Glial activation in pain and emotional processing regions in the nitroglycerin mouse model of chronic migraine.

OBJECTIVE: Our aim was to survey astrocyte and microglial activation across four brain regions in a mouse model of chronic migraine. BACKGROUND: Chronic migraine is a leading cause of disability, with higher rates in females. The role of central nervous system neurons and glia in migraine pathophysiology is not fully elucidated. Preclinical studies have shown abnormal glial activation in the trigeminal nucleus caudalis of male rodents. No current reports have investigated glial activation in both sexes in other important brain regions involved with the nociceptive and emotional processing of pain. METHODS: The mouse nitroglycerin model of migraine was used, and nitroglycerin (10 mg/kg) or vehicle was administered every other day for 9 days. Prior to injections on days 1, 5, and 9, cephalic allodynia was determined by periorbital von Frey hair testing. Immunofluorescent staining of astrocyte marker, glial fibrillary protein (GFAP), and microglial marker, ionized calcium binding adaptor molecule 1 (Iba1), in male and female trigeminal nucleus caudalis, periaqueductal gray, somatosensory cortex, and nucleus accumbens was completed. RESULTS: Behavioral testing demonstrated increased cephalic allodynia in nitroglycerin- versus vehicle-treated mice. An increase in the percent area covered by GFAP+ cells in the trigeminal nucleus caudalis and nucleus accumbens, but not the periaqueductal gray or somatosensory cortex, was observed in response to nitroglycerin. No significant differences were observed for Iba1 staining across brain regions. We did not detect significant sex differences in GFAP or Iba1 quantification. CONCLUSIONS: Immunohistochemical analysis suggests that, at the time point tested, immunoreactivity of GFAP+ astrocytes, but not Iba1+ microglia, changes in response to chronic migraine-associated pain. Additionally, there do not appear to be significant differences between males and females in GFAP+ or Iba1+ cells across the four brain regions analyzed.

Financial Stressors for Parents of Children and Emerging Adults With Congenital Heart Disease: A Qualitative Study.

INTRODUCTION: Congenital heart disease (CHD) is the most prevalent congenital disability globally. This study aimed to describe parents' perspectives on financial stressors related to having a child with CHD using a descriptive qualitative approach. METHOD: Qualitative data were obtained from parents of children with CHD in a cross-sectional web-based survey study. Iterative data analysis was used to develop essential themes that enabled a rich description of 147 parents' perspectives. RESULTS: Parents identified five financial stressors: perpetual worries about health insurance, facing the dilemma of "making too much money," struggling to balance work, worrying over having an emerging adult with CHD, and constant constraints because of financial needs. DISCUSSION: As experts in pediatric care, pediatric advanced practice providers need to work with policymakers to provide further financial assistance and sufficient insurance coverage for families that struggle to balance finances for the whole family and children with CHD.

Assessment of endoscope reprocessing at World Gastroenterology Organisation training centers using adenosine triphosphate testing.

Background and study aims Adequacy of endoscope disinfection in resource-limited settings is unknown. Adenosine triphosphate (ATP) testing is useful for evaluation of endoscope reprocessing, and ATP <200 relative light units (RLUs) after manual endoscope cleaning has been associated with adequacy of endoscope disinfection. Methods Consecutive endoscopes undergoing reprocessing at five World Gastroenterology Organisation (WGO) training centers underwent ATP testing before and after an on-site educational intervention designed to optimize reprocessing practices. Results A total of 343 reprocessing cycles of 65 endoscopes were studied. Mean endoscope age was 5.3 years (range 1-13 years). Educational interventions, based on direct observation of endoscope reprocessing practices at each site, included refinements in pre-cleaning, manual cleaning, high-level disinfection, and endoscope drying and storage. The percentage of reprocessing cycles with post-manual cleaning ATP ≧200 decreased from 21.4% prior to educational intervention to 14.8% post-intervention ( P =0.11). In multivariable logistic modelling, gastroscopes were significantly less likely (odds ratio [OR] 0.04, 95% confidence interval [CI] 0.01-0.19; P <0.001) than colonoscopes to achieve post-manual cleaning ATP < 200. No other factor (educational intervention, study site, endoscope age) was significantly associated with improved outcomes. Endoscope ID was not significantly associated with ATP values, and sites that performed manual versus automated HLD did not have significantly different likelihood of post-manual cleaning ATP <200 (OR 1.18, 95% CI 0.56-2.50; P =0.67). Conclusions In resource-limited settings, approximately 20% of endoscope reprocessing cycles may result in inadequate disinfection. This was not significantly improved by a comprehensive educational intervention. Alternative approaches to endoscope reprocessing are needed.

Cognitive, Disability, and Treatment Outcome Implications of Symptom-Based Phenotyping in Late-Life Depression.

OBJECTIVE: Late-life depression is associated with substantial heterogeneity in clinical presentation, disability, and response to antidepressant treatment. We examined whether self-report of severity of common symptoms, including anhedonia, apathy, rumination, worry, insomnia, and fatigue were associated with differences in presentation and response to treatment. We also examined whether these symptoms improved during treatment with escitalopram. DESIGN: Eighty-nine older adults completed baseline assessments, neuropsychological testing and providing self-reported symptom and disability scales. They then entered an 8-week, placebo-controlled randomized trial of escitalopram, and self-report scales were repeated at the trial's end. Raw symptom scale scores were combined into three standardized symptom phenotypes and models examined how symptom phenotype severity was associated with baseline measures and depression improvement over the trial. RESULTS: While rumination/worry appeared independent, severity of apathy/anhedonia and fatigue/insomnia were associated with one another and with greater self-reported disability. Greater fatigue/insomnia was also associated with slower processing speed, while rumination/worry was associated with poorer episodic memory. No symptom phenotype severity score predicted a poorer overall response to escitalopram. In secondary analyses, escitalopram did not improve most phenotypic symptoms more than placebo, aside for greater reductions in worry and total rumination severity. CONCLUSION: Deeper symptom phenotype characterization may highlight differences in the clinical presentation of late-life depression. However, when compared to placebo, escitalopram did not improve many of the symptoms assessed. Further work is needed to determine whether symptom phenotypes inform longer-term course of illness, and which treatments may best benefit specific symptoms.

Person with Heart Failure and Care Partner Dyads: Current Knowledge, Challenges, and Future Directions: State-of-the-Art Review.

Over the past decade, there has been substantial growth in heart failure (HF) research that focuses on persons with HF and their care partners (family members or other close friends that provide unpaid support) as an interdependent team, or care dyad. In this state-of-the-art review, we use a dyadic lens to identify and summarize current research on HF care dyads, from qualitative studies, to nonexperimental quantitative studies, to randomized controlled trials. Although much work has been done, this literature is younger and less well-developed than care dyad literatures from other conditions (eg, cancer, Alzheimer's disease). We discuss the substantial challenges and limitations in this body of work, with an eye toward addressing common issues that impact rigor. We also look toward future directions, and discuss the promise dyadic research holds for improving patient, care partner, and relationship health.

Migraine and peripheral pain models show differential alterations in neuronal complexity.

OBJECTIVE: Our laboratory has recently shown that there is a decrease in neuronal complexity in head pain processing regions in mouse models of chronic migraine-associated pain and aura. Importantly, restoration of this neuronal complexity corresponds with anti-migraine effects of known and experimental pharmacotherapies. The objective of the current study was to expand this work and examine other brain regions involved with pain or emotional processing. We also investigated the generalizability of our findings by analyzing neuronal cytoarchitectural changes in a model of complex regional pain syndrome (CRPS), a peripheral pain disorder. METHODS: We used the nitroglycerin (NTG) model of chronic migraine-associated pain in which mice receive 10 mg/kg NTG every other day for 9 days. Cortical spreading depression (CSD), a physiological corelate of migraine aura, was evoked in anesthetized mice using KCl. CRPS was induced by tibial fracture followed by casting. Neuronal cytoarchitecture was visualized with Golgi stain and analyzed with Simple Neurite Tracer. RESULTS: In the NTG model, we previously showed decreased neuronal complexity in the trigeminal nucleus caudalis (TNC) and periaqueductal gray (PAG). In contrast, we found increased neuronal complexity in the thalamus and no change in the amygdala or caudate putamen in this study. Following CSD, we observed decreased neuronal complexity in the PAG, in line with decreases in the somatosensory cortex and TNC reported with this model previously. In the CRPS model there was decreased neuronal complexity in the hippocampus, as reported by others; increased complexity in the PAG; and no change within the somatosensory cortex. CONCLUSIONS: Collectively these results demonstrate that alterations in neuronal complexity are a feature of both chronic migraine and chronic CRPS. However, each type of pain presents a unique cytoarchitectural signature, which may provide insight on how these pain states differentially transition from acute to chronic conditions.

The Effects of Exercise-Based Interventions on Fluid Overload Symptoms in Patients with Heart Failure: A Systematic Review and Meta-Analysis.

Patients with heart failure are subjected to a substantial burden related to fluid overload symptoms. Exercise can help the lymphatic system function more effectively to prevent fluid build-up in tissues and interstitium, thus potentially mitigating the symptoms due to fluid overload. The objective of this systematic review was to examine the effects of exercise-based interventions on fluid overload symptoms among patients with heart failure. MEDLINE, Embase, Cochrane Library, and CINAHL databases were systematically searched for relevant studies published from inception to August 2021. We included randomized controlled trials that compared exercise-based interventions of different modalities and usual medical care for adult patients with heart failure and reported the effects of interventions on any symptoms related to fluid overload. A random-effects meta-analysis was used to estimate the effectiveness, and a subgroup analysis and univariate meta-regression analysis were used to explore heterogeneity. Seventeen studies covering 1086 participants were included. We found robust evidence indicating the positive effect of exercises in dyspnea relief (SMD = -0.48; 95%CI [-0.76, -0.19]; p = 0.001); the intervention length also influenced the treatment effect (ß = 0.033; 95%CI [0.003, 0.063]; p = 0.04). Initial evidence from existing limited research showed that exercise-based intervention had positive effect to alleviate edema, yet more studies are needed to verify the effect. In contrast, the exercise-based interventions did not improve fatigue compared with usual care (SMD = -0.27; 95%CI [-0.61, 0.06]; p = 0.11). Findings regarding the effects of exercises on bodily pain, gastro-intestinal symptoms, and peripheral circulatory symptoms were inconclusive due to limited available studies. In conclusion, exercise-based interventions can be considered as an effective nonpharmacological therapy for patients with heart failure to promote lymph flow and manage fluid overload symptoms. Exercise-based interventions seem to have very limited effect on fatigue. More research should investigate the mechanism of fatigue related to heart failure. Future studies with high methodological quality and comprehensive assessment of symptoms and objective measure of fluid overload are warranted.

The role of generalized linear models in handling cost and count data.

Scientists from nursing and allied health disciplines frequently examine data with complex distributions. Key examples include data on cost that typically are skewed, and count data like the number of hospitalizations that regularly have greater variation than expected and a majority of observations at zero. Common approaches to handling complex data involve transformations that can interfere with interpretation, or force-fitting of data into linear or logistic regression. In this article, worked examples of generalized linear models, which allow for flexibility in the distribution of data, involving cost and count outcomes, are presented to help expose researchers to their nuances.

Neuronal complexity is attenuated in preclinical models of migraine and restored by HDAC6 inhibition.

Migraine is the sixth most prevalent disease worldwide but the mechanisms that underlie migraine chronicity are poorly understood. Cytoskeletal flexibility is fundamental to neuronal-plasticity and is dependent on dynamic microtubules. Histone-deacetylase-6 (HDAC6) decreases microtubule dynamics by deacetylating its primary substrate, α-tubulin. We use validated mouse models of migraine to show that HDAC6-inhibition is a promising migraine treatment and reveal an undiscovered cytoarchitectural basis for migraine chronicity. The human migraine trigger, nitroglycerin, produced chronic migraine-associated pain and decreased neurite growth in headache-processing regions, which were reversed by HDAC6 inhibition. Cortical spreading depression (CSD), a physiological correlate of migraine aura, also decreased cortical neurite growth, while HDAC6-inhibitor restored neuronal complexity and decreased CSD. Importantly, a calcitonin gene-related peptide receptor antagonist also restored blunted neuronal complexity induced by nitroglycerin. Our results demonstrate that disruptions in neuronal cytoarchitecture are a feature of chronic migraine, and effective migraine therapies might include agents that restore microtubule/neuronal plasticity.

Migraines are a common brain disorder that affects 14% of the world's population. For many people the main symptom of a migraine is a painful headache, often on one side of the head. Other symptoms include increased sensitivity to light or sound, disturbed vision, and feeling sick. These sensory disturbances are called aura and they often occur before the headache begins. One particularly debilitating subset of migraines are chronic migraines, in which patients experience more than 15 headache days per month. Migraine therapies are often only partially effective or poorly tolerated, making it important to develop new drugs for this condition, but unfortunately, little is known about the molecular causes of migraines. To bridge this gap, Bertels et al. used two different approaches to cause migraine-like symptoms in mice. One approach consisted on giving mice nitroglycerin, which dilates blood vessels, produces hypersensitivity to touch, and causes photophobia in both humans and mice. In the second approach, mice underwent surgery and potassium chloride was applied onto the dura, a thick membrane that surrounds the brain. This produces cortical spreading depression, an event that is linked to migraine auras and involves a wave of electric changes in brain cells that slowly propagates across the brain, silencing brain electrical activity for several minutes. Using these approaches, Bertels et al. studied whether causing chronic migraine-like symptoms in mice is associated with changes in the structures of neurons, focusing on the effects of migraines on microtubules. Microtubules are cylindrical protein structures formed by the assembly of smaller protein units. In most cells, microtubules assemble and disassemble depending on what the cell needs. Neurons need stable microtubules to establish connections with other neurons. The experiments showed that provoking chronic migraines in mice led to a reduction in the numbers of connections between different neurons. Additionally, Bertels et al. found that inhibiting HDAC6 (a protein that destabilizes microtubules) reverses the structural changes in neurons caused by migraines and decreases migraine symptoms. The same effects are seen when a known migraine treatment strategy, known as CGRP receptor blockade, is applied. These results suggest that chronic migraines may involve decreased neural complexity, and that the restoration of this complexity by HDAC6 inhibitors could be a potential therapeutic strategy for migraine.

Academy of Nutrition and Dietetics: Revised 2020 Standards of Practice and Standards of Professional Performance for Registered Dietitian Nutritionists (Competent, Proficient, and Expert) in Intellectual and Developmental Disabilities.

Intellectual and developmental disabilities (IDD) encompass both intellectual disabilities (ID) and developmental disabilities (DD). In 2016, 7.37 million people in the United States and 200 million worldwide were identified with an ID or DD. Approximately 1 in 6 (17.8%) children have been identified with a DD in the United States, which is up from 16.2% in 2009-2011. Globally, 52.9 million children from birth to 5 years of age have been identified with a DD. Registered dietitian nutritionists (RDNs) have an important role in the treatment of this population, as optimizing nutrition status improves cognition and quality of life. The Behavioral Health Nutrition Dietetic Practice Group, with guidance from the Academy of Nutrition and Dietetics Quality Management Committee, has revised the Standards of Practice (SOP) and Standards of Professional Performance (SOPP) for RDNs in Intellectual and Developmental Disabilities for 3 levels of practice-competent, proficient, and expert. The SOP uses the Nutrition Care Process and clinical workflow elements for care of individuals with an ID or DD. The SOPP describes 6 domains that focus on professionalism. Indicators outlined in the SOP and SOPP depict how these standards apply to practice. The SOP and SOPP are complementary resources for RDNs caring for individuals with an ID or DD. The SOP and SOPP are intended to be used by RDNs for self-evaluation to assure competent practice and for determining potential education and training needs for advancement to a higher practice level in a variety of settings.

Axonal blockage with microscopic magnetic stimulation.

Numerous neurological dysfunctions are characterized by undesirable nerve activity. By providing reversible nerve blockage, electric stimulation with an implanted electrode holds promise in the treatment of these conditions. However, there are several limitations to its application, including poor bio-compatibility and decreased efficacy during chronic implantation. A magnetic coil of miniature size can mitigate some of these problems, by coating it with biocompatible material for chronic implantation. However, it is unknown if miniature coils could be effective in axonal blockage and, if so, what the underlying mechanisms are. Here we demonstrate that a submillimeter magnetic coil can reversibly block action potentials in the unmyelinated axons from the marine mollusk Aplysia californica. Using a multi-compartment model of the Aplysia axon, we demonstrate that the miniature coil causes a significant local depolarization in the axon, alters activation dynamics of the sodium channels, and prevents the traveling of the invading action potentials. With improved biocompatibility and capability of emitting high-frequency stimuli, micro coils provide an interesting alternative for electric blockage of axonal conductance in clinical settings.

Alteration of immune markers in a group of melancholic depressed patients and their response to electroconvulsive therapy.

BACKGROUND: Immune system dysfunction is implicated in the pathophysiology of major depression, and is hypothesized to normalize with successful treatment. We aimed to investigate immune dysfunction in melancholic depression and its response to ECT. METHODS: 55 melancholic depressed patients and 26 controls participated. 33 patients (60%) were referred for ECT. Blood samples were taken at baseline, one hour after the first ECT session, and 48h after ECT series completion. RESULTS: At baseline, melancholic depressed patients had significantly higher levels of the pro-inflammatory cytokine IL-6, and lower levels of the regulatory cytokine TGF-ß than controls. A significant surge in IL-6 levels was observed one hour after the first ECT session, but neither IL-6 nor TGF-ß levels normalized after completion of ECT series. Seventy per cent (n=23) of ECT recipients showed clinical response and 42% (n=10) reached remission. Neither IL-6 nor TGF-ß changes correlated with clinical improvement following ECT. No significant changes in IL-10, TNF-α and CRP levels were found in relation to melancholia or response to ECT. LIMITATIONS: As a naturalistic study, some potential confounders could not be eliminated or controlled, including medication use. CONCLUSIONS: Melancholic depressed patients demonstrated a peripheral increase in IL-6 and reduction in TGF-ß, which did not normalize despite clinical response to ECT. These findings may be consistent with emerging hypotheses of the role of inflammation in mediating neurotrophin expression. The implications of chronic inflammation in the melancholic depressed population for future medical health, particularly cardiovascular risk, are largely unknown and warrant further investigation.

The Application of Digital Pathology to Improve Accuracy in Glomerular Enumeration in Renal Biopsies.

BACKGROUND: In renal biopsy reporting, quantitative measurements, such as glomerular number and percentage of globally sclerotic glomeruli, is central to diagnostic accuracy and prognosis. The aim of this study is to determine the number of glomeruli and percent globally sclerotic in renal biopsies by means of registration of serial tissue sections and manual enumeration, compared to the numbers in pathology reports from routine light microscopic assessment. DESIGN: We reviewed 277 biopsies from the Nephrotic Syndrome Study Network (NEPTUNE) digital pathology repository, enumerating 9,379 glomeruli by means of whole slide imaging. Glomerular number and the percentage of globally sclerotic glomeruli are values routinely recorded in the official renal biopsy pathology report from the 25 participating centers. Two general trends in reporting were noted: total number per biopsy or average number per level/section. Both of these approaches were assessed for their accuracy in comparison to the analogous numbers of annotated glomeruli on WSI. RESULTS: The number of glomeruli annotated was consistently higher than those reported (p<0.001); this difference was proportional to the number of glomeruli. In contrast, percent globally sclerotic were similar when calculated on total glomeruli, but greater in FSGS when calculated on average number of glomeruli (p<0.01). The difference in percent globally sclerotic between annotated and those recorded in pathology reports was significant when global sclerosis is greater than 40%. CONCLUSIONS: Although glass slides were not available for direct comparison to whole slide image annotation, this study indicates that routine manual light microscopy assessment of number of glomeruli is inaccurate, and the magnitude of this error is proportional to the total number of glomeruli.

Morphometry Predicts Early GFR Change in Primary Proteinuric Glomerulopathies: A Longitudinal Cohort Study Using Generalized Estimating Equations.

OBJECTIVE: Most predictive models of kidney disease progression have not incorporated structural data. If structural variables have been used in models, they have generally been only semi-quantitative. METHODS: We examined the predictive utility of quantitative structural parameters measured on the digital images of baseline kidney biopsies from the NEPTUNE study of primary proteinuric glomerulopathies. These variables were included in longitudinal statistical models predicting the change in estimated glomerular filtration rate (eGFR) over up to 55 months of follow-up. RESULTS: The participants were fifty-six pediatric and adult subjects from the NEPTUNE longitudinal cohort study who had measurements made on their digital biopsy images; 25% were African-American, 70% were male and 39% were children; 25 had focal segmental glomerular sclerosis, 19 had minimal change disease, and 12 had membranous nephropathy. We considered four different sets of candidate predictors, each including four quantitative structural variables (for example, mean glomerular tuft area, cortical density of patent glomeruli and two of the principal components from the correlation matrix of six fractional cortical areas-interstitium, atrophic tubule, intact tubule, blood vessel, sclerotic glomerulus, and patent glomerulus) along with 13 potentially confounding demographic and clinical variables (such as race, age, diagnosis, and baseline eGFR, quantitative proteinuria and BMI). We used longitudinal linear models based on these 17 variables to predict the change in eGFR over up to 55 months. All 4 models had a leave-one-out cross-validated R2 of about 62%. CONCLUSIONS: Several combinations of quantitative structural variables were significantly and strongly associated with changes in eGFR. The structural variables were generally stronger than any of the confounding variables, other than baseline eGFR. Our findings suggest that quantitative assessment of diagnostic renal biopsies may play a role in estimating the baseline risk of succeeding loss of renal function in future clinical studies, and possibly in clinical practice.

The TRAIL receptor agonist drozitumab targets basal B triple-negative breast cancer cells that express vimentin and Axl.

Previously, we found that GST-tagged tumor necrosis factor-related apoptosis inducing ligand preferentially killed triple-negative breast cancer (TNBC) cells with a mesenchymal phenotype by activating death receptor 5 (DR5). The purpose of this study was to explore the sensitivity of breast cancer cell lines to drozitumab, a clinically tested DR5-specific agonist; identify potential biomarkers of drozitumab-sensitive breast cancer cells; and determine if those biomarkers were present in tumors from patients with TNBC. We evaluated viability, caspase activity, and sub-G1 DNA content in drozitumab-treated breast cancer cell lines and we characterized expression of potential biomarkers by immunoblot. Expression levels of vimentin and Axl were then explored in 177 TNBC samples from a publically available cDNA microarray dataset and by immunohistochemistry (IHC) in tumor tissue samples obtained from 53 African-American women with TNBC. Drozitumab-induced apoptosis in mesenchymal TNBC cell lines but not in cell lines from other breast cancer subtypes. The drozitumab-sensitive TNBC cell lines expressed the mesenchymal markers vimentin and Axl. Vimentin and Axl mRNA and protein were expressed in a subset of human TNBC tumors. By IHC, ~15 % of TNBC tumors had vimentin and Axl expression in the top quartile for both. These findings indicate that drozitumab-sensitive mesenchymal TNBC cells express vimentin and Axl, which can be identified in a subset of human TNBC tumors. Thus, vimentin and Axl may be useful to identify TNBC patients who would be most likely to benefit from a DR5 agonist.

Recovery and concordance in a secure forensic psychiatry hospital - the self rated DUNDRUM-3 programme completion and DUNDRUM-4 recovery scales.

BACKGROUND: Detention in a secure forensic psychiatric hospital may inhibit engagement and recovery. Having validated the clinician rated DUNDRUM-3 (programme completion) and DUNDRUM-4 (recovery) in a forensic hospital, we set out to draft and validate scales measuring the same programme completion and recovery items that patients could use to self-rate. Based on previous work, we hypothesised that self-rating scores might be predictors of objective progress including conditional discharge. We hypothesised also that the difference between patients' and clinicians' ratings of progress in treatment and other factors relevant to readiness for discharge (concordance) would diminish as patients neared discharge. We hypothesised also that this difference in matched scores would predict objective progress including conditional discharge. METHOD: In a prospective naturalistic observational cohort study in a forensic hospital, we examined whether scores on the self-rated DUNDRUM-3 programme completion and DUNDRUM-4 recovery scales or differences between clinician and patient ratings on the same scales (concordance) would predict moves between levels of therapeutic security and conditional discharge over the next twelve months. RESULTS: Both scales stratified along the recovery pathway of the hospital, but clinician ratings matched the level of therapeutic security more accurately than self ratings. The clinician rated scales predicted moves to less secure units and to more secure units and predicted conditional discharge but the self-rated scores did not. The difference between clinician and self-rated scores (concordance) predicted positive and negative moves and conditional discharge, but this was not always an independent predictor as shown by regression analysis. In regression analysis the DUNDRUM-3 predicted moves to less secure places though the HCR-20 C & R score dominated the model. Moves back to more secure places were predicted by lack of concordance on the DUNDRUM-4. Conditional discharge was predicted predominantly by the DUNDRUM-3. CONCLUSIONS: Patients accurately self-rate relative to other patients however their absolute ratings were consistently lower (better) than clinicians' ratings and were less accurate predictors of outcomes including conditional discharge. Quantifying concordance is a useful part of the recovery process and predicts outcomes but self-ratings are not accurate predictors.

Brightfield proximity ligation assay reveals both canonical and mixed transforming growth factor-ß/bone morphogenetic protein Smad signaling complexes in tissue sections.

Transforming growth factor-ß (TGF-ß) is an important regulator of cellular homeostasis and disease pathogenesis. Canonical TGF-ß signaling occurs through Smad2/3-Smad4 complexes; however, recent in vitro studies suggest that elevated levels of TGF-ß may activate a novel mixed Smad complex (Smad2/3-Smad1/5/9), which is required for some of the pro-oncogenic activities of TGF-ß. To determine if mixed Smad complexes are evident in vivo, we developed antibodies that can be used with a proximity ligation assay to detect either canonical or mixed Smad complexes in formalin-fixed paraffin-embedded sections. We demonstrate high expression of mixed Smad complexes in the tissues from mice genetically engineered to express high levels of TGF-ß1. Mixed Smad complexes were also prominent in 15-16 day gestation mouse embryos and in breast cancer xenografts, suggesting important roles in embryonic development and tumorigenesis. In contrast, mixed Smad complexes were expressed at extremely low levels in normal adult mouse tissue, where canonical complexes were correspondingly higher. We show that this methodology can be used in archival patient samples and tissue microarrays, and we have developed an algorithm to quantitate the brightfield read-out. These methods will allow quantitative analysis of cell type-specific Smad signaling pathways in physiological and pathological processes.

PLCE1 mRNA and protein expression and survival of patients with esophageal squamous cell carcinoma and gastric adenocarcinoma.

BACKGROUND: Germline genetic variants in PLCE1 (10q23) have demonstrated consistent associations with risk of esophageal squamous cell carcinoma (ESCC) and gastric cancer among Chinese. We evaluated PLCE1 mRNA and protein expression in paired tumor-normal tissues, and their relationship with survival. METHODS: PLCE1 mRNA was profiled using three probes in the Affymetrix GeneChip U133 for paired tumor-normal tissues of ESCC (n = 132), gastric cardia adenocarcinoma (GCA, n = 62), and gastric noncardia adenocarcinoma (GNCA, n = 72). We used immunohistochemistry to detect PLCE1 protein on slides from tissue microarrays in paired tumor-normal tissues of ESCC (n = 303), and tumors of GCA (n = 298) and GNCA (n = 124). RESULTS: Compared with normal tissues, PLCE1 mRNA expression was significantly reduced in ESCC tumors (P = 0.03, probe_205112_at), as well as in GCA and GNCA tumors (P < 0.0001, each probe). Protein expression was nonsignificantly reduced in ESCC tumors (P = 0.51). Increased tumor-normal mRNA fold change (probe_205112_at) was associated with longer survival in ESCC (9.6 months for highest vs. lowest quartile; Ptrend = 0.02). Increased mRNA tumor-normal fold change (probe_205111_at) was associated with longer survival for GCA (10.7 months for highest quartile; Ptrend = 0.04), but not for GNCA cases (P = 0.72). Similar to mRNA, elevated tumor-normal fold change for protein in ESCC was also associated with improved survival (8.1 months for highest quartile; Ptrend = 0.04). CONCLUSIONS: Dysregulated PLCE1 mRNA expression was observed for both ESCC (one probe only) and GCA tumors, and the altered PLCE1 expression seems to be associated with cancer prognosis. IMPACT: A potential role for PLCE1 in the early detection and/or therapy of ESCC and GCA warrants further investigation.

Profiling of phospho-AKT, phospho-mTOR, phospho-MAPK and EGFR in non-small cell lung cancer.

Activation of numerous pathways has been documented in non-small cell lung cancer (NSCLC). Epidermal growth factor receptor (EGFR) has emerged as a common therapeutic target. The mitogen-activated protein kinase (MAPK) and AKT signaling pathways are downstream of EGFR and deregulated via genetic and epigenetic mechanisms in many human cancers. We evaluated selected markers in the EGFR pathway with reference to outcome. Tissues from 220 cases of NSCLC patients presented in a tissue microarray were assayed with immunohistochemistry for phosphorylated AKT, phosphorylated MAPK, phosphorylated mTOR, and EGFR and then quantified by automated image analysis. Individually, the biomarkers did not predict. Combined as ratios, p-mTOR/p-AKT, and p-MAPK/EGFR function as prognostic markers of survival (p=0.008 and p=0.029, respectively), however, no significance was found after adjustment (p=0.221, p=0.103). The sum of these ratios demonstrates a stronger correlation with survival (p<0.001) and remained statistically significant after adjustment (p=0.026). The algebraic combination of biomarkers offer the capacity to understand factors that predict outcome better than current approaches of evaluating biomarkers individually or in pairs. Our results show the sum of p-mTOR/p-AKT and p-MAPK/EGFR is a potential predictive marker of survival in NSCLC patients.