Articles - Research

Sex Differences in Chondrocyte Maturation in the Mandibular Condyle from a Decreased Occlusal Loading Model

J. Chen et al. — Fri, 03 May 2013 14:17:08 PDT

Temporomandibular joint disorders (TMDs) predominantly afflict women of childbearing age. Defects in mechanical loading-induced temporomandibular joint (TMJ) remodeling are believed to be a major etiological factor in the development of TMD. The goal of this study was to determine if there are sex differences in CD-1 and C57BL/6 mice exposed to a decreased occlusal loading TMJ remodeling model. Male and female CD-1 and C57BL/6 mice, 21 days old, were each divided into two groups. They were fed either a normal pellet diet (normal loading) or a soft diet and had their incisors trimmed out of occlusion (decreased occlusal loading) for 4 weeks. The mandibular condylar cartilage was evaluated by histology, and the subchondral bone was evaluated by micro-CT analysis. Gene expression from both was evaluated by real-time PCR analysis. In both strains and sexes of mice, decreased occlusal loading caused similar effects in the subchondral bone, decreases in bone volume and total volume compared with their normal loading controls. However, in both strains, decreased occlusal loading caused a significant decrease in the expression of collagen type II (Col2) and Sox9 only in female mice, but not in male mice, compared with their normal loading controls. Decreased occlusal loading causes decreased bone volume in both sexes and a decrease in early chondrocyte maturation exclusively in female mice.

High Sensitivity Carbon Nanotube Based Electrochemiluminescence Sensor Array

James F. Rusling — Fri, 03 May 2013 13:33:19 PDT

Ink jet printed carbon nanotube forest arrays capable of detecting picomolar concentrations of immunoglobulin G (IgG) using electrochemiluminescence (ECL) are described. Patterned arrays of vertically aligned single walled carbon nanotube (SWCNT) forests were printed on indium tin oxide (ITO) electrodes. Capture anti-IgG antibodies were then coupled through peptide bond formation to acidic functional groups on the vertical nanotubes. IgG immunoassays were performed using silica nano particles (Si NP) functionalized with the ECL luminophore [Ru(bpy)₂ PICH₂]²⁺], and IgG labelled G1.5 acid terminated PAMAM dendrimers. PAMAM is poly(amido amine), bpy is 2,2′-bipyridyl and PICH₂ is (2-(4-carboxyphenyl)imidazo[4,5-f][1,10]phenanthroline). The carboxyl terminal of [Ru(bpy)₂ PICH₂]²⁺ (fluorescence lifetime ≈682 ± 5 ns) dye was covalently coupled to amine groups on the 800 nm diameter silica spheres in order to produce significant ECL enhancement in the presence of sodium oxalate as co-reactant in PBS at pH 7.2). Significantly, this SWCNT-based sensor array shows a wide linear dynamic range for IgG coated spheres (10⁶ to 10¹² spheres) corresponding to IgG concentrations between 20 pM and 300 nM. A detection limit of 1.1 ± 0.1 pM IgG is obtained under optimal conditions.

Expansion of the Eukaryotic Proteome by Alternative Splicing

Brenton R. Graveley — Fri, 03 May 2013 12:11:41 PDT

The collection of components required to carry out the intricate processes involved in generating and maintaining a living, breathing and, sometimes, thinking organism is staggeringly complex. Where do all of the parts come from? Early estimates stated that about 100,000 genes would be required to make up a mammal; however, the actual number is less than one-quarter of that, barely four times the number of genes in budding yeast. It is now clear that the ‘missing’ information is in large part provided by alternative splicing, the process by which multiple different functional messenger RNAs, and therefore proteins, can be synthesized from a single gene.

Orthogonal Activation of the Reengineered A3 Adenosine Receptor (Neoceptor) Using Tailored Nucleoside Agonists

Tatiana Sonin et al. — Fri, 03 May 2013 12:02:48 PDT

An alternative approach to overcome the inherent lack of specificity of conventional agonist therapy can be the reengineering of the GPCRs and their agonists. A reengineered receptor (neoceptor) could be selectively activated by a modified agonist, but not by the endogenous agonist. Assisted by rhodopsin-based molecular modeling, we pinpointed mutations of the A₃ adenosine receptor (AR) for selective affinity enhancement following complementary modifications of adenosine. Ribose modifications examined included, at 3′: amino, aminomethyl, azido, guanidino, ureido; and at 5′: uronamido, azidodeoxy. N⁶-variations included: 3-iodobenzyl, 5-chloro-2-methyloxybenzyl, and methyl. An N⁶-3-iodobenzyl-3′-ureido adenosine derivative 10 activated phospholipase C in COS-7 cells (EC₅₀=0.18 μM) or phospholipase D in chick primary cardiomyocytes mediated by a mutant (H272E), but not the wild-type, A₃AR. The affinity enhancements for 10 and the corresponding 3′-acetamidomethyl analogue 6 were >100-fold and >20-fold, respectively. 10 concentration-dependently protected cardiomyocytes transfected with the neoceptor against hypoxia. Unlike 10, adenosine activated the wild-type A₃AR (EC₅₀ of 1.0 μM), but had no effect on the H272E mutant A₃AR (100 μM). Compound 10 was inactive at human A₁, A_2A, and A_2BARs. The orthogonal pair comprising an engineered receptor and a modified agonist should be useful for elucidating signaling pathways and could be therapeutically applied to diseases following organ-targeted delivery of the neoceptor gene.

Biochemical Applications of Ultrathin Films of Enzymes, Polyions and DNA

James F. Rusling et al. — Fri, 03 May 2013 11:55:42 PDT

This feature article summarizes recent applications of ultrathin films of enzymes and DNA assembled layer-by-layer (LbL). Using examples mainly from our own research, we focus on systems developed for biocatalysis and biosensors for toxicity screening. Enzyme–poly(l-lysine) (PLL) films, especially when stabilized by crosslinking, can be used for biocatalysis at unprecedented high temperatures or in acidic or basic solutions on electrodes or sub-micron sized beads. Such films have bright prospects for chiral synthesis and biofuel cells. Excellent bioactivity and retention of enzyme structure in these films facilitates their use in detailed kinetic studies. Biosensors and arrays employing DNA–enzyme films show great promise in predicting genotoxicity of new drug and chemical product candidates. These devices combine metabolic biocatalysis, reactive metabolite–DNA reactions, and DNA damage detection. Catalytic voltammetry or electrochemiluminescence (ECL) can be used for high throughput arrays utilizing multiple LbL “spots” of DNA, enzyme and metallopolymer. DNA–enzyme films can also be used to produce nucleobase adduct toxicity biomarkers for detection by LC-MS. These approaches provide valuable high throughput tools for drug and chemical product development and toxicity prediction.

Microsome Biocolloids for Rapid Drug Metabolism and Inhibition Assessment by LC-MS

Besnik Bajrami et al. — Fri, 03 May 2013 11:40:04 PDT

Rat liver microsomes attached to nanoparticles were used for LC-MS studies of CYP3A and 2E1 enzymes in metabolism of N-nitroso compounds. Using these biocolloids, turnover rates were measured within 2 min. Inhibitor IC₅₀ values for ketoconazole (KET) and 4-methylpyrazole (4-MEP) were estimated.

Defining the Developmental Parameters of Temper Loss in Early Childhood: Implications for Developmental Psychopathology

Heide Hullsiek et al. — Mon, 29 Apr 2013 04:08:56 PDT

Abstract

Background

Temper modulation problems are both a hallmark of early childhood and a common mental health concern. Thus, characterizing specific behavioral manifestations of temper loss along a dimension from normative misbehaviors to clinically significant problems is an important step toward identifying clinical thresholds.

Methods

Parent-reported patterns of temper loss were delineated in a diverse community sample of preschoolers (n = 1,490). A developmentally sensitive questionnaire, the Multidimensional Assessment of Preschool Disruptive Behavior (MAP-DB), was used to assess temper loss in terms of tantrum features and anger regulation. Specific aims were: (a) document the normative distribution of temper loss in preschoolers from normative misbehaviors to clinically concerning temper loss behaviors, and test for sociodemographic differences; (b) use Item Response Theory (IRT) to model a Temper Loss dimension; and (c) examine associations of temper loss and concurrent emotional and behavioral problems.

Results

Across sociodemographic subgroups, a unidimensional Temper Loss model fit the data well. Nearly all (83.7%) preschoolers had tantrums sometimes but only 8.6% had daily tantrums. Normative misbehaviors occurred more frequently than clinically concerning temper loss behaviors. Milder behaviors tended to reflect frustration in expectable contexts, whereas clinically concerning problem indicators were unpredictable, prolonged, and/or destructive. In multivariate models, Temper Loss was associated with emotional and behavioral problems.

Conclusions

Parent reports on a developmentally informed questionnaire, administered to a large and diverse sample, distinguished normative and problematic manifestations of preschool temper loss. A developmental, dimensional approach shows promise for elucidating the boundaries between normative early childhood temper loss and emergent psychopathology.

The Impact of Contingency Management on Quality of Life Among Cocaine Abusers With and Without Alcohol Dependence

Nancy M. Petry et al. — Thu, 11 Apr 2013 06:44:57 PDT

The use of quality of life measures (QOL) in substance abuse treatment research is important because it may lead to a broader understanding of patients’ health status and effects of interventions. Despite the high rates of comorbid cocaine and alcohol use disorders, little is known about the QOL of this population, and even less about the impact of an efficacious behavioral treatment, contingency management (CM), on QOL. In this study, data from three clinical trials were retrospectively analyzed to examine QOL in outpatient cocaine abusers with and without alcohol dependence (AD) and the impact of CM on QOL over time as a function of AD status. Patients were randomized to standard care (n = 115) or standard care plus CM (n = 278) for 12 weeks. QOL was assessed at baseline and Months 1, 3, 6, and 9. At treatment initiation, AD patients had lower QOL total scores and they scored lower on several subscale scores than those without AD. CM treatment was associated with improvement in QOL regardless of AD status. These data suggest that CM produces benefits that go beyond substance abuse outcomes, and they support the use of QOL indices to capture information related to treatment outcomes.

Under-treatment of Depression in Older Persons

Lisa C. Barry — Tue, 09 Apr 2013 14:30:34 PDT

Abstract

Background

Due to the cross-sectional design of most existing studies, longitudinal characterization of treatment for depression in older persons is largely unknown.

Method

754 men and women (aged 70+ years) underwent monthly assessments of mental health professional use and 18-month assessments of antidepressant medication use and depressive symptoms over 9 years. Scores of ≥20 on the Center for Epidemiological Studies-Depression (CES-D) scale denoted depression. We evaluated trends in depression treatment over time in the entire sample and among the depressed participants. Using generalized linear models, we determined characteristics associated with receiving treatment for depression in these groups and among those with persistent depression.

Results

During the 9-year follow-up period (1998–2007), 339 (45.0%) of the participants reported depression treatment. Over time, antidepressant use alone decreased (p trend <0.001) while treatment with both antidepressants and a mental health professional increased (p trend = 0.002). Of the 286 (27.9%) depressed participants, between 43% and 69% did not receive depression treatment during any 18-month interval. 30.5% of the 121 participants with persistent depression did not receive treatment during the study period. Increasing number of years of education, decreasing cognitive status score, and being physically frail were associated with a higher likelihood of receiving treatment in all models.

Limitations

Pre-baseline depression, pre-baseline treatment, and indication for treatment were unavailable.

Conclusions

Our findings indicate that the profile of treatment for depression in older persons has changed over time, that depressed older persons, including those with persistent depression, are under-treated, and that patient characteristics influence receipt of treatment.

Bevacizumab and Breast Cancer: What Does the Future Hold?

Christina E. Stevenson — Tue, 09 Apr 2013 14:21:14 PDT

Breast cancer is a major health concern for many women, but despite the current standard therapies, many women still die of metastatic disease. Angiogenesis has been evaluated as a possible target for therapy and bevacizumab (Avastin^®, Genentech/Roche, CA, USA), a monoclonal antibody against VEGF-A, has been developed to target this. Current clinical trials utilizing bevacizumab have shown an increase in progression-free survival, but this has not translated to an increase in overall survival in breast cancer patients. In this article, we summarize the currently published trials utilizing bevacizumab in the treatment of breast cancer and describe various methods of measuring angiogenesis in vitro and in vivo. We also describe the related process of lymphangiogenesis, as this may contribute to the mechanism of cancer progression and may be a potential target for therapy in the future. Understanding these processes may help us develop new treatments for breast cancer.

Essential Features and Rational Design of CRISPR RNAs That Function With the Cas RAMP Module Complex to Cleave RNAs

Sara Olson et al. — Tue, 09 Apr 2013 14:12:39 PDT

Small RNAs target invaders for silencing in the CRISPR-Cas pathways that protect bacteria and archaea from viruses and plasmids. The CRISPR RNAs (crRNAs) contain sequence elements acquired from invaders that guide CRISPR-associated (Cas) proteins back to the complementary invading DNA or RNA. Here, we have analyzed essential features of the crRNAs associated with the Cas RAMP module (Cmr) effector complex, which cleaves targeted RNAs. We show that Cmr crRNAs contain an 8-nucleotide 5’ sequence tag (also found on crRNAs associated with other CRISPR-Cas pathways) that is critical for crRNA function and can be used to engineer crRNAs that direct cleavage of novel targets. We also present data that indicates that the Cmr complex cleaves an endogenous complementary RNA in Pyrococcus furiosus, providing direct in vivo evidence of RNA targeting by the CRISPR-Cas system. Our findings indicate that the CRISPR RNA-Cmr protein pathway may be exploited to cleave RNAs of interest.

Virtual Cell: Computational Tools for Modeling in Cell Biology

James C. Schaff et al. — Tue, 09 Apr 2013 13:58:50 PDT

The Virtual Cell (VCell) is a general computational framework for modeling physico-chemical and electrophysiological processes in living cells. Developed by the National Resource for Cell Analysis and Modeling at the University of Connecticut Health Center, it provides automated tools for simulating a wide range of cellular phenomena in space and time, both deterministically and stochastically. These computational tools allow one to couple electrophysiology and reaction kinetics with transport mechanisms, such as diffusion and directed transport, and map them onto spatial domains of various shapes, including irregular three-dimensional geometries derived from experimental images. In this article, we review new robust computational tools recently deployed in VCell for treating spatially resolved models.

Identifying Provider Beliefs Related to Contingency Management Adoption Using the Contingency Management Beliefs Questionnaire

Carla J. Rash et al. — Tue, 09 Apr 2013 13:47:52 PDT

Abstract

Background

Contingency management (CM) is a widely recognized empirically-supported addiction treatment; however, dissemination and adoption of CM into routine clinical practice has been slow. Assessment of beliefs about CM may highlight key barriers and facilitators of adoption and inform dissemination efforts. In the present study, we developed a 35-item questionnaire (Contingency Management Beliefs Questionnaire; CMBQ) assessing CM beliefs and examined the relation of these beliefs to clinician characteristics and clinical practices.

Methods

The web-based study was completed by 617 substance abuse treatment providers. We examined the factor structure using exploratory factor analysis (EFA) in a randomly selected half-sample (n =318) and evaluated the generalizability of the solution using confirmatory factor analysis (CFA) in the second half-sample (n = 299).

Results

EFA results suggested a 3-factor solution with 32 items retained; factors represented general barriers, training-related barriers, and pro-CM items. CFA results supported the solution, and reliability was good within each half-sample (α = .88 and 0.90). Therapeutic approach, years experience in addictions field, perception of CM’s research support, prior CM training, and CM adoption interest were significantly associated with the factors.

Conclusions

Overall, participants viewed CM favorably yet endorsed barriers, indicating a need for more extensive and targeted response to the most common misperceptions in dissemination efforts.

The Role of IFNAR and MyD88 Signaling in Induction of IL-15 Expression In Vivo

Leo Lefrancois et al. — Tue, 09 Apr 2013 13:42:16 PDT

Interleukin-15 (IL-15) plays a multifaceted role in immune homeostasis, but the unreliability of IL-15 detection has stymied exploration of IL-15 regulation in vivo. To visualize IL-15 expression, we created a transgenic mouse expressing emerald-GFP (EmGFP) under IL-15 promoter control. EmGFP/IL-15 was prevalent in innate cells including dendritic cells (DCs), macrophages, and monocytes. However, DC subsets expressed varying levels of EmGFP/IL-15 with CD8⁺ DCs constitutively expressing EmGFP/IL-15 and CD8⁻ DCs expressing low EmGFP/IL-15 levels. Virus infection resulted in IL-15 upregulation in both subsets. By crossing the transgenic mice to mice deficient in specific elements of innate signaling, we found a cell-intrinsic dependency of DCs and Ly6C⁺ monocytes on IFNAR expression for EmGFP/IL-15 upregulation after VSV infection. In contrast, myeloid cells did not require the expression of MyD88 to upregulate EmGFP/IL-15 expression. These findings provide evidence of previously unappreciated regulation of IL-15 expression in myeloid lineages during homeostasis and following infection.

YKL-40 Genetic Polymorphisms and the Risk of Liver Disease Progression in Patients with Advanced Fibrosis Due to Chronic Hepatitis C

Herbert L. Bonkovsky — Tue, 09 Apr 2013 13:12:46 PDT

Abstract

Background/Aims

The aim of this study was to explore the association of a functional YKL-40 promoter polymorphism (rs4950928) with baseline disease stage, response to antiviral therapy, and risk of liver disease progression in a group of patients with chronic hepatitis C (CHC).

Methods

YKL-40 promoter polymorphisms were determined in 456 HALT-C Trial patients with bridging fibrosis or cirrhosis entering a prerandomization lead-in peginterferon/ribavirin 24-week treatment phase and in 462 patients followed for a mean of 3.8 years after randomization to maintenance peginterferon or observation.

RESULTS

Mean patient age was 49.5 years, 70.4% were male, and 71.2% were Caucasian. The 17% frequency of the YKL-40 minor allele (T) was similar to that reported in the general population. YKL-40 genotype was associated significantly with baseline serum YKL-40 levels but was not associated with the likelihood of a virological response following 24 to 48 weeks of peginterferon/ribavirin therapy. Serum YKL-40 levels remained significantly lower during follow-up in the randomized TT homozygotes compared to CT heterozygotes and CC homozygotes (p <0.001). Despite this association, YKL-40 genotype was not associated with the risk of clinical or histologic liver disease progression.

CONCLUSIONS

A reduced frequency of the protective YKL-40 promoter polymorphism was not observed in the HALT-C Trial patient population. The absence of an association between YKL-40 promoter polymorphisms and baseline liver-disease severity as well as with the risk of liver-disease progression over time suggests that this polymorphism is not associated with disease progression in CHC patients with established fibrosis.

Pharmacological and Therapeutic Effects of A3 Adenosine Receptor (A3AR) Agonists

Bruce T. Liang — Tue, 09 Apr 2013 12:58:34 PDT

The G_i-coupled A₃ adenosine receptor (A₃AR) mediates anti-inflammatory, anticancer and anti-ischemic protective effects. The receptor is overexpressed in inflammatory and cancer cells, while low expression is found in normal cells, rendering the A₃AR as a potential therapeutic target. Highly selective A₃AR agonists have been synthesized and molecular recognition in the binding site has been characterized. The present review summarizes preclinical and clinical human studies demonstrating that A₃AR agonists induce specific anti-inflammatory and anticancer effects via a molecular mechanism that entails modulation of the Wnt and the NF-κB signal transduction pathways. Currently, A₃AR agonists are being developed for the treatment of inflammatory diseases including rheumatoid arthritis and psoriasis; ophthalmic diseases such as dry eye syndrome and glaucoma; liver diseases such as hepatocellular carcinoma and hepatitis.

Contingency Management Delivered by Community Therapists in Outpatient Settings

Nancy M. Petry et al. — Tue, 09 Apr 2013 12:45:33 PDT

Abstract

Background

Few community-based clinicians have been trained to deliver contingency management (CM) treatments, and little data exist regarding the efficacy of CM when administered by clinicians.

Method

Fifteen clinicians from four intensive outpatient treatment programs received training in CM. Following a didactics seminar and a period in which clinicians delivered CM to pilot patients while receiving weekly supervision, clinicians treated 43 patients randomized to standard care or CM, without supervision. In both treatment conditions, urine and breath samples were collected up to twice weekly for 12 weeks, and CM patients earned the opportunity to win prizes ranging in value from $1 to $100 for submitting drug-free samples. Primary treatment outcomes were sessions attended, unexcused absences, longest continuous period of abstinence, and proportion of negative samples submitted.

Results

All therapists completed the training and supervision phase, and 10 treated randomized patients. Patients randomized to CM achieved significantly greater durations of abstinence than patients randomized to standard care (5.0 ± 3.8 versus 2.6 ± 3.7 weeks) and had fewer unexcused absences (4.3 ± 1.2 versus 8.1 ± 5.4), but proportion of negative samples submitted and attendance did not differ significantly between groups. Therapist adherence and competence in CM delivery decreased when supervision was no longer provided, and competence in CM delivery was associated with duration of abstinence achieved and attendance.

Conclusions

Community-based clinicians can effectively administer CM, and outcomes relate to competence in CM delivery. These data call for further training and supervision of community clinicians in this evidence-based treatment.

Electrophysiological Responses to Threat in Youth with and without Posttraumatic Stress Disorder

Damion J. Grasso — Tue, 09 Apr 2013 12:36:16 PDT

The current study was designed to examine event-related brain potentials and autonomic responses to pictures indicating threat, relative to non-threat, and acoustic startle reflexes in traumatized youth diagnosed with PTSD, relative to non-exposed children, before and after receiving psychotherapy. Children in the control group were individually yoked and demographically matched to the PTSD group. Both groups displayed enhanced late positive potentials and more prolonged heart rate deceleration to pictures indicating threat, relative to non-threat, and larger skin conductance responses to pictures indicating threat, relative to non-threat, at time one. At time two, controls appeared to habituate, as reflected by an overall attenuated skin conductance response, whereas the PTSD group showed little change. Across time points the PTSD group exhibited greater acoustic startle reflexes than the control group. Psychotherapy and symptom reduction was not associated with electrophysiology. Drawing from the adult literature, this study was an attempt to address the scarcity of research examining electrophysiological irregularities in childhood PTSD. The overall results suggest that children and adolescents allocate more attention to threat-related stimuli regardless of PTSD status, and exaggerated startle and a possible failure to habituate skin conductance responses to threat-related stimuli in youth with versus without PTSD.

Risk Factors for Alcohol Problems in Victims of Partner Violence

Howard A. Tennen — Sat, 06 Apr 2013 09:12:21 PDT

Despite the high prevalence of alcohol problems and disorders among women who experience intimate partner violence (IPV), factors related to current alcohol use are understudied. We examined current risk factors for alcohol problems among 143 substance-using, IPV-exposed women recruited from an urban community from 2007-2010. PTSD symptom severity was associated with alcohol-related problems and a positive alcohol screen; physical IPV severity was related to alcohol dependence. Posthoc analyses revealed that PTSD symptom severity mediated relationships between physical IPV severity and hazardous, harmful, and dependent drinking. Focusing on managing PTSD symptoms and physical IPV in community-based interventions may halt the progression from alcohol use to dependence.

Immunocytochemistry at the Electron-Microscopic Level

David S. Papermaster — Fri, 05 Apr 2013 09:23:38 PDT

Since the early slxtles, a number of reports have been published concerning immunocytochemical techniques for the detection of intracellular constituents at the electron microscope level. The· initial studies were made possible by the introduction by Singer of ferritin-tagged antibodies which could be visualized in the electron microscope. From all the procedures described later, immunocytochemical methods at the electron microscope can be classified into two main categories: 1) preembedding staining and 2) postembedding staining techniques.