpora lutea in a Meishan × Yorkshire cross

Previously genomic scans revealed quantitative trait loci (QTL) on porcine Chromosome 8 (SSC8) as significantly affecting the number of corpora lutea (CL) in swine. In one study, statistical evidence for the putative QTL was found in the chromosomal region defined by the microsatellites (MS) SW205, SW444, SW206, and SW29. A Yeast Artificial Chromosome library was screened by using the corresponding primers for clones containing these MS by PCR. From five positive YAC clones, 10 additional MS were isolated and mapped to SSC8 with the INRA-University of Minnesota porcine Radiation Hybrid (IMpRH) panel. The genetic map position of the QTL has been refined by addition of these 10 markers. The QTL evaluation included pedigrees of F₂-intercross Meishan × Yorkshire design, with phenotypic data of 108 F₂ female offspring and genotypic data for 29 MS markers on SSC8. The analysis was performed by using the least squares regression method. The calculated QTL effect for CL obtained by the multilocus least squares method showed a maximum test statistic (F value = 13.98) at position 99 cM between three MS derived from YACs containing SW205 and SW1843 spanning an interval of 7.1 cM. The point-wise (nominal) P-value was 5.21 × 10⁻⁶ corresponding to a genome-wide P-value of 0.009. The additive QTL effect explained 17.4% of the phenotypic variance. Received: 23 December 2000 / Accepted: 07 May 2001     

Delayed-Type Hypersensitivity to Metals of Environmental Burden in Patients with Takotsubo Syndrome – Is There a Clinical Relevance?

ObjectiveTakotsubo syndrome (TS) is a heart condition characterised by a sudden transient left ventricular dysfunction; its pathophysiology is probably associated with elevated levels of catecholamines but the exact mechanism is not known as yet. Literature and clinical experience suggest that TS affects persons with various comorbidities. This pilot work aims to evaluate the frequency of comorbidities with potential pathological immune reactivity, and to evaluate the potential association between TS and hypersensitivity to metals assessed by LTT-MELISA®.ConclusionOur work shows that conditions with pathological immune reactivity occur frequently in TS patients, and our data suggest a possible association between TS and hypersensitivity to metals (mercury in particular) evaluated by LTT-MELISA®. We also suggest that apart from the triggering stress factor, potential existence of other serious conditions should be considered when taking medical history of TS patients.     

Characterization of a transient unfolding intermediate in a core mutant of γS-crystallin

In many age-related and neurological diseases, formerly native proteins aggregate via formation of a partially unfolded intermediate. γS-Crystallin is a highly stable structural protein of the eye lens. In the mouse Opj cataract, a non-conservative F9S mutation in the N-terminal domain core of γS allows the adoption of a native fold but renders the protein susceptible to temperature- and concentration-dependent aggregation, including fibril formation. Hydrogen/deuterium exchange and denaturant unfolding studies of this mutant protein (Opj) have suggested the existence of a partially unfolded intermediate in its aggregation pathway. Here, we used NMR and fluorescence spectroscopy to obtain evidence for this intermediate. In 3.5 M urea, Opj forms a stable and partially unfolded entity that is characterized by an unstructured N-terminal domain and a largely intact C-terminal domain. Under physiologically relevant conditions, Carr-Purcell-Meiboom-Gill T₂-relaxation dispersion experiments showed that the N-terminal domain residues were in conformational exchange with a loosely structured intermediate with a population of 1-2%, which increased with temperature. This provides direct evidence for a model in which proteins of native fold can explore an intermediate state with an increased propensity for formation of aggregates, such as fibrils. For the crystallins, this shows how inherited sequence variants or environmentally induced modifications can destabilize a well-folded protein, allowing the formation of intermediates able to act as nucleation sites for aggregation and the accumulation of light-scattering centers in the cataractous lens.     

A comparison of ten digestive enzymes reveals a lack of chitinase in a phasmid and the loss of two β-glucanases in a mantid

In all developmental stages, the phasmid Peruphasma schultei (Conle & Hennemann, 2005) is an obligate herbivore, whereas the mantid Hierodula membranacea (Burmeister, 1838) is an obligatory carnivore. In P. schultei, the luminal activity of all enzymes is approxximately 50% in the crop and 50% in the midgut, which corresponds to the approximate 50 : 50 ratio of volumes of these two regions. These ratios would be expected in insects with a constant feeding rate on an unvaried diet. The enzyme activity and volume ratios in Hierodula membranacea vary considerably because of the irregular feeding habits. These differences in activity ratios between phasmids and mantids are not associated with the obligate phytophagous or carnivorous diet. The ratio of membrane bound to luminal aminopeptidases and disaccharidases in the midgut of both species are not significantly different and are within the normal range of other paurometabolous insects. Cellobiase and other plant cell wall digesting enzymes, laminarinase and cellobiase, are present in the phasmid but totally lacking in the mantid. The obligate carnivorous feeding habits of mantids could represent a selective factor leading to the loss of the ability to produce β-glucanases. Chitinase is a moulting enzyme in all insects, whereas, in H. membranacea, chitinase also occurs as a luminal digestive enzyme. This modified enzyme function requires production and secretion in another tissue, namely the midgut.