Increased Expression of Cell-Cell Signaling Genes by Stimulated Mononuclear Leukocytes in Patients with Previous Atherothrombotic Stroke A Whole Genome Expression Profile Study

Background/Aims: Inflammation plays an important role in atherosclerosis and stroke. Acute infections are recognized as trigger factors for ischemic stroke. Methods: In this whole genome expression profile study of 15 patients and 15 control subjects, we tested the hypothesis that patients with a history of atherothrombotic stroke show enhanced transcription of inflammatory genes in circulating leukocytes. RNA from unstimulated or lipopolysaccharide (LPS)-stimulated peripheral blood mononuclear cells (PBMCs) was analyzed with Affymetrix U133A GeneChips using a pooling design. Expression of single genes and functional groups of genes was analyzed by global statistical tests. Results: A total of 10,197 probe sets showed positive calls. After correction for multiple testing no single probe set revealed significant differences either without or with LPS stimulation. However, significant global expression differences were found upon LPS stimulation for the group of genes that are involved in cell-cell signaling. Conclusion: LPS stimulation of PBMCs, a condition mimicking bacterial infection, induces differential expression of a group of cell-cell signaling genes in patients with previous atherothrombotic stroke. This finding can be caused by genetic differences between both groups, but acquired risk factors, medication and technical factors may also have contributed to the result. Copyright (C) 2009 S. Karger AG, Base

Contextual factors and programme theories associated with implementing blue prescription programmes: a systematic realist review

Nature-based social prescribing such as “blue prescription” promotes public health and health improvement of individuals with long-term health conditions. However, there is limited evidence explaining the relationship of contexts, mechanisms, and outcomes of implementing blue prescription programmes (BPPs) in health and social care settings that could inform policy and practice. We conducted a systematic realist review by searching PubMed, Web of Science, PsycInfo, Scopus, MEDLINE, and CINAHL for articles published in English between January 2000 and June 2022 about health and social care professionals providing referral to or prescription of blue space activities (e.g., swimming, fishing, surfing, etc.) with health-related outcomes. Components and descriptions of BPP implementation were extracted and used to develop themes of contextual factors used to develop programme theories and a logic model demonstrating the mechanisms of BPP implementation. Sixteen studies with adequate to strong quality were included from 8,619 records. After participating in BPPs referred to or prescribed by health and social care professionals, service users had improvements in their physical, cognitive (mental), social health, and proenvironmental knowledge. Service user-related contextual factors were referral information, free equipment, transportation, social support, blue space environments, and skills of service providers. Programme-related contextual factors were communication, multistakeholder collaboration, financing, and adequate service providers. Programme theories on service user enrolment, engagement, adherence, communication protocols, and programme sustainability explain the mechanisms of BPP implementation. BPPs could promote health and wellbeing if contextual factors and programme theories associated with service users’ characteristics and programme delivery are considered in the design, delivery, and evaluation of BPPs. Our study was registered with PROSPERO (CRD42020170660)

Seasonal Variation of the Effect of Extremely Diluted Agitated Gibberellic Acid (10e-30) on Wheat Stalk Growth: A Multiresearcher Study

The influence of a homeopathic high dilution of gibberellic acid on wheat growth was studied at different seasons of the year. Seedlings were allowed to develop under standardized conditions for 7 days; plants were harvested and stalk lengths were measured. The data obtained confirm previous findings, that ultrahigh diluted potentized gibberellic acid affects stalk growth. Furthermore, the outcome of the study suggests that experiments utilizing the bioassay presented should best be performed in autumn season. In winter and spring, respectively, no reliable effects were found

Nos2 Inactivation Promotes the Development of Medulloblastoma in Ptch1+/− Mice by Deregulation of Gap43–Dependent Granule Cell Precursor Migration

Medulloblastoma is the most common malignant brain tumor in children. A subset of medulloblastoma originates from granule cell precursors (GCPs) of the developing cerebellum and demonstrates aberrant hedgehog signaling, typically due to inactivating mutations in the receptor PTCH1, a pathomechanism recapitulated in Ptch1+/− mice. As nitric oxide may regulate GCP proliferation and differentiation, we crossed Ptch1+/− mice with mice lacking inducible nitric oxide synthase (Nos2) to investigate a possible influence on tumorigenesis. We observed a two-fold higher medulloblastoma rate in Ptch1+/− Nos2−/− mice compared to Ptch1+/− Nos2+/+ mice. To identify the molecular mechanisms underlying this finding, we performed gene expression profiling of medulloblastomas from both genotypes, as well as normal cerebellar tissue samples of different developmental stages and genotypes. Downregulation of hedgehog target genes was observed in postnatal cerebellum from Ptch1+/+ Nos2−/− mice but not from Ptch1+/− Nos2−/− mice. The most consistent effect of Nos2 deficiency was downregulation of growth-associated protein 43 (Gap43). Functional studies in neuronal progenitor cells demonstrated nitric oxide dependence of Gap43 expression and impaired migration upon Gap43 knock-down. Both effects were confirmed in situ by immunofluorescence analyses on tissue sections of the developing cerebellum. Finally, the number of proliferating GCPs at the cerebellar periphery was decreased in Ptch1+/+ Nos2−/− mice but increased in Ptch1+/− Nos2−/− mice relative to Ptch1+/− Nos2+/+ mice. Taken together, these results indicate that Nos2 deficiency promotes medulloblastoma development in Ptch1+/− mice through retention of proliferating GCPs in the external granular layer due to reduced Gap43 expression. This study illustrates a new role of nitric oxide signaling in cerebellar development and demonstrates that the localization of pre-neoplastic cells during morphogenesis is crucial for their malignant progression

Agonist-Independent Interactions between β-Arrestins and Mutant Vasopressin Type II Receptors Associated with Nephrogenic Syndrome of Inappropriate Antidiuresis

Nephrogenic syndrome of inappropriate antidiuresis is a recently identified genetic disease first described in two unrelated male infants with severe symptomatic hyponatremia. Despite undetectable arginine vasopressin levels, patients have inappropriately concentrated urine resulting in hyponatremia, hypoosmolality, and natriuresis. It was found that each infant had a different mutation of the vasopressin type II receptor (V2R) at codon 137 where arginine was converted to cysteine or leucine (R137C or R137L), resulting in constitutive signaling. Interestingly, a missense mutation at the same codon, converting arginine to histidine (R137H), leads to the opposite disease phenotype with a loss of the kidney’s ability to concentrate urine resulting in nephrogenic diabetes insipidus. This mutation is associated with impaired signaling, although whether this is predominantly due to impaired trafficking to the plasma membrane, agonist-independent internalization, or G protein uncoupling is currently unclear. Using bioluminescence resonance energy transfer and confocal microscopy, we demonstrate that both V2R-R137C and V2R-R137L mutants interact with β-arrestins in an agonist-independent manner resulting in dynamin-dependent internalization. This phenotype is similar to that observed for V2R-R137H, which is intriguing considering that it is accompanied by constitutive rather than impaired signaling. Consequently, it would seem that agonist-independent internalization per se is unlikely to be the major determinant of impaired V2R-R137H signaling. Our findings indicate that the V2R-R137C and V2R-R137L mutants traffic considerably more efficiently to the plasma membrane than V2R-R137H, identifying this as a potentially important mutation-dependent difference affecting V2R function

Helix I of β-Arrestin Is Involved in Postendocytic Trafficking but Is Not Required for Membrane Translocation, Receptor Binding, and Internalization

{beta}-arrestins bind to phosphorylated, seven-transmembrane-spanning, G protein-coupled receptors (GPCRs), including the type 1 angiotensin II receptor (AT1R), to promote receptor desensitization and internalization. The AT1 R is a class B GPCR that recruits both {beta}-arrestin1 and {beta}-arrestin2, forming stable complexes that cotraffic to deep-core endocytic vesicles. {beta}-Arrestins contain one amphipathic and potentially amphitropic (membrane-targeting) {alpha}-helix (helix I) that may promote translocation to the membrane or influence receptor internalization or trafficking. Here, we investigated the trafficking and function of {beta}-arrestin1 and {beta}-arrestin2 mutants bearing substitutions in both the hydrophobic and positively charged faces of helix I. The level of expression of these mutants and their cytoplasmic localization (in the absence of receptor activation) was similar to wild-type {beta}-arrestins. After angiotensin II stimulation, both wild-type and {beta}-arrestin mutants translocated to the cell membrane, although recruitment was weaker for mutants of the hydrophobic face of helix I. For all {beta}-arrestin mutants, the formation of deep-core vesicles was less observed compared with wild-type {beta}-arrestins. Furthermore, helix I conjugated to green fluorescent protein is not membrane-localized, suggesting that helix I, in isolation, is not amphitropic. Bioluminescence resonance energy transfer analysis revealed that both wild-type and {beta}-arrestin mutants retained a capacity to interact with the AT1R, although the interaction with the mutants was less stable. Finally, wild-type and mutant {beta}-arrestins fully supported receptor internalization in human embryonic kidney cells and mouse embryonic fibroblasts deficient in {beta}-arrestin1 and -2. Thus, helix I is implicated in postmembrane trafficking but is not strongly amphitropic

Application of G Protein-Coupled Receptor-Heteromer Identification Technology to Monitor β-Arrestin Recruitment to G Protein-Coupled Receptor Heteromers

Understanding the role of G protein-coupled receptor (GPCR; also known as a 7 transmembrane receptor) heteromerization in the physiology and pathophysiology of cellular function has now become a major research focus. However, there is currently a lack of cell-based assays capable of profiling the specific functional consequences of heteromerization in a ligand-dependent manner. Understanding the pharmacology specifically associated with heteromer function in contrast to monomer or homomer function enables the so-called biochemical fingerprints of the receptor heteromer to be ascertained. This is the first step in establishing the physiological relevance of heteromerization, the goal of everyone in the field, as these fingerprints can then be utilized in future endeavors to elucidate heteromer function in native tissues. The simple, robust, ligand-dependent methodology described in this study utilizes a novel configuration of components of a proximity-based reporter system. This is exemplified by the use of bioluminescence resonance energy transfer due to the advantages of real-time live cell monitoring of proximity specifically between the heteromer complex and a protein that is recruited in a ligand-dependent manner, in this case, β-arrestin 2. Further, the demonstration of Z′-factor values in excess of 0.6 shows the potential of the method for screening compounds for heteromer-selective or biased activity. Three previously characterized GPCR heteromers, the chemokine receptor heteromers CCR2-CCR5 and CCR2-CXCR4, as well as the angiotensin II receptor type 1-bradykinin receptor type 2 heteromer, have been used to illustrate the profiling capability and specificity of the GPCR heteromer identification technology