Reliability and Responsiveness of Clinical and Endoscopic Outcome Measures in Crohn's Disease.

BACKGROUND: Regulatory guidance for Crohn's disease trials recommends coprimary efficacy end points that evaluate both symptoms and mucosal inflammation. We aimed to characterize the operating properties of commonly used disease activity assessments alone and in combination. METHODS: Endoscopic and clinical data were available for 129 participants from the Study of Biologic and Immunomodulator Naïve Patients in Crohn's Disease trial. Readers scored the Simple Endoscopic Score for Crohn's Disease and the Crohn's Disease Endoscopic Index of Severity using standardized conventions. Index reliability was determined using intraclass correlation coefficients. Index responsiveness was assessed using standardized effect sizes based upon treatment assignment. Outcomes were evaluated for optimal sensitivity to treatment effect. RESULTS: Substantial inter-rater reliability was observed when the Simple Endoscopic Score for Crohn's Disease and Crohn's Disease Endoscopic Index of Severity were used as continuous measures (intraclass correlation coefficient, 0.64; 95% confidence interval [CI], 0.50-0.73; and 0.62 95% CI, 0.36-0.77) compared with moderate reliability when dichotomized (0.46; 95% CI, 0.26-0.65; and 0.51; 95% CI, 0.00-0.78). The Simple Endoscopic Score for Crohn's Disease, Crohn's Disease Endoscopic Index of Severity, patient-reported outcome-2, and Crohn's Disease Activity Index were similarly responsive (standardized effect size, 0.43, 95% CI, 0.05-0.81; 0.38, 95% CI, 0.0-0.76; 0.53, 95% CI, 0.15-0.91). A composite outcome of Crohn's Disease Activity Index score <150 and Crohn's Disease Endoscopic Index of Severity score <6 was most sensitive to treatment effect (28.9%; 95% CI, 11.0%-46.8%; P = .003). CONCLUSION: Endoscopic indices were more reliable as continuous measures. Composite outcomes including endoscopy improved sensitivity to treatment effect.

This study largely supports current regulatory guidance for Crohn's disease trials recommending coprimary efficacy end points evaluating both symptoms and mucosal inflammation. Continuous endoscopic measures are most reliable and improve sensitivity to treatment effect when employed in composite outcomes.

A personalized probabilistic approach to ovarian cancer diagnostics.

OBJECTIVE: The identification/development of a machine learning-based classifier that utilizes metabolic profiles of serum samples to accurately identify individuals with ovarian cancer. METHODS: Serum samples collected from 431 ovarian cancer patients and 133 normal women at four geographic locations were analyzed by mass spectrometry. Reliable metabolites were identified using recursive feature elimination coupled with repeated cross-validation and used to develop a consensus classifier able to distinguish cancer from non-cancer. The probabilities assigned to individuals by the model were used to create a clinical tool that assigns a likelihood that an individual patient sample is cancer or normal. RESULTS: Our consensus classification model is able to distinguish cancer from control samples with 93% accuracy. The frequency distribution of individual patient scores was used to develop a clinical tool that assigns a likelihood that an individual patient does or does not have cancer. CONCLUSIONS: An integrative approach using metabolomic profiles and machine learning-based classifiers has been employed to develop a clinical tool that assigns a probability that an individual patient does or does not have ovarian cancer. This personalized/probabilistic approach to cancer diagnostics is more clinically informative and accurate than traditional binary (yes/no) tests and represents a promising new direction in the early detection of ovarian cancer.

Population genomic structure of a widespread, urban-dwelling mammal: The eastern grey squirrel (Sciurus carolinensis).

Urbanization is a persistent and widespread driver of global environmental change, potentially shaping evolutionary processes due to genetic drift and reduced gene flow in cities induced by habitat fragmentation and small population sizes. We tested this prediction for the eastern grey squirrel (Sciurus carolinensis), a common and conspicuous forest-dwelling rodent, by obtaining 44K SNPs using reduced representation sequencing (ddRAD) for 403 individuals sampled across the species' native range in eastern North America. We observed moderate levels of genetic diversity, low levels of inbreeding, and only a modest signal of isolation-by-distance. Clustering and migration analyses show that estimated levels of migration and genetic connectivity were higher than expected across cities and forested areas, specifically within the eastern portion of the species' range dominated by urbanization, and genetic connectivity was less than expected within the western range where the landscape is fragmented by agriculture. Landscape genetic methods revealed greater gene flow among individual squirrels in forested regions, which likely provide abundant food and shelter for squirrels. Although gene flow appears to be higher in areas with more tree cover, only slight discontinuities in gene flow suggest eastern grey squirrels have maintained connected populations across urban areas in all but the most heavily fragmented agricultural landscapes. Our results suggest urbanization shapes biological evolution in wildlife species depending strongly on the composition and habitability of the landscape matrix surrounding urban areas.

The efficacy and safety of tranexamic acid utilization in total ankle arthroplasty: a systematic review and meta-analysis.

INTRODUCTION: There is still a lack of information on the role of Tranexamic acid (TXA) in total ankle arthroplasty (TAA). The purpose of this study is to comprehensively review, consolidate, and analyze findings from existing research on the effectiveness and safety of TXA in TAA. MATERIALS AND METHODS: The comprehensive literature review was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) using PubMed, Embase, Web of Science, and Cochrane databases, for original, English-language studies investigating the efficacy and safety of TXA in TAA, through February 2023. Evaluated data for the meta-analysis included estimated blood loss (EBL), change in perioperative hemoglobin, need for transfusion, and complications including DVT/PE, and wound complications. RESULTS: A total of nine studies were included in this study. In total, 450 TAA were included, with 244 receiving TXA (54.2%) and 206 not receiving TXA (45.8%). TXA in TAA significantly decreased EBL. A significantly lower rate of wound complications in the TXA group with the relative risk (RR) of 0.51. We classified wound complications into wound infection and delayed wound healing/dehiscence. A significant decrease in the rate of wound infection and a tendency showing a decrease in the rate of delayed wound healing/dehiscence in the TXA group were noted: the RR of 0.29, and 0.63, respectively. TXA did not increase the incidence of DVT/PE following TAA. CONCLUSIONS: In conclusion, the utilization of TXA during TAA demonstrated a statistically significant reduction in EBL and relative risk for wound complications. However, further RCTs with larger sample sizes will be necessary to establish a more robust conclusion regarding the efficacy and safety of TXA in TAA. LEVEL OF EVIDENCE: Level III, systematic review and meta-analysis.

Overdiagnosis of Newborn Hyperbilirubinemia: A Natural Experiment in Quality Improvement Fundamentals.

Two hospitals noted increased newborn hyperbilirubinemia coinciding with an undisclosed total serum bilirubin (TSB) assay change. Clinicians rapidly applied quality improvement methodologies to ascertain increased jaundice evaluations, readmissions, and possible safety issues. Methods: In January 2020, 2 hospitals (A and B) transitioned to a new method of measuring TSB using a new clinical chemistry analyzer (Siemens Atellica CH), which measured TSB by vanadate oxidase assay instead of the previous diazo assay. Five affiliated hospitals (C-G) continued to utilize the diazo assay. This natural experiment led to a comparison of data across the 7 hospitals. We analyzed: (1) TSB levels, (2) hospital hyperbilirubinemia readmissions, and (3) paired TSB measurements comparing the diazo assay and vanadate oxidase method. Results: Compared to the 2019 baseline, Hospitals A and B had a significant increase in TSBs ≥17.0 mg/dl and TSBs ≥20 mg/dl in 2020; Hospitals C-G did not. Readmissions for phototherapy significantly increased in hospitals A and B in 2020 compared to 2019. Paired blood samples showed bias-elevated TSBs by vanadate assay compared to the diazo method. By 2021, the laboratory resumed processing TSB samples by diazo assay, and the frequency of elevated TSBs and hyperbilirubinemia readmissions returned to 2019 levels. Conclusions: Factitious TSB elevation related to an assay change significantly increased newborn hyperbilirubinemia evaluations and phototherapy readmissions. Imbedded quality improvement methodologies of careful structure, process, and outcomes review hastened resolution.

The Clinical Significance of Genetic Variation in Ovarian Cancer.

Genetic variation is a well-known contributor to the onset and progression of cancer. The goal of this study is to provide a comprehensive examination of the nucleotide and chromosomal variation associated with the onset and progression of serous ovarian cancer. Using a variety of computational and statistical methods, we examine the exome sequence profiles of genetic variants present in the primary tumors of 432 ovarian cancer patient samples to compute: (1) the tumor mutational burden for all genes and (2) the chromosomal copy number alterations associated with the onset/progression of ovarian cancer. Tumor mutational burden is reduced in the late vs. early stages, with the highest levels being associated with loss-of-function mutations in DNA-repair genes. Nucleotide variation and copy number alterations associated with known cancer driver genes are selectively favored over ovarian cancer development. The results indicate that genetic variation is a significant contributor to the onset and progression of ovarian cancer. The measurement of the relative levels of genetic variation associated with individual ovarian cancer patient tumors may be a clinically valuable predictor of potential tumor aggressiveness and resistance to chemotherapy. Tumors found to be associated with high levels of genetic variation may help in the clinical identification of high-risk ovarian cancer patients who could benefit from more frequent monitoring.

Standardizing Scoring Conventions for Crohn's Disease Endoscopy: An International RAND/UCLA Appropriateness Study.

BACKGROUND AND AIMS: Endoscopic assessment of disease activity is integral for evaluating treatment response in patients with Crohn's disease (CD). We aimed to define appropriate items for evaluating endoscopic activity and conventions for consistent endoscopic scoring rules in CD. METHODS: A 2-round modified RAND/University of California at Los Angeles Appropriateness Method study was conducted. A panel of 15 gastroenterologists used a 9-point Likert scale to rate the appropriateness of statements pertaining to the Simple Endoscopic Score for CD, Crohn's Disease Endoscopic Index of Severity, and additional items relevant to endoscopy scoring in CD. Each statement was voted as appropriate, uncertain, or inappropriate based on the median panel rating and presence of disagreement. RESULTS: Panelists voted that it is appropriate for all ulcers to contribute to endoscopic scoring in CD, including aphthous ulcers, ulcerations at a surgical anastomosis, and anal canal ulcers (scored in the rectum). Endoscopic healing should reflect an absence of ulcers. Narrowing should be defined as a clear decrease in luminal diameter; stenosis should be defined by an impassable narrowing, and if occurring at the junction of 2 segments, scored in the distal segment. Scarring and inflammatory polyps were considered inappropriate for including in the affected area score. The optimal method for defining ulcer depth remains uncertain. CONCLUSIONS: We outlined scoring conventions for the Simple Endoscopic Score for CD and Crohn's Disease Endoscopic Index of Severity, noting that both scores have limitations. Therefore, we identified priorities for future research and steps for developing and validating a more representative endoscopic index in CD.

Correlating mechanical and gene expression data on the single cell level to investigate metastatic phenotypes.

Stiffness has been observed to decrease for many cancer cell types as their metastatic potential increases. Although cell mechanics and metastatic potential are related, the underlying molecular factors associated with these phenotypes remain unknown. Therefore, we have developed a workflow to measure the mechanical properties and gene expression of single cells that is used to generate large linked-datasets. The process combines atomic force microscopy to measure the mechanics of individual cells with multiplexed RT-qPCR gene expression analysis on the same single cells. Surprisingly, the genes that most strongly correlated with mechanical properties were not cytoskeletal, but rather were markers of extracellular matrix remodeling, epithelial-to-mesenchymal transition, cell adhesion, and cancer stemness. In addition, dimensionality reduction analysis showed that cell clustering was improved by combining mechanical and gene expression data types. The single cell genomechanics method demonstrates how single cell studies can identify molecular drivers that could affect the biophysical processes underpinning metastasis.

Characteristics of Marginalized Populations Seeking Acupuncture: Findings from an Observational Study of an Inner-City Community Health Service.

Introduction: Marginalized populations experience health inequities and are often underserved within existing health systems. Australian marginalized populations and their use of complementary medicine, including acupuncture, lack investigation. We have collected information on the health-seeking behaviors of marginalized individuals who utilize an acupuncture service within a community-based integrative health setting. Methods: A secondary analysis of pre-existing data involving the linking of three datasets. Information was collected across four domains: health characteristics, socio-demographics, health services utilization, and vulnerability markers. Bivariate analysis using Fisher's exact and chi-square tests additional to logistic regression analysis were conducted to determine the characteristics of the study population. All data once analyzed were then presented as a cumulative statistic. Results: Study participants (n = 42) included 28% of individuals with reported histories of homelessness (n = 12) and 32% with a history of psychological trauma (n = 13). Eighty-three percent (n = 31) of the population sought acupuncture to manage pain and 91% (n = 36) for musculoskeletal conditions. Sixty-three percent (n = 24) reported a mental health diagnosis, most commonly depression (n = 18). Participants were most likely to engage with three other health services within the study setting, in addition to acupuncture. Participants with an illicit substance abuse problem were 12 times more likely to seek a greater number of acupuncture treatments, and individuals with trauma histories were twice as likely to attend the acupuncture clinic eight or more times. Discussion: Our study findings suggest a robust level of engagement with acupuncture treatment among the target population and a willingness to engage with integrative health services when barriers to uptake such as accessibility and affordability are removed. Findings support current evidence pertaining to acupuncture's use as an adjunct to managing pain in marginalized populations as well as a perceived acceptability and feasibility of the integration of acupuncture within conventional health settings. There is the further observation that acupuncture in a group setting is suitable for a marginalized population and an interest in commitment to treatment among individuals with substance abuse problems.

Integration of molecular modelling and in vitro studies to inhibit LexA proteolysis.

Introduction: As antibiotic resistance has become more prevalent, the social and economic impacts are increasingly pressing. Indeed, bacteria have developed the SOS response which facilitates the evolution of resistance under genotoxic stress. The transcriptional repressor, LexA, plays a key role in this response. Mutation of LexA to a non-cleavable form that prevents the induction of the SOS response sensitizes bacteria to antibiotics. Achieving the same inhibition of proteolysis with small molecules also increases antibiotic susceptibility and reduces drug resistance acquisition. The availability of multiple LexA crystal structures, and the unique Ser-119 and Lys-156 catalytic dyad in the protein enables the rational design of inhibitors. Methods: We pursued a binary approach to inhibit proteolysis; we first investigated ß-turn mimetics, and in the second approach we tested covalent warheads targeting the Ser-119 residue. We found that the cleavage site region (CSR) of the LexA protein is a classical Type II ß-turn, and that published 1,2,3-triazole compounds mimic the ß-turn. Generic covalent molecule libraries and a ß-turn mimetic library were docked to the LexA C-terminal domain using molecular modelling methods in FlexX and CovDock respectively. The 133 highest-scoring molecules were screened for their ability to inhibit LexA cleavage under alkaline conditions. The top molecules were then tested using a RecA-mediated cleavage assay. Results: The ß-turn library screen did not produce any hit compounds that inhibited RecA-mediated cleavage. The covalent screen discovered an electrophilic serine warhead that can inhibit LexA proteolysis, reacting with Ser-119 via a nitrile moiety. Discussion: This research presents a starting point for hit-to-lead optimisation, which could lead to inhibition of the SOS response and prevent the acquisition of antibiotic resistance.

Ethnic disparities in mortality and group-specific risk factors in the UK Biobank.

Despite a substantial overall decrease in mortality, disparities among ethnic minorities in developed countries persist. This study investigated mortality disparities and their associated risk factors for the three largest ethnic groups in the United Kingdom: Asian, Black, and White. Study participants were sampled from the UK Biobank (UKB), a prospective cohort enrolled between 2006 and 2010. Genetics, biological samples, and health information and outcomes data of UKB participants were downloaded and data-fields were prioritized based on participants with death registry records. Kaplan-Meier method was used to evaluate survival differences among ethnic groups; survival random forest feature selection followed by Cox proportional-hazard modeling was used to identify and estimate the effects of shared and ethnic group-specific mortality risk factors. The White ethnic group showed significantly worse survival probability than the Asian and Black groups. In all three ethnic groups, endoscopy and colonoscopy procedures showed significant protective effects on overall mortality. Asian and Black women show lower relative risk of mortality than men, whereas no significant effect of sex was seen for the White group. The strongest ethnic group-specific mortality associations were ischemic heart disease for Asians, COVID-19 for Blacks, and cancers of respiratory/intrathoracic organs for Whites. Mental health-related diagnoses, including substance abuse, anxiety, and depression, were a major risk factor for overall mortality in the Asian group. The effect of mental health on Asian mortality, particularly for digestive cancers, was exacerbated by an observed hesitance to answer mental health questions, possibly related to cultural stigma. C-reactive protein (CRP) serum levels were associated with both overall and cause-specific mortality due to COVID-19 and digestive cancers in the Black group, where elevated CRP has previously been linked to psychosocial stress due to discrimination. Our results point to mortality risk factors that are group-specific and modifiable, supporting targeted interventions towards greater health equity.

Existing Bowel Preparation Quality Scales Are Reliable in the Setting of Centralized Endoscopy Reading.

BACKGROUND: Development of bowel preparation products has been based upon colon cleansing rating by a local endoscopist. It is unclear how bowel preparation scales perform when centrally evaluated. AIMS: To evaluate the reliability of bowel preparation quality scales when assessed by central readers. METHODS: Four central readers evaluated 52 videos in triplicate, 2 weeks apart, during the entire endoscopic procedure (insertion/withdrawal of the colonoscope) and exclusively on colonoscope withdrawal using the Boston Bowel Preparation Scale (BBPS), Chicago Bowel Preparation scale, Harefield Cleansing Scale, Ottawa Bowel Preparation Quality Scale (OBPQS), Aronchick score, a visual analogue scale, and additional items proposed in a modified Research and Development/University of California Los Angeles appropriateness process. Reliability was assessed with intraclass correlation coefficients. RESULTS: Intraclass correlation coefficients (95% confidence interval) for inter-rater reliability of the quality scales ranged from 0.51 to 0.65 (consistent with moderate to substantial inter-rater reliability) during the entire procedure. Corresponding intraclass correlation coefficients for intra-rater reliability ranged from 0.69 to 0.77 (consistent with substantial intra-rater reliability). Reliability was highest in the right colon and lowest in the left colon. No differences were observed in reliability when assessed for the procedure overall (insertion/withdrawal) relative to assessment on withdrawal alone. CONCLUSION: All five bowel preparation quality scales had moderate to substantial inter-rater reliability. Panelists considered the Aronchick score too simplistic for clinical trials and recognized that assessment of residual fluid in the Ottawa Bowel Preparation Quality Scale was not amenable to central assessment.

Revisiting the automaticity of reading: Electrophysiological recordings show that stroop words capture spatial attention.

Interference in the Stroop task is reduced when the word and color patch are placed at different locations and is diluted further by the presence of another distractor that is response neutral. Such dilution indicates that reading is not independent of an observer's attentional focus and thus is not a fully automatic process. So where does reading fall on the automaticity continuum? To address this question, we sought to determine whether an irrelevant word that appears abruptly in the field of view invariably draws attention to its location or whether observers can successfully ignore it while identifying a centrally presented target. In two experiments, electrical brain activity was recorded while healthy young adults participated in nonintegrated Stroop tasks. Irrelevant color words appearing randomly to the left or right of a target shape elicited an event-related potential component that reflects the spatial focusing of attention (posterior contralateral N2; N2pc). This N2pc was observed when participants discriminated the color of the target and when they discriminated the shape of the target. These findings demonstrate that color words reflexively capture spatial attention even when their meaning is unrelated to the task at hand. We conclude that although reading is not fully automatic, skilled readers cannot ignore words that appear abruptly in their field of view. (PsycInfo Database Record (c) 2023 APA, all rights reserved).

Difficulty suppressing visual distraction while dual tasking.

Human beings must often perform multiple tasks concurrently or in rapid succession. Laboratory research has revealed striking limitations in the ability to dual task by asking participants to identify two target objects that are inserted into a rapid stream of irrelevant items. Under a variety of conditions, identification of the second target (T2) is impaired for a short period of time following presentation of the first target (T1). Several theories have been developed to account for this "attentional blink" (AB), but none makes a specific prediction about how processing of T1 might impact an observer's ability to ignore a salient distractor that accompanies T2. Using event-related potentials (ERPs) to track target and distractor processing, we show that healthy young adults are capable of suppressing a salient visual-search distractor (D2) while dual tasking (as measured by the PD component, which has been associated with suppression) but struggle to do so shortly after the appearance of T1. In fact, the impairment was more severe for distractor processing than it was for target processing (as measured by the N2pc component). Whereas, the T2-elicited N2pc was merely delayed during the AB, the distractor PD was reduced in magnitude and was found to be statistically absent. We conclude that the inhibitory control processes that are typically engaged to prevent distraction are unavailable while an observer is busy processing a target that appeared earlier.

Is Electroacupuncture Contraindicated for Preventing and Treating Chemotherapy-Induced Peripheral Neuropathy?

Background: Chemotherapy-induced peripheral neuropathy (CIPN) is a common side-effect of several drugs used to combat cancer. Thus, researchers have sought better treatments for and prevention of CIPN, such as electroacupuncture (EA). Some trials show EA worsens or prolongs CIPN pain and recommend against further studies on this. This narrative review explores EA for preventing or treating CIPN, comparing positive and negative outcomes. Methods: PubMed, ScienceDirect, and Google Scholar were searched for electroacupuncture, CIPN, and peripheral neuropathy. A snowballing method was used to find systematic reviews and studies in systematic reviews. Results: Seven English-language trials were found on using EA for preventing or treating CIPN. In 3 prevention studies, 1 had significant benefits, 1 had modest benefits, and 1 had worse pain in an EA group at follow-up, compared to sham controls. In 4 treatment studies, 2 had significant benefits, 1 had no difference from 3 controls, and 1 had sham control was superior to verum EA. Conclusions: Most of the studies were limited by small sample sizes, and some studies used EA protocols and treatment doses (frequency and total number of sessions) that were potentially suboptimal. The quantity and quality of the studies are insufficient to draw firm conclusions on effectiveness and safety. More studies must test optimal EA protocols and treatment dosages. It is inappropriate to say that EA is not recommended for CIPN prevention or treatment, because there is no robust evidence to justify this. Generally, research has found benefits and no harms.

Attentional enhancement predicts individual differences in visual working memory under go/no-go search conditions.

Attention-control processes transfer relevant information to visual working memory (WM) and prevent irrelevant information from consuming WM resources. Although event-related potentials (ERPs) have revealed attention-control processes associated with enhancement of relevant stimuli (targets) and suppression of irrelevant stimuli (distractors), only the suppressive processes have been found to predict WM capacity. We hypothesised a link between target-enhancement processes and WM capacity would be revealed in a task that requires more control than the conventional visual search paradigms used to study target selection. Here, participants searched for a pop-out target on Go trials and withheld responses on an equal number of randomly intermixed No-Go trials, depending on the colour of the stimulus array. Magnitudes of ERP indices associated with target enhancement (the singleton detection positivity, SDP, and N2pc) were positively correlated with individual differences in WM capacity. These relationships vanished when participants searched for the pop-out target on every trial, regardless of stimulus-array colour. Inhibitory processes associated with suppressing distractors (PD) and withholding responses (no-go P3) on No-Go trials did not predict WM capacity. These findings indicate that target-enhancement mechanisms control access to WM in search tasks that require dynamic control and disconfirm the view that the gateway to WM is entirely inhibitory by nature.

Synthesis, Biological Activity and Molecular Docking of Chimeric Peptides Targeting Opioid and NOP Receptors.

Recently, mixed opioid/NOP agonists came to the spotlight for their favorable functional profiles and promising outcomes in clinical trials as novel analgesics. This study reports on two novel chimeric peptides incorporating the fragment Tyr-c[D-Lys-Phe-Phe]Asp-NH2 (RP-170), a cyclic peptide with high affinity for µ and κ opioid receptors (or MOP and KOP, respectively), conjugated with the peptide Ac-RYYRIK-NH2, a known ligand of the nociceptin/orphanin FQ receptor (NOP), yielding RP-170-RYYRIK-NH2 (KW-495) and RP-170-Gly3-RYYRIK-NH2 (KW-496). In vitro, the chimeric KW-496 gained affinity for KOP, hence becoming a dual KOP/MOP agonist, while KW-495 behaved as a mixed MOP/NOP agonist with low nM affinity. Hence, KW-495 was selected for further in vivo experiments. Intrathecal administration of this peptide in mice elicited antinociceptive effects in the hot-plate test; this action was sensitive to both the universal opioid receptor antagonist naloxone and the selective NOP antagonist SB-612111. The rotarod test revealed that KW-495 administration did not alter the mice motor coordination performance. Computational studies have been conducted on the two chimeras to investigate the structural determinants at the basis of the experimental activities, including any role of the Gly3 spacer.

In vitro sepsis induces Nociceptin/Orphanin FQ receptor (NOP) expression in primary human vascular endothelial but not smooth muscle cells.

Sepsis is a dysregulated host response to infection that can cause widespread effects on other organs including cardiovascular depression, hypotension and organ failure. The receptor for Nociceptin/Orphanin FQ (N/OFQ), NOP is expressed on immune cells and these cells can release the peptide. Exogenous N/OFQ can dilate blood vessels and this peptide is increased in animal and human sepsis. We hypothesise that NOP receptors are present on vascular endothelial cells and therefore provide the target for released N/OFQ to cause vasodilation and hence hypotension. Using human umbilical vein endothelial cells (HUVEC) and human vascular smooth muscle cells (HVSMC) freshly prepared from umbilical cords and up to passage 4, we assessed NOP mRNA expression by Polymerase Chain Reaction (PCR), NOP surface receptor expression using a fluorescent NOP selective probe (N/OFQATTO594) and NOP receptor function with N/OFQ stimulated ERK1/2 phosphorylation. As an in vitro sepsis mimic we variably incubated cells with 100ng/ml Lipopolysaccharide and Peptidoglycan G (LPS/PepG). HUVECs express NOP mRNA and this was reduced by ~80% (n = 49) after 24-48 hours treatment with LPS/PepG. Untreated cells do not express surface NOP receptors but when treated with LPS/PepG the reduced mRNA was translated into protein visualised by N/OFQATTO594 binding (n = 49). These NOP receptors in treated cells produced an N/OFQ (1µM) driven increase in ERK1/2 phosphorylation (n = 20). One (of 50) HUVEC lines expressed NOP mRNA and receptor protein in the absence of LPS/PepG treatment. In contrast, HVSMC expressed NOP mRNA and surface receptor protein (n = 10) independently of LPS/PepG treatment. These receptors were also coupled to ERK1/2 where N/OFQ (1µM) increased phosphorylation. Collectively these data show that an in vitro sepsis mimic (LPS/PepG) upregulates functional NOP expression in the vascular endothelium. Activation of these endothelial receptors as suggested from in vivo whole animal work may contribute to the hypotensive response seen in sepsis. Moreover, blockade of these receptors might be a useful adjunct in the treatment of sepsis.