Polyamine depletion induces G1 and S phase arrest in human retinoblastoma Y79 cells

<p>Abstract</p> <p>Background</p> <p>Polyamines and ornithine decarboxylase (ODC) are essential for cell proliferation. DL-α-difluoromethylornithine (DFMO), a synthetic inhibitor of ODC, induces G₁arrest through dephosphorylation of retinoblastoma protein (pRb). The effect of DFMO on cell growth of pRb deficient cells is not known. We examined the effects of DFMO on pRb deficient human retinoblastoma Y79 cell proliferation and its molecular mechanism.</p> <p>Methods</p> <p>Using cultured Y79 cells, the effects of DFMO were studied by using polyamine analysis, western blot, gel shift, FACS and promoter analysis.</p> <p>Results</p> <p>DFMO suppressed the proliferation of Y79 cells, which accumulated in the G1 and S phase. DFMO induced p27/Kip1 protein expression, p107 dephosphorylation and accumulation of p107/E2F-4 complex in Y79 cells.</p> <p>Conclusion</p> <p>These results indicate that p107 dephosphorylation and accumulation of p107/E2F-4 complex is involved in G₁and S phase arrest of DFMO treated Y79 cells.</p

An Angiotensin II Type 1 Receptor Blocker Prevents Renal Injury via Inhibition of the Notch Pathway in Ins2 Akita Diabetic Mice

Recently, it has been reported that the Notch pathway is involved in the pathogenesis of diabetic nephropathy. In this study, we investigated the activation of the Notch pathway in Ins2 Akita diabetic mouse (Akita mouse) and the effects of telmisartan, an angiotensin II type1 receptor blocker, on the Notch pathway. The intracellular domain of Notch1 (ICN1) is proteolytically cleaved from the cell plasma membrane in the course of Notch activation. The expression of ICN1 and its ligand, Jagged1, were increased in the glomeruli of Akita mice, especially in the podocytes. Administration of telmisartan significantly ameliorated the expression of ICN1 and Jagged1. Telmisartan inhibited the angiotensin II-induced increased expression of transforming growth factor β and vascular endothelial growth factor A which could directly activate the Notch signaling pathway in cultured podocytes. Our results indicate that the telmisartan prevents diabetic nephropathy through the inhibition of the Notch pathway

Functional localization and effective connectivity of cortical theta and alpha oscillatory activity during an attention task

Objectives: The aim of this paper is to investigate cortical electric neuronal activity as an indicator of brain function, in a mental arithmetic task that requires sustained attention, as compared to the resting state condition. The two questions of interest are the cortical localization of different oscillatory activities, and the directional effective flow of oscillatory activity between regions of interest, in the task condition compared to resting state. In particular, theta and alpha activity are of interest here, due to their important role in attention processing. Methods: We adapted mental arithmetic as an attention ask in this study. Eyes closed 61-channel EEG was recorded in 14 participants during resting and in a mental arithmetic task (“serial sevens subtraction”). Functional localization and connectivity analyses were based on cortical signals of electric neuronal activity estimated with sLORETA (standardized low resolution electromagnetic tomography). Functional localization was based on the comparison of the cortical distributions of the generators of oscillatory activity between task and resting conditions. Assessment of effective connectivity was based on the iCoh (isolated effective coherence) method, which provides an appropriate frequency decomposition of the directional flow of oscillatory activity between brain regions. Nine regions of interest comprising nodes from the dorsal and ventral attention networks were selected for the connectivity analysis. Results: Cortical spectral density distribution comparing task minus rest showed significant activity increase in medial prefrontal areas and decreased activity in left parietal lobe for the theta band, and decreased activity in parietal-occipital regions for the alpha1 band. At a global level, connections among right hemispheric nodes were predominantly decreased during the task condition, while connections among left hemispheric nodes were predominantly increased. At more detailed level, decreased flow from right inferior frontal gyrus to anterior cingulate cortex for theta, and low and high alpha oscillations, and increased feedback (bidirectional flow) between left superior temporal gyrus and left inferior frontal gyrus, were observed during the arithmetic task. Conclusions: Task related medial prefrontal increase in theta oscillations possibly corresponds to frontal midline theta, while parietal decreased alpha1 activity indicates the active role of this region in the numerical task. Task related decrease of intracortical right hemispheric connectivity support the notion that these nodes need to disengage from one another in order to not interfere with the ongoing numerical processing. The bidirectional feedback between left frontal-temporal-parietal regions in the arithmetic task is very likely to be related to attention network working memory function. Significance: The methods of analysis and the results presented here will hopefully contribute to clarify the roles of the different EEG oscillations during sustained attention, both in terms of their functional localization and in terms of how they integrate brain function by supporting information flow between different cortical regions. The methodology presented here might be clinically relevant in evaluating abnormal attention function

Evaluation of focal damage in the retinal pigment epithelium layer in serous retinal pigment epithelium detachment

The purpose of this study was to evaluate focal damage in the retinal pigment epithelium (RPE) layer in serous retinal pigment epithelium detachment (PED) with multi-contrast optical coherence tomography (OCT), which is capable of simultaneous measurement of OCT angiography, polarization-sensitive OCT and standard OCT images. We evaluated 37 eyes with age-related macular degeneration that had serous PED. Focal RPE damage was indicated by hyper-transmission beneath the RPE-Bruch’s membrane band in standard OCT images. Distribution of RPE melanin was calculated using the dataset from multi-contrast OCT. Twenty-four points with hyper-transmission were detected in 21 of the 37 eyes. Standard OCT images failed to show disruption of the RPE-Bruch’s membrane band at 5 of the 24 hyper-transmission points. Conversely, multi-contrast OCT images clearly showed melanin defects in the RPE-Bruch’s membrane band at all points. Areas of melanin defects with disruption of the RPE-Bruch’s membrane band were significantly larger than those without disruption. The volume of intraretinal hyper-reflective foci was significantly larger in eyes with hyper-transmission than that in eyes without hyper-transmission. Multi-contrast OCT is more sensitive than standard OCT for displaying changes at the RPE-Bruch’s membrane band when there are small areas of RPE damage

The whole blood transcriptional regulation landscape in 465 COVID-19 infected samples from Japan COVID-19 Task Force

「コロナ制圧タスクフォース」COVID-19患者由来の血液細胞における遺伝子発現の網羅的解析 --重症度に応じた遺伝子発現の変化には、ヒトゲノム配列の個人差が影響する--. 京都大学プレスリリース. 2022-08-23.Coronavirus disease 2019 (COVID-19) is a recently-emerged infectious disease that has caused millions of deaths, where comprehensive understanding of disease mechanisms is still unestablished. In particular, studies of gene expression dynamics and regulation landscape in COVID-19 infected individuals are limited. Here, we report on a thorough analysis of whole blood RNA-seq data from 465 genotyped samples from the Japan COVID-19 Task Force, including 359 severe and 106 non-severe COVID-19 cases. We discover 1169 putative causal expression quantitative trait loci (eQTLs) including 34 possible colocalizations with biobank fine-mapping results of hematopoietic traits in a Japanese population, 1549 putative causal splice QTLs (sQTLs; e.g. two independent sQTLs at TOR1AIP1), as well as biologically interpretable trans-eQTL examples (e.g., REST and STING1), all fine-mapped at single variant resolution. We perform differential gene expression analysis to elucidate 198 genes with increased expression in severe COVID-19 cases and enriched for innate immune-related functions. Finally, we evaluate the limited but non-zero effect of COVID-19 phenotype on eQTL discovery, and highlight the presence of COVID-19 severity-interaction eQTLs (ieQTLs; e.g., CLEC4C and MYBL2). Our study provides a comprehensive catalog of whole blood regulatory variants in Japanese, as well as a reference for transcriptional landscapes in response to COVID-19 infection

DOCK2 is involved in the host genetics and biology of severe COVID-19

「コロナ制圧タスクフォース」COVID-19疾患感受性遺伝子DOCK2の重症化機序を解明 --アジア最大のバイオレポジトリーでCOVID-19の治療標的を発見--. 京都大学プレスリリース. 2022-08-10.Identifying the host genetic factors underlying severe COVID-19 is an emerging challenge. Here we conducted a genome-wide association study (GWAS) involving 2, 393 cases of COVID-19 in a cohort of Japanese individuals collected during the initial waves of the pandemic, with 3, 289 unaffected controls. We identified a variant on chromosome 5 at 5q35 (rs60200309-A), close to the dedicator of cytokinesis 2 gene (DOCK2), which was associated with severe COVID-19 in patients less than 65 years of age. This risk allele was prevalent in East Asian individuals but rare in Europeans, highlighting the value of genome-wide association studies in non-European populations. RNA-sequencing analysis of 473 bulk peripheral blood samples identified decreased expression of DOCK2 associated with the risk allele in these younger patients. DOCK2 expression was suppressed in patients with severe cases of COVID-19. Single-cell RNA-sequencing analysis (n = 61 individuals) identified cell-type-specific downregulation of DOCK2 and a COVID-19-specific decreasing effect of the risk allele on DOCK2 expression in non-classical monocytes. Immunohistochemistry of lung specimens from patients with severe COVID-19 pneumonia showed suppressed DOCK2 expression. Moreover, inhibition of DOCK2 function with CPYPP increased the severity of pneumonia in a Syrian hamster model of SARS-CoV-2 infection, characterized by weight loss, lung oedema, enhanced viral loads, impaired macrophage recruitment and dysregulated type I interferon responses. We conclude that DOCK2 has an important role in the host immune response to SARS-CoV-2 infection and the development of severe COVID-19, and could be further explored as a potential biomarker and/or therapeutic target

Polyamine depletion induces Gand S phase arrest in human retinoblastoma Y79 cells-3

<p><b>Copyright information:</b></p><p>Taken from "Polyamine depletion induces Gand S phase arrest in human retinoblastoma Y79 cells"</p><p>http://www.cancerci.com/content/8/1/2</p><p>Cancer Cell International 2008;8():2-2.</p><p>Published online 21 Jan 2008</p><p>PMCID:PMC2259317.</p><p></p>) treated with 5 mM DFMO for 0, 24, 48 h or 5 mM DFMO and 20 μM putrescine for 48 h were immunoprecipitated with anti E2F-4 antibody and immunoblotted with anti p107 anitibody. B: Cell lysates (100 μg) were immunoprecipitated with anti E2F-1, 2, 3, and 5 antibodies and immunoblotted with anti p107 anitibody (upper panels) or with anti E2F-1, 2, 3, and 5 antibodies (lower panels). C: Cell lysates (100 μg) were immunoprecipitated with anti C-Myc antibody and immunoblotted with anti p107 anitibody (left panel). The lysate of Y79 cells treated with DFMO for 24 h were immunoprecipitated with anti C-Myc antibody and immunoblotted with anti C-Myc antibody (right panel). D: Cell lysates (100 μg) were immunoprecipitated with anti p107 antibody and immunoblotted with anti B-Myb anitibody (left panel). The lysate of Y79 cells treated with DFMO for 24 h were immunoprecipitated with anti p107 antibody and immunoblotted with anti p107 antibody (right panel). E: Cell lysates (100 μg) were immunoprecipitated with anti MCM7 antibody and immunoblotted with anti p107 anitibody

Polyamine depletion induces Gand S phase arrest in human retinoblastoma Y79 cells-4

<p><b>Copyright information:</b></p><p>Taken from "Polyamine depletion induces Gand S phase arrest in human retinoblastoma Y79 cells"</p><p>http://www.cancerci.com/content/8/1/2</p><p>Cancer Cell International 2008;8():2-2.</p><p>Published online 21 Jan 2008</p><p>PMCID:PMC2259317.</p><p></p>eated Y79 cells and E2F binding oligonucleotides. Lanes 1–4 : Nuclear extracts from Y79 cells treated with 5 mM DFMO for 0, 24, 48 h or 5 mM DFMO and 20 μM putrescine for 48 h were incubated as described under Experimental. Lanes 5–7 : Nuclear extracts from Y79 cells treated with 5 mM DFMO for 24 h were preincubated with control IgG, E2F-4 antibody, and p107 antibody, respectively. Lanes 8, 9 : Nuclear extracts from Y79 cells treated with 5 mM DFMO for 24 h were incubated with 50 ng of unlabeled competitor (WT and MT, respectively). B: One μg of pE2WTx4-Luc and 10 ng of pRL-CMV were co-transfected into Y79 cells (5 × 10) using FuGene™ Transfection Reagent. After 24 h culture media were changed to RPMI 1640 media with or without 5 mM DFMO or 5 mM DFMO and 20 μM putrescine. Cell lysates were prepared at 24 h, 48 h and 72 h after the addition of DFMO, and examined for luciferase activity to monitor the promoter activity. Significantly different compared to control cells: *p < 0.0