title: PARK7/DJ-1 deficiency modulates microglial activation in response to LPS-induced inflammation

city: Luxembourg

first: Arnaud

last: Muller

address: 84 Val Fleuri

department: DONC

institute: LIH

summary: Background: Specific microglia responses are thought to contribute to the development and progression of neurodegenerative diseases, including Parkinson’s disease (PD). However, the phenotypic acquisition of microglial cells and their role during the underlying neuroinflammatory processes remain largely elusive. Here, according to the multiple-hit hypothesis, which stipulates that PD etiology is determined by a combination of genetics and various environmental risk factors, we investigate microglial transcriptional programs and morphological adaptations under PARK7/DJ-1 deficiency, a genetic cause of PD, during lipopolysaccharide (LPS)-induced inflammation. Methods: Using a combination of single-cell RNA-sequencing, bulk RNA-sequencing, multicolor flow cytometry and immunofluorescence analyses, we comprehensively compared microglial cell phenotypic characteristics in PARK7/DJ-1 knock-out (KO) with wildtype littermate mice following 6- or 24-hour intraperitoneal injection with LPS. In a translational approach, we conducted corresponding analyses in human PARK7/DJ-1 mutant induced pluripotent stem cell (iPSC)-derived microglia and murine bone marrow-derived macrophages (BMDMs). Results: By excluding the contribution of other immune brain resident and peripheral cells, we show that microglia acutely isolated from PARK7/DJ-1 KO mice display a distinct phenotype, specially related to type II interferon and DNA damage response signaling, when compared with wildtype microglia, in response to LPS. We also detected discrete signatures in human PARK7/DJ-1 mutant iPSC-derived microglia and BMDMs from PARK7/DJ-1 KO mice. These specific transcriptional signatures were reflected at the morphological level, with microglia in LPS-treated PARK7/DJ-1 KO mice showing a less amoeboid cell shape compared to wildtype mice, both at 6 and 24 hours after acute inflammation, as also observed in BMDMs. Conclusions: Taken together, our results show that, under inflammatory conditions, PARK7/DJ-1 deficiency skews microglia towards a distinct phenotype characterized by downregulation of genes involved in type II interferon signaling and a less prominent amoeboid morphology compared to wildtype microglia. These findings suggest that the underlying oxidative stress associated with the lack of PARK7/DJ-1 affects microglia neuroinflammatory responses, which may play a causative role in PD onset and progression.

overall_design: To investigate the transcriptional changes of microglia from PARK7/DJ-1 deficient mice with and without LPS-induced inflammation, we injected LPS or saline vehicle i.p. and 6h after we sacrificed the mice and extracted microglia. Cells were extracted via magnetic-activated cell sorting followed by CD11b enrichment and subsequently RNA were extracted and sequenced.

first: Frida

last: Lind-Holm Mogensen


title: FOXO-inhibited  iPSC-derived cardiomyocytes compared against DMSO vehicle controls.

status: Public on Aug 09 2024

city: Cambridge

first: Nivedhitha

last: Velayutham

state: Massachusetts

address: 7 Divinity Ave

department: Stem Cell and Regenerative Biology

country: USA

institute: Harvard University

type: Expression profiling by high throughput sequencing

summary: Human induced pluripotent stem cell (iPSC)-derived cardiomyocytes cultured in 3-dimensional suspension culture were treated with AS1842856 FOXO-inhibitor (n=3 independent wells) or DMSO vehicle control (n=3 independent wells). Total RNA was isolated from each well and utilized for RNA sequencing.

overall_design: Human iPSC-derived cardiomyocyte 3D cultures were treated with AS1842856 (1µM) FOXO-inhibitor or DMSO (0.2%) vehicle control, for 5 days starting at Day 9 of differentiation. They were maintained in RPMI 1640 media supplemented with B27 and Insulin. UCSD142i-86-1 iPSC line (female 116 donor, fibroblast-derived) was obtained from WiCell and utilized following institutional guidelines.

first: Richard

middle: T

last: Lee


title: Human liver sinusoidal endothelial cells support the development of functional human pluripotent stem cell-derived Kupffer cells

city: London

first: Tallulah

middle: Suzanne

last: Andrews

state: Ontario

address: 1151 Richmond St.

country: Canada

institute: University of Western Ontario

summary: In mice, Kupffer cells (KCs) first derive from yolk sac (YS) hematopoietic progenitors prior to the emergence of the hematopoietic stem cell. To investigate human KC ontogeny, we recapitulated YS hematopoiesis from human pluripotent stem cells (hPSCs) and transplanted derivative macrophage progenitors into NSG mice previously humanized with hPSC-liver sinusoidal endothelial cells (LSECs). We demonstrate that hPSC-LSECs facilitate stable hPSC-YS-macrophage engraftment for at least 7 weeks. Single cell RNA sequencing of engrafted YS-macrophages revealed a homogenous MARCO-expressing KC gene signature and low expression of inflammatory macrophage genes. In contrast, human cord blood (CB)-derived macrophages generated grafts that contain multiple hematopoietic lineages in addition to KCs. Functional analyses showed that the engrafted KCs actively perform phagocytosis and erythrophagocytosis in vivo. Taken together, these findings demonstrate that it is possible to generate human KCs from hPSCs and show that the equivalent of the human YS hematopoietic programs represent enriched sources of progenitors for this lineage.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: hPSC-derived macrophages were FACS sorted from mice with humanized LSECs 7 weeks following transplantation, single-cell RNAseq was performing using 10X Chromium 3' scRNAseq (V3 chemistry). Reads were mapped to human & mouse genomes using STARsolo.

first: Gregory

last: Kent

first: Michael

middle: H

last: Atkins

first: Bryan

last: Lung

first: Adele

last: Nikitina

first: Jamie

last: Kwan

first: Sonya

middle: A

last: MacParland

first: Gordon

middle: M

last: Keller

first: Blair

middle: K

last: Gage


title: QTL Mapping and Bulk Segregant Analysis identifies CO2 tolerance genes associated with virulence in the global pathogen Cryptococcus neoformans

city: Iowa City

first: Damian

middle: J

last: Krysan

state: IA

address: 25 South Grand, 2040 ML

department: Division of Pediatrics & Infectious Disease

institute: University of Iowa Hospitals and Clinics

summary: Cryptococcus neoformans is a ubiquitous free-living soil yeast and opportunistic pathogen that causes ~223,100 cases of cryptococcal meningitis per year, killing over 180,000 people. The pathogenicity of C. neoformans relies on its adaptation to the host conditions. An important difference between its natural environment and the mammalian host is the concentration of CO2. CO2 levels in the host fluctuate around 5%, which is ~125-fold higher than in ambient air. We recently found that while clinical isolates are tolerant to host levels ofCO2, many environmental isolates are CO2-sensitive and virulence-attenuated in animal models. The genetic basis responsible for cryptococcal adaptation to high levels of CO2 is unknown. Here, we utilized quantitative trait loci (QTL) mapping with 374 progeny from a cross between a CO2-tolerant clinical isolate and a CO2-sensitive environmental isolate to identify genetic regions regulating CO2 tolerance. To identify specific quantitative trait genes (QTGs), we applied fine mapping through backcrossing and bulk segregant analysis coupled with pooled genome sequencing of near-isogenic progeny but with distinct tolerance levels to CO2. The roles of the identified QTGs in CO2 tolerance were verified by targeted gene deletion. We further demonstrated that virulence levels among near-isogenic strains in a murine model of cryptococcosis correlate with their levels of CO2 tolerance. Moreover, we discovered that sensitive strains may adapt in vivo to become more tolerant to increased CO2 levels and more virulent. These findings highlight the underappreciated role of the host CO2 tolerance and its importance in the ability of an opportunistic environmental pathogen to cause disease.

overall_design: To investigate the effects of host levels of CO2 compared to enviornmental levels of CO2 on Cryptococcus neoformans gene expression in environmental isolates A1-84-14 and A7-35-23 compared to a clinical isolate and reference strain, H99 at 24 hours, in biological triplicate.

first: Benjamin

last: Chadwick

first: Xiaofeng

last: Xie

first: Laura

middle: C

last: Ristow

first: Xiaorong

last: Lin


title: Genome-wide 5mC and 5hmC patterns determine unique transcriptional signatures,regulators and exon inclusion of neural cell types in mouse brain

city: Blacksburg

first: Michelle

middle: Lynne

last: Olsen

state: VA

address: 970 Washington Street SW

department: School of Neuroscience

institute: Virginia Tech

type: Methylation profiling by high throughput sequencing

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Genome-wide 5mC and 5hmC patterns determine unique transcriptional signatures,regulators and exon inclusion of neural cell types in mouse brain [RNA-Seq]

summary: The DNA modifications, 5-methylcytosine (5mc) and 5-hydroxymethylcytosine (5hmc), represent powerful epigenetic regulators of temporal and spatial gene expression across unique brain cell types in health and disease. Yet, how the cooperation of these genome-wide, epigenetic marks vary across different neural cell populations serve to determine unique transcriptional signatures and alternative splicing has not been evaluated. Here we applied Nanopore sequencing of native DNA to obtain a complete, genome wide, single-base resolution atlas of 5mc and 5hmc modifications in neurons, astrocytes and microglia in the mouse cortex (over 40 million CpG sites quantified, 99% genome coverage). Cell-type specific RNA sequencing was performed in tandem to evaluate the transcript expression of all coding genes. In each cell type evaluated integrating the DNA methylation status with coding gene expression revealed 5hmc positively correlated with gene expression. We identified that 5hmC was enriched in astrocytes and demonstrated 100% positive correlation with previously described and novel astrocyte specific cell markers. As reported, 5mC methylation in promoters negatively correlated with gene expression across cell types and this modification was highest in microglia. Our analysis also identified several novel transcription factors which cooperate with 5mC and 5hmC methylation to regulate exon inclusion across all cell populations. Finally, we provide this quantitative, genome-wide, base resolution DNA methylation data as an interactive, online resource (NAM-ME, Neuronal, Astrocyte, Microglia Methylome) to serve as a benchmark dataset for those interested in the methylome landscape in pre-clinical murine models in health and disease.

overall_design: To investigate the gene expression difference between different neural cells, we isolated neurons, astrocytes and microglia from P28 WT male mouse cortices with the magnetic cell isolation method. RNA was extracted to do RNA sequencing with the polyA enrichment method.

middle: L


title: Epigenetic modifiers MMSET and EZH2 physically interact and cooperate to support Multiple Myeloma pathophysiology

city: Montpellier

first: Emmanuel

last: Varlet

address: 141 rue de la Cardonille

country: France

institute: CNRS

type: Genome binding/occupancy profiling by high throughput sequencing


title: Epigenetic modifiers MMSET and EZH2 physically interact and cooperate to support Multiple Myeloma pathophysiology [RNA-seq]

summary: Multiple myeloma (MM) is a malignant B cell dyscrasia characterized by the accumulation of clonal plasma cells (PC) within the bone marrow. Epigenetic factors are involved in MM initiation, progression, and occurrence of chemoresistance. Among them EZH2, the Polycomb Repressive Complex 2 (PRC2) catalytic subunit and NSD2 (MMSET), the target oncogene of t(4;14) primary translocation, are both associated with poor prognosis values and contribute to MM disease. In this study, we identified a physical interaction between EZH2 and MMSET in MM. We used MMSET-depleted MM cells and MAK-683, an allosteric inhibitor of EZH2, to disrupt this interaction and understand its implication in MM biology and resistance to anti-MM drugs. Through its interaction with EZH2, MMSET regulates PRC2 nuclear localization in MM cells, promotes a particularly poor prognosis in MM patients, and modulates the expression of tumor-suppressor genes involved in p53 pathway. We also demonstrated that MAK-683 can interfere with this interaction and synergizes with conventional drugs used in MM treatment: Melphalan, an alkylating agent triggering DNA Damage Response (DDR); and Panobinostat, a Histone De-Acetylase (HDAC) inhibitor directedly regulating p53 acetylation and stability.

overall_design: To investigate to cooperative function of EZH2-MMSET complex in the regulation of MM cells, we established two MMSET-CRISPR cell lines (MMSET-CRISPR-1 and MMSET-CRISPR-2) in which MMSET has been depleted by a CRISPR-Cas9 system. We then performed a comparative gene expression profiling analysis using data obtained from RNA-Seq of our two MMSET-CRISPR cell lines and of XG-7 parental cell line.

first: Sara

last: Ovejero

first: Elina

last: Alaterre

first: Hugues

last: de Boussac

first: Laure

last: Vincent

first: Guillaume

last: Cartron

first: Charles

last: Herbaux

first: Caroline

last: Bret

first: Laurie

last: Herviou

first: Anne

last: Martinez

first: Christophe

last: Hirtz

first: Aurore

last: Attina

first: Jerome

last: Vialaret

first: Giacomo

last: Cavalli

last: Moreaux


title: Epigenetic modifiers MMSET and EZH2 physically interact and cooperate to support Multiple Myeloma pathophysiology [ChIP-seq]

overall_design: To investigate to cooperative function of EZH2-MMSET complex in the regulation of MM cells, we established two MMSET-CRISPR cell lines (MMSET-CRISPR-1 and MMSET-CRISPR-2) in which MMSET has been depleted by a CRISPR-Cas9 system. We then performed Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for EZH2 and histone modification H3K27me3 in XG7, MMSET-CRISPR-1 and MMSET-CRISPR-2 cells.


title: The molecular mechanism responsible for HbSC retinopathy may depend on the action of angiogenesis-related genes ROBO1 and SLC38A5.

city: Campinas

first: Mônica

last: Barbosa de Melo

state: Sao Paulo

address: Avenue Candido Rondon, 400. Cidade Universitária

department: Center for Molecular Biology and Genetic Engineering - CBMEG

country: Brazil

institute: Campinas University

summary: Purpose: HbSC disease, a less severe form of sickle cell disease, affects the retina more frequently and patients have higher rates of proliferative retinopathy that can progress to vision loss. This study aimed to identify differences in the expression of endothelial cell-derived molecules associated with pathophysiology of proliferative sickle cell retinopathy (PSCR).

overall_design: mRNA sequencing (RNAseq) was used to compare the gene expression profile of circulating endothelial colony-forming cells from patients with SC hemoglobinopathy and proliferative retinopathy (n=5), versus SC patients without retinopathy (n=3).

first: Sueli

last: da Silva Costa

first: Mônica

middle: B

last: de Melo


title: Molecular differentiation between complete and incomplete responders to neoadjuvant therapy in rectal cancer

city: Birmingham

first: Fengyuan

last: Huang

state: AL

address: University Blvd

department: Genetics

institute: University of Alabama at Birmingh

summary: Neoadjuvant chemoradiotherapy (nCRT) is the standard treatment for locally advanced rectal cancer, but less than half of patients have a clinical complete response to this treatment. To define the molecular features that are associated with response to neoadjuvant therapy, we generated and collected genomic and transcriptomic data from 712 cancers prior to treatment from our own data and from publicly available data. We found that patients with a complete response have decreased risk of local recurrence and future metastasis. We identified multiple differences in DNA mutations and transcripts between complete and incomplete responders. Complete responder tumors have a higher tumor mutation burden and many more significant co-occurring mutations than the incomplete responder tumors. In addition, mutations in DNA repair genes (across multiple mechanisms of repair) were enriched in complete responders and they also had lower expression of these genes indicating that defective DNA repair is associated with complete response to nCRT. Using logistic regression, we identified three significant predictors of complete response: tumor size, mutations within specific network genes, and the existence of three or more concurrent mutations. In tumors that do not completely respond to nCRT, abnormal cell-cell interaction and increased cancer associated fibroblasts were associated with recurrence. Additionally,gene expression analysis identified a subset of immune hot tumors with worse outcomes and upregulation of immune checkpoint proteins. Overall, our study provides a comprehensive understanding of the molecular features associated with response to nCRT and the molecular differences in non-responder tumors that later reoccur. This knowledge may provide critical insight into the development of precision therapy for rectal cancer.

overall_design: To investigate the transcriptomic differences between complete and incomplete responders to neoadjuvant therapy in rectal cancer,  we collected pre-treatment biopsy samples and isolated RNA from the samples. We then perfored gene expression profiling analysis using data obtained from RNA-seq of 15 samples from 7 patients.

first: Karin

last: Hardiman

first: Zechen

last: Chong

first: Marshall

last: McLeod

first: Regin

last: Irwin

first: Mary

last: Smithson

first: Zongliang

last: Yue

first: Min

last: Gao


title: Epigenetic and transcriptional alterations with Kmt2c and Kmt2d loss in urothelial cells

city: New York

first: Naitao

last: Wang

state: NY

address: 1275 York Avenue

department: HOPP

institute: Memorial Sloan Kettering Cancer Center

summary: KMT2C and KMT2D are two of the most frequently mutated genes in bladder cancer and in histologically normal urothelium. In this study, we developed mouse models to investigate the molecular mechanism of Kmt2c/d loss in urothelial tumorigenesis.

overall_design: Comparision of epigenome and transcriptome between WT and Kmt2c/d knockout mouse bladder urothelail cells


title: Single cell RNA sequencing of human induced pluripotent stem cell-derived cardiomyocytes with or without small molecule activation of FOXO signaling

first: Jessica

last: Garbern

state: MA

summary: We treated differentiated iPSC-derived cardiomyocytes with a small molecule FOXO activator to to evaluate potential genes that might facilitate cardiomyocyte maturation.

overall_design: iPSC-derived cardiomyocytes were treated with the FOXO activator, LOM612 (5 µM) or DMSO (control) for 48 hours prior to sample harvest.

first: Hannah

last: Elwell

first: Richard

last: Lee


title: Sphingosine-1 phosphate receptor 1 (S1PR1) is crucial for maintaining the stem cell properties of acute myeloid leukemia stem cells

city: Beijing

first: Yuqing

address: NO.11 Xizhimen South St, Xicheng district, Beijing

department: Institute of Hematology

country: China

institute: Peking University People’s Hospital

summary: To identify the most important gene(s) for maintaining the stem cell properties of leukemia stem cells (LSCs), we analyzed the transcriptional profiles of LSC and non-LSC subsets from acute leukemia patients and identified S1PR1 as a novel biomarker of LSCs. Then to understand the underlying mechanisms of S1PR1-dependent stemness maintenance, we performed gene expression profiling analyses comparing CD34+S1PR1+ and CD34+S1PR1- cell fractions from AML patients and healthy donors; we also determined the global transcriptional response of KG1 cells to an inhibitor of S1PR1 (FTY720) at two concentrations.

overall_design: Data set 1:   Three acute myeloid leukemia (AML) samples (patient #M1, #M2, and #M3) were sorted into paired LSC (CD34+CD38-) and non-LSC (CD34+CD38+, CD34-CD38+, and CD34-CD38-) fractions by flow cytometry. RNA-seq profiling was performed on the paired fractions for each individual.     Data set 2:  Three Philadelphia chromosome-positive B cell acute lymphoblastic leukemia (ALL) samples (patient #L1, #L2, and #L3) were sorted into paired LSC (CD34+CD38-CD58-) and non-LSC (CD34+CD38+CD58+, CD34+CD38+CD58-, and CD34+CD38-CD58+) fractions by flow cytometry. RNA-seq profiling was performed on the paired fractions for each individual.     Data set 3:  RNA-seq profiling was performed on CD34+S1PR1+ (labeled as P_pos) and CD34+S1PR1- (labeled as P_neg) bone marrow mononuclear cell fractions from AML patients (patient #9 and #10) purified by flow cytometry.     Data set 4:  RNA-seq profiling was performed on CD34+S1PR1+ (labeled as D_pos) and CD34+S1PR1- (labeled as D_neg) bone marrow mononuclear cell fractions from healthy donors (donor #1 and #2) purified by flow cytometry.     Data set 5:  RNA-seq profiling was performed on KG1 cells that were pre-treated with FTY720 at 3 μM (labeled as F3) or 4 μM (labeled as F4) for 24 h or sham treatment (labeled as F0) without FTY720.

first: Yue

last: Ren

first: Xiaojun


title: Genome-wide 5mC and 5hmC patterns determine unique transcriptional signatures,regulators and exon inclusion of neural cell types in mouse brain [Nanopore]

first: Michelle

last: Olsen

overall_design: To investigate the methylation difference between different neural cells, we isolated neurons, astrocytes and microglia from P28 WT male mouse cortices with the magnetic cell isolation method. High molecular weight DNA was extracted to do Nanopore.


title: Gene expression analysis in GATA4fl/flVillin cre+/cre- jejunum

city: Corvallis

first: Andrey

last: Morgun

state: OR

address: 1601 SW Jefferson Way

institute: Oregon State University

summary:
Whole genome transcriptional profiling was performed on conventional and germ free GATA4fl/flVillin cre+ and cre- littermate control mice to determine
the role of GATA4 and the microbiota in mediating gene expression changes in the intestine.

overall_design: Total RNA was extracted from the jejunum of SPF/GF GATA4fl/flVillin cre+ and cre- littermate control mice and whole genome transcriptional profiling was performed using Illumina MouseRef-8v2 arrays.

first: Andriy

middle: R

last: Rodrigues


title: Conserved 5-methyluridine tRNA modification modulates ribosome translocation

status: Public on Aug 08 2024

city: Ann Arbor

first: Rachel

middle: Orion

last: Niederer

state: MI

address: 1150 W. Medical Center Dr

department: Biological Chemistry

institute: University of Michigan

summary: While the centrality of post-transcriptional modifications to RNA biology has long been acknowledged, the function of the vast majority of modified sites remains to be discovered. Illustrative of this, there is not yet a discrete biological role assigned for one the most highly conserved modifications, 5-methyluridine at position 54 in tRNAs (m5U54). Here, we uncover contributions of m5U54 to both tRNA maturation and protein synthesis. Our mass spectrometry analyses demonstrate that cells lacking the enzyme that installs m5U in the T-loop (TrmA inE. coli, Trm2 inS. cerevisiae) exhibit altered tRNA modifications patterns. Furthermore, m5U54 deficient tRNAs are desensitized to small molecules that prevent translocationin vitro.This finding is consistent with our observations that, relative to wild-type cells,trm2Δ cell growth and transcriptome-wide gene expression are less perturbed by translocation inhibitors. Together our data suggest a model in which m5U54 acts as an important modulator of tRNA maturation and translocation of the ribosome during protein synthesis.

overall_design: To investigate the impact of m5U modifications on expression we knocked out the methyltransferase Trm2 and then did RNA-seq on both WT and KO cells in the presence or absence of the elongation inhibitor hygromycin B. All experiments were performed in biological duplicate

first: Joshua

middle: D

last: Jones

first: Monika

last: Franco

last: Giles

first: Daniel

middle: E

last: Eyler

first: Mehmet

last: Tardu

first: Tyler

last: Smith

first: Laura

last: Snyder

first: Yury

middle: S

last: Polikanov

first: Robert

last: Kennedy

middle: O

first: Kristin

last: Koutmou


title: Defining the nature of human pluripotent stem cell-derived interneurons via single cell analysis

city: Santa Clara

first: Marcos

last: Otero-Garcia

state: CA

address: 2255 Martin Ave suite D

institute: Rebus Biosystems

summary: sc-RNA-Seq profiling of human brain nuclei

overall_design: Human brain cortex nuclei were isolated and subjected to sc-RNA-seq

first: William

last: Lowry

first: Inma

last: Cobos

first: Kathrin

last: Plath

last: Otero

first: Tom

last: Allison


title: Single-cell gene expression profile of urogenital sinus from neonatal mice at postnatal day 0

city: shanghai

first: Jian

last: Wang

address: yueyang road 320

institute: Shanghai Institutes for Biological Sciences CAS

summary: The identity, fate and function of these intermediate cells in prostate development are not well understood. Through single-cell RNA-seq analysis on neonatal urogenital sinus tissue, we identified intermediate cells exhibiting stem cell potential.

overall_design: Urogenital sinus of 9 wild-type neonatal mice at postnatal day 0 were dissociated and used for scRNAseq using 10x Genomics.

first: Xiaoyu

last: Zhang

first: Wangxin

last: Guo


title: Deciphering epigenetic and transcriptional alterations after Kmt2c/2d knock-out in mouse bladder urothelial cells

first: Naitao

summary: KMT2C and KMT2D are two of the most frequently mutated genes in histologically normal human bladder urothelium. In this study, we sought to model Kmt2c/d LOF mutations and investigate the biological mechanism of Kmt2c/d LOF mutations in bladder cancer development.

overall_design: Comparision of epigenome and transcriptome between control and Kmt2c/d knock-out mouse bladder urothelail cells

first: Yu

last: Chen


title: Repurposing CD19-directed immunotherapies for pediatric t(8;21) acute myeloid leukemia

city: Utrecht

first: Joost

last: Koedijk

address: Heidelberglaan 25

country: Netherlands

institute: Princess Máxima Center for Pediatric Oncology

summary: In contrast to patients with B cell precursor acute lymphoblastic leukemia (BCP-ALL), patients with acute myeloid leukemia (AML) have not yet benefited from recent advances in targeted immunotherapy. Repurposing immunotherapies that have been successfully used to target other hematological malignancies could, in case of a shared target antigen, represent a promising opportunity to expand the immunotherapeutic options for AML. Here, we evaluated the expression of CD19 in a large pediatric AML cohort, assessed the ex vivo AML killing efficacy of CD19-directed immunotherapies, and characterized the bone marrow immune microenvironment in pediatric AML, BCP-ALL, and non-leukemic controls. Out of 167 newly diagnosed de novo pediatric AML patients, 18 patients (11%) had CD19+ AML, with 61% carrying the translocation t(8;21)(q22;q22). Among CD19+ samples, we observed a continuum of CD19 expression levels on AML cells. In individuals exhibiting unimodal and high CD19 expression, the antigen was consistently present on nearly all CD34+CD38- and CD34+CD38+ subpopulations. In ex vivo AML-T cell co-cultures, blinatumomab demonstrated substantial AML killing, with an efficacy similar to BCP-ALL. In addition, CAR T cells could effectively eliminate CD19+ AML cells ex vivo. Furthermore, our immunogenomic assessment of the bone marrow immune microenvironment of newly diagnosed pediatric t(8;21) AML revealed that T- and NK cells had a less exhausted and senescent phenotype in comparison to pediatric BCP-ALL. Altogether, our study underscores the promise of CD19-directed immunotherapies for the treatment of pediatric CD19+ AML.

overall_design:
In this study, we compared the bulk RNA-seq gene expression profiles of pediatric t(8;21) AML patients (n=10), non-t(8;21) pediatric AML (n=30), non-leukemic controls (n=4; derived from another publication: PMID 37908360), and pediatric B cell precursor acute lymphoblastic leukemia patients (n=209). To acquire the gene expression profiles from these patient samples, total RNA was isolated from viably frozen bone marrow mononuclear cells (BMMCs; derived from BM aspirates), RNA-seq libraries were generated from 300 ng RNA, and sequenced with NovaSeq 6000 (2x150 bp; Illumina). After pre-processing, RNA-sequencing yielded 80-100 million raw reads. Reads were aligned to GRCh38 and gencode version 31. Downstream analysis was done as shown in our preprint: https://www.biorxiv.org/content/10.1101/2024.04.19.590200v1.

***************************************************************
Raw data not allowed to make the data publicly available due to informed consent restrictions; contact authors for these data
***************************************************************

first: Barneh

last: Farnaz

first: Koedijk

last: Joost

first: Wijnen

last: Noa

first: Heidenreich

last: Olaf


title: Epigenetic marks correlated with differential parental genome contributions to plant zygotic transcriptomes

city: Irapuato

first: Charles

middle: Stewart

last: Gillmor

state: Guanajuato

address: Libramiento Norte Carretera Leon Km 9.6

department: LANGEBIO

country: Mexico

institute: CINVESTAV

summary: We profile allele specific expression of transcripts in hybrid embryos of Arabidopsis at seven developmental stages, revealing dynamic parental contributions and ecotype-dependent transcript abundances.

overall_design: RNA-seq of  hybrid Arabidopsis embryos derived from reciprocal crosses of the Tsu-1 and Col-0 accessions, at seven developmental stages.

first: Jaime

last: Alaniz

first: Stewart

first: Daoquan

last: Xiang

first: Raju

last: Datla


title: Combined with Gefitinib Overcomes Immunosuppression Induced by long-term Glutamine blockade in Bladder Cancer

city: Qingdao

first: Guofeng

last: Ma

address: Jiangsu Road

institute: Affiliated Hospital of Qingdao University

summary: Glutamine is a major energy source for tumor cells and blocking glutamine metabolism is being investigated as a promising strategy for cancer therapy. However, the antitumor effect of glutamine blockade in bladder cancer remains unclear, necessitating further investigation. Here, we demonstrated that glutamine metabolism was involved in the malignant progression of bladder cancer. Treatment with the glutamine antagonist 6-Diazo-5-oxo-L-norleucine (DON) inhibited bladder cancer cells in vitro in several ways. In addition, we observed a remarkable inhibition of tumor growth in bladder cancer-bearing mice using JHU083, a prodrug that was designed to prevent DON-induced toxicity. However, the antitumor immune effect of T cells changed from activation to inhibition as the administrated time extended. We found that both in vitro treatment with DON and in vivo prolonged administration of JHU083 led to the upregulation of PD-L1 in bladder cancer cells. Mechanistically, glutamine blockade up-regulates PD-L1 expression in bladder cancer cells by accumulating ROS, subsequently activating the EGFR/ERK/C-Jun signaling pathway. Combinatorial treatment with JHU083 and gefitinib reversed the up-regulation of PD-L1 in bladder cancer cells induced by prolonged glutamine blockade. This reversal resulted in the alleviation of T-cell immunosuppression and a significant improvement in therapeutic outcome. These preclinical findings suggested that broad targeting of glutamine metabolism could represent a viable therapeutic strategy for bladder cancer, with the potential for further enhancement through combined treatment with gefitinib.

overall_design: The bladder cancer cell line T24 was treated with DON or cultured under normal conditions for 48 h. Differential genes between the two groups were detected by RNA sequencing.

first: Huiqing

last: Jia

first: Zhiqiang

last: Li

first: Xiangyan


title: Effect of melatonin during LPS-induced inflammation on RAW 264.7 cells

city: Shanghai

first: Yuying

last: Cai

state: Please Select

address: 450 Tengyue Road, Shanghai 200090, China

institute: Yangpu Hospital, Tongji University

summary: Melatonin is a powerful anti-inflammatory agent and antioxidant. To explore specific anti-inflammatory mechanisms, LPS was used to intervene in RAW 264.7 cells to construct an inflammation model. Then, cells were pretreated with melatonin at concentrations of 10-3M to 10-7M, respectively. The above cells were subjected to DRUG-seq sequencing to explore the specific mechanism of melatonin anti-inflammation.

overall_design: To investigate the specific mechanism of melatonin anti-inflammation, RAW 264.7 cells were divided into four groups: control, LPS, melatonin and LPS+melatonin groups. Among them, the control group was intervened with solvent only. the LPS group was treated with 10µg/mL of LPS. In order to investigate the concentration effect of melatonin, the melatonin group and LPS+melatonin group could be divided into five subgroups. Among them, the melatonin group was intervened with melatonin at concentrations of 10-7 M, 10-6 M, 10-5 M, 10-4 M, and 10-3 M. The RAW 264.7 cells in the LPS+melatonin group were pre-treated with a concentration gradient of melatonin for 30 min before the addition of LPS. the above cells were treated for 12 hrs and then collected for the next sequencing step.

first: Xin

first: Yaping

last: Jiang

first: Yihui


title: Combination of oncolytic Maraba virus with immune checkpoint blockade overcomes therapy resistance in an immunologically cold model of advanced melanoma with dysfunctional T cell receptor signalling

first: Anton

last: Patrikeev

state: London

address: Chester Beatty Laboratories, 237 Fulham Rd.

department: Targeted Therapy

country: United Kingdom

institute: The Institute of Cancer Research

summary: Oncolytic viruses are promising immunotherapeutic agents, but their efficacy, particularly following intra-tumoural injection, in relation to the size of the targeted tumour, is unclear. Here we show that an oncolytic rhabdovirus, maraba, activates an immune response in human as well as mouse pre-clinical systems, which targets viral as well as tumour-associated antigens. The addition of immune checkpoint blockade to virotherapy is apparent only on the treatment of larger tumours, which have an inherently more immunosuppressive tumour microenvironment, including skewing of T cell receptor engagement with antigen from conventional to regulatory CD4+ cells. Maraba converts the immunologically cold tumour to hot, thus priming the tumour microenvironment for effective αPD1 combination therapy. This study supports the use of maraba oncolytic virotherapy in combination with immune checkpoint inhibitors in melanoma and highlights the impact of the tumour burden on the immune microenvironment and benefit of single agent and combination treatment.

overall_design: To investigate therapeutic effects of Maraba oncolytic viral treatment, we used five- to six-week-old female C57Bl/6 mice (Charles River, UK) or Nr4a3 Tocky mice (C57Bl/6 background), inoculated with 4434 thyroid cancer murine melanoma tumour cells.


title: Age-Related Retinal Degeneration Resulting from the Deletion of Shp2 Tyrosine Phosphatase in Photoreceptor

city: Oklahoma City

first: Rahul

last: Rajala

state: Oklahoma

address: 825 NE 13th Street

department: Cardiovascular Biology

institute: Oklahoma Medical Research Foundation

summary: Shp2, a critical SH2-domain-containing tyrosine phosphatase, is essential for cellular regulation and implicated in metabolic disruptions, obesity, diabetes, Noonan syndrome, LEOPARD syndrome, and cancers. This study focuses on Shp2 in rod photoreceptor cells, revealing its enrichment, particularly in rods. Deletion of Shp2 in rods leads to age-dependent photoreceptor degeneration. Shp2 targets occludin (OCLN), a tight junction protein, and its deletion reduces OCLN expression in the retina and retinal pigment epithelium (RPE). Isolation of actively translating mRNAs from rods lacking Shp2, followed by RNA sequencing, reveals alterations in cell cycle regulation. Altered retinal metabolism is observed in retinal cells lacking Shp2. Our studies indicate that Shp2 is crucial for maintaining the structure and function of photoreceptors.

overall_design: Total tissue lysate and Rod-TRAP RNASeq data were collected from the retinas of 2-week-old littermate control mice (RiboTag;Shp2fl/+;i75-Rod-Cre) and experimental rod photoreceptor Shp2 knockout mice (RiboTag;Shp2fl/fl;i75-Rod-Cre).

first: Ammaji

first: Mohd

last: Bhat

first: Mark

last: Eminhizer

first: Jeff

last: Hao

first: Jianhai

last: Du

middle: V


title: GNAS knockout potentiates HDAC3 inhibition through viral mimicry-related interferon responses in lymphoma

city: Toronto

first: Robert

last: Kridel

state: ON

address: 101 College St

institute: Princess Margaret Cancer Centre – University Health Network

summary:
Despite selective HDAC3 inhibition showing promise in a subset of lymphomas with CREBBP mutations, wild-type tumors generally exhibit resistance. Here, using unbiased genome-wide CRISPR screening, we identify GNAS knockout (KO) as a sensitizer of resistant lymphoma cells to HDAC3 inhibition. Mechanistically, GNAS KO-induced sensitization is independent of the canonical G-protein activities but unexpectedly mediated by viral mimicry-related interferon (IFN) responses, characterized by TBK1 and IRF3 activation, double-stranded RNA formation, and transposable element (TE) expression. GNAS KO additionally synergizes with HDAC3 inhibition to enhance CD8+ T cell-induced cytotoxicity. Moreover, we observe in human lymphoma patients that low GNAS expression is associated with high baseline TE expression and upregulated IFN signaling and shares common disrupted biological activities with GNAS KO in histone modification, mRNA processing, and transcriptional regulation. Collectively, our findings establish an unprecedented link between HDAC3 inhibition and viral mimicry in lymphoma. We suggest low GNAS expression as a potential biomarker that reflects viral mimicry priming for enhanced response to HDAC3 inhibition in the clinical treatment of lymphoma,
especially the CREBBP wild-type cases.

overall_design: Three SU-DHL-4-derived cell lines (one WT and two GNAS knockout) were treated with HDAC3 inhibitor RGFP966 (5 µM) or DMSO. Two biological replicates were included for each condition. In total, there are six conditions and 12 samples.

first: Michael

middle: Y

last: He

first: Ting

last: Liu

first: Zhibin

last: Lu


title: ATM-Dependent Phosphorylation of Nemo SQ Motifs is Dispensable for Nemo-Mediated Gene Expression Changes in Response to DNA Double Strand Breaks

city: Philadelphia

first: Craig

last: Bassing

address: 3501 Civic Center Blvd

institute: Children's Hospital of Philadelphia

summary: In response to DNA double strand breaks (DSBs), the ATM kinase activates NF-κB factors to stimulate gene expression changes that promote survival and allow time for cells to repair damage. In cell lines, ATM can activate NF-κB transcription factors via two independent, convergent mechanisms. One is ATM-mediated phosphorylation of nuclear NF-κB essential modulator (Nemo) protein, which leads to monoubiquitylation and export of Nemo to the cytoplasm where it engages the IκB kinase (IKK) complex to activate NF-κB. Another is DSB-triggered migration of ATM into the cytoplasm where it promotes monoubiquitylation of Nemo and resulting IKK-mediated activation of NF-κB. ATM has many other functions in the DSB response beyond activation of NF-κB, and Nemo activates NF-κB downstream of diverse stimuli, including developmental or proinflammatory stimuli such as lipopolysaccharides (LPS). To elucidate the in vivo role of DSB-induced, ATM-dependent changes in expression of NF-κB-responsive genes, we generated mice expressing phosphomutant Nemo protein lacking consensus SQ sites for phosphorylation by ATM or related kinases. We demonstrate that these mice are viable/healthy, fertile, and exhibit overall normal B and T lymphocyte development. Moreover, treatment of their B lineage cells with LPS induces normal NF-κB-regulated gene expression changes. Furthermore, in marked contrast to results from a pre-B cell line, primary B lineage cells expressing phosphomutant Nemo treated with the genotoxic drug etoposide induce normal ATM- and Nemo-dependent changes in expression of NF-κB-regulated genes. Our data demonstrate that ATM-dependent phosphorylation of Nemo SQ motifs in vivo is dispensable for DSB-signaled changes in expression of NF-κB-regulated genes.

overall_design: RNA-Seq was performed on isolated CD19+ splenocytes from WT, Nemo3SAΔT, or Mb1cre+Nemoflox (MN) mice

first: Rebecca

last: Glynn


title: Overcoming ABCB1 mediated multidrug resistance in castration resistant prostate cancer

city: Gainesville

first: Alexander

last: Ishov

address: 2033 Mowry Rd

institute: University of Florida

summary: The few available therapies for mCRPC patients include taxanes docetaxel (DTX) and cabazitaxel (CBZ). However, development of resistance limits their clinical use. PCa that relapses after hormonal therapies, referred to as castration resistant prostate cancer (CRPC), often presents with metastases (mCRPC) that are the major cause of mortality. The few available therapies for mCRPC patients include taxanes docetaxel (DTX) and cabazitaxel (CBZ). However, development of resistance limits their clinical use. To study taxanes resistance, we produced CBZ resistant C4-2B cells (RC4-2B) and documented resistance to both CBZ and DTX in cell culture and in 3D prostaspheres settings. RNAseq identified increased expression of ABCB1 in RC4-2B, that was confirmed by immunoblotting and immunofluorescent analysis. ABCB1-specific inhibitor elacridar reversed CBZ and DTX resistance in RC4-2B cells, confirming ABCB1-mediated resistance mechanism.

overall_design: To study taxanes resistance in CRPC, we have produced CBZ-resistant mCRPC C4-2B cell line following the next experimental procedure: after 72 h of CBZ treatment, cells were recovered for one week, re-plated, and the cycle was repeated with increased concentration of CBZ starting at 0.1nM and ending at 20nM.  After 4 months, we completed the production of CBZ resistant subclone RC4-2B. We characterized CBZ resistance of RC4-2B cells using metabolic viability (Alamar Blue) and colony formation assays. These complementary methods revealed that IC50 for CBZ is ~10x higher in RC4-2B cells compared to the parental C4-2B cells. To understand mechanisms of taxanes resistance, we performed transcriptome analysis in C4-2B and RC4-2B cells.

first: Viacheslav

last: Morozov

first: Sadia

last: Sarwar


title: Multi-omic signatures of sarcoidosis and progression in bronchoalveolar lavage cells

city: Aurora

first: Ivana

last: Yang

state: CO

institute: University of Colorado - Anschutz Medical Campus

summary: We profiled DNA methylation, mRNA expression, and miRNA expression from bronchoalveolar lavage cells obtained from 64 sarcoidosis subjects and 16 healthy controls.

overall_design:
Bronchoalveolar lavage cells were collected from 64 sarcoidosis subjects (26 with progressive disease based on clinical characteristics, 38 with non-progressive disease) and 16 healthy control subjects

***************************************************************
Raw sequencing files aren't provided due to patient privacy concerns, as patient genotypes can be inferred from sequencing
***************************************************************

first: Iain

last: Konigsberg

first: Nancy

middle: W

last: Lin

first: Lisa

middle: A

last: Maier


title: RNA-seq analysis of AtrR-regulated gene expression in Aspergillus fumigatus cells in response to voriconazole.

city: Iowa City

first: Scott

last: Moye-Rowley

state: Iowa

address: 6-530 Bowen Science Bldg./Dept. of Mol Physio

department: Molecular Physiology & Biophysics

institute: University of Iowa

summary: Aspergillus fumigatus transcription factor AtrR is a critical determinant of the azole resistance phenotype of this organism. AtrR positively regulates expression of a range of genes involved in azole resistance including the ATP-binding cassette transporter-encoding locus abcG1 (cdr1B/abcC) as well as the gene that encodes the enzymatic target of azole drugs cyp51A. Homology searches of A. fumigatus AtrR against a range of fungal species identified highly conserved 25 amino acid region in the carboxy-terminus. To determine the contribution of this region to AtrR function, we prepared a mutant from of the atrR that lacked this region (Δ855-879). We compared the response of an isogenic wild-type strain to a strain expressing the Δ855-879 AtrR derivative using RNA-seq, both in the absence and the presence of voriconazole challenge. The resulting data indicate that this 25 amino acid segment of AtrR is required for expression of some but not all AtrR target genes.

overall_design: This experiment was designed to compare the voriconazole-induced transcriptome that was altered by loss of the transcription factor AtrR or the presence of a mutant form of AtrR that lacked a 25 amino acid domain from its C-terminus that is conserved across fungi. Introduction of this mutant form of AtrR required replacement of the native 3’ untranslated region with an element from the cyc1 gene and was linked to a phleomycin resistance gene. To make a wild-type control for this new derivative of atrR, we introduced this same construct into the native atrR locus (atrR-phleoR). This allowed us to confidently compare the level of wild-type AtrR transcription driven in the presence of this atrR-phleoR derivative compared to the Δ855-879 atrR-phleoR locus. We compared transcription of isogenic wild-type and atrRD strains to assess the voriconazole-induced transcription in the presence or absence of AtrR.

first: Shivani

last: Ror

last: Stamnes

first: W


title: Antihypertensive potential of Pistacia lentiscus var. Chia: Molecular Insights and Therapeutic Implications.

city: Athens

first: Dimitris

last: Valakos

address: Soranou Ephessiou 4

department: Center of Basic Research

country: Greece

institute: Biomedical Reasearch Foundation Academy of Athens

summary: Hypertension poses a significant global health burden and is closely associated with cardiovascular morbidity. Pistacia lentiscus var. chia, commonly known as mastic, has emerged as a potential nutraceutical, due to its diverse pharmacological properties, including antioxidant and vasodilatory effects. Yet, its impact on hypertension in translational in vivo models and on hypertension-associated abnormalities in circulation, vasculature and kidneys have not been investigated. Herein we sought to decipher the putative antihypertensive potential of mastic, focusing on vascular and renal endothelium in clinically relevant in vivo models of essential hypertension. Two translational in vivo models of hypertension induced by Angiotensin II and Deoxycorticosterone acetate (DOCA)-High Salt in male rats, were utilized. Mastic administration at a therapeutic dose (220mg/kg, per os) was administered daily for four weeks, after the establishment of hypertension. Non-invasive blood pressure measurements and whole blood RNA sequencing and metabolomics were performed. Real-time PCR and Western blot analysis were conducted in kidney and aorta, to decipher its underlying antihypertensive mechanism. Mastic administration significantly reduced systolic, diastolic, and mean blood pressure in both in vivo hypertensive models. RNA sequencing revealed distinct gene expression profiles, with modulation of immune-related pathways in the Angiotensin II model and metabolic pathways in the DOCA-HS model. Additionally, RNA sequencing analysis revealed that mastic downregulated Rhob, Jun, Txnip and Egr1 mRNA expression in the circulation, paving towards a putative antioxidant and endothelial-targeted mechanism. Metabolomic analysis identified lysophosphatidylinositol upregulation as a consistent effect of mastic in both in vivo models, being implicated with improved endothelial homeostasis. RT-PCR analysis in the kidneys and aorta, showed that mastic increased endothelial markers such as Vegfa and Egr-1 mRNA expression in both experimental models. Concerning the endothelial homeostasis, mastic additionally increased renal nitric oxide synthase phosphorylation in both in vivo models of hypertension. This study highlights the potential of mastic as a natural adjuvant therapy for hypertension. The findings shed new light on its molecular mechanisms of action, providing insights into its protective impact on renal endothelium.

overall_design: RNA-sequencing

first: Dimitrios

first: Panayiotis

last: Efentakis

first: Georgios

last: Dedousis

first: Ioanna

last: Andreadou


title: Restoring Bone Regeneration in Critical Size Defects through Local Delivery of Angiogenic Factors

city: St Louis

first: Jie

last: Shen

state: MO

address: 425 S. Euclid Ave

institute: Washington University in St Louis

summary: Critical size bone defects represent a significant challenge worldwide, often leading to persistent pain and physical disability that profoundly impact patients’ quality of life and mental well-being.To address the intricate and complex repair processes involved in these defects, we performed single-cell RNA sequencing and revealednotable shifts in cellular populations within regenerative tissue. Specifically, we observed a decrease in progenitor lineage cells and endothelial cells, coupled with an increase in fibrotic lineage cells and pro-inflammatory cells within regenerative tissue. Furthermore, our analysis of differentially expressed genes and associated signaling pathway at the single-cell level highlighted impaired angiogenesis as a central pathway in critical size bone defects, notably influenced by reduction of Spp1 and Cxcl12 expression. This deficiency was particularly pronounced in progenitor lineage cells and myeloid lineage cells, underscoring its significance in the regeneration process. In response to these findings, we developed an innovative approach to enhance bone regeneration in critical size bone defects. Our fabrication process involves the integration of electrospun PCL fibers with electrosprayed PLGA microspheres carrying Spp1 and Cxcl12. This design allows for the gradual release of Spp1 and Cxcl12 in vitro and in vivo. To evaluate the efficacy of our approach, we applied locally delivered Spp1 and Cxcl12 embedded PCL scaffolds in a murine model of critical size bone defects. Our results demonstrated restored angiogenesis, accelerated bone regeneration, alleviated pain responses and improved mobility in treated mice.

overall_design: The regenerative tissues from the defect gap were harvested at 3 weeks post-surgery and dissociated in 2 mg/mL Collagenase P and 2 mg/mL pronase at room temperature for 1 hour. The erythrocytes were removed using anti-GYPA and Streptavidin Microbubbles. Subsequently, the cell suspension was filtered through a 30 μm strainer to yield a single-cell suspension. Two sets of single-cell libraries were prepared: one from cells derived from a 1 mm bone defect (control, n=3) and the other from cells obtained from a 3 mm bone defect (nonunion, n=3). For each sample, ten thousand cells were loaded for processed to generate single-cell mRNA libraries using the Chromium Single Cell 3′ kit (v3.1 Chemistry, 10× Genomics Inc). These libraries were barcoded, purified, and sequenced in a 2 × 150-bp paired-end configuration on an Illumina NovaSeq platform.


title: Multi-omic signatures of sarcoidosis and progression in bronchoalveolar lavage cells [DNAm]

type: Methylation profiling by genome tiling array

overall_design: Bronchoalveolar lavage cells were collected from 64 sarcoidosis subjects (26 with progressive disease based on clinical characteristics, 38 with non-progressive disease) and 16 healthy control subjects


title: miR-182, miR-221 and miR-222 are potential urinary extracellular vesicle biomarkers for canine urothelial carcinoma

city: Cambridge

first: Lajos

last: Kalmar

address: Tennis Court Road

institute: MRC Toxicology Unit, University of cambridge

type: Non-coding RNA profiling by high throughput sequencing

summary: Urothelial carcinoma (UC) is the most common tumour type in canine bladder cancer. Current diagnostic methods are technically challenging or can lack specificity, hence there is a need for novel biomarkers of UC. To this end, we analysed the microRNA (miRNA) cargo of extracellular vesicles (EVs) from urine samples of dogs with UC to identify miRNAs that could be utilised as biomarkers. Urine was fractionated using ultrafiltration combined with size-exclusion chromatography and small RNA sequencing analysis was performed on both the EV enriched and (EV free) protein fractions. A greater number of candidate miRNA biomarkers were detected in the EV fraction than the protein fraction, and further validation using droplet digital PCR (ddPCR) was performed on the EV enriched fraction of a second cohort of dogs with bladder cancer which validated three miRNAs as candidate biomarkers; miR-182, miR-221 and miR-222 as being significantly overrepresented in dogs with UC when compared with healthy dogs and dogs with urinary tract infections. Pathway analysis confirmed that these three miRNAs are involved in cancer. In addition, their potential downstream gene targets were predicted and PIK3R1, a well-known oncogene is likely to be a shared target between miRNA-182 and miRNA-221/222. In summary, this study highlights the potential of urinary EV-associated miRNAs as a source of biomarkers for the diagnosis of canine UC.

overall_design: To find new biomarkers for canine urithelial carcinoma from extracellular vesicle and protein SEC fraction samples isolated from dog urine. Sample groups include urothelial carcinoma (UC), urinary tract infection (UTI) and healty control (HC) samples. Eleven samples were analysed as replicates and two libraries were perepared from same RNA.

first: Jenni

last: Karttunen

first: Andrew

last: Grant

first: Jason

last: Ying

first: Sarah

last: Stewart

first: Xiaonan

last: Wang

first: Fiona

last: Karet Frankl

first: Tim

last: Williams


title: Enterovirus-A71 Preferentially Infects and Replicates in Human Motor Neurons, Inducing Neurodegeneration by Ferroptosis

city: Singapore

first: Yingying

last: Zeng

state: --- Select a state ---

address: 61 Biopolis Drive

country: Singapore

institute: Institute of Molecular and Cell Biology

summary: Enterovirus 71 (EV71) causes Hand, Foot, and Mouth Disease and has been clinically associated with neurological complications. However, there is a lack of relevant models to elucidate the neuropathology of EV71 and its mechanism, as the current models mainly utilize animal model or immortalized cell lines. In this study, we established a human motor neuron model for EV71 infection.

overall_design: scRNA-seq were performed in hESC-derived spinal neuronal cultures after Mock-infected or EV71 (S41) infected

middle: H

last: Chooi

first: Y

last: Loh

first: S

last: Ng


title: PGC-1α Agonism Induces Fetal Hemoglobin and Exerts Anti-Sickling Effects in Sickle Cell Disease

city: Boston

first: Single Cell

last: Sequencing Core

address: 72 E. Concord St

institute: BU Single Cell Sequencing Core

summary: Sickle cell disease is a growing health burden afflicting millions around the world. Clinical observation and laboratory studies have shown that the severity of sickle cell disease is ameliorated in individuals who have elevated levels of fetal hemoglobin. However, pharmacologic induction of fetal hemoglobin sufficient to diminish clinical severity in sickle patients has been challenging. We recently found that up-regulation of PGC-1α can induce fetal hemoglobin synthesis in human primary erythroblasts. Here, we report that a small molecular compound SR-18292 increases PGC-1α leading to enhanced fetal hemoglobin expression in human erythroid cells or β-YAC and sickle cell disease mice. Sickled red blood cells are significantly reduced and disease complications are alleviated in SR-18292-treated sickle mice. SR-18292, or agents in its class, could be a promising therapeutic for sickle cell disease.

overall_design: The objective of the study was to identify a potent small molecule targeting PGC-1α (SR-18292) and evaluate whether SR-18292 can induce fetal hemoglobin synthesis in vitro and in vivo and reduce sickle cell disease-related symptoms in humanized SCD mice. In vitro, we measured the protein levels and mRNA levels of PGC-1α and γ-globin in human CD34+ cells treated with SR-18292 at different dosages as well as F-cells. In vivo, we evaluate the fetal hemoglobin induction in two mouse models: a model is β-YAC mice (a generous gift from Dr. Stuart Orkin's group at Harvard University), which have the entire human β-globin gene cluster knocked in; the other is sickle cell disease mice model, which carry the human sickle hemoglobin gene and present many phenotypic similarities. For the mouse study, we also measured the protein levels and mRNA levels of PGC-1α and γ-globin, as well as the F-cells. And in the sickle cell disease mice model, we also checked the SCD-related symptoms after SR-18292 treatment. All animal  experiments were performed in strict accordance with the ethical guideline and were approved by Boston University Institutional Animal Care & Use Committee (IACUC). For all experiments, the number of replicates, statistical analysis and P value are reported in the figure legends or the following detailed experiments description.

first: Yanan

last: Sun


title: Effects of Teriparatide on miRNA Profile in Different Treatments of Mesenchymal Stem Cells

city: Ljubljana

first: Lucija

middle: Ana

last: Vrščaj

address: Aškerčeva cesta 7

department: Faculty of Pharmacy

country: Slovenia

institute: University of Ljubljana

summary: Teriparatide, a synthetic peptide, mimics the initial 34 amino acids of the natural human parathyroid hormone (PTH). Studies suggest that PTH reduces osteoblast cell death and activates dormant bone-lining cells. Additionally, it hinders the conversion of mesenchymal stem cells (MSCs) into adipocytes while promoting their differentiation into osteoblasts. However, previous research has also revealed PTH's detrimental effects on bone, demonstrating its catabolic nature. These effects vary depending on the type of treatment; intermittent therapy may enhance osteoblast formation, while continuous treatment may inhibit it. To understand the underlying mechanism of this disparity, a thorough grasp of signaling pathways and various mechanisms governing bone cell activity and overall bone regeneration is essential. Despite significant progress, the exact mechanism by which teriparatide influences bone development and equilibrium remains partially understood. Our study aimed to unravel teriparatide's bone-related mechanisms through microRNAs (miRNAs). Human mesenchymal stem cells underwent treatment with teriparatide at various stages of their transformation into osteoblasts, and samples were gathered at the end of the process (day 21).

overall_design:
To elucidate the effects of different PTH treatments we treated MSCs through the process of osteogenic differentiation.
We then performed differential miRNA analysis on treated MSCs after 21 days.
Differential miRNA expression analysis of treated MSCs versus untreated cells.

last: Vrscaj

first: Janja

last: Marc

first: Barbara

last: Ostanek


title: Impact of Nfat5 on normoxia- and hypoxia-exposed mouse lung endothelial cells

city: Heidelberg

first: Thomas

last: Korff

address: INF 326

department: Institute of Physiology and Pathophysiology

country: Germany

institute: Heidelberg University

summary: Chronic hypoxia causes detrimental structural alterations in the lung, which are partially dependent on stress responses of the endothelium. In this context, we revealed that hypoxia-exposed murine lung endothelial cells (MLEC) activate nuclear factor of activated T-cells 5 (NFAT5/TonEBP) - a transcription factor that adjusts the cellular transcriptome to cope with multiple environmental stressors. Here, we studied the impact of NFAT5 on hypoxia-induced gene expression in MLEC by comparing the transcriptome of control and Nfat5-deficient MLEC, which were  isolated from mouse lungs after exposure to normoxia and hypoxia for 7 days and processed for scRNA seq.

overall_design: Nfat5fl/fl mice were crossed with Cdh5-CreERT2 mice and genetic ablation (deletion of loxP site-flanked exon 4 of Nfat5) of Nfat5 (Nfat5(EC)-/-) in the corresponding offspring was induced at 10-12 weeks of age by i.p. administration of 1 mg tamoxifen per day for 5 consecutive days or miglyol as solvent control (Nfat5fl/fl). Mice were used in experiments after a recovery period of 2-3 weeks. Mice were exposed to normobaric hypoxia for 7 days by placing them in a hypoxia chamber (A-Chamber/ProOx O2 controller, Biospherix, USA) with free access to drinking water and food (humidity: 50-65%, temperature: 21-23°C) with 10% oxygen and 90% nitrogen. Mouse lungs were digested by applying the Lung Dissociation Kit (Miltenyi Biotec, Bergisch Gladbach, Germany) following manufacturer’s instruction. Single-cell suspensions were prepared by passing the digestion mixture through 18G and 19G cannula syringes and filtered through a 100 µm cell strainer. Erythrocytes were lysed after the addition of ACK buffer (154.4 mM ammonium chloride, 10 mM potassium bicarbonate, and 97.3 μM EDTA tetrasodium salt). CD31high cells were enriched by magnetic activated cell sorting (MACS). To exclude non-(vascular) endothelial cell populations, cell suspensions were treated with the following antibodies: anti-PDPN-AlexaFluor® 488 (eBioscience, Frankfurt am Main, Germany, 53–5381, RRID: AB_1106990), anti-LYVE1-AlexaFluor® 488 (eBioscience, 53–0443, RRID:AB_1633415), anti-CD45-FITC (BD Biosciences, Heidelberg, Germany, 553080, RRID:AB_394609), anti-LY76-FITC (BD Biosciences, 561032, RRID:AB_396936) and anti-EPCAM-FITC (eBioscience, 11-5791, RRID:AB_ 11151709) for 30 min at 4°C in PBS containing 5% FCS. Vascular endothelial cells were simultaneously labelled with anti-CD31-APC (BD Biosciences, 551262, RRID: AB_398497) and anti-CD146-PE-Cy7 (BioLegend, San Diego, CA, USA, 134713, RRID: AB_2563108). Dead cells were excluded by propidium iodide (PI, 1:2000) staining. CD45- LYVE1- LY76‐ PDPN- EPCAM‐ PI‐ CD31+ CD146+ cells were sorted using a BD FACSAria Fusion Flow Cytometer (BD Biosciences, Heidelberg, Germany). Immediately  after sorting, cells were frozen in DMSO-supplemented MLEC medium and shipped to Single Cell Discoveries (Netherlands) for scRNAseq analysis. Upon arrival, cells were thawed, checked for viability (trypan blue), filtered and processed for scRNAseq analysis (~2-5,000 cells/sample) according to the indexing protocol CG000315. Sequencing was performed on Illumina Novaseq 6000 (167 GB, 2x50, S1, HMF, up to 50,000 reads/cell). Data analysis BCL files resulting from sequencing were transformed into FASTQ files using 10x Genomics Cell Ranger mkfastq. FASTQ files were mapped using Cell Ranger count. During sequencing, read 1 was assigned 28 base pairs, and was used to identify the Illumina library barcode, cell barcode and UMI. Read 2 was used to map to the mouse reference transcriptome (mm10). Filtering of empty barcodes was done by using Cell Ranger. Each sample represents the MLEC population isolated from one mouse lung.

first: Hebatullah

last: Laban


title: Non-Invasive, Label-free Image Approaches to Predict Multimodal Molecular Markers in Pluripotency Assessment

city: Tokyo

first: Ryutaro

last: Akiyoshi

address: 2-9-32 Nakacho, Musashino-shi

country: Japan

institute: Yokogawa Electric Corporation

summary: We detail an innovative non-invasive technique to predict gene and protein expression in pluripotent stem cells using advanced bright-field microscopy. The method employs machine learning algorithms to classify cells based on brightfield images, avoiding the need for traditional staining and manual annotation. Our approach uses DeepLearning technology to predict the gene expression (qPCR, RNA-seq) and protein expression (immunostaining, Flowcytometry) of cells from brightfield microscopy images. It provides a robust tool for non-destructive and continuous monitoring of the pluripotency status of stem cells, which will greatly advance regenerative medicine. It will be an approach that will contribute significantly to the manufacturing process of cellular products, especially where non-destructive and continuous monitoring is required.

overall_design:
Four different culture conditions were used for following iPSC maintenance and propagation. (1) Control: Cultured as in the general iPSC maintenance culture method.　(2) Low nutrient: Cultured in inactivated medium (StemFit medium with heat treatment at 56°C for 30 min). (3) Differentiation: Cultured using differentiation medium (DMEM medium supplemented with 10% FBS, 1% MEM NEAA, 1% GlutaMax). (4) Physical stimulas: iPS cells were suspended by pipetting 20 times at passaging, and then cultured in a general way. Microscopic imaging was performed on days 1 and 4 under these four conditions, and various evaluations (qPCR, Flow Cytometry, immunocytochemistry, and RNA-seq) were performed on the cells after the imaging on day 5. Four conditions were placed in the same 6-well plate, one well per condition, and three plates were prepared in the same way (N=3).RNA-seq data were tied to each culture condition and named as follows. (1) control: YOK1, (2)Low nutrient: YOK2, (3)Differentiation: YOK3, (4)Physical stimulas: YOK4
YOK1-1 represents the first RNA-seq data of the YOK1 condition, for which data were acquired at N = 3.

first: Ayako

last: Yachie

first: Takeshi

last: Hase

first: Mayuri

last: Sathiyananthavel


title: Gene expression profile at single cell level of T cells from PBMC

city: Seoul

first: Cho-Rong

last: Lee

address: 1, Gwanak-ro

country: South Korea

institute: Seoul national university

summary: We used single cell RNA sequencing to analyze the diversity of T cells in PBMC

overall_design: T cells from healthy donor and patient were isolated by FACS using scRNA-seq

first: Sung-Gyoo

last: Park


title: Human α-synuclein overexpression upregulates SKOR1 in a rat model of simulated nigrostriatal aging

city: Cork

first: Gerard

last: O'Keeffe

address: Western Gateway Building, Western Road

department: Anatomy and Neuorscience

country: Ireland

institute: University College Cork

summary: Parkinson’s disease is characterised by progressive loss of dopaminergic (DA) neurons from the substantia nigra (SN) and intraneuronal accumulation of α-synuclein (αSyn) protein. Age is the biggest risk factor for Parkinson’s and has been suggested to create a vulnerable pre-parkinsonian state, but the drivers of this association are unclear. It is known that aging leads to increased αSyn expression in DA neurons and it is possible that this may alter molecular processes that are central to maintaining nigrostriatal integrity. To investigate this, adult female Sprague-Dawley rats received unilateral intranigral injection of adeno-associated viral (AAV) vector carrying wild-type human αSyn (AAV-αSyn) or control vector (AAV-Null). AAV-αSyn induced no detrimental effects on motor behaviour, but there was expression of human wild-type αSyn throughout the midbrain and ipsilateral striatum at 20 weeks post-surgery. Microarray analysis revealed that the gene most-upregulated in the ipsilateral SN of the AAV-αSyn group was the SKI Family Transcriptional Corepressor 1 (SKOR1). Bioenergetic state analysis of mitochondrial function found that SKOR1 overexpression reduced the maximum rate of cellular respiration. Furthermore, SKOR1 impaired neurite growth to the same extent as α-synuclein, and inhibited BMP-SMAD-dependent transcription, a pathway that promotes DA neuron survival and growth. Given the normal influence of ageing on DA neuron loss in human SN, the extent to which αSyn upregulates SKOR1 may influence whether an individual undergoes normal nigrostriatal ageing or reaches a threshold for prodromal PD. This provides new insight into mechanisms through which age-related changes may influence molecular mechanisms controlling neuronal integrity across the lifespan.

overall_design: Twenty adult female Sprague‐Dawley rats (Envigo, UK) were assigned to either an AAV-Null (n=10) or AAV-αSyn (n=10) group and received unilateral stereotaxic injection of either AAV6-Null (5.3×109 vg/µL) or AAV6-αSyn (5.2×1010 vg/µL) (Vector Biosystems Inc) into the substantia nigra at coordinates: AP ‐5.3, ML ± 2.0, DV ‐7.2 relative to bregma. At 20 weeks post-surgery, animals were euthanised by decapitation under isoflurane anaesthesia (5% in O2).

first: Noelia

last: Morales-Prieto

first: Louise

middle: M

last: Collins

first: Aideen

last: Sullivan


title: Multi-omic signatures of sarcoidosis and progression in bronchoalveolar lavage cells [miRNA]

first: Ivana

last: Yang

first: Iain

last: Konigsberg

first: Nancy

last: Lin

first: Lisa

last: Maier


title: Paternal methamphetamine exposure differentially affects first and second generations in mice

city: Shinagawa-ku

first: Sakiko

last: Munetomo-Aoki

state: Tokyo

address: 1-5-8, Hatanodai

department: School of Pharmacy

institute: Showa University Graduate School

summary: To determine whether paternal methamphetamine exposure alters the development, behavior and gene expression of first (F1) and second (F2) generations. We administrated methamphetamine (5 mg/kg) or saline (10 ml/kg) to sire (F0) and performed gene expression profiling analysis using data obtained from RNA-seq of striatum of 7-week-old male F1 mice. Male F1 mice were mated with untreated female mice to obtain F2 mice. Male F1 mice were mated with female mice to obtain F2 mice. RNA-seq of striatum of 7-week-old male F2 mice was also analyzed in the same way.

overall_design: Comparative gene expression profiling analysis of 7-week-old striatum RNA-seq data from male F1 mice born to methamphetamine or saline-treated sires. The same analysis of male F2 mice obtained by mating male F1 mice with untreated females.

first: Asuka

last: Kaizaki-Mitsumoto

first: Ryota

last: Nakano

first: Satoshi

last: Numazawa


title: Effect of PFKP deletion on gene expression in laryngeal carcinoma cells LIU-LSC-1

city: Hefei

first: Yue

last: Zhu

address: The First Affiliated Hospital of Anhui Medical University, Hefei, Anhui, China

summary: To investigate the role of PFKP in laryngea cancer, we established a LIU-LSC-1 cell line with PFKP knocked down by lentivirus technique. We then performed gene expression profiling targeting the RNA-seq data of the control and PFKP knockdown groups.

overall_design: Gene expression profile analysis of RNA-seq data from control group (shNC) and PFKP knockdown group (shPFKP) in LIU-LSC-1 cells.

first: Weiwei

last: Liu

first: Yehai


title: Vascularized Macrophage-Islet Organoids to Model Immune-Mediated Pancreatic beta cell Pyroptosis upon Viral Infection [snATACseq.VMI]

city: New York

first: Shuibing

last: Chen

address: A 827B, 1300 York Ave

department: Surgery

institute: Weill Cornell Medical College

summary: Immune cells play critical roles in causing host damage in various diseases, including infectious diseases. However, there is a paucity of human models to study immune-mediated host damage. Here, we systematically analyzed the changes in immune cell composition for COVID-19 pancreatic autopsy samples using the GeoMx spatial RNA and protein platform and found accumulation of proinflammatory macrophages. Single cell RNA-seq analysis of human islets upon SARS-CoV-2 or Coxsackievirus B4 (CVB4) infection identified the activation of proinflammatory macrophages and pyroptosis in β cells. To distinguish whether the virus or proinflammatory macrophages mediated β cell pyroptosis, we developed human pluripotent stem cell (hPSC)-derived vascularized macrophage-islet (VMI) organoids, containing pancreatic endocrine cells, endothelial cells, and macrophages. Both pancreatic β cells and endothelial cells in VMI organoids demonstrated improved marker expression and function compared to separately cultured cells. Additionally, proinflammatory macrophages in VMI organoids were found to induce β cell pyroptosis. Further mechanistic studies identified TNFSF12-TNFRSF12A as a contributor to proinflammatory macrophage-mediated β cell pyroptosis. Together, this study established a hPSC-derived VMI organoid as a valuable tool for studying immune cell-mediated host damage and uncovered previous unknown mechanism of β cell damage in viral infection.

overall_design: We conducted RNA-seq analysis of unstimulated or LPS and IFN-g induced hESC-derived macrophages to identify the expression of pro-inflammatory genes. Then, we performed single cell RNA-seq analysis of human islets upon SARS-CoV-2 or Coxsackievirus B4 (CVB4) infection to identify the activation of proinflammatory macrophages and pyroptosis in β cells. To distinguish whether the virus or proinflammatory macrophages mediated β cell pyroptosis, we developed human pluripotent stem cell (hPSC)-derived vascularized macrophage-islet (VMI) organoids, containing pancreatic endocrine cells, endothelial cells, and macrophages and performed scRNA-seq and snATAC-seq.

first: Liuliu

first: Yuling

last: Han

first: Tuo

last: Zhang


title: Methazolamide can treat atherosclerosis

city: Qingdao

first: XIAOTIAN

last: CHANG

state: Shandong

address: 1677 Wutaishan road

institute: The Affiliated Hospital of Qingdao University

summary: Methoxazolamide (MTZ) has been confirmed to treat atherosclerosis (AS) by inhibiting calcification in aortic tissues. This study investigated the therapeutic mechanism. In this study, ApoE-/- mice were fed a high-fat diet to establish an AS model and then treated with MTZ. The aortic tissues were analyzed using single-cell sequencing. MTZ significantly increased the proportions of B-1/MZB cells, CD8+CD122+ Treg-like cell. Those cells can exert atheroprotective role by suppressing immune response and proinflammation. Meanwhile, MTZ decreased the proportions of nonclassical CD14+CD16++ monocytes and Spp1+ macrophages. Those cells can exert atherogenic role by suppressing proinflammation, calcification and tissue remodeling. The result suggested that MTZ is a potential anti-AS drug that regulates proportions of immunosuppressive cells and proinflammatory cells as well as their calcification-stimulating gene expression.

first: Xiaotian

last: Chang

first: Kehua

last: Fang


title: The effects of inhaled corticosteroids on healthy airways

city: Oxford

first: Timothy

last: Hinks

state: United Kingdom

address: John Radcliffe Hospital, Headley Way, Oxford

department: NDM Experimental Medicine - New College

institute: University of  Oxford

summary: The effects of inhaled corticosteroids (ICS) on healthy airways are poorly defined. Our objective was to delineate the effects of ICS on gene expression in healthy airways, without confounding caused by changes in disease-related genes and disease-related alterations in ICS-responsiveness. We performed a randomised open-label bronchoscopy study of high dose ICS therapy in 30 healthy adult volunteers randomised 2:1 to i) fluticasone propionate 500 mcg bd or ii) no treatment (control), for 4 weeks. Laboratory staff were blinded to allocation. Biopsies and brushings were analysed by bulk RNA sequencing. ICS treatment upregulated 72 genes in brushings and 53 genes in biopsies, and downregulated 82 genes in brushings and 416 genes in biopsies. The most downregulated genes in both tissues were canonical markers of type-2 inflammation (FCER1A, CPA3, IL33, CLEC10A, SERPINB10 and CCR5), T cell-mediated adaptive immunity (TARP, TRBC1, TRBC2, PTPN22, TRAC, CD2, CD8A, HLA-DQB2, CD96, PTPN7), B cell immunity (CD20, immunoglobulin heavy and light chains), and innate immunity, including CD48, Hobit, RANTES, Langerin and GFI1. An IL-17-dependent gene signature was not upregulated by ICS. In healthy airways, 4-week ICS exposure reduces gene expression related to both innate and adaptive immunity, and reduces markers of type-2 inflammation. This implies that homeostasis in health involves tonic type-2 signalling in the airway mucosa, which is exquisitely sensitive to ICS.

overall_design:
Randomised open-label bronchoscopy study of high dose ICS therapy in 30 healthy adult volunteers randomised 2:1 to i) fluticasone propionate 500 mcg bd or ii) no treatment, for 4 weeks.RNAseq was performed on bronchial brushes and biopsies pre-treatment or control observation, and 4 weeks later
NOTE FROM SUBMITTER: human derived data, we would request not to submit the raw fastq sequences for privacy protection..

first: Emanuele

last: Marchi

first: Matthew

last: Richardson

first: Latifa

last: Chachi

first: Fiona

last: Symon

first: Poojitha

last: Rajasekar

first: Rachel

last: Clifford

first: Beverley

last: Hargadon

first: Cary

last: Austin

first: Salman

last: Siddiqui

first: Jordan

last: Mar

first: Joseph

last: Arron

first: David

last: Choy

first: Peter

last: Bradding


title: BRD4 inhibitor GNE987 exerts anti-cancer effects in Osteosarcoma

city: Suzhou

first: Zimu

last: Zhang

state: Jiangsu

address: Zhongnan Street

institute: Children’s Hospital of Soochow University

summary: GNE987 is a newly developed von Hippel-Lindau tumor suppressor (VHL)-based pan-BET-targeting PROTAC, which can bind to the target proteins (BET proteins, including BRD2, BRD3, and BRD4) and recruit them to the ubiquitin/proteasome system for selective degradation. In the our study, ChIP-Seq analysis was performed to explore the effect of GNE987 on osteosarcoma cells.

overall_design: Genome binding/occupancy profiling of histone H3K27ac in osteosarcoma cells were generated by ChIP-Seq using DNBSEQ-G400

first: Wu

last: Di

first: Pan

last: Jian


title: Zebrafish genetic model of accelerated muscle atrophy associated with Fbxo32/Atrogin-1 expression

city: Portland

first: Thomas

middle: Dexter

last: Morse

state: ME

address: 154 New Island Ave

institute: MDIBL

summary: The degenerative loss of muscle associated with aging leading to muscular atrophy is called sarcopenia. Currently, practicing regular physical exercise is the only efficient way to delay the onset of sarcopenia. To identify therapeutic targets to suppress the symptoms of aging, there is a need for in vivo model organisms of accelerated muscle degeneration and atrophy. The zebrafish harbors hallmarks of aging, notably mitochondrial dysfunction, telomeres shortening and accumulation of senescent cells. Unfortunately, zebrafish aged slowly, and no specific zebrafish models of accelerated muscle atrophy associated with aging or sarcopenia are currently available. We have developed a new genetic tool to efficiently accelerate muscle fibers degeneration and muscle tissue atrophy in zebrafish larvae and adults. We used a gain-of-function strategy with a molecule that has been shown to be necessary and sufficient to induce muscle atrophy and a sarcopenia phenotype in mammals: Atrogin1 (also named Fbxo32). We report the generation, validation, and characterization of a zebrafish genetic model of accelerated muscle atrophy, the sarcofish. We demonstrated that specific expression of Atrogin1 in skeletal muscle tissue induces a muscle atrophic phenotype associated with locomotion dysfunctions in both larvae and adult fish. We identified that degradation of the myosin light chain as an event occurring before the degeneration of muscle fibers. Biological processes associated with muscle aging such as inflammation, proteolysis, apoptosis and stress response are upregulated in the sarcofish. Surprisingly, we observed a strong correlation between muscle fibers degeneration and a reduction of neuromuscular junctions in the peripheral nervous system, as well as neuronal cell bodies in the spinal cord, suggesting that muscle atrophy could underly a neurodegenerative phenotype in the central nervous system. Finally, while sarcofish larvae harbor regeneration of muscle fibers, spinal neurons and recover locomotive functions, adult sarcofish have impaired regenerative capacities as observed in mammals during muscle aging. In the future, the sarcofish could open new strategies in the fight against the sarcopenia disease to test therapeutic molecules aiming to treat or alleviate the symptoms of muscle aging.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for zebrafish sarcopinia phenotype (Fbxo32 over-expressed) vs wild-type at time points 6d and 8d post induced muscle attrophy

first: Romain

last: Menard

first: Elena

last: Morin

first: Marko

last: Pende

first: Caroline

last: Halluin

first: Dexter

first: Frederic

last: Bonnet

first: Jijia

last: Li

first: Laetitia

last: Lancelot

first: Prayag

last: Murawala

first: Cedric

last: Dray

first: Jean-Philippe

last: Pradere

last: Madelaine


title: Circ_0008146 exacerbates ferroptosis via regulating miR-342-5p/ACSL4 axis after cerebral ischemia-reperfusion in mice

city: Nanjing

first: Qiang

last: Peng

address: No. 68 Changle Road

department: Neurology

institute: Nanjing First Hospital, Nanjing Medical University

summary:
Background: Characterized by high incidence, mutilation, and death rate, acute ischemic stroke (AIS) has threatened people’s health seriously. Therefore, there is an urge need for early diagnosis and effective treatment of AIS. The purpose of this study was to clarify the regulatory role and potential molecular mechanism of miR-342-5p and circRNA_0008146 in AIS.
Materials and Methods: Small RNA sequencing analysis was performed to screen differentially expressed miRNAs between the AIS and control group. Middle cerebral artery occlusion/reperfusion (MCAO/R) models were constructed in C57BL/6J mice. The circRNA_0008146 and miRNA expression levels were detected by quantitative real-time PCR. Assay kits were used to determine Fe2+ levels and assess a battery of oxidative stress indicators, including ROS, MDA, LPO, SOD, and GSH/GSSG. The content of ACSL4 was measured by Western blot. The neurobehavioral manifestation was evaluated using the modified Neurological Severity (mNSS) Score, rotarod test, and corner test. TTC staining was employed to detect cerebral infarction volume. Nissl staining was adapted to detect histological change and neuronal damage.
Results: RNA sequencing analysis revealed miR-342-5p was significantly downregulated in the plasma of AIS patients. MiR-342-5p inhibited oxidative stress and RSL3-induced ferroptosis after cerebral ischemic-reperfusion injury in vivo by targeting ferroptosis-related gene ACSL4. CircRNA_0008146 acted as a sponge of miR-342-5p, and overexpression of circRNA_0008146 increased brain damage in stroke mice. CircRNA_0008146 contributed to ferroptosis in cerebral infraction via sponging miR-342-5p to regulate ACSL4.
Conclusion: CircRNA_0008146/miR-342-5p/ACSL4 axis regulate ferroptosis after cerebral ischemia-reperfusion, which may be a potential therapeutic target for ischemic stroke.

overall_design: miRNA profiles of plasma from 2 acute ischemic stroke patients and 2 healthy controls


title: Analysis of transcriptome in hypoxia-exposed lungs upon KO of Nfat5 in ECs

city: Mannheim

first: Carolina

last: Delatorre

address: Theodor-Kutzer-Ufer 1-3

institute: University Heidelberg

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Bromodomain inhibitor PFI-3 induces cellular plasticity and enhances iPSC reprogramming efficiency by targeting extracellular matrix

city: Durham

first: James

last: Ward

state: NC

address: 111 TW Alexander Dr

department: Integrative Bioinformatics

institute: NIEHS

summary: The study assessed fibroblast reprogramming into induced pluripotent stem cells (iPS), and the effects on efficiency of iPS reprogramming by treatment with 10-uM PFI-3. Samples were collected across multiple time points during reprogramming: day 0, 3, 5, 7, 14, and day 21, each in the presence or absence of PFI-3 treatment, with n=3 biological replicates.

overall_design: Human dermal fibroblasts were studied in two independent experimental batches. Treatment with iPS reprogramming media was started at day 0, as described in Yang et al 2023. Samples were also treated with either vehicle control, or 10-uM working concentration of PFI-3. Each treatment time was produced as n=3 biological replicate.

first: Jun

last: Yang

first: Harriet

middle: K

last: Kinyamu

first: Erica

last: Scappini

first: Trevor

last: Archer


title: Immune Editing Roles of SIGLEC15 in Tumor Microenvironment [scRNA-seq]

city: Shanghai

first: Zheng

last: Chen

state: Shanghai

address: No.180 Fenglin Road, Xuhui District

department: Department of Liver Surgery and Transplantation, Liver Cancer Institute

institute: Zhongshan Hospital Fudan University

summary: SIGLEC15 in tumor microenvironment could dampen cytotoxic functions of T cells, and we blocked SIGLEC15 with mono-antibody to examine functional changes of cell populations  in comparison to treatment of IgG.

overall_design: Percutaneous tumor grafts from C57 mice, treated by IgG (n=3) or SIGLEC15 (n=3) antibody, were trimmed and digested for cell pelletes, and CD45 positive selection magnetic beads were used to enrich CD45+ cells. After examination of cell viability (>90%), CD45+ cells and unsorted cell pelletes were loaded for sample preparation at the ratio of 1:1.


title: Vascularized Macrophage-Islet Organoids to Model Immune-Mediated Pancreatic beta cell Pyroptosis upon Viral Infection [scRNAseq.VMI]

first: Shuibing

state: NY

first: Liuliu

first: Yuling

last: Han

first: Tuo

last: Zhang


title: CRISPR-Cas9 knockout screen informs efficient reduction of the Komagataella phaffii secretome

city: Cambridge

first: Charles

middle: Arthur

last: Whittaker

state: MA

institute: Koch Institute

summary: The yeast Komagataella phaffii is a promising alternative host for manufacturing of therapeutic proteins. Deletion of unneeded endogenous proteins could increase the secreted titer of recombinant proteins by redirecting cellular resources. Genetic engineering in non-model hosts is hampered by limited annotation of genes, especially essential genes. In this study, we identified the set of endogenous secreted proteins in K. phaffii and attempted to disrupt these genes. We designed, transformed, and sequenced a pooled CRISPR-Cas9 knockout library to determine which genes are essential. With this knowledge, we rapidly disrupted up to 9 consecutive genes in K. phaffii. Engineered strains exhibited a ~20x increase in the production of human serum albumin and a 2x increase in the production of a monoclonal antibody. The pooled CRISPR-Cas9 library and knowledge of gene essentiality reported here will facilitate future efforts to engineer K. phaffii for production of other recombinant therapeutic proteins and enzymes.

overall_design: This experiment is a head to head comparison of the transcriptomes of two yeast strains. The strains are compared during expression of two proteins, human serum albumin (HSA) and trastuzumab.

first: Neil

middle: C

last: Dalvie

first: Timothy

last: Lorgeree

first: Yuchen

first: Sergio

last: Rodriguez-Aponte

first: John

middle: J

last: Clark

first: Amanda

last: del Rosario

first: Kerry

last: Love

first: J


title: Analysis of SKMEL-103, AsPC1 and RH-36 cells after treatment with BAY 11-7082

city: Birmimgham

first: Romi

last: Gupta

state: Alabama

address: 720, 20th street south

institute: University of Alabama at Birmingham

summary: To identify the gene expression changes in NRAS mutant cell line SKMEL-103, KRAS mutant cell line AsPC1 and HRAS mutant cell line RH-36 upon BAY 11-7082 treatment, we analyzed these cell line with either control DMSO or BAY 11-7082 treatment via RNA sequencing.

overall_design: SKMEL-103, AsPC1 and RH-36 cells were plated, samples were either treated with DMSO or BAY 11-7082 (5-micromolar for 48 hours), cells were harvested and were used to prepare the total RNA, followed by RiboRNA depletion with random priming and RNASeq Illumina Hi-Seq 2500 platform, with three biological replicates for each sample.


title: Vascularized Macrophage-Islet Organoids to Model Immune-Mediated Pancreatic beta cell Pyroptosis upon Viral Infection


title: Vascularized Macrophage-Islet Organoids to Model Immune-Mediated Pancreatic beta cell Pyroptosis upon Viral Infection [bulkRNAseq]


title: Vascularized Macrophage-Islet Organoids to Model Immune-Mediated Pancreatic beta cell Pyroptosis upon Viral Infection [scRNAseq.islets]


title: Activation of skeletal ChREBP increases IMFlevel in mice and Broiler Chickens

city: Nanning of Guangxi

first: peng

last: wang

address: Xixiangtang

institute: Guangxi University

summary: Intramuscular fat (IMF) content is one of the key factors affecting meat quality. Carbohydrate response element binding protein (ChREBP) can promote glucose metabolism and fat synthesis by activating the expression of glycolysis and lipogenesis-related genes, but its function and regulatory mechanism in IMF are still unclear. This study evaluated the correlation between TG content and ChREBP expression level in muscles of different species to reveal the relationship between ChREBP and IMF. RNA-seq analysis showed that overexpressing ChREBP in C2C12 cells can significantly up-regulate fatty acid synthesis pathways, while significantly down-regulate the expression levels of muscle development pathways and related genes. In vivo, we found that overexpression of ChREBP or activation by fructose significantly increased the triglyceride content of the tibialis anterior muscle (TA) and the IMF content of yellow feather chicken leg and breast muscle in mice. And lipidomics data also found that feeding fructose can change the lipid composition of yellow feather broiler breast muscle and improve the flavor substances. This study demonstrated that ChREBP is a key gene regulating IMF deposition, providing a new target for genetic selection and nutritional regulation of IMF content.

overall_design: Comparative gene expression profiling of RNA-seq data from C2C12 cells overexpressing ChREBP.

first: Peng

last: Wang

first: Haihan

last: Xiao

first: Tian

last: Wu

first: Ziyi

last: Song


title: Tumor cell-intrinsic Piezo2 drives radioresistance by impairing CD8+ T cell stemness maintenance

first: Wenwen

last: Li

address: Room 1004, West #5 Building, 280 South Chongqing Road, Shanghai 200025, China

institute: Shanghai Institute of Immunology; Department of Immunology and Microbiology, Shanghai Jiao Tong University School of Medicine

summary: Piezo2 impedes stemness maintenance of CD8+ T cells by downregulating IL-15 production after radiotherapy, thereby leading to cancer radioresistance.

overall_design: WT-MC38 (CRISPR-Ctrl) or Piezo2-KO MC38 (Piezo2-gRNA1) tumor cells were subjected to single fraction of X-ray (30 Gy). 60 hours later, total RNA from samples was extracted using Trizol Reagent following standard protocols and mRNA sequencing was performed.


title: Integrin αV is a prognostic marker for cutaneous squamous cell carcinoma relapse

city: New Brunswick

first: Victoria

last: da Silva-Diz

address: 195 Little Albany Street

institute: Rutgers University

summary: Early cutaneous squamous cell carcinomas (cSCCs) show epithelial differentiation features and good prognosis, whereas advanced cSCCs present mesenchymal traits and are associated with tumor relapse, metastasis, and poor survival. Prognostic biomarkers for cSCC relapse must therefore be found that accurately predict the clinical course of the disease, since established markers are suboptimal. Using mouse models of cSCC progression, we showed that the emergence of epithelial plastic cancer cells with a hybrid epithelial/mesenchymal phenotype promotes tumor progression to a mesenchymal state. These cells can be identified early on by the expression of integrin αV (ITGAV). Analysis of ITGAV expression in a cohort of primary cSCCs from patients provided prognostic information about the risk of relapse beyond current histopathological parameters. Together, our findings provide an opportunity to clinically implement ITGAV by standard immunodetection approaches and thereby improve patient stratification and therapeutic management.

overall_design: Excised mouse WD-SCCs and MD/PD-SCCs were mechanically minced and enzymatically digested overnight at 37ºC with collagenase type I and dispase. Cell suspensions were filtered and depleted of red blood cells and  endothelial cells. Then, full epithelial cancer cells expressing EpCAM and alpha 6-integrin (a6-integrin+/CD45-/EpCAM+ cells), were isolated by flow cytometry-sorter from 3 independent WD-SCCs, and EpCAM+ plastic cancer cells were isolated from 3 independent MD/PD-SCCs. RNA extraction of isolated cells and hybridization on Affymetrix microarrays was carried out to compare whole gene profiling of these populations of cancer cells.

first: Marta

last: Lopez-Cerda

first: Laura

last: Lorenzo-Sanz

first: Sandra

last: Llop

first: Rosa

last: Penin

first: Josep

last: Oriol Bermejo

first: Juan

last: Martin-Liberal

first: Purificación

last: Muñoz


title: Effect of Transcriptional-like Effectors  (TALE) on Nicotiana benthamiana leaves

city: Valencia

first: Jose

last: Gadea

address: Ingeniero Fausto Elio

department: Stress

country: Spain

institute: IBMCP

summary: To investigate gene expression upon inoculation of Nicotiana benthamiana leaves with a TALE inducing cell death

overall_design: Comparative gene expression profiling between TALE50, TALE 55 (inducing cell death) and TALE3 (not inducing cell death) at 36h after infiltration


title: Transcriptional responses to arbuscular mycorrhizal symbiosis development are conserved in the liverwort Marchantia paleacea

first: Uta

last: Paszkowski

address: 93 Lawrence Weaver Rd

department: Plant Sciences

institute: University of Cambridge

summary: The mutualistic arbuscular mycorrhizal (AM) symbiosis arose in land plants more than 450 million years ago. This symbiosis is still widely found across  major land plant lineages, including bryophytes. Despite its broad taxonomic distribution, little is known about the molecular components underpinning symbiosis outside of flowering plants. Here, we demonstrate that a broad AM genetic programme is conserved amongst land plants. In this study, we characterised the dynamic response of the liverwort Marchantia paleacea to Rhizophagus irregularis colonization by time-resolved transcriptomics across three stages of symbiosis. Comparative analysis of transcriptional responses to symbiosis in the liverwort M. paleacea and the legume Medicago truncatula further revealed evolutionarily conserved expression patterns for genes underpinning pre-symbiotic signalling, intracellular colonization and nutrient exchange. This study demonstrates that the genetic machinery regulating key aspects of symbiosis in plant hosts is largely conserved and coregulated across land plants.

overall_design: To understand the molecular changes associated with development of AM symbiosis in Marchantia paleacea, we characterised the transcriptional response of the liverwort at five, eight- and eleven-weeks post-inoculation (WPI) with the AM fungus Rhizophagus irregularis. For every time point, transcript levels of genes in mock-inoculated (control) thalli were compared to colonised M. paleacea thalli.

first: Mara

last: Sgroi

first: Karina

last: Jimenez

first: Armando

last: Bravo


overall_design: MC38-WT tumor cells were subjected to single fraction of X-ray (30 Gy). 60 hours later, total RNA from samples was extracted using Trizol Reagent following standard protocols and mRNA sequencing was performed.


title: Analysis of transcriptome in hypoxia(7d)-exposed lungs upon KO of Nfat5 in ECs

first: Carolina

last: Delatorre

summary: Chronic hypoxic stress stimulates lung endothelial cells to promote vascular remodeling processes, which - in the long run - increase the resistance of pulmonary arteries. While several molecular determinants promoting these maladaptive changes have been delineated, their transcriptional regulation is not well studied. In this context, we revealed that hypoxia activates nuclear factor of activated T-cells 5 (NFAT5/TonEBP) in murine lung endothelial cells (MLECs) - a transcription factor that regulates the adjustment of the cellular transcriptome to cope with osmotic, biomechanical or metabolic environmental stressors. Here, we studied the functional relevance of NFAT5 for the control of endothelial hypoxic stress responses in the lung. Genetic ablation of Nfat5 in endothelial cells did not evoke any obvious phenotypic alterations under normoxia. However, microarray-based transcriptome analyses of lung tissue revealed significant alterations 7 but not 21 days after exposure to normobaric hypoxia (10% O2).

overall_design: RNA was isolated from lungs of 4 male/female Nfat5fl/fl (control) and Nfat5(EC)-/- (EC-specific KO) mice exposed to hypoxia for 7 days.  Lung preparations containing contaminating tissue (e.g. thymus) or showing inadequate (expression) responses to hypoxia were excluded from further analyses.

first: Thomas

last: Korff


title: Analysis of transcriptome in normoxia/hypoxia(21d)-exposed lungs upon KO of Nfat5 in ECs

overall_design: RNA was isolated from lungs of 4 male/female Nfat5fl/fl (control) and Nfat5(EC)-/- (EC-specific KO) mice exposed to normoxia and hypoxia for 21 days. Lung preparations containing contaminating tissue (e.g. thymus) or showing inadequate (expression) responses to hypoxia were excluded from further analyses.