True Biological Effect Research Articles

Massive false-positive gene–gene interactions by Rothman’s additive model

Rothman’s additive interaction model based on additivity of the relative risks of two disease-risk factors has frequently been used to investigate gene–gene interactions for various traits, especially for rheumatic diseases...

A systematic review and meta-analysis of the impact of mineralocorticoid receptor antagonists on glucose homeostasis.

Spironolactone, a nonselective mineralocorticoid receptor antagonist (MRA), may have a deleterious effect on glycemia. The objective of this review was to assess current knowledge on MRAs' influence (spironolactone, eplerenone, and canrenone) on glucose homeostasis and the risk of diabetes. A systematic review was conducted using the Medline database on articles published from 1946 to January 2017 that studied the effects of MRAs on any glucose-related endpoints, without any restrictions regarding the participants' characteristics.Study design, patient population, dose and duration of intervention, and the quantitative results on glycemic markers were extracted, interpreted for result synthesis, and evaluated for sources of bias. From the articles included in the qualitative analysis, a select number were used in a meta-analysis on studies having measured glycated hemoglobin (HbA1c) or risk of diabetes. Seventy-two articles were selected from the Medline database and references of articles. Results on spironolactone were heterogeneous, but seemed to be disease-specific. A potential negative effect on glucose regulation was mainly observed in heart failure and diabetes trials, while a neutral or positive effect was detected in diseases characterized by hyperandrogenism, and inconclusive for hypertension. Interpretation of data from heart failure trials was limited by the small number of studies. From a meta-analysis of 12 randomized controlled studies evaluating spironolactone's impact on HbA1c in diabetic patients, spironolactone had a nonsignificant effect in parallel-group studies (mean difference 0.03 [-0.20;0.26]), but significantly increased HbA1c in crossover studies (mean difference 0.24 [0.18;0.31]). Finally, eplerenone did not seem to influence glycemia, while limited data indicated that canrenone may exert a neutral or beneficial effect.The studies had important limitations regarding study design, sample size, duration of follow-up, and choice of glycemic markers. Spironolactone may induce disease-specific and modest alterations on glycemia. It is uncertain whether these effects are transient or not. Data from the most extensively studied population, individuals with diabetes, do not support a long-term glycemic impact in these patients. Further prospective studies are necessary to establish spironolactone's true biological effects and their clinical implications.

Correlation and cluster analysis of immunomodulatory drugs based on cytokine profiles

Drug discovery is a constant struggle to overcome hurdles posed by the complexity of biological systems. One of these hurdles is to find and understand the molecular target and the biological mechanism of action. Although the molecular target has been determined, the true biological effect may be unforeseen also for well-established drugs. Hence, there is a need for novel ways to increase the knowledge of the biological effects of drugs in the developmental process. In this study, we have determined cytokine profiles for 26 non-biological immunomodulatory drugs or drug candidates and used these profiles to cluster the compounds according to their effect in a preclinical ex vivo culture model of arthritis. This allows for prediction of functions and drug target of a novel drug candidate based on profiles obtained in this study. Results from the study showed that the JAK inhibitors tofacitinib and ruxolitinib formed a robust cluster and were found to have a distinct cytokine profile compared to the other drugs. Another robust cluster included the calcineurin inhibitors cyclosporine A and tacrolimus and the protein kinase inhibitors fostamatinib disodium and sotrastaurin acetate, which caused a strong overall inhibition of the cytokine production. The results of this methodology indicate that cytokine profiles can be used to provide a fingerprint-like identification of a drug as a tool to benchmark novel drugs and to improve descriptions of mode of action.

Statistical Analysis of Zebrafish Locomotor Behaviour by Generalized Linear Mixed Models

Upon a drastic change in environmental illumination, zebrafish larvae display a rapid locomotor response. This response can be simultaneously tracked from larvae arranged in multi-well plates. The resulting data have provided new insights into neuro-behaviour. The features of these data, however, present a challenge to traditional statistical tests. For example, many larvae display little or no movement. Thus, the larval responses have many zero values and are imbalanced. These responses are also measured repeatedly from the same well, which results in correlated observations. These analytical issues were addressed in this study by the generalized linear mixed model (GLMM). This approach deals with binary responses and characterizes the correlation of observations in the same group. It was used to analyze a previously reported dataset. Before applying the GLMM, the activity values were transformed to binary responses (movement vs. no movement) to reduce data imbalance. Moreover, the GLMM estimated the variations among the effects of different well locations, which would eliminate the location effects when two biological groups or conditions were compared. By addressing the data-imbalance and location-correlation issues, the GLMM effectively quantified true biological effects on zebrafish locomotor response.

Reply to Urinary toxicity after stereotactic body radiotherapy: The boy who cried wolf?

We thank Kishan et al for their insightful comments regarding our analysis of the utilization of and toxicity after stereotactic body radiotherapy (SBRT) for localized prostate cancer.1 The authors raise concerns regarding the selection and interpretation of toxicity endpoints, particularly the composite endpoint of urinary incontinence. Our definitions of urinary incontinence and nonincontinence toxicity were derived from prior population-based studies evaluating radiation therapy for localized prostate cancer.2 These studies and ours used both International Classification of Diseases, Ninth Revision (ICD-9) diagnosis codes and Current Procedural Terminology, 4th Edition (CPT-4) procedural codes to capture all outcomes. In light of the aforementioned commentary, we performed a sensitivity analysis separately stratifying toxicity outcomes according to ICD-9 and CPT-4 codes (diagnostic vs therapeutic). According to ICD-9 diagnosis codes alone, SBRT has a higher rate of urinary incontinence at 2 years than conventional intensity-modulated radiation therapy (IMRT; 6.3% vs 5.9%; P < .001). Likewise, SBRT has a higher rate of urinary incontinence–related diagnostic procedures at 2 years (20.5% vs 16.7%; P < .001). When outcomes were stratified according to therapeutic procedures, there was no difference between SBRT and IMRT at 2 years (the absolute incidence was not reportable because of the cell size; P = .21). How can we interpret these findings? Kishan et al note that the absolute rates of urinary incontinence reported herein are higher than the incidence of severe toxicity reported in prior clinical trials, and they suggest that this may be an artifact of code selection. However, the use of diagnostic procedures as a surrogate for urinary incontinence–related toxicity may better capture true clinical toxicity because these procedures signal the presence of symptoms prompting an evaluation. Furthermore, even when diagnosis codes alone are examined and this results in lower absolute toxicity rates consistent with prior studies, SBRT demonstrates a higher incidence of incontinence. The authors suggest that this difference may result from a detection bias because men undergoing SBRT were often enrolled in clinical trials with rigorous protocols for the identification of toxicity. To our knowledge, there is no research to demonstrate different practice patterns concerning the diagnosis and treatment of subjects enrolled in clinical trials. Even so, a true biological effect cannot be ruled out. Although these data demonstrate a statistically significant difference in urinary toxicity after SBRT versus IMRT, the clinical significance of these findings, particularly over the long term, warrants further investigation. A higher demand for incontinence-related procedures after SBRT may be driven by greater patient bother. Furthermore, although we found that SBRT's 1-year overall costs were lower than IMRT's costs, more diagnostic procedures following SBRT may ultimately generate greater costs in the long term. Likewise, longer follow-up is needed to determine whether differences in toxicity persist over time. Ultimately, we agree that only randomized controlled data will truly elucidate the risk of toxicity after SBRT. In the interim, we aim neither to cry wolf nor to be the canary in the coal mine. Rather, we hope to shed light on the ongoing utilization and outcomes of SBRT to inform current clinical practice, in the context of our study design, until randomized data are available. No specific funding was disclosed. Jim C. Hu belongs to the Speakers' Bureau for Intuitive Surgical and Genomic Health. Joshua A. Halpern, MD, MS Department of Urology Weill Cornell Medicine New York, New York Art Sedrakyan, MD, PhD Department of Healthcare Policy and Research Weill Cornell Medicine New York, New York Wei-Chun Hsu, MS Department of Healthcare Policy and Research Weill Cornell Medicine New York, New York Jim C. Hu, MD, MPH Department of Urology Weill Cornell Medicine New York, New York

From Function to Phenotype: Impaired DNA Binding and Clustering Correlates with Clinical Severity in Males with Missense Mutations in MECP2.

Mutations in the MECP2 gene cause Rett syndrome (RTT). MeCP2 binds to chromocentric DNA through its methyl CpG-binding domain (MBD) to regulate gene expression. In heterozygous females the variable phenotypic severity is modulated by non-random X-inactivation, thus making genotype-phenotype comparisons unreliable. However, genotype-phenotype correlations in males with hemizygousMECP2 mutations can provide more accurate insights in to the true biological effect of specific mutations. Here, we compared chromatin organization and binding dynamics for twelve MeCP2 missense mutations (including two novel and the five most common MBD missense RTT mutations) and identifiedacorrelation with phenotype in hemizygous males. We observed impaired interaction of MeCP2-DNA for mutations around the MBD-DNA binding interface, and defective chromatin clustering for distal MBD mutations. Furthermore, binding and mobility dynamics show a gradient of impairment depending on the amino acid properties and tertiary structure within the MBD. Interestingly, a wide range of phenotypic/clinical severity, ranging from neonatal encephalopathy to mild psychiatric abnormalities were observed and all are consistent with our functional/molecular results. Overall, clinical severity showed a direct correlation with the functional impairment of MeCP2. These mechanistic and phenotypic correlations of MeCP2 mutations will enable improved and individualized diagnostics, and may lead to personalized therapeutic interventions.

Parsing psychosis subtypes through investigations of rare genetic variants

Parsing psychosis subtypes through investigations of rare genetic variants

Endotoxin contamination: a key element in the interpretation of nanosafety studies.

The study of toxicity and potential risks of engineered nanoparticles is of particular importance in nanomedicine. Endotoxin, a common contaminant of bacterial origin, has biological effects that can mask the true biological effects of nanoparticles, if its presence is overlooked. In this review, we report the features of nanoparticle contamination by endotoxin, and the different biological effects of endotoxin-contaminated nanoparticles. We will describe different methods for endotoxin detection applied to nanoparticles, and discuss their pros and cons. Eventually, we describe various methods for eliminating endotoxin contamination in nanoparticles. Although there is no universal technique for efficiently removing endotoxin from nanoparticles, specific solutions can be found case by case, which can allow us to perform nanosafety studies in biologically relevant conditions.

Circulation : Clinical Summaries

<i>Circulation</i> : Clinical Summaries

Cerebrovascular Diseases in Workers at Mayak PA: The Difference in Radiation Risk between Incidence and Mortality.

A detailed analysis of cerebrovascular diseases (CeVD) for the cohort of workers at Mayak Production Association (PA) is presented. This cohort is especially suitable for the analysis of radiation induced circulatory diseases, due to the detailed medical surveillance and information on several risk factors. The risk after external, typically protracted, gamma exposure is analysed, accounting for potential additional internal alpha exposure. Three different endpoints have been investigated: incidence and mortality from all cerebrovascular diseases and incidence of stroke. Particular emphasis was given to the form of the dose-response relationship and the time dependence of the radiation induced risk. Young attained age was observed to be an important, aggravating modifier of radiation risk for incidence of CeVD and stroke. For incidence of CeVD, our analysis supports a dose response sub-linear for low doses. Finally, the excess relative risk per dose was confirmed to be significantly higher for incidence of CeVD compared to CeVD mortality and incidence of stroke. Arguments are presented for this difference to be based on a true biological effect.

Meat Intake and Reproductive Parameters Among Young Men

In the United States, anabolic sex steroids are administered to cattle for growth promotion. There is concern regarding the reproductive consequences of this practice in men who eat beef. We investigated whether meat consumption was associated with semen quality parameters and reproductive hormone levels in young men. Semen samples were obtained from 189 men aged 18-22 years. Diet was assessed with a previously validated food frequency questionnaire. We used linear regression to analyze the cross-sectional associations of meat intake with semen quality parameters and reproductive hormones while adjusting for potential confounders. There was an inverse relation between processed red meat intake and total sperm count. The adjusted relative differences in total sperm counts for men in increasing quartiles of processed meat intake were 0 (ref), -3 (95% confidence interval = -67 to 37), -14 (-82 to 28), and -78 (-202 to -5) million (test for trend, P = 0.01). This association was strongest among men with abstinence time less than 2 days and was driven by a strong inverse relation between processed red meat intake and ejaculate volume (test for trend, P = 0.003). In our population of young men, processed meat intake was associated with lower total sperm count. We cannot distinguish whether this association is because of residual confounding by abstinence time or represents a true biological effect.

Testing the Sensitivity of Tract-Based Spatial Statistics to Simulated Treatment Effects in Preterm Neonates

Early neuroimaging may provide a surrogate marker for brain development and outcome after preterm birth. Tract-Based Spatial Statistics (TBSS) is an advanced Diffusion Tensor Image (DTI) analysis technique that is sensitive to the effects of prematurity and may provide a quantitative marker for neuroprotection following perinatal brain injury or preterm birth. Here, we test the sensitivity of TBSS to detect diffuse microstructural differences in the developing white matter of preterm infants at term-equivalent age by modelling a ‘treatment’ effect as a global increase in fractional anisotropy (FA). As proof of concept we compare these simulations to a real effect of increasing age at scan. 3-Tesla, 15-direction diffusion tensor imaging (DTI) was acquired from 90 preterm infants at term-equivalent age. Datasets were randomly assigned to ‘treated’ or ‘untreated’ groups of increasing size and voxel-wise increases in FA were used to simulate global treatment effects of increasing magnitude in all ‘treated’ maps. ‘Treated’ and ‘untreated’ FA maps were compared using TBSS. Predictions from simulated data were then compared to exemplar TBSS group comparisons based on increasing postmenstrual age at scan. TBSS proved sensitive to global differences in FA within a clinically relevant range, even in relatively small group sizes, and simulated data were shown to predict well a true biological effect of increasing age on white matter development. These data confirm that TBSS is a sensitive tool for detecting global group-wise differences in FA in this population.

FRI0099 Obese patients with rheumatoid arthritis have reduced response rates to biologic anti-rheumatic agents: a comparison between european and american ra patients

BackgroundObesity in RA patients has been associated with an increased risk of comorbidity, total joint replacement, pain, disability, high medical costs and impaired quality of life. One study has suggested...

Causal Genomic and Epigenomic Network Analysis emerges as a New Generation of Genetic Studies of Complex Diseases.

Causal Genomic and Epigenomic Network Analysis emerges as a New Generation of Genetic Studies of Complex Diseases.

Meta‐analysis: peri‐operative anti‐TNFα treatment and post‐operative complications in patients with inflammatory bowel disease

The impact of peri-operative use of TNFα antagonists on post-operative complications such as infection and wound healing is controversial. To conduct a systematic review and meta-analysis to assess the impact of peri-operative use of TNFα antagonists on post-operative complications such as infection and wound healing in patients with inflammatory bowel disease (IBD). A literature search identified studies that investigated post-operative outcomes in patients with IBD using TNFα antagonists. The primary outcome was the rate of post-operative infectious complications. Secondary outcomes included the rates of non-infectious complications and total complications. Odds ratios (OR) with 95% confidence intervals (CI) are reported. Overall, 18 studies with 4659 participants were eligible for inclusion. Patients with IBD using preoperative anti-TNFα therapies had significant increases in post-operative infectious [OR 1.56 (95% CI, 1.09-2.24)], non-infectious [OR 1.57 (95% CI, 1.14-2.17)] and total complications [OR 1.73 (95% CI, 1.23-2.43)]. Studies limited to patients with Crohn's disease demonstrated a statistically significant increase in infectious (OR 1.93, 95% CI 1.28-2.89) and total (OR 2.19, 95% CI 1.69-2.84) complications, and a trend towards increase in non-infectious complications (OR 1.73, 95% CI 0.94-3.17). Studies of patients with ulcerative colitis did not demonstrate significant increases in infectious (OR 1.39, 95% CI 0.56-3.45), non-infectious (OR 1.40, 95% CI 0.68-2.85), or total complications (OR 1.10, 95% CI 0.81-1.47). Anti-TNFα therapies appear to increase the risk of post-operative complications. The increase in risk is small, and may well reflect residual confounding rather than a true biological effect. Nevertheless, physicians should exercise caution when continuing biological therapies during the peri-operative period.

The complexity of nanoparticle dissolution and its importance in nanotoxicological studies

Dissolution of nanoparticles (NPs) is an important property that alters their abundance and is often a critical step in determining safety of nanoparticles. The dissolution status of the NPs in exposure media (i.e. whether they remain in particulate form or dissolve — and to what extent), strongly affects the uptake pathway, toxicity mechanisms and the environmental compartment in which NPs will have the highest potential impact. A review of available dissolution data on NPs demonstrates there is a range of potential outcomes depending on the NPs and the exposure media. For example two nominally identical nanoparticles, in terms of size and composition, could have totally different dissolution behaviours, subject to different surface modifications. Therefore, it is imperative that toxicological studies are conducted in conjunction with dissolution of NPs to establish the true biological effect of NPs and hence, assist in their regulation.

Induction of Caspase-2 Activation by a DNA Enzyme Evokes Tumor Cell Apoptosis

Caspase-2 is an enigmatic caspase that is now increasingly being associated with certain types of cell death in cells exposed to cytotoxic agents. It is now known that in some cases of cell stress, such as DNA damage, activation of this caspase is triggered, sometimes in the absence of activation of both the intrinsic and extrinsic pathways of apoptosis. Part of the reason for this enigma has been lack of a suitable stimulus for this caspase, and with the discovery of DNAzyme 13 (Dz13), a potent oligonucleotide-based caspase-2 activator, much more can now be elucidated. For instance, one thing that could be unraveled is whether caspase-8 and Fas (CD95)-associated protein with death domain are indeed involved in caspase-2 activation as part of the death-inducing signaling complex. It is also becoming apparent that this enigmatic caspase may be important in the mechanisms behind which chemotherapeutic agents inhibit tumor cell growth. A better understanding of the true biological effects of this enzyme may indeed lead to more effective ways of managing tumors clinically. This review article briefly examines the different compounds capable of inducing activation of caspase-2 and proposes Dz13 as one that will be valuable for evaluation of the biological functions of caspase-2.

TP53 Arg72Pro Polymorphism and Colorectal Cancer Risk: A Systematic Review and Meta-Analysis

The TP53 rs1042522 polymorphism (c.215C>G, Arg72Pro) has been extensively investigated as a potential risk factor for colorectal cancer, but the results have thus far been inconclusive. We searched multiple electronic databases to identify studies investigating the association between the Arg72Pro polymorphism and colorectal cancer. Individual study odds ratios (OR) and their confidence intervals were estimated using allele-frequency, recessive, and dominant genetic models. Summary ORs where estimated using random effects models. We identified 23 eligible case-control studies, investigating 6,514 cases and 9,334 controls. There was significant between-study heterogeneity for all genetic models. The control group in one of the studies was not in Hardy-Weinberg equilibrium; only three studies reported that genotyping was blinded to case/control status and five studies used tumor tissue for case genotyping. Overall, we did not identify any association between rs1042522 and colorectal cancer risk under an allele-frequency comparison (OR, 0.99; 95% confidence interval, 0.89-1.09). Likewise, no association was evident under dominant or recessive models. Studies using tumor tissue for case genotyping found a protective effect for the Pro allele, compared with studies using somatic DNA (P(interaction) = 0.03). Results were also inconsistent between different genotyping methods (P(interaction) = 0.03). We did not identify an association between TP53 rs1042522 and colorectal cancer. Published results seem to be driven by technical artifacts rather than true biological effects. Future genetic association studies should use more rigorous genotyping methods and avoid the use of tumor tissue as a source of DNA to prevent genotype misclassification due to loss of heterozygosity.

Transcriptional effects of transfection: the potential for misinterpretation of gene expression data generated from transiently transfected cells

Transfection is used to introduce a gene of interest into a cell. To interpret the downstream results, understanding which effects are the true biological responses to the gene and which, if any, are off-target effects can be difficult. In order to discriminate true biological effects from off-target effects, we transfected a breast cancer cell line, MCF7, with a vector encoding either a reporter gene or the identical vector without the reporter gene insert. Both resulted in similar numbers of differentially expressed transcripts, suggesting that very few of the responses were directly due to the introduction of the reporter gene. We postulate that many differentially expressed transcripts are the result of the introduction of foreign DNA, as the biological processes associated with these genes are primarily associated with an immune response to a viral infection. Interestingly, different transfection reagents resulted in > 10-fold difference in the number of differentially expressed transcripts. This suggests the importance of testing multiple reagents and selecting the best transfection reagent along with the appropriate vector within the context of the experimental model system to ensure that the majority of the observed responses are biological effects of the gene of interest and not based on a particular transfection process used.

Does nitrogen concentration affect relative uptake rates of nitrate, ammonium, and glycine?

AbstractPlant roots are exposed to a variety of nitrogen forms (e.g., nitrate, ammonium, amino acids) and take up these forms at different rates. Many studies have investigated whether plants prefer nitrate, ammonium, or amino acids; but studies may not be comparable because they used substrate concentrations between 100 and 2000 μmol L–1. This study tests the hypothesis that substrate concentrations from 10 to 1750 μmol L–1 affect plant preference for N forms. Nitrogen uptake by the herb Ocimum basilicum and the evergreen tree Eucalyptus regnans was examined by placing roots of intact seedlings in equimolar mixtures of nitrate, ammonium, and glycine in which one of the N forms was 15N‐labelled (and 13C‐labelled in the case of glycine). In both species, preference for N forms was affected by substrate concentration. At 10 μmol L–1 (O. basilicum) or 10 and 50 μmol L–1 (E. regnans), rates of N uptake did not differ among N forms. At substrate concentrations of 50 μmol L–1 and greater O. basilicum took up ammonium the fastest, glycine the slowest, and nitrate at an intermediate rate. At substrate concentrations from 100 to 1750 μmol L–1, E. regnans took up ammonium the fastest with glycine and nitrate taken up at slower rates. The absence of significant differences at lower concentrations was a true biological effect rather than a function of larger relative errors. This study demonstrates that substrate concentration has a large effect on plant preference for N forms, and sounds a warning for studies of N nutrition that do not consider the concentration‐dependence of plant preference for N forms.