Search

Electron pressure profile and transport behaviors in ion Bernstein waves (IBW) heated plasmas were investigated at three frequencies (24, 27, and 30 MHz) in the Hefei Tokamak-7. Both on-axis and off-axis electron heating with global peaked and locally steepened electron pressure profiles were realized, if the ion cyclotron resonant layer was properly selected. Improvement of the IBW-heated plasma confinement was observed except in the 24 MHz case. Reduction of the local electron heat transport was evident from sawtooth heat pulse propagation. High performance plasma volume was extended in the IBW off-axis heating mode, which resulted in improvement of the global plasma confinement. These studies demonstrate that IBW is a potentially powerful tool for active control of plasma profiles and transport. © 2003 American Institute of Physics. [ABSTRACT FROM AUTHOR]

Objective: To examine the associations between components of social networks and health-related quality of life (HRQoL) in community-dwelling old adults in urban areas in China. Study design: Data from the 4th National Household Health Survey (NHHS) in China, conducted in 2008, were used. HRQoL of respondents aged ≥15 years was assessed using EQ-5D in the NHHS. Methods: The sample for the current analysis included 9833 old adults aged ≥60 years. Multiple linear and logistic regression models were used to assess the associations between indicators of social network and HRQoL. Results: Approximately 6% of the respondents saw their children once a year or less, and approximately 1% reported that they had no children. Thirteen percent of the sample seldom contacted their neighbours and seldom met with relatives or friends; approximately 62% seldom attended social gatherings. The five dimensions of HRQoL (mobility, self-care, usual activities, pain/discomfort and anxiety/depression) were reported to be moderate or severe in 14.5%, 9.4%, 12.6%, 18.3% and 9.3% of the sample, respectively. The mean visual analogue scale (VAS) score and EQ-5D index using the time trade-off method was 70.96 [standard deviation (SD) 14.79] and 0.869 (SD 0.163), respectively. After adjusting for potential confounding variables, old adults with weaker social networks were more likely to report problems on EQ-5D dimensions, lower VAS scores and lower EQ-5D indexes. Conclusions: For old adults living in urban communities in China, increased social participation has a positive effect on various dimensions of HRQoL. There is a need for policy considerations that will improve integration of community-level public resources in order to encourage frequent social interaction among old adults, and promote health and social care as a whole. [ABSTRACT FROM AUTHOR]

Aim: To determine the optic disc size in the adult Chinese population in an urban and a rural region of Beijing. Methods: The population based, cross sectional cohort study included 4439 subjects out of 5324 subjects invited to participate (response rate 83.4%). It was divided into a rural part (1973 (44.4%) subjects) and an urban part (2466 (55.6%) subjects). Mean age was 56.2 (SD 10.6) years (range 40–101 years). Colour optic disc photographs were morphometrically examined. Main outcome measure was optic disc area. Results: Optic disc photographs were available for 4027 (90.7%) subjects. Mean optic disc area measured 2.65 (0.57) mm² (range 1.03 mm²–7.75 mm²). Optic disc area was significantly (p<0.001) correlated with myopic refractive error, with a steep decrease in optic disc area from high myopia to the mid-range of refractive error, a slightly horizontal course in the refractive error range between -8 dioptres and +4 dioptres, and a further decrease in optic disc area towards higher hyperopia. Optic disc area was not related to age (p=0.14) or sex (p=0.93) (optic disc area, males: 2.65 (0.56) mm² versus females: 2.65 (0.57) mm²). ‘Microdiscs’ may be defined as smaller than 1.51 mm², and ‘macrodiscs’ as larger than 3.79 mm². Conclusions: Compared with data of preceding studies, mean optic disc size is larger in Chinese people than in white people. In Chinese people highly hyperopic eyes have significantly smaller optic discs, and highly myopic eyes have significantly larger optic discs than emmetropic eyes. [ABSTRACT FROM AUTHOR]

Full impedance matrices of the method of moments (MoM) type have been decomposed into sparse matrices in the application of on-surface measured equation of invariance (OSMEI) [1], [2]. The objectives in [1] and [2] are to find continuous current distributions on scatterers. Thus, the concept of metron and measure is justified under the postulate that the local, relation between fields and current densities is an invariant of the excitation. In this paper, we have extended the concept to dipole arrays, where the objective is to find the discrete driving point currents. Numerical results show that using that windowed metrons presented in this paper, the sparsity rate of the sparse matrices can be kept within 1%, while the E- and H-plane patterns are still in good agreement with the MoM results except for the farthest side lobes of the H-plane patterns. [ABSTRACT FROM AUTHOR]

Reports on a molecular phylogenetic analysis of the Ixodes ricinus complex. Distribution of the Ixodes ricinus, which is a group of ticks, in almost all geographic regions of the world; Transmission of the tick species by lyme borreliosis spirochetes; Sequencing of two fragments of mitochondrial 16S ribosomal deoxyribonucleic acid from 11 species in the Ixodes ricinus complex.

Background: Problematic alcohol use is a recognized risk factor for suicidal thoughts and behaviors. Individuals who misuse alcohol and those experiencing suicidal ideation (SI) often exhibit elevated impulsivity, suggesting that impulsivity may be an underlying mechanism. Impulsivity fluctuates considerably within individuals over short periods, particularly among those engaged in episodic heavy drinking, college students, and those with SI. While impulsivity is associated with suicidal thinking at a trait level, its relationship with SI on a momentary level remains relatively unexplored. This study aimed to (a) replicate cross-sectional findings linking impulsivity and SI in those with problematic alcohol use, and (b) extend these findings to examine the momentary associations between impulsivity and SI within this population., Method: Two independent undergraduate samples were utilized: the first cross-sectional (n = 653) and the second employing ecological momentary assessment (n = 56). All participants screened for problematic alcohol or substance use., Results: Our results show a unique association between the impulsivity facet of negative urgency and SI. Results also demonstrate an interactive effect of negative affect and "spur of the moment decisions" in predicting concurrent SI, as well as an interaction between negative affect and "saying things without thinking" in prospectively predicting SI., Conclusions: Findings highlight the saliency of negative urgency, both at the trait and state level in the prediction of SI. Although in need of replication, results indicate that specific aspects of momentary impulsivity may be uniquely linked with SI, and that this relationship may be context dependent., (© 2024 Wiley Periodicals LLC.)

Introduction: The transition from suicidal thoughts to behaviors often involves considering the consequences of suicide as part of the decision-making process. This study explored the relationship between this consideration process and the decision to either abort or carry out a suicide attempt., Methods: Among inpatients with a suicide-related event in the past 2 weeks (suicide attempt n = 30 or aborted attempt n = 16), we assessed the degree to which they considered six domains of consequences, the impact of these considerations on their inclination to attempt suicide, and the duration of their decision-making., Results: All the participants who aborted and 87% of those who attempted considered consequences of suicide. Participants who aborted took longer to progress through decision-making stages and considered more suicide-hindering factors, especially interpersonal ones, though these differences were no longer significant after correction. Group status moderated the relationship between the balance of suicide-facilitating and suicide-hindering considerations and decision-making duration. Considering the consequences of suicide more favorably was related to a shorter ideation-to-action period before a suicide attempt and a longer ideation period before aborting an attempt., Conclusion: This study highlights the complexity of suicide decision-making and its role in better understanding the progression from ideation to action., (© 2024 The Author(s). Suicide and Life‐Threatening Behavior published by Wiley Periodicals LLC on behalf of American Association of Suicidology.)

Importance: While the association between cross-sectional measures of social isolation and adverse health outcomes is well established, less is known about the association between changes in social isolation and health outcomes., Objective: To assess changes of social isolation and mortality, physical function, cognitive function, cardiovascular disease (CVD), and stroke., Design, Setting, and Participants: In a cohort design, social isolation changes in 4 years and subsequent risk of mortality and other outcomes were assessed using the 13 649 eligible Health and Retirement Study (HRS) respondents from the 2006 to 2020 waves. Data were analyzed from October 11, 2023, to April 26, 2024., Exposure: The main exposure was the change in social isolation measured by the Steptoe 5-item Social Isolation Index from the initial assessment to a second assessment conducted 4 years later. Participants were classified into decreased isolation, stable, or increased isolation groups, stratified by their baseline isolation status., Main Outcomes and Measures: The primary outcomes were mortality, self-reported dependencies in activities of daily living, Alzheimer disease and Alzheimer disease-related dementia, CVD, and stroke. Dementia, CVD, and stroke were assessed using HRS-linked Medicare records. Incidence rates (IRs) of each group were estimated and a Cox proportional hazards regression model was used, with inverse-probability treatment weighting to adjust for confounders., Results: Among 13 649 participants (mean [SD] age at baseline, 65.3 [9.5] years; 8011 [58.7%] women) isolated at baseline, those with increased isolation had higher mortality (n = 693; IR = 68.19; 95% CI, 60.89-76.36 per 1000 person-years) than those who were stable (n = 1796; IR = 44.02; 95% CI, 40.47-47.88 person-years) or had decreased isolation (n = 2067; IR = 37.77; 95% CI, 34.73-41.09 person-years) isolation. Increased isolation was associated with higher risks of mortality (adjusted hazard ratio [AHR], 1.29; 95% CI, 1.09-1.51), disability (AHR, 1.35; 95% CI, 1.09-1.67), and dementia (AHR, 1.40; 95% CI, 1.02-1.93) compared with stable isolation. Similar findings were observed among socially nonisolated participants at baseline., Conclusions and Relevance: In this cohort study, increased isolation was associated with elevated risks of mortality, disability, and dementia, irrespective of baseline isolation status. These results underscore the importance of interventions targeting the prevention of increased isolation among older adults to mitigate its adverse effects on mortality, as well as physical and cognitive function decline.

Charcot-Marie-Tooth (CMT) disease is a neuromuscular disorder affecting the peripheral nervous system. The diagnostic yield in demyelinating CMT (CMT1) is typically ∼80-95%, of which at least 60% is due to the PMP22 gene duplication. The remainder of CMT1 is more genetically heterogeneous. We used whole exome and whole genome sequencing data included in the GENESIS database to investigate novel causal genes and mutations in a cohort of ∼2,670 individuals with CMT neuropathy. A recurrent heterozygous missense variant p.Thr1424Met in the recently described CMT gene ITPR3, encoding IP3R3 (inositol 1,4,5-trisphosphate receptor 3) was identified. This previously reported p.Thr1424Met change was present in 33 affected individuals from nine unrelated families from multiple populations, representing an unusual recurrence rate at a mutational hotspot, strengthening the gene-disease relationship (GnomADv4 allele frequency 1.76e-6). Sanger sequencing confirmed the co-segregation of the CMT phenotype with the presence of the mutation in autosomal dominant and de novo inheritance patterns, including a four-generation family with multiple affected second-degree cousins. Probands from all families presented with slow nerve conduction velocities, matching the diagnostic category of CMT1. Remarkably, we observed a uniquely variable clinical phenotype for age at onset and phenotype severity in p.Thr1424Met carrying patients, even within families. Finally, we present data supportive of a dominant-negative effect of the p.Thr1424Met mutation with associated changes in protein expression in patient-derived cells., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Charcot-Marie-Tooth disease type 1A (CMT1A) is a demyelinating peripheral neuropathy caused by the duplication of peripheral myelin protein 22 (PMP22), leading to muscle weakness and loss of sensation in the hands and feet. A recent case-only genome-wide association study of CMT1A patients conducted by the Inherited Neuropathy Consortium identified a strong association between strength of foot dorsiflexion and variants in signal induced proliferation associated 1 like 2 (SIPA1L2), indicating that it may be a genetic modifier of disease. To validate SIPA1L2 as a candidate modifier and to assess its potential as a therapeutic target, we engineered mice with deletion of exon 1 (including the start codon) of the Sipa1l2 gene and crossed them to the C3-PMP22 mouse model of CMT1A. Neuromuscular phenotyping showed that Sipa1l2 deletion in C3-PMP22 mice preserved muscular endurance assayed by inverted wire hang duration and changed femoral nerve axon morphometrics such as myelin thickness. Gene expression changes suggest involvement of Sipa1l2 in cholesterol biosynthesis, a pathway that is also implicated in C3-PMP22 mice. Although Sipa1l2 deletion did impact CMT1A-associated phenotypes, thereby validating a genetic interaction, the overall effect on neuropathy was mild., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Association of Neuropathologists, Inc. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Substantial evidence suggests a role for immunotherapy in treating Alzheimer's disease (AD). While the precise pathophysiology of AD is incompletely understood, clinical trials of antibodies targeting aggregated forms of β amyloid (Aβ) have shown that reducing amyloid plaques can mitigate cognitive decline in patients with early-stage AD. Here, we describe what we believe to be a novel approach to target and degrade amyloid plaques by genetically engineering macrophages to express an Aβ-targeting chimeric antigen receptor (CAR-Ms). When injected intrahippocampally, first-generation CAR-Ms have limited persistence and fail to significantly reduce plaque load, which led us to engineer next-generation CAR-Ms that secrete M-CSF and self-maintain without exogenous cytokines. Cytokine secreting "reinforced CAR-Ms" have greater survival in the brain niche and significantly reduce plaque load locally in vivo. These findings support CAR-Ms as a platform to rationally target, resorb, and degrade pathogenic material that accumulates with age, as exemplified by targeting Aβ in AD.

Expansions of tandem repeats (TRs) cause approximately 60 monogenic diseases. We expect that the discovery of additional pathogenic repeat expansions will narrow the diagnostic gap in many diseases. A growing number of TR expansions are being identified, and interpreting them is a challenge. We present RExPRT (Repeat EXpansion Pathogenicity pRediction Tool), a machine learning tool for distinguishing pathogenic from benign TR expansions. Our results demonstrate that an ensemble approach classifies TRs with an average precision of 93% and recall of 83%. RExPRT's high precision will be valuable in large-scale discovery studies, which require prioritization of candidate loci for follow-up studies., (© 2024. The Author(s).)

Objective: To identify tools that predict the risk of complications for patients presenting to an outpatient clinic or an emergency department (ED) with influenza-like illness., Methods: We searched Medline, Embase, Cochrane Library and CINAHL from inception to July 2023. We included articles reporting on the derivation or validation of a score or algorithm used to stratify the risk of hospitalization or mortality among patients with influenza-like illness in the ED or outpatient clinic., Results: Twelve articles reporting on eight scores and six predictive models were identified. For predicting the need for hospitalization, the area under the curve (AUC) of the PMEWS and the CURB-65 ranged respectively from 0.76 to 0.94, and 0.65 to 0.88. The Community Assessment Tool had an AUC of 0.62. For predicting inpatient mortality, AUC was 0.66 for PMEWS and 0.79 for CURB-65, 0.79 for the SIRS criteria and 0.86 for the qSOFA score. Two scores were developed without external validation during the Covid-19 pandemic. The CovHos score and the Canadian Covid discharge score had an AUC ranged from 0.70 to 0.91. The predictive models performed adequately (AUC from 0.76 to 0.92) but will require external validation for clinical use. Tool diversity and study population heterogeneity precluded meta-analysis., Conclusion: Although the CURB, PMEWS and qSOFA scores appear to predict accurately the risk of complications of influenza-like illness, none were reliable enough to justify their widespread ED use. Refinement of an existing tool or development of a new tool to optimize the management of these patients is needed., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:Simon Berthelot reports article publishing charges was provided by Quebec Ministry of the Economy and Innovation. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors. Published by Elsevier Ltd.)

Conditional gene regulation in Drosophila through binary expression systems like the LexA-LexAop system provides a superb tool for investigating gene and tissue function. To increase the availability of defined LexA enhancer trap insertions, we present molecular, genetic, and tissue expression studies of 301 novel Stan-X LexA enhancer traps derived from mobilization of the index SX4 line. This includes insertions into distinct loci on the X, II, and III chromosomes that were not previously associated with enhancer traps or targeted LexA constructs, an insertion into ptc, and seventeen insertions into natural transposons. A subset of enhancer traps was expressed in CNS neurons known to produce and secrete insulin, an essential regulator of growth, development, and metabolism. Fly lines described here were generated and characterized through studies by students and teachers in an international network of genetics classes at public, independent high schools, and universities serving a diversity of students, including those underrepresented in science. Thus, a unique partnership between secondary schools and university-based programs has produced and characterized novel resources in Drosophila, establishing instructional paradigms devoted to unscripted experimental science., Competing Interests: Conflicts of interest statement The author(s) declare no conflict of interest., (© The Author(s) 2023. Published by Oxford University Press on behalf of The Genetics Society of America.)

Background: Most children with asthma have poor inhaler technique, with detrimental morbidity effects. Guidelines recommend clinicians provide inhaler education at every opportunity, yet resources are limited. A low-cost, technology-based intervention-Virtual Teach-to-Goal (V-TTG)-was developed to deliver tailored inhaler technique education with high fidelity., Objective: To evaluate whether V-TTG leads to less inhaler misuse among children with asthma who are hospitalized vs brief intervention (BI, reading steps aloud)., Methods: A single-center randomized controlled trial of V-TTG vs BI was conducted with 5-to-10-year-old children with asthma hospitalized between January 2019 and February 2020. Inhaler technique was assessed pre- and post-education using 12-step validated checklists (misuse: < 10 steps correct)., Results: Among 70 children enrolled, mean age was 7.8 years (SD = 1.6). Most (86%) were Black. Most had an emergency department visit (94%) or hospitalization (90%) in the previous year. At baseline, nearly all children misused inhalers (96%). The proportion of children with inhaler misuse decreased significantly in V-TTG (100%→74%, P = .002) and BI (92%→69%, P = .04) groups, with no difference between groups at both time points (P = .2 and .9). On average, children performed 1.5 more steps correctly (SD = 2.0), with greater improvement with V-TTG (mean [SD] = 1.7 [1.6]) vs BI (mean [SD] = 1.4 [2.3]), though not significant (P = .6). Concerning pre and post technique, older children were significantly more likely than younger children to show more correct steps (mean change = 1.9 vs 1.1, P = .002)., Conclusion: A technology-based intervention for tailored inhaler education led to improved technique among children, similarly to reading steps aloud. Older children saw greater benefits. Future studies should evaluate the V-TTG intervention across diverse populations and disease severities to identify the greatest impact., Clinical Trial Registration: NCT04373499., (Copyright © 2023 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Background: The current methods of evaluating cognitive functioning typically rely on a single time point to assess and characterize an individual's performance. However, cognitive functioning fluctuates within individuals over time in relation to environmental, psychological, and physiological contexts. This limits the generalizability and diagnostic utility of single time point assessments, particularly among individuals who may exhibit large variations in cognition depending on physiological or psychological context (eg, those with type 1 diabetes [T1D], who may have fluctuating glucose concentrations throughout the day)., Objective: We aimed to report the reliability and validity of cognitive ecological momentary assessment (EMA) as a method for understanding between-person differences and capturing within-person variation in cognition over time in a community sample and sample of adults with T1D., Methods: Cognitive performance was measured 3 times a day for 15 days in the sample of adults with T1D (n=198, recruited through endocrinology clinics) and for 10 days in the community sample (n=128, recruited from TestMyBrain, a web-based citizen science platform) using ultrabrief cognitive tests developed for cognitive EMA. Our cognitive EMA platform allowed for remote, automated assessment in participants' natural environments, enabling the measurement of within-person cognitive variation without the burden of repeated laboratory or clinic visits. This allowed us to evaluate reliability and validity in samples that differed in their expected degree of cognitive variability as well as the method of recruitment., Results: The results demonstrate excellent between-person reliability (ranging from 0.95 to 0.99) and construct validity of cognitive EMA in both the sample of adults with T1D and community sample. Within-person reliability in both samples (ranging from 0.20 to 0.80) was comparable with that observed in previous studies in healthy older adults. As expected, the full-length baseline and EMA versions of TestMyBrain tests correlated highly with one another and loaded together on the expected cognitive domains when using exploratory factor analysis. Interruptions had higher negative impacts on accuracy-based outcomes (β=-.34 to -.26; all P values <.001) than on reaction time-based outcomes (β=-.07 to -.02; P<.001 to P=.40)., Conclusions: We demonstrated that ultrabrief mobile assessments are both reliable and valid across 2 very different clinic versus community samples, despite the conditions in which cognitive EMAs are administered, which are often associated with more noise and variability. The psychometric characteristics described here should be leveraged appropriately depending on the goals of the cognitive assessment (eg, diagnostic vs everyday functioning) and the population being studied., (©Shifali Singh, Roger Strong, Irene Xu, Luciana M Fonseca, Zoe Hawks, Elizabeth Grinspoon, Lanee Jung, Frances Li, Ruth S Weinstock, Martin J Sliwinski, Naomi S Chaytor, Laura T Germine. Originally published in the Journal of Medical Internet Research (https://www.jmir.org), 02.06.2023.)

Background: We aimed to develop an autophagy-related prognostic model with single-cell RNA sequencing (ScRNA-Seq) data for hepatocellular carcinoma (HCC) patients., Methods: ScRNA-Seq datasets of HCC patients were analyzed by Seurat. The expression of genes involved in canonical and noncanonical autophagy pathways in scRNA-seq data was also compared. Cox regression was applied to construct an AutRG risk prediction model. Subsequently, we examined the characteristics of AutRG high-risk and low-risk group patients., Results: Six major cell types (hepatocytes, myeloid cells, T/NK cells, B cells, fibroblast cells, and endothelial cells) were identified in the scRNA-Seq dataset. The results showed that most of the canonical and noncanonical autophagy genes were highly expressed in hepatocytes, with the exception of MAP 1LC3B, SQSTM1, MAP 1LC3A, CYBB, and ATG3. Six AutRG risk prediction models originating from different cell types were constructed and compared. The AutRG prognostic signature (GAPDH, HSP90AA1, and TUBA1C) in endothelial cells had the best overall performance for predicting the overall survival of HCC patients, with 1-year, 3-year, and 5-year AUCs equal to 0.758, 0.68, and 0.651 in the training cohort and 0.760, 0.796, and 0.840 in the validation cohort, respectively. The different tumor mutation burden, immune infiltration, and gene set enrichment characteristics of the AutRG high-risk and low-risk group patients were identified., Conclusion: We constructed an endothelial cell-related and autophagy-related prognostic model of HCC patients using the ScRNA-Seq dataset for the first time. This model demonstrated the good calibration ability of HCC patients and provided a new understanding of the evaluation of prognosis., (© 2023 Journal of Gastroenterology and Hepatology Foundation and John Wiley & Sons Australia, Ltd.)

Background: We sought to determine whether management of LDL-C following invasive angiography and assessment by fractional flow reserve (FFR) differs between those with obstructive vs non-obstructive CAD., Methods: Retrospective study of 721 patients undergoing coronary angiography involving assessment by FFR between 2013 and 2020 at a single academic center. Groups with obstructive vs non-obstructive CAD by index angiographic and FFR findings were compared over 1 year of follow-up., Results: Based on index angiographic and FFR findings, 421 (58%) patients had obstructive CAD vs 300 (42%) with non-obstructive CAD, mean (±SD) age 66±11 years, 217 (30%) women, and 594 (82%) white. There was no difference in baseline LDL-C. At 3-months follow-up, LDL-C was lower than baseline in both groups, with no between group difference. In contrast, at 6-months, median (Q1, Q3) LDL-C was significantly higher in non-obstructive vs obstructive CAD (LDL-C 73 (60, 93) vs 63 (48, 77) mg/dL, respectively ( p  = 0.003), ( p  = 0.001 in multivariable linear regression)). At 12-months, LDL-C remained higher in non-obstructive vs obstructive CAD (LDL-C 73 (49, 86) vs 64 (48, 79) mg/dL, respectively, although not statistically significant ( p  = 0.104)). The rate of high-intensity statin use was lower among those with non-obstructive CAD vs obstructive CAD at all time points ( p  < 0.05)., Conclusions: After coronary angiography involving FFR, there is intensification of LDL-C lowering at 3-months follow-up in both obstructive and non-obstructive CAD. However, by 6-months follow-up LDL-C is significantly higher among those with non-obstructive CAD vs obstructive CAD. Following coronary angiography involving FFR, patients with non-obstructive CAD may benefit from greater attention to LDL-C lowering to reduce residual ASCVD risk., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors. Published by Elsevier B.V.)

Diabetes mellitus (DM) is a metabolic disorder characterized by chronic hyperglycemia as a result of insufficient insulin levels and/or impaired function as a result of autoimmune destruction or insulin resistance. While Type 1 DM (T1DM) and Type 2 DM (T2DM) occur through different pathological processes, both result in β-cell destruction and/or dysfunction, which ultimately lead to insufficient β-cell mass to maintain normoglycemia. Therefore, therapeutic agents capable of inducing β-cell proliferation is crucial in treating and reversing diabetes; unfortunately, adult human β-cell proliferation has been shown to be very limited (~0.2% of β-cells/24 h) and poorly responsive to many mitogens. Furthermore, diabetogenic insults result in damage to β cells, making it ever more difficult to induce proliferation. In this review, we discuss β-cell mass/proliferation pathways dysregulated in diabetes and current therapeutic agents studied to induce β-cell proliferation. Furthermore, we discuss possible combination therapies of proliferation agents with immunosuppressants and antioxidative therapy to improve overall long-term outcomes of diabetes.

Sickle cell anemia (SCA) is characterized by decreased red blood cell (RBC) deformability due to polymerization of deoxygenated hemoglobin, leading to abnormal mechanical properties of RBC, increased cellular adhesion, and microcirculatory obstruction. Prior work has demonstrated that NO• influences RBC hydration and deformability and is produced at a basal rate that increases under shear stress in normal RBC. Nevertheless, the origin and physiological relevance of nitric oxide (NO•) production and scavenging in RBC remains unclear. We aimed to assess the basal and shear-mediated production of NO• in RBC from SCA patients and control (CTRL) subjects. RBCs loaded with a fluorescent NO• detector, DAF-FM (4-Amino-5-methylamino- 2',7'-difluorofluorescein diacetate), were imaged in microflow channels over 30-min without shear stress, followed by a 30-min period under 0.5Pa shear stress. We utilized non-specific nitric oxide synthase (NOS) blockade and carbon monoxide (CO) saturation of hemoglobin to assess the contribution of NOS and hemoglobin, respectively, to NO• production. Quantification of DAF-FM fluorescence intensity in individual RBC showed an increase in NO• in SCA RBC at the start of the basal period; however, both SCA and CTRL RBC increased NO• by a similar quantity under shear. A subpopulation of sickle-shaped RBC exhibited lower basal NO• production compared to discoid RBC from SCA group, and under shear became more circular in the direction of shear when compared to discoid RBC from SCA and CTRL, which elongated. Both CO and NOS inhibition caused a decrease in basal NO• production. Shear-mediated NO• production was decreased by CO in all RBC, but was decreased by NOS blockade only in SCA. In conclusion, total NO• production is increased and shear-mediated NO• production is preserved in SCA RBC in a NOS-dependent manner. Sickle shaped RBC with inclusions have higher NO• production and they become more circular rather than elongated with shear., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Fuchs endothelial corneal dystrophy (FECD) is characterized by a progressive loss of corneal endothelial cells (CECs) and an abnormal accumulation of extracellular matrix in Descemet's membrane leading to increased thickness and formation of excrescences called guttae. Extracellular matrix homeostasis is modulated by an equilibrium between matrix metalloproteinases (MMPs) and their endogenous tissue inhibitors (TIMPs). This study aimed to investigate MMPs and TIMPs profile in FECD, taking into account cell morphology. Populations of FECD and healthy CECs were cultured and their conditioned media collected for analysis. The presence of proteases in the conditioned media was studied using a semi-quantitative proteome profiler array, and MMPs levels were assessed using quantitative assays (ELISA and quantitative antibody array). MMP activity was determined by zymography and fluorometry. The expression pattern of the membrane type 1-MMP (MT1-MMP, also known as MMP-14) was examined by immunofluorescence on ex vivo FECD and healthy explants of CECs attached to Descemet's membrane. Finally, MMPs and TIMPs protein expression was compared to gene expression obtained from previously collected data. FECD and healthy CEC populations generated cultures of endothelial, intermediate, and fibroblastic-like morphology. Various MMPs (MMP-1, -2, -3, -7, -8, -9, -10, and -12) and TIMPs (TIMP-1 to -4) were detected in both FECD and healthy CECs culture supernatants. Quantitative assays revealed a decrease in MMP-2 and MMP-10 among FECD samples. Both these MMPs can degrade the main extracellular matrix components forming guttae (fibronectin, laminin, collagen IV). Moreover, MMPs/TIMPs ratio was also decreased among FECD cell populations. Activity assays showed greater MMPs/Pro-MMPs proportions for MMP-2 and MMP-10 in FECD cell populations, although overall activities were similar. Moreover, the analysis according to cell morphology revealed among healthy CECs, both increased (MMP-3 and -13) and decreased (MMP-1, -9, -10, and -12) MMPs proteins along with increased MMPs activity (MMP-2, -3, -9, and -10) in the fibroblastic-like subgroup when compared to the endothelial subgroup. However, FECD CECs did not show similar behaviors between the different morphology subgroups. Immunostaining of MT1-MMP on ex vivo FECD and healthy explants revealed a redistribution of MT1-MMP around guttae in FECD explants. At the transcriptional level, no statistically significant differences were detected, but cultured FECD cells had a 12.2-fold increase in MMP1 and a 4.7-fold increase in TIMP3. These results collectively indicate different, and perhaps pathological, MMPs and TIMPs profile in FECD CECs compared to healthy CECs. This is an important finding suggesting the implication of MMPs and TIMPs in FECD pathophysiology., (Copyright © 2021. Published by Elsevier Ltd.)

The NRF transcription factors NRF1, NRF2, and NRF3, are a subset of Cap'n'collar transcriptional regulators which modulate the expression of genes harboring antioxidant-response element (ARE) sequences within their genomic loci. Despite the emerging physiological importance of NRF family members, the repertoire of their genetic targets remains incompletely defined. Here we use RNA-sequencing-based transcriptional profiling and quantitative proteomics to delineate the overlapping and differential genetic programs effected by the three NRF transcription factors. We then create consensus target gene sets regulated by NRF1, NRF2, and NRF3 and define the integrity of these gene sets for probing NRF activity in mammalian cell culture and human tissues. Together, our data provide a quantitative assessment of how NRF family members sculpt proteomes and transcriptomes, providing a framework to understand the critical physiological importance of NRF transcription factors and to establish pharmacologic approaches for therapeutically activating these transcriptional programs in disease.

For the advancement of porcine xenotransplantation for clinical use in type 1 diabetes mellitus, the concerns of a sustainable and safe digestion enzyme blend must be overcome. Incorporating good manufacturing practices (GMP) can facilitate this through utilizing GMP-grade enzymes. In conjunction, still taking into account the cost-effectiveness, a wide concern. We evaluated how GMP-grade enzyme blends impact our piglet islets and their long-term effects.  Preweaned porcine islets (PPIs) were isolated from 8- to 10-day-old pigs. Digestion enzyme blends, collagenase type V (Type V), collagenase AF-1 GMP-grade with collagenase NB 6 GMP-grade (AF-1 and NB 6), and collagenase AF-1 GMP-grade with collagenase neutral protease AF GMP-grade (AF-1 and NP AF) were compared. Islet quality control assessments, islet yield, viability, and function, were performed on days 3 and 7, and cell content was performed on day 7.  GMP-grade AF-1 and NB 6 (17,209 ± 2,730 islet equivalent per gram of pancreatic tissue [IE/g] on day 3, 9,001 ± 1,034 IE/g on day 7) and AF-1 and NP AF (17,214 ± 3,901 IE/g on day 3, 8,833 ± 2,398 IE/g on day 7) showed a significant increase in islet yield compared to Type V (4,618 ± 1,240 IE/g on day 3, 1,923 ± 704 IE/g on day 7). Islet size, viability, and function showed comparable results in all enzyme blends. There was no significant difference in islet cellular content between enzyme blends.  This study demonstrated a comparison of GMP-grade collagenase enzyme blends and a standard crude collagenase enzyme in preweaned-aged porcine, a novel topic in this age. GMP-grade enzyme blends of AF-1 and NB 6 and AF-1 and NP AF resulted in substantially higher yields and as effective PPIs compared to Type V. In the long run, considering costs, integrity, and sustainability, GMP-grade enzyme blends are more favorable for clinical application due to high reproducibility in comparison to undefined manufacturing processes of standard enzymes.

The corneal endothelium forms a leaky barrier between the corneal stroma and the aqueous humor of the anterior chamber. This cell monolayer maintains the corneal stroma in a state of relative dehydration, a process called deturgescence, which is required in order to obtain corneal stromal transparency. Endothelial dysfunctions lead to visual impairment that ultimately can only be treated surgically via the corneal transplantation of a functional endothelium. Shortages of corneas suitable for transplantation has motivated research toward new alternatives involving in vitro corneal endothelial cell (CEC) expansion.This chapter describes current methods that allow isolate and culture CECs. In brief, Descemet membrane is peeled out of the cornea and digested in order to obtain CECs. Cells are then seeded and cultured.

Hypoxia is commonly found in cancers. Hypoxia, due to the lack of oxygen (O 2 ) as the electron recipient, causes inefficient electron transfer through the electron transport chain at the mitochondria leading to accumulation of reactive oxygen species (ROS) which could create irreversible cellular damages. Through hypoxia-inducible factor 1 (HIF-1) which elicits various molecular events, cells are able to overcome low O 2 . Knowledge about the new molecular mechanisms governed by HIF-1 is important for new therapeutic interventions targeting hypoxic tumors. Using hepatocellular carcinoma (HCC) as a model, we revealed that the HIF-1 and the Notch signaling pathways cross-talk to control mitochondrial biogenesis of cancer cells to maintain REDOX balance. From transcriptome sequencing, we found that HEY1, a transcriptional repressor, in the NOTCH pathway was consistently induced by hypoxia in HCC cell lines. We identified a strong hypoxia response element (HRE) in HEY1 by chromatin immunoprecipitation (ChIP) and luciferase reporter assays. Transcriptome and ChIP sequencing further identified PINK1, a gene essential for mitochondrial biogenesis, as a novel transcriptional target of HEY1. HCC cells with HEY1 knockdown re-expressed PINK1. HEY1 and PINK1 expressions inversely correlated in human HCC samples. Overexpression of HEY1 and under-expression of PINK1 were detected in human HCC and associated with poor clinical outcomes. Functionally, we found that overexpression of HEY1 or knockdown of PINK1 consistently reduced mitochondrial cristae, mitochondrial mass, oxidative stress level, and increased HCC growth.

A major goal of T1D research is to develop new approaches to increase β -cell mass and control autoreactive T cell responses. GABA A -receptors (GABA A -Rs) are promising drug targets in both those regards due to their abilities to promote β -cell replication and survival, as well as inhibit autoreactive T cell responses. We previously showed that positive allosteric modulators (PAMs) of GABA A -Rs could promote rat β -cell line INS-1 and human islet cell replication in vitro . Here, we assessed whether treatment with alprazolam, a widely prescribed GABA A -R PAM, could promote β -cell survival and replication in human islets after implantation into NOD/scid mice. We observed that alprazolam treatment significantly reduced human islet cell apoptosis following transplantation and increased β -cell replication in the xenografts. Evidently, the GABA A -R PAM works in conjunction with GABA secreted from β -cells to increase β -cell survival and replication. Treatment with both the PAM and GABA further enhanced human β -cell replication. Alprazolam also augmented the ability of suboptimal doses of GABA to inhibit antigen-specific T cell responses in vitro . Thus, combined GABA A -R agonist and PAM treatment may help control inflammatory immune responses using reduced drug dosages. Together, these findings suggest that GABA A -R PAMs represent a promising drug class for safely modulating islet cells toward beneficial outcomes to help prevent or reverse T1D and, together with a GABA A -R agonist, may have broader applications for ameliorating other disorders in which inflammation contributes to the disease process.

Achilles tendon vibration generates proprioceptive information that is incongruent with the actual body position; it alters the perception of body orientation leading to a vibration-induced postural response. When a person is standing freely, vibration of the Achilles tendon shifts the internal representation of the verticality backward thus the vibration-induced postural response realigned the whole body orientation with the shifted subjective vertical. Because utricular otoliths information participates in the creation of the internal representation of the verticality, changing the natural orientation of the head-neck system during Achilles tendon vibration could alter the internal representation of the earth vertical to a greater extent. Consequently, it was hypothesized that compared to neutral head-neck orientation, alteration in the head-neck orientation should impair balance control immediately after Achilles tendon vibration onset or offset (i.e., sensory transition) as accurate perception of the earth vertical is required. Results revealed that balance control impairment was observed only immediately following Achilles tendon vibration offset; both groups with the head-neck either extended or flexed showed larger body sway (i.e., larger root mean square scalar distance between the center of pressure and center of gravity) compared to the group with the neutral head-neck orientation. The fact that balance control was uninfluenced by head-neck orientation immediately following vibration onset suggests the error signal needs to accumulate to a certain threshold before the internal representation of the earth vertical becomes incorrect., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Transforming growth factor β (TGF-β) physiologically induces Epstein-Barr virus (EBV) lytic infection by activating the expression of EBV's latent-lytic switch BZLF1 gene. Liang et al. (J. Biol. Chem. 277:23345-23357, 2002) previously identified a Smad-binding element (SBE) within the BZLF1 promoter, Zp; however, it accounts for only 20 to 30% of TGF-β-mediated activation of transcription from Zp. Here, we identified additional factors responsible for the rest of this activation. The incubation of EBV-positive MutuI cells with a TGF-β neutralizing antibody or inhibitors of the TGF-β type I receptor (TβRI) or Smad3 eliminated the TGF-β-induced reactivation of EBV. The coexpression of Smad2, Smad3, and Smad4 together with a constitutively active form of TβRI induced 15- to 25-fold transcription from Zp in gastric carcinoma AGS cells. By electrophoretic mobility shift assays, we identified four additional Smad-binding elements, named SBE2 to SBE5. Substitution mutations in individual SBEs reduced Smad-mediated activation of Zp by 20 to 60%; together, these mutations essentially eliminated it. Chromatin immunoprecipitation assays confirmed that Smad4 newly bound the Zp region of the EBV genome following the incubation of MutuI cells with TGF-β. SBE2 overlaps the ZEB-binding ZV silencing element of Zp. Depending upon posttranslational modifications, Smad4 either competed with ZEB1 for binding or formed a complex with ZEB1 on the Zp ZV element in a cell-free assay system. In transiently transfected cells, exogenously expressed ZEB1 inhibited Smad-mediated transcriptional activation from Zp. We conclude that TGF-β induces EBV lytic reactivation via the canonical Smad pathway by activating BZLF1 gene expression through multiple SBEs acting in concert.

An RRLC method capable of simultaneous identification and rapid quantification of six biologically active compounds (salidroside, tyrosol, rosarin, rosavin, rosin, rosiridin) in Rhodiola rosea L. and two active compounds (eleutheroside B and eleutheroside E) in Eleutherococcus senticosus Maxim. was developed. The chromatographic analyses were performed on a reversed phase Phenomenex C18 (2)-HST column at 40°C with a neutral mobile phase (purified water and acetonitrile) gradient system at a flow rate of 1.0ml/min and UV detection at 205 and 220nm simultaneously. Baseline separation of eight active compounds was achieved within 8min. This developed method provides good linearity (R>0.9997), precision (RSD<1.99%) and recovery of the bioactive compounds. The RRLC method developed is capable of controlling the quality of R. rosea and E. senticosus raw herbs, commercial extracts, as well as polyherbal formulations containing R. rosea and E. senticosus as ingredients. This RRLC method is accurate and sensitive; in addition, it greatly increases sample analysis throughput with reduced analysis time, which is suitable for routine quality control analysis., (Copyright © 2011 Elsevier B.V. All rights reserved.)

A simple, sensitive and reliable reversed phase Rapid Resolution Liquid Chromatography (RRLC) method was developed and validated for six biologically active compounds (salidroside, tyrosol, rosarin, rosavin, rosin and rosiridin) in Rhodiola rosea L. roots and powder extracts. The method uses a Phenomenex C18 (2)-HST column at 40 degrees C with a neutral gradient system mobile phase (H20 and acetonitrile), a flow rate of 1.0 mL/min, and UV detection wavelengths set at 205 and 254 nm, simultaneously. Baseline separation of the six active compounds was achieved within 8 minutes. The average percentages of rosavins (rosarin, rosavin, and rosin) in authentic R. rosea roots and root powder extracts were quantitatively determined and a characteristic R. rosea roots RRLC profile was established. The RRLC method is accurate and sensitive; in addition, it effectively increases the sample analysis throughput compared with conventional HPLC.

Shuang-Huang-Lian (SHL) is a traditional Chinese formula which comprises of three medicinal herbs: Flos Lonicerae, Radix Scutellariae and Fructus Forsythiae, and is commonly used to treat acute upper respiratory tract infection, acute bronchitis and light pneumonia. A simple, reliable and reproducible rapid resolution liquid chromatography (RRLC) method was developed for the quality control of SHL preparations, which baseline separates the major bioactive compounds within 6min. The method uses a C18-HST column (2.5μm, 100mm×3.0mm) kept at 40°C. The mobile phases consist of 0.1% phosphoric acid aqueous solution and acetonitrile. Flow rate is 1.0ml/min and UV detection is performed at 327nm from 0 to 4min and 229nm from 4 to 7min. This method was further validated according to the ICH guidelines. Eight batches of commercial SHL preparations obtained from different pharmaceutical manufacturers as well as individual herbs were examined and their chromatographic profiles were compared. The stability test revealed that chlorogenic acid is stable only at acidic pH, and hence it is necessary to further evaluate and optimize the preparatory procedures and storage conditions for commercial SHL preparations., (Copyright © 2010 Elsevier B.V. All rights reserved.)

We previously showed that the cellular proteins ZEB1 and ZEB2/SIP1 both play key roles in regulating the latent-lytic switch of Epstein-Barr Virus (EBV) by repressing BZLF1 gene expression. We investigated here the effects of cellular microRNA (miRNA) 200 (miR200) family members on the EBV infection status of cells. We show that miR200b and miR429, but not miR200a, can induce EBV-positive cells into lytic replication by downregulating expression of ZEB1 and ZEB2, leading to production of infectious virus. The levels of miR200 family members in EBV-infected cells strongly negatively correlated with the levels of the ZEBs (e.g., -0.89 [P < 0.001] for miR429 versus ZEB1) and positively correlated with the degree of EBV lytic gene expression (e.g., 0.73 [P < 0.01] for miR429 versus BZLF1). The addition of either miR200b or miR429 to EBV-positive cells led to EBV lytic reactivation in a ZEB-dependent manner; inhibition of these miRNAs led to decreased EBV lytic gene expression. The degree of latent infection by an EBV mutant defective in the primary ZEB-binding site of the EBV BZLF1 promoter was not affected by the addition of these miRNAs. Furthermore, EBV infection of primary blood B cells led to downregulation of these miRNAs and upregulation of ZEB levels. Thus, we conclude that miRNAs 200b and 429 are key regulators via their effects on expression of ZEB1 and ZEB2 of the switch between latent and lytic infection by EBV and, therefore, potential targets for development of new lytic induction therapeutics with which to treat patients with EBV-associated malignancies.

For most systems in biology, a large body of literature exists that describes the complexity of the system based on experimental results. Manual review of this literature to extract targeted information into biological databases is difficult and time consuming. To address this problem, we developed PubSearch and PubFetch, which store literature, keyword, and gene information in a relational database, index the literature with keywords and gene names, and provide a Web user interface for annotating the genes from experimental data found in the associated literature. A set of protocols is provided in this unit for installing, populating, running, and using PubSearch and PubFetch. In addition, we provide support protocols for performing controlled vocabulary annotations. Intended users of PubSearch and PubFetch are database curators and biology researchers interested in tracking the literature and capturing information about genes of interest in a more effective way than with conventional spreadsheets and lab notebooks.

Controlled vocabularies are increasingly used by databases to describe genes and gene products because they facilitate identification of similar genes within an organism or among different organisms. One of The Arabidopsis Information Resource's goals is to associate all Arabidopsis genes with terms developed by the Gene Ontology Consortium that describe the molecular function, biological process, and subcellular location of a gene product. We have also developed terms describing Arabidopsis anatomy and developmental stages and use these to annotate published gene expression data. As of March 2004, we used computational and manual annotation methods to make 85,666 annotations representing 26,624 unique loci. We focus on associating genes to controlled vocabulary terms based on experimental data from the literature and use The Arabidopsis Information Resource-developed PubSearch software to facilitate this process. Each annotation is tagged with a combination of evidence codes, evidence descriptions, and references that provide a robust means to assess data quality. Annotation of all Arabidopsis genes will allow quantitative comparisons between sets of genes derived from sources such as microarray experiments. The Arabidopsis annotation data will also facilitate annotation of newly sequenced plant genomes by using sequence similarity to transfer annotations to homologous genes. In addition, complete and up-to-date annotations will make unknown genes easy to identify and target for experimentation. Here, we describe the process of Arabidopsis functional annotation using a variety of data sources and illustrate several ways in which this information can be accessed and used to infer knowledge about Arabidopsis and other plant species.

Arabidopsis thaliana is the most widely-studied plant today. The concerted efforts of over 11 000 researchers and 4000 organizations around the world are generating a rich diversity and quantity of information and materials. This information is made available through a comprehensive on-line resource called the Arabidopsis Information Resource (TAIR) (http://arabidopsis.org), which is accessible via commonly used web browsers and can be searched and downloaded in a number of ways. In the last two years, efforts have been focused on increasing data content and diversity, functionally annotating genes and gene products with controlled vocabularies, and improving data retrieval, analysis and visualization tools. New information include sequence polymorphisms including alleles, germplasms and phenotypes, Gene Ontology annotations, gene families, protein information, metabolic pathways, gene expression data from microarray experiments and seed and DNA stocks. New data visualization and analysis tools include SeqViewer, which interactively displays the genome from the whole chromosome down to 10 kb of nucleotide sequence and AraCyc, a metabolic pathway database and map tool that allows overlaying expression data onto the pathway diagrams. Finally, we have recently incorporated seed and DNA stock information from the Arabidopsis Biological Resource Center (ABRC) and implemented a shopping-cart style on-line ordering system.

The Arabidopsis Information Resource (TAIR; http://arabidopsis.org) provides an integrated view of genomic data for Arabidopsis thaliana. The information is obtained from a battery of sources, including the Arabidopsis user community, the literature, and the major genome centers. Currently TAIR provides information about genes, markers, polymorphisms, maps, sequences, clones, DNA and seed stocks, gene families and proteins. In addition, users can find Arabidopsis publications and information about Arabidopsis researchers. Our emphasis is now on incorporating functional annotations of genes and gene products, genome-wide expression, and biochemical pathway data. Among the tools developed at TAIR, the most notable is the Sequence Viewer, which displays gene annotation, clones, transcripts, markers and polymorphisms on the Arabidopsis genome, and allows zooming in to the nucleotide level. A tool recently released is AraCyc, which is designed for visualization of biochemical pathways. We are also developing tools to extract information from the literature in a systematic way, and building controlled vocabularies to describe biological concepts in collaboration with other database groups. A significant new feature is the integration of the ABRC database functions and stock ordering system, which allows users to place orders for seed and DNA stocks directly from the TAIR site.

Peptide nucleic acids (PNA) are nucleic acid analogues containing neutral amide backbone, forming stable and tight complexes with complementary DNA/RNA. However, it is unclear whether unmodified PNA can efficiently penetrate neuronal tissue in order to act as antisense reagent. Here we show that intrathecal (i.t.) injection of an unmodified antisense PNA complementary to the rat galanin receptor type 1 (GalR1) mRNA is able to block the inhibitory effect of i.t. administered galanin on spinal nociceptive transmission. Autoradiographic ligand binding studies using [125I]galanin show that the unmodified PNA is able to reduce the density of galanin binding sites in the dorsal horn. Thus, unmodified PNA applied i.t. appears to function as an effective antisense reagent in rat spinal cord in vivo.

SH3 domains are a conserved feature of many nonreceptor protein tyrosine kinases, such as Hck, and often function in substrate recruitment and regulation of kinase activity. SH3 domains modulate kinase activity by binding to polyproline helices (PPII helix) either intramolecularly or in target proteins. The preponderance of bimolecular and distal interactions between SH3 domains and PPII helices led us to investigate whether proximal placement of a PPII helix relative to an SH3 domain would result in tight, intramolecular binding. We have fused the PPII helix region of human GAP to the C-terminus of Hck SH3 and expressed the recombinant fusion protein in Escherichia coli. The fusion protein, SH3Hck : PPIIhGAP, folded spontaneously into a structure in which the PPII helix was bound intramolecularly to the hydrophobic crevice of the SH3 domain. The SH3Hck : PPIIhGAP fusion protein is useful for investigating SH3: PPII helix interactions, for studying concepts in protein folding and design, and may represent a protein structural motif that is widely distributed in nature.

Nociceptin and its receptor are present in dorsal spinal cord, indicating a possible role for this peptide in pain transmission. The majority of functional studies using behavioral and electrophysiological studies have shown that nociceptin applied at spinal level produces antinociception through pre- and post-synaptic mechanisms. The spinal inhibitory effect of nociceptin is not sensitive to antagonists of opioid receptors such as naloxone. Thus, nociceptin-induced antinociception is mediated by a novel mechanism independent of activation of classic opioid receptors. This has raised the possibility that agonists of the nociceptin receptor may represent a novel class of analgesics. Supporting this hypothesis, several groups have shown that intrathecal nociceptin alleviated hyperalgesic and allodynic responses in rats after inflammation or partial peripheral nerve injury. Electrophysiological studies have also indicated that the antinociceptive potency of spinal nociceptin is maintained or enhanced after nerve injury. It is concluded that the predominant action of nociceptin in the spinal cord appears to be inhibitory. The physiological role of nociceptin in spinal nociceptive mechanisms remains to be defined. Moreover, further evaluation of nociceptin as a new analgesic calls the development of non-peptide brain penetrating agents.

We studied the effects of intrathecal (i.t.) nocistatin, a peptide identified from the precursor of orphanin FQ/nociceptin (OFQ) on the spinal nociceptive flexor reflex in decerebrate, spinalized, unanesthetized rats and its interaction with i.t. OFQ. Nocistatin induced a moderate, non-dose-dependent facilitation of the flexor reflex without producing reflex depression whereas i.t. OFQ induced a biphasic dose-dependent facilitatory and inhibitory effect. The facilitatory effect of low dose (0.55 pmol) OFQ was significantly increased by nocistatin. On the other hand, the duration, but not magnitude, of reflex depression induced by a high (550 pmol) dose of OFQ was significantly shortened by 5.5 nmol nocistatin. Thus, nocistatin interacts with OFQ in a complex fashion, increasing excitation and reducing inhibition. No evidence was obtained for an antinociceptive effect of nocistatin in rat spinal cord.

We have examined the effects of intrathecal (i.t.) administration of [Leu31,Pro34]-neuropeptide Y (NPY) or NPY-(13-36), selective agonists of NPY Y1 or Y2 receptors, respectively, on the excitability of the flexor reflex in normal rats and after unilateral transection of the sciatic nerve. In rats with intact and sectioned sciatic nerves, i.t. [Leu31,Pro34]-NPY induced a similar biphasic effect on the flexor reflex with facilitation at low doses and facilitation followed by depression at high doses. In contrast, i.t. NPY-(13-36) only facilitated the flexor reflex in normal rats, and at high dose it caused ongoing discharges in the electromyogram. NPY-(13-36) caused dose-dependent depression of the flexor reflex in rats after sciatic nerve transection, in addition to its facilitatory effect. Topical application of [Leu31,Pro34]-NPY or NPY-(13-36) caused a moderate and brief reduction in spinal cord blood flow. No difference was noted between the vasoconstrictive effect of [Leu31,Pro34]-NPY and NPY-(13-36). It is suggested that activation of Y1 receptors may be primarily responsible for the reflex depressive effect of i.t. neuropeptide Y in rats with intact sciatic nerves, whereas both Y1 and Y2 receptors may be involved in mediating the depressive effect of NPY after axotomy., (Copyright 1999 Elsevier Science B.V.)