CD69 is a TGF-β/1α,25-dihydroxyvitamin D3 target gene in monocytes

CD69 is a transmembrane lectin that can be expressed on most hematopoietic cells. In monocytes, it has been functionally linked to the 5-lipoxygenase pathway in which the leukotrienes, a class of highly potent inflammatory mediators, are produced. However, regarding CD69 gene expression and its regulatory mechanisms in monocytes, only scarce data are available. Here, we report that CD69 mRNA expression, analogous to that of 5-lipoxygenase, is induced by the physiologic stimuli transforming growth factor-β (TGF-β) and 1α,25-dihydroxyvitamin D3 (1α,25(OH)2D3) in monocytic cells. Comparison with T- and B-cell lines showed that the effect was specific for monocytes. CD69 expression levels were increased in a concentration-dependent manner, and kinetic analysis revealed a rapid onset of mRNA expression, indicating that CD69 is a primary TGF-β/1α,25(OH)2D3 target gene. PCR analysis of different regions of the CD69 mRNA revealed that de novo transcription was initiated and proximal and distal parts were induced concomitantly. In common with 5-lipoxygenase, no activation of 0.7 kb or ~2.3 kb promoter fragments by TGF-β and 1α,25(OH)2D3 could be observed in transient reporter assays for CD69. Analysis of mRNA stability using a transcription inhibitor and a 3′UTR reporter construct showed that TGF-β and 1α,25(OH)2D3 do not influence CD69 mRNA stability. Functional knockdown of Smad3 clearly demonstrated that upregulation of CD69 mRNA, in contrast to 5-LO, depends on Smad3. Comparative studies with different inhibitors for mitogen activated protein kinases (MAPKs) revealed that MAPK signalling is involved in CD69 gene regulation, whereas 5-lipoxygenase gene expression was only partly affected. Mechanistically, we found evidence that CD69 gene upregulation depends on TAK1-mediated p38 activation. In summary, our data indicate that CD69 gene expression, conforming with 5-lipoxygenase, is regulated monocyte-specifically by the physiologic stimuli TGF-β and 1α,25(OH)2D3 on mRNA level, although different mechanisms account for the upregulation of each gene

Advancing Equity in Accessibility and Travel Experiences: The Role of Gender and Identity

(c)1036342 (wo) 25Gender can have a significant influence on people\u2019s behaviors and experiences. Hence, excluding gender diversity in transportation research and practices can result in biased or incomplete understandings of issues and perceptions about transportation and quality of life. This study examined whether and how gender, in a broader sense, can result in distinctly different activity-travel patterns and subjective well-being (SWB) outcomes using survey data. The study reviewed existing literature and found that gender was not binary meaning that some gender identities were not solely female or male. The literature also indicated a person\u2019s gender typically intersected with their other social identities such as race and family type and created unique needs and experiences. To address the complex nature of gender, the team collected new data using the Daynamica smartphone application and included specific questions concerning (1) participants\u2019 gender identities and attitudes toward gender roles, (2) their share of household-supporting tasks in 14-day travel diaries, and (3) their emotions during trips and activity participation. The team used 2021 Daynamica survey data and 2019 Travel Behavior Inventory data from the Metropolitan Council to extract activity-travel patterns before and after COVID-19. The team associated these patterns with participants\u2019 gender and other identities and SWB outcomes through visual explorations and statistical analysis. The findings suggested the importance of capturing the complex, intersectional nature of gender, confirmed the persistent existence of gender differences in transportation needs, experiences, and SWB outcomes in Minnesota

Perioperative mental health intervention for depression and anxiety symptoms in older adults study protocol: Design and methods for three linked randomised controlled trials

INTRODUCTION: Preoperative anxiety and depression symptoms among older surgical patients are associated with poor postoperative outcomes, yet evidence-based interventions for anxiety and depression have not been applied within this setting. We present a protocol for randomised controlled trials (RCTs) in three surgical cohorts: cardiac, oncological and orthopaedic, investigating whether a perioperative mental health intervention, with psychological and pharmacological components, reduces perioperative symptoms of depression and anxiety in older surgical patients. METHODS AND ANALYSIS: Adults ≥60 years undergoing cardiac, orthopaedic or oncological surgery will be enrolled in one of three-linked type 1 hybrid effectiveness/implementation RCTs that will be conducted in tandem with similar methods. In each trial, 100 participants will be randomised to a remotely delivered perioperative behavioural treatment incorporating principles of behavioural activation, compassion and care coordination, and medication optimisation, or enhanced usual care with mental health-related resources for this population. The primary outcome is change in depression and anxiety symptoms assessed with the Patient Health Questionnaire-Anxiety Depression Scale from baseline to 3 months post surgery. Other outcomes include quality of life, delirium, length of stay, falls, rehospitalisation, pain and implementation outcomes, including study and intervention reach, acceptability, feasibility and appropriateness, and patient experience with the intervention. ETHICS AND DISSEMINATION: The trials have received ethics approval from the Washington University School of Medicine Institutional Review Board. Informed consent is required for participation in the trials. The results will be submitted for publication in peer-reviewed journals, presented at clinical research conferences and disseminated via the Center for Perioperative Mental Health website. TRIAL REGISTRATION NUMBERS: NCT05575128, NCT05685511, NCT05697835, pre-results

Repeated amphetamine treatment induces neurite outgrowth and enhanced amphetamine-stimulated dopamine release in rat pheochromocytoma cells (PC12 cells) via a protein kinase C- and mitogen activated protein kinase-dependent mechanism

Repeated intermittent treatment with amphetamine (AMPH) induces both neurite outgrowth and enhanced AMPH-stimulated dopamine (DA) release in PC12 cells. We investigated the role of protein kinases in the induction of these AMPH-mediated events by using inhibitors of protein kinase C (PKC), mitogen activated protein kinase (MAP kinase) or protein kinase A (PKA). PKC inhibitors chelerythrine (100 nm and 300 nm), Ro31-8220 (300 nm) and the MAP kinase kinase inhibitor, PD98059 (30 µm) inhibited the ability of AMPH to elicit both neurite outgrowth and the enhanced AMPH-stimulated DA release. The direct-acting PKC activator, 12- O -tetradecanoyl phorbol 13-acetate (TPA, 250 nm) mimicked the ability of AMPH to elicit neurite outgrowth and enhanced DA release. On the contrary, a selective PKA inhibitor, 100 µm Rp-8-Br-cAMPS, blocked only the development of AMPH-stimulated DA release but not the neurite outgrowth. Treatment of the cells with acute AMPH elicited an increase in the activity of PKC and MAP kinase but not PKA. These results demonstrated that AMPH-induced increases in MAP kinase and PKC are important for induction of both the enhancement in transporter-mediated DA release and neurite outgrowth but PKA was only required for the enhancement in AMPH-stimulated DA release. Therefore the mechanisms by which AMPH induces neurite outgrowth and the enhancement in AMPH-stimulated DA release can be differentiated.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/66040/1/j.1471-4159.2003.02127.x.pd

Differential Epigenetic Regulation of TOX Subfamily High Mobility Group Box Genes in Lung and Breast Cancers

Aberrant cytosine methylation affects regulation of hundreds of genes during cancer development. In this study, a novel aberrantly hypermethylated CpG island in cancer was discovered within the TOX2 promoter. TOX2 was unmethylated in normal cells but 28% lung (n = 190) and 23% breast (n = 80) tumors were methylated. Expression of two novel TOX2 transcripts identified was significantly reduced in primary lung tumors than distant normal lung (p<0.05). These transcripts were silenced in methylated lung and breast cancer cells and 5-Aza-2-deoxycytidine treatment re-expressed both. Extension of these assays to TOX, TOX3, and TOX4 genes that share similar genomic structure and protein homology with TOX2 revealed distinct methylation profiles by smoking status, histology, and cancer type. TOX was almost exclusively methylated in breast (43%) than lung (5%) cancer, whereas TOX3 was frequently methylated in lung (58%) than breast (30%) tumors. TOX4 was unmethylated in all samples and showed the highest expression in normal lung. Compared to TOX4, expression of TOX, TOX2 and TOX3 in normal lung was 25, 44, and 88% lower, respectively, supporting the premise that reduced promoter activity confers increased susceptibility to methylation during lung carcinogenesis. Genome-wide assays revealed that siRNA-mediated TOX2 knockdown modulated multiple pathways while TOX3 inactivation targeted neuronal development and function. Although these knockdowns did not result in further phenotypic changes of lung cancer cells in vitro, the impact on tissue remodeling, inflammatory response, and cell differentiation pathways suggest a potential role for TOX2 in modulating tumor microenvironment

Global Developmental Gene Expression and Pathway Analysis of Normal Brain Development and Mouse Models of Human Neuronal Migration Defects

Heterozygous LIS1 mutations are the most common cause of human lissencephaly, a human neuronal migration defect, and DCX mutations are the most common cause of X-linked lissencephaly. LIS1 is part of a protein complex including NDEL1 and 14-3-3ε that regulates dynein motor function and microtubule dynamics, while DCX stabilizes microtubules and cooperates with LIS1 during neuronal migration and neurogenesis. Targeted gene mutations of Lis1, Dcx, Ywhae (coding for 14-3-3ε), and Ndel1 lead to neuronal migration defects in mouse and provide models of human lissencephaly, as well as aid the study of related neuro-developmental diseases. Here we investigated the developing brain of these four mutants and wild-type mice using expression microarrays, bioinformatic analyses, and in vivo/in vitro experiments to address whether mutations in different members of the LIS1 neuronal migration complex lead to similar and/or distinct global gene expression alterations. Consistent with the overall successful development of the mutant brains, unsupervised clustering and co-expression analysis suggested that cell cycle and synaptogenesis genes are similarly expressed and co-regulated in WT and mutant brains in a time-dependent fashion. By contrast, focused co-expression analysis in the Lis1 and Ndel1 mutants uncovered substantial differences in the correlation among pathways. Differential expression analysis revealed that cell cycle, cell adhesion, and cytoskeleton organization pathways are commonly altered in all mutants, while synaptogenesis, cell morphology, and inflammation/immune response are specifically altered in one or more mutants. We found several commonly dysregulated genes located within pathogenic deletion/duplication regions, which represent novel candidates of human mental retardation and neurocognitive disabilities. Our analysis suggests that gene expression and pathway analysis in mouse models of a similar disorder or within a common pathway can be used to define novel candidates for related human diseases

Identification of Retinoic Acid in a High Content Screen for Agents that Overcome the Anti-Myogenic Effect of TGF-Beta-1

Transforming growth factor beta 1 (TGF-β1) is an inhibitor of muscle cell differentiation that is associated with fibrosis, poor regeneration and poor function in some diseases of muscle. When neutralizing antibodies to TGF-β1 or the angiotensin II inhibitor losartan were used to reduce TGF-β1 signaling, muscle morphology and function were restored in mouse models of Marfan Syndrome and muscular dystrophy. The goal of our studies was to identify additional agents that overcome the anti-myogenic effect of TGF-β1.A high-content cell-based assay was developed in a 96-well plate format that detects the expression of myosin heavy chain (MHC) in C2C12 cells. The assay was used to quantify the dose-dependent responses of C2C12 cell differentiation to TGF-β1 and to the TGF-β1 Type 1 receptor kinase inhibitor, SB431542. Thirteen agents previously described as promoting C2C12 differentiation in the absence of TGF-β1 were screened in the presence of TGF-β1. Only all-trans retinoic acid and 9-cis retinoic acid allowed a maximal level of C2C12 cell differentiation in the presence of TGF-β1; the angiotensin-converting enzyme inhibitor captopril and 10 nM estrogen provided partial rescue. Vitamin D was a potent inhibitor of retinoic acid-induced myogenesis in the presence of TGF-β1. TGF-β1 inhibits myoblast differentiation through activation of Smad3; however, retinoic acid did not inhibit TGF-β1-induced activation of a Smad3-dependent reporter gene in C2C12 cells.Retinoic acid alleviated the anti-myogenic effect of TGF-β1 by a Smad3-independent mechanism. With regard to the goal of improving muscle regeneration and function in individuals with muscle disease, the identification of retinoic acid is intriguing in that some retinoids are already approved for human therapy. However, retinoids also have well-described adverse effects. The quantitative, high-content assay will be useful to screen for less-toxic retinoids or combinations of agents that promote myoblast differentiation in the presence of TGF-β1

Three-Dimensional Regulation of Radial Glial Functions by Lis1-Nde1 and Dystrophin Glycoprotein Complexes

Lis1-Nde1 integrates cerebral cortical neurogenesis with neuronal migration by stabilizing the basal-lateral surface of radial glial cells