Distinct Gene Expression Patterns Research Articles

Comparable CD8+ T-cell responses to SARS-CoV-2 vaccination in single-cell transcriptomics of recently allogeneic transplanted patients and healthy individuals.

Despite extensive research on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccination responses in healthy individuals, there is comparatively little known beyond antibody titers and T-cell responses in the vulnerable cohort of patients after allogeneic hematopoietic stem cell transplantation (ASCT).In this study, we assessed the serological response and performed longitudinal multimodal analyses including T-cell functionality and single-cell RNA sequencing combined with T cell receptor (TCR)/B cell receptor (BCR) profiling in the context of BNT162b2 vaccination in ASCT patients. In addition, these data were compared to publicly available data sets of healthy vaccinees.Protective antibody titers were achieved in 40% of patients. We identified a distorted B- and T-cell distribution, a reduced TCR diversity, and increased levels of exhaustion marker expression as possible causes for the poorer vaccine response rates in ASCT patients. Immunoglobulin heavy chain gene rearrangement after vaccination proved to be highly variable in ASCT patients. Changes in TCRα and TCRβ gene rearrangement after vaccination differed from patterns observed in healthy vaccinees. Crucially, ASCT patients elicited comparable proportions of SARS-CoV-2 vaccine-induced (VI) CD8+T-cells, characterized by a distinct gene expression pattern that is associated with SARS-CoV-2 specificity in healthy individuals.Our study underlines the impaired immune system and thus the lower vaccine response rates in ASCT patients. However, since protective vaccine responses and VICD8+T-cells can be induced in part of ASCT patients, our data advocate early posttransplant vaccination due to the high risk of infection in this vulnerable group.

(059) Epigenetic Profiles of Men with Peyronie’s Disease

Abstract Introduction Peyronie’s Disease (PD) can significantly impact sexual function. A hereditary predisposition to PD suggests that genetic variants may exist, and gene expression may modulate the penetrance of the disease. Identification of changes in epigenetic or gene expression profiles would facilitate a deeper understanding of the disease and provide insight for more targeted therapies. Objective Our objective is to determine if predisposition to PD is correlated with epigenetic variations and/or gene expression changes, as indicated by DNA methylation assessment and RNA sequencing. Methods Penile tunica albuginea samples were obtained from men with PD undergoing penile prosthesis placement. Penile sample from these men were not from fibrotic plaques. Control samples were collected from men with erectile dysfunction (ED) without PD undergoing penile prosthesis placement. DNA methylation analyses were performed on homogenized penile tissue samples via an Illumina Human MethylationEPIC BeadChip v2 array. Using the minfi package in R, beta values were produced for all 936,990 CpG sites for each sample and SWAN normalization was applied. Differentially methylated regions (DMR) were found via USEQ with a threshold Wilcoxon FDR score of 40. Gene expression levels were examined in PD and ED samples via RNA-sequencing. Gene expression data was compared against epigenetic data to determine if any epigenetics changes identified affected gene expression profiles. Results A total of 24 tissue samples were collected: 12 ED and 12 PD. USEQ yielded 36 total DMRs with a FDR score of 40 or greater (p-value<0.0001). Of these 36 DMRs, a Stanford GREAT analysis revealed 60 total gene-region associations and five implicated biological processes: anterior/posterior pattern specification, chordate embryonic development, embryo development (ending in birth or egg hatching), somitogenesis, and pattern specification process. Furthermore, differential gene expression analysis identified 76 genes with significant expression changes between PD and ED groups, of which 17 genes were upregulated while 59 were downregulated. Long noncoding RNAs and the keratocan gene showed notable expression differences between PD and ED groups. GO enrichment analysis revealed significant terms related to DNA binding and protein binding. None of the differentially regulated genes overlapped with identified DMRs. Conclusions This study identified distinct gene expression and DNA methylation patterns between PD and ED groups, despite PD samples not being taken from plaque regions. Upregulated genes in PD included long noncoding RNAs and the keratocan gene, suggesting their involvement in disease pathogenesis. Long noncoding RNA overexpression may serve a regulatory function in gene expression or cell processes while overexpression of the keratocan gene may promote collagen accumulation and remodeling, leading to fibrous plaque formation in the affected area. Likewise, DMRs and their region-gene associations suggest a role in early embryonic development for pathogenesis. Although RNA-seq did not indicate any significant differentially expressed genes that matched with gene-region associations correlated with DMRs, the significance and quantity of DMRs between ED and PD sample groups is indicative of affected biological processes that manifest in distinct pathophysiologies. Further research is needed to investigate if differentially regulated genes exist in DMRs that exhibited changes but did not meet the criteria threshold for inclusion here. Disclosure Any of the authors act as a consultant, employee or shareholder of an industry for: Paterna bio, Firmtech, Turtle health, Maximus, Carrot, Contraline, endo pharmaceuticals, inherent biosciences, techfields inc, macro trials, alto neurosciences, curebase, vivify medical.

Identification and molecular characterization of genes modulating progression of an oocyte from M-I to M-II in rat ovary.

To achieve oocyte competence for successful fertilization, bidirectional communication between oocyte and granulosa cells is crucial. The acquisition of meiotic competency in oocyte is facilitated by various regulatory genes however, expression pattern of these genes is not well documented during meiotic transition from Metaphase-I to Metaphase-II stage. Therefore, the present research analyzed the expression pattern of regulatory genes that are involved in the transition from M-I to M-II stages in rat oocyte. The analysis of the data was conducted by applying an array of bioinformatic tools. The investigation of gene group interactions was carried out by employing the STRING database, which relies on co-expression information. The gene ontology (GO) analysis was performed utilizing the comparative GO database. Functional annotation forGO and pathway enrichment analysis were performed for genes involved in networking.The GOobtained through computational simulations was subsequently validated using quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis. The findings of our study suggest that there is a distinct gene expression pattern in both the oocyte and granulosa cells. This pattern indicates that oocyte-secreted factors, such as BMP15 and GDF9, play a crucial role in regulating the progression of the meiotic cell cycle from the M-I to M-II stages. We have also examined the level of mRNA expression of genes including CYP11A1, CYP19A1, and STAR, which are crucial for the steroidogenesis. It is fascinating to observe that the oscillatory pattern of specific key genes may hold significance in the process of in vitro oocyte maturation, specifically during the transition from the M-I to M-II stage. It might be useful for determining biomarker genes and potential pathways that play a role in attaining oocyte competency, thereby aiding in the assessment of oocyte quality for the purpose of achieving successful fertilization.

Phylogenetic and functional analysis of tiller angle control homeologs in allotetraploid cotton.

Plants can adapt their growth to optimize light capture in competitive environments, with branch angle being a crucial factor influencing plant phenotype and physiology. Decreased branch angles in cereal crops have been shown to enhance productivity in high-density plantings. The Tiller Angle Control (TAC1) gene, known for regulating tiller inclination in rice and corn, has been found to control branch angle in eudicots. Manipulating TAC1 in field crops like cotton offers the potential for improving crop productivity. Using a homolog-based methodology, we examined the distribution of TAC1-related genes in cotton compared to other angiosperms. Furthermore, tissue-specific qPCR analysis unveiled distinct expression patterns of TAC1 genes in various cotton tissues. To silence highly expressed specific TAC1 homeologs in the stem, we applied CRISPR-Cas9 gene editing and Agrobacterium-mediated transformation, followed by genotyping and subsequent phenotypic validation of the mutants. Gene duplication events of TAC1 specific to the Gossypium lineage were identified, with 3 copies in diploid progenitors and 6 copies in allotetraploid cottons. Sequence analysis of the TAC1 homeologs in Gossypium hirsutum revealed divergence from other angiosperms with 1-2 copies, suggesting possible neo- or sub-functionalization for the duplicated copies. These TAC1 homeologs exhibited distinct gene expression patterns in various tissues over developmental time, with elevated expression of A11G109300 and D11G112200, specifically in flowers and stems, respectively. CRISPR-mediated loss of these TAC1 homeologous genes resulted in a reduction in branch angle and altered petiole angles, and a 5 to 10-fold reduction in TAC1 expression in the mutants, confirming their role in controlling branch and petiole angles. This research provides a promising strategy for genetically engineering branch and petiole angles in commercial cotton varieties, potentially leading to increased productivity.

P119 Single-cell RNA sequencing in Inflammatory Bowel Disease in patients with Primary Sclerosing Cholangitis: a distinct form of colitis

Abstract Background Primary sclerosing cholangitis (PSC) is an inflammatory disorder of the bile ducts, in which Inflammatory bowel disease (IBD) concomitantly occurs (PSC-IBD). Substantial differences in clinical presentation are observed between PSC-IBD and ulcerative colitis (UC). In this study we aim to find distinct pathomechanisms for PSC-IBD using single-cell RNA sequencing. Methods Forty-seven colonic samples of PSC-IBD (n=24), UC (n=18) and non-IBD subjects (n=5) were collected. Whole biopsies were dissociated into single cells using collagenase digestion. Library preparation was done with 10x Genomics and sequencing was performed on an MGI2000 sequencer. Then, differential abundance, differential expression, and cell-cell interaction analyses were performed. Stainings for DUOX2 and HLA-DR were performed on paraffin embedded colon mucosal slides. Results In total, 71,798 high quality cells identified 54 distinct cell types, including a new type of epithelial cells: DUOX2+ enterocytes. DUOX2+ enterocytes are almost exclusively present in diseased inflamed colon and can act as antigen presenting cells through the expression of MHC2 on its surface. A general shift in cell type composition upon inflammation was observed, comparable between PSC and UC samples. However, an increased abundance was seen for stem cells in non-inflamed (NI) PSC, and not in UC-NI, compared to healthy controls. Additionally, we found distinct gene expression patterns between PSC and UC inflammation: while activation of inflammatory monocytes was observed in PSC inflammation, several types of fibroblasts were activated upon UC inflammation. Conclusion We describe DUOX2+ enterocytes, which may play a role in both PSC and UC colitis through antigen presentation. Furthermore, we show that intestinal inflammation in PSC-IBD, as compared to UC, is characterized by distinct mucosal cell composition and cell-specific gene expression patterns. PSC colitis, and not UC colitis, is driven by activation of inflammatory monocytes, with antigen presentation through HLA-DRB1, and activation of CD4+ T cells. Additionally, an increased abundance of stem cells in PSC-NI could be related with the increased colorectal cancer risk in PSC-IBD.

Genome-wide identification of TPS and TPP genes in cultivated peanut (Arachis hypogaea) and functional characterization of AhTPS9 in response to cold stress.

Trehalose is vital for plant metabolism, growth, and stress resilience, relying on Trehalose-6-phosphate synthase (TPS) and Trehalose-6-phosphate phosphatase (TPP) genes. Research on these genes in cultivated peanuts (Arachis hypogaea) is limited. This study employed bioinformatics to identify and analyze AhTPS and AhTPP genes in cultivated peanuts, with subsequent experimental validation of AhTPS9's role in cold tolerance. In the cultivated peanut genome, a total of 16 AhTPS and 17 AhTPP genes were identified. AhTPS and AhTPP genes were observed in phylogenetic analysis, closely related to wild diploid peanuts, respectively. The evolutionary patterns of AhTPS and AhTPP genes were predominantly characterized by gene segmental duplication events and robust purifying selection. A variety of hormone-responsive and stress-related cis-elements were unveiled in our analysis of cis-regulatory elements. Distinct expression patterns of AhTPS and AhTPP genes across different peanut tissues, developmental stages, and treatments were revealed, suggesting potential roles in growth, development, and stress responses. Under low-temperature stress, qPCR results showcased upregulation in AhTPS genes (AhTPS2-5, AhTPS9-12, AhTPS14, AhTPS15) and AhTPP genes (AhTPP1, AhTPP6, AhTPP11, AhTPP13). Furthermore, AhTPS9, exhibiting the most significant expression difference under cold stress, was obviously induced by cold stress in cultivated peanut, and AhTPS9-overexpression improved the cold tolerance of Arabidopsis by protect the photosynthetic system of plants, and regulates sugar-related metabolites and genes. This comprehensive study lays the groundwork for understanding the roles of AhTPS and AhTPP gene families in trehalose regulation within cultivated peanuts and provides valuable insights into the mechanisms related to cold stress tolerance.

Clinical and molecular profiling of human visceral adipose tissue reveals impairment of vascular architecture and remodeling as an early hallmark of dysfunction

Adipose tissue dysfunction is more related to insulin resistance than body mass index itself and an alteration in adipose tissue function is thought to underlie the shift from metabolically healthy to unhealthy obesity. Herein, we performed a clustering analysis that revealed distinct visceral adipose tissue gene expression patterns in patients with obesity at distinct stages of metabolic dysregulation. We have built a cross-sectional cohort that aims at reflecting the evolution of the metabolic sequelae of obesity with the main objective to map the sequential events that play a role in adipose tissue dysfunction from the metabolically healthy (insulin-sensitive) state to several incremental degrees of metabolic dysregulation, encompassing insulin resistance establishment, pre-diabetes, and type 2 diabetes. We found that insulin resistance is mainly marked by the downregulation of adipose tissue vasculature remodeling-associated gene expression, suggesting that processes like angiogenesis and adaptative expansion/retraction ability suffer early dysregulation. Prediabetes was characterized by compensatory growth factor-dependent signaling and increased response to hypoxia, while type 2 diabetes was associated with loss of cellular response to insulin and hypoxia and concomitant upregulation of inflammatory markers. Our findings suggest a putative sequence of dysregulation of biological processes that is not linear and has multiple distinct phases across the metabolic dysregulation process, ultimately culminating in the climax of adipose tissue dysfunction in type 2 diabetes. Several studies have addressed the transcriptomic changes in adipose tissue of patients with obesity. However, to the best of our knowledge, this is the first study unraveling the potential molecular mechanisms associated with the multi-step evolution of adipose tissue dysfunction along the metabolic sequelae of obesity.

A transcriptomic analysis of skeletal muscle tissues reveals promising candidate genes and pathways accountable for different daily weight gain in Hanwoo cattle

Cattle traits like average daily weight gain (ADG) greatly impact profitability. Selecting based on ADG considering genetic variability can lead to economic and genetic advancements in cattle breeding. This study aimed to unravel genetic influences on ADG variation in Hanwoo cattle at the skeletal muscle transcriptomic level. RNA sequencing was conducted on longissimus dorsi (LD), semimembranosus (SB), and psoas major (PM) muscles of 14 steers assigned to same feed, grouped by low (≤ 0.71 kg) and high (≥ 0.77 kg) ADG. At P ≤ 0.05 and log2fold > 1.5, the distinct pattern of gene expression was identified with 184, 172, and 210 differentially expressed genes in LD, SB, and PM muscles, respectively. Tissue-specific responses to ADG variation were evident, with myogenesis and differentiation associated JAK-STAT signaling pathway and prolactin signaling pathways enriched in LD and SB muscles, while adipogenesis-related PPAR signaling pathways were enriched in PM muscle. Key hub genes (AXIN2, CDKN1A, MYC, PTGS2, FZD5, SPP1) were upregulated and functionally significant in muscle growth and differentiation. Notably, DPP6, CDKN1A, and FZD5 emerged as possible candidate genes linked to ADG variation. These findings enhance our understanding of genetic factors behind ADG variation in Hanwoo cattle, illuminating skeletal muscle mechanisms influencing ADG.

Distinct patterns of proteostasis network gene expression are associated with different prognoses in melanoma patients

The proteostasis network (PN) is a collection of protein folding and degradation pathways that spans cellular compartments and acts to preserve the integrity of the proteome. The differential expression of PN genes is a hallmark of many cancers, and the inhibition of protein quality control factors is an effective way to slow cancer cell growth. However, little is known about how the expression of PN genes differs between patients and how this impacts survival outcomes. To address this, we applied unbiased hierarchical clustering to gene expression data obtained from primary and metastatic cutaneous melanoma (CM) samples and found that two distinct groups of individuals emerge across each sample type. These patient groups are distinguished by the differential expression of genes encoding ATP-dependent and ATP-independent chaperones, and proteasomal subunits. Differences in PN gene expression were associated with increased levels of the transcription factors, MEF2A, SP4, ZFX, CREB1 and ATF2, as well as markedly different survival outcomes. However, surprisingly, similar PN alterations in primary and metastatic samples were associated with discordant survival outcomes in patients. Our findings reveal that the expression of PN genes demarcates CM patients and highlights several new proteostasis sub-networks that could be targeted for more effective suppression of CM within specific individuals.

Gene Expression Profiles of HSP70 and HSP90 Genes and Biochemical Responses in Juvenile Abalone, Haliotis diversicolor squamata in Response to Total Suspended Solids

Prokaryotic and eukaryotic organisms contain heat shock proteins (HSPs), crucial for rapid response to environmental stress. However, their specific roles in different stress conditions are not fully understood. This study investigated HSP70 and HSP90 expression in H. diversicolor squamata, using qRT-PCR. Data analysis employed SPSS, including t-tests and ANOVA, with significance set at P < 0.05. Results showed distinct expression patterns of HSP genes under varying TSS levels. Both HSP70 and HSP90 mRNA levels significantly increased in response to TSS stressors, with HSP70 exhibiting the highest sensitivity to TSS changes. The duration and amount of TSS exposure influenced gene transcripts, particularly notable at 12 hours and 150 mgL-1 concentration. These findings suggest HSP genes play a role in cellular stress responses to environmental stimuli. HSP70 and HSP90, sensitive to TSS stress, can serve as biomarkers for assessing stress levels from TSS exposure and detecting TSS contamination in abalone farming.

Comprehensive Analysis of the Immunosuppressive Function of Regulatory T Cells in Human Hepatocellular Carcinoma Tissues.

Immune-based therapies are commonly employed to combat hepatocellular carcinoma (HCC). However, the presence of immune-regulating elements, especially regulatory T cells (Tregs), can dramatically impact the treatment efficacy. A deeper examination of the immune-regulation mechanisms linked to these inhibitory factors and their impact on HCC patient outcomes is warranted. We employed multicolor fluorescence immunohistochemistry (mIHC) to stain Foxp3, cytokeratin, and nuclei on an HCC tissue microarray (TMA). Leveraging liver cancer transcriptome data from TCGA, we built a prognostic model focused on Treg-associated gene sets and represented it with a nomogram. We then sourced liver cancer single-cell RNA sequencing data (GSE140228) from the GEO database, selectively focusing on Treg subsets, and conducted further analyses, including cell-to-cell communication and pseudo-time trajectory examination. Our mIHC results revealed a more substantial presence of Foxp3+Tregs in HCC samples than in adjacent normal tissue samples (P < .001). An increased presence of Foxp3+Tregs in HCC samples correlated with unfavorable patient outcomes (HR = 1.722, 95% CI:1.023-2.899, P = .041). The multi-factorial prognosis model we built from TCGA liver cancer data highlighted Tregs as a standalone risk determinant for predicting outcomes (HR = 3.84, 95% CI:2.52-5.83, P < .001). Re-analyzing the scRNA-seq dataset (GSE140228) showcased distinctive gene expression patterns in Tregs from varying tissues. Interactions between Tregs and other CD4+T cell types were predominantly governed by the CXCL13/CXCR3 signaling pathway. Communication pathways between Tregs and macrophages primarily involved MIF-CD74/CXCR4, LGALS9/CD45, and PTPRC/MRC1. Additionally, macrophages could influence Tregs via HLA-class II and CD4 interactions. An elevated presence of Tregs in HCC samples correlated with negative patient outcomes. Elucidating the interplay between Tregs and other immune cells in HCC could provide insights into the modulatory role of Tregs within HCC tissues.

Unveiling the cellular landscape: insights from single-cell RNA sequencing in multiple myeloma.

The aim of this research was to gain a thorough understanding of the processes involved in cell communication and discover potential indicators for treating multiple myeloma (MM) through the use of single-cell RNA sequencing (scRNA-seq). And explored the expression of multiple myeloma-related subgroups on metal ion-related pathways to explore the relationship between MM and metal ions. We performed a fair examination using single-cell RNA sequencing on 32 bone marrow specimens collected from 22 individuals at different points of MM advancement and 9 individuals without any health issues. To analyze the scRNA-seq data, we employed advanced computational algorithms, including Slingshot, Monocle2, and other methodologies. Specifically, Slingshot and Monocle2 enabled us to simulate the biological functionalities of different cell populations and map trajectories of cell developmental pathways. Additionally, we utilized the UMAP algorithm, a powerful dimension reduction technique, to cluster cells and identify genes that were differentially expressed across clusters. Our study revealed distinct gene expression patterns and molecular pathways within each patient, which exhibited associations with disease progression. The analysis provided insights into the tumor microenvironment (TME), intra- and inter-patient heterogeneity, and cell-cell interactions mediated by ligand-receptor signaling. And found that multiple myeloma-related subgroups were expressed higher levels in MMP and TIMP pathways, there were some associations. Our study presents a fresh perspective for future research endeavors and clinical interventions in the field of MM. The identified gene expression patterns and molecular pathways hold immense potential as therapeutic targets for the treatment of multiple myeloma. The utilization of scRNA-seq technology has significantly contributed to a more precise understanding of the complex cellular processes and interactions within MM. Through these advancements, we are now better equipped to unravel the underlying mechanisms driving the development and progression of this complex disease.

Genetic and transcriptomic analyses of diffuse large B-cell lymphoma patients with poor outcomes within two years of diagnosis

Despite the improvements in clinical outcomes for DLBCL, a significant proportion of patients still face challenges with refractory/relapsed (R/R) disease after receiving first-line R-CHOP treatment. To further elucidate the underlying mechanism of R/R disease and to develop methods for identifying patients at risk of early disease progression, we integrated clinical, genetic and transcriptomic data derived from 2805 R-CHOP-treated patients from seven independent cohorts. Among these, 887 patients exhibited R/R disease within two years (poor outcome), and 1918 patients remained in remission at two years (good outcome). Our analysis identified four preferentially mutated genes (TP53, MYD88, SPEN, MYC) in the untreated (diagnostic) tumor samples from patients with poor outcomes. Furthermore, transcriptomic analysis revealed a distinct gene expression pattern linked to poor outcomes, affecting pathways involved in cell adhesion/migration, T-cell activation/regulation, PI3K, and NF-κB signaling. Moreover, we developed and validated a 24-gene expression score as an independent prognostic predictor for treatment outcomes. This score also demonstrated efficacy in further stratifying high-risk patients when integrated with existing genetic or cell-of-origin subtypes, including the unclassified cases in these models. Finally, based on these findings, we developed an online analysis tool (https://lymphprog.serve.scilifelab.se/app/lymphprog) that can be used for prognostic prediction for DLBCL patients.

Patient-derived scaffolds representing breast cancer microenvironments influence chemotherapy responses in adapted cancer cells consistent with clinical features

BackgroundThe tumor microenvironment clearly influences cancer progressing properties but less is known about how individual cancer microenvironments potentially moderate cancer treatment effects. By cultivating and treating cancer cell lines in patient-derived scaffolds (PDS), the impact of specific characteristics of individual cancer microenvironments can be incorporated in human-like growth modelling and cancer drug treatment testing.MethodsPDSs from 78 biobanked primary breast cancer samples with known patient outcomes, were prepared and repopulated with donor breast cancer cell lines, followed by treatment with 5-fluorouracil or doxorubicin after cellular adaption to the various microenvironments. Cancer cell responses to the treatments were monitored by RNA-analyses, highlighting changes in gene sets representative for crucial tumor biological processes such as proliferation, cancer stem cell features, differentiation and epithelial-to-mesenchymal transition.ResultsThe chemotherapy treatments induced distinct gene expression patterns in adapted cancer cells with clusters of similar treatment responses depending on the patient-derived cancer microenvironment used as growth substrate. The doxorubicin treatment displayed a favorable gene signature among surviving cancer cells with low proliferation (MKI67) and pluripotency features (NANOG, POU5F1), in comparison to 5-fluorouracil showing low proliferation but increased pluripotency. Specific gene changes monitored post-treatment were also significantly correlated with clinical data, including histological grade (NANOG), lymph node metastasis (SLUG) and disease-free patient survival (CD44).ConclusionsThis laboratory-based treatment study using patient-derived scaffolds repopulated with cancer cell lines, clearly illustrates that the human cancer microenvironment influences chemotherapy responses. The differences in treatment responses defined by scaffold-cultures have potential prognostic and treatment predictive values.

Parental sex-dependent effects of either maternal or paternal eNOS deficiency on the offspring's phenotype without transmission of the parental eNOS deficiency to the offspring.

Background: Preclinical animal studies and clinical studies indicate that both maternal as well as paternal genetic alterations/gene defects might affect the phenotype of the next-generation without transmissions of the affected gene. Currently, the question of whether the same genetic defect present in the mother or father leads to a similar phenotype in the offspring remains insufficiently elucidated. Methods: In this head-to-head study, we crossbred female and male mice with heterozygous endothelial eNOS knockout (eNOS+/-) with male and female wild-type (wt) mice, respectively. Subsequently, we compared the phenotype of the resulting wt offspring with that of wt offspring born to parents with no eNOS deficiency. Results: Wt female offspring of mothers with heterozygous eNOS showed elevated liver fat accumulation, while wt male offspring of fathers with heterozygous eNOS exhibited increased fasting insulin, heightened insulin levels after a glucose load, and elevated liver glycogen content. By quantitative mass-spectrometry it was shown that concentrations of six serum metabolites (lysoPhosphatidylcholine acyl C20:3, phosphatidylcholine diacyl C36:2, phosphatidylcholine diacyl C38:1, phosphatidylcholine acyl-alkyl C34:1, phosphatidylcholine acyl-alkyl C36:3, and phosphatidylcholine acyl-alkyl C42:5 (PC ae C42:5) as well as four liver carbon metabolites (fructose 6-phosphate, fructose 1,6-bisphosphate, glucose 6-phosphate and fumarate) were different between wt offspring with eNOS+/- mothers and wt offspring with eNOS+/- fathers. Importantly, fumarate was inversely correlated with the liver fat accumulation in female offspring with eNOS+/- mothers and increased liver glycogen in offspring of both sexes with eNOS+/- fathers. The qRT-PCR results revealed that the gene expression patterns were different between wt offspring with eNOS+/- mothers and those offspring with eNOS+/- fathers. Different gene expression patterns were correlated with different observed phenotypic changes in male/female offspring born to mothers or fathers with a heterozygous eNOS genotype. Conclusion: The identical parental genetic alteration (heterozygous eNOS deficiency), without being passed on to the offspring, results in distinct metabolic, liver phenotype, and gene expression pattern variations depending on whether the genetic alteration originated from the father or the mother.

High-Throughput Transcriptomics Differentiates Toxic versus Non-Toxic Chemical Exposures Using a Rat Liver Model.

To address the challenge of limited throughput with traditional toxicity testing, a newly developed high-throughput transcriptomics (HTT) platform, together with a 5-day in vivo rat model, offers an alternative approach to estimate chemical exposures and provide reasonable estimates of toxicological endpoints. This study contains an HTT analysis of 18 environmental chemicals with known liver toxicity. They were evaluated using male Sprague Dawley rats exposed to various concentrations daily for five consecutive days via oral gavage, with data collected on the sixth day. Here, we further explored the 5-day rat model to identify potential gene signatures that can differentiate between toxic and non-toxic liver responses and provide us with a potential histopathological endpoint of chemical exposure. We identified a distinct gene expression pattern that differentiated non-hepatotoxic compounds from hepatotoxic compounds in a dose-dependent manner, and an analysis of the significantly altered common genes indicated that toxic chemicals predominantly upregulated most of the genes and several pathways in amino acid and lipid metabolism. Finally, our liver injury module analysis revealed that several liver-toxic compounds showed similarities in the key injury phenotypes of cellular inflammation and proliferation, indicating potential molecular initiating processes that may lead to a specific end-stage liver disease.

Deciphering temporal gene expression dynamics in multiple coral species exposed to heat stress: Implications for predicting resilience

Coral reefs are facing unprecedented threats due to global climate change, particularly elevated sea surface temperatures causing coral bleaching. Understanding coral responses at the molecular level is crucial for predicting their resilience and developing effective conservation strategies. In this study, we conducted a comprehensive gene expression analysis of four coral species to investigate their long-term molecular response to heat stress. We identified distinct gene expression patterns among the coral species, with laminar corals exhibiting a stronger response compared to branching corals. Heat shock proteins (HSPs) showed an overall decreasing expression trend, indicating the high energy cost associated with sustaining elevated HSP levels during prolonged heat stress. Peroxidases and oxidoreductases involved in oxidative stress response demonstrated significant upregulation, highlighting their role in maintaining cellular redox balance. Differential expression of genes related to calcium homeostasis and bioluminescence suggested distinct mechanisms for coping with heat stress among the coral species. Furthermore, the impact of heat stress on coral biomineralization varied, with downregulation of carbonic anhydrase and skeletal organic matrix proteins indicating reduced capacity for biomineralization in the later stages of heat stress. Our findings provide insights into the molecular mechanisms underlying coral responses to heat stress and highlight the importance of considering species-specific responses in assessing coral resilience. The identified biomarkers may serve as indicators of heat stress and contribute to early detection of coral bleaching events. These findings contribute to our understanding of coral resilience and provide a basis for future research aimed at enhancing coral survival in the face of climate change.

Integration of Transcriptomic Features to Improve Prognosis Prediction of Pediatric Acute Myeloid Leukemia With KMT2A Rearrangement.

Lysine methyltransferase 2A-rearranged acute myeloid leukemia (KMT2A-r AML) is a special entity in the 2022 World Health Organization classification of myeloid neoplasms, characterized by high relapse rate and adverse outcomes. Current risk stratification was established on the treatment response and translocation partner of KMT2A. To study the transcriptomic feature and refine the current stratification of pediatric KMT2A-r AML, we analyzed clinical and RNA sequencing data of 351 patients. By implementing least absolute shrinkage and selection operator algorithm, we identified 7 genes (KIAA1522, SKAP2, EGFL7, GAB2, HEBP1, FAM174B, and STARD8) of which the expression levels were strongly associated with outcomes. We then developed a transcriptome-based score, dividing patients into 2 groups with distinct gene expression patterns and prognosis, which was further validated in an independent cohort and outperformed the LSC17 score. We also found cell cycle, oxidative phosphorylation, and metabolism pathways were upregulated in patients with inferior outcomes. By integrating clinical characteristics, we proposed a simple-to-use prognostic scoring system with excellent discriminability, which allowed us to distinguish allogeneic hematopoietic stem cell transplantation candidates more precisely. In conclusion, pediatric KMT2A-r AML is heterogenous on transcriptomic level and the newly proposed scoring system combining clinical characteristics and transcriptomic features can be instructive in clinical routines.

Abstract A052: Comprehensive EBV-specific T cell profiling and multi-omics analysis of nasopharyngeal carcinoma CD8 T cells reveals novel cancer-associated phenotype

Abstract Nasopharyngeal carcinoma (NPC) is a high mortality, Epstein-Barr virus (EBV)-driven cancer with limited treatment options. Although EBV is highly antigenic and detectable in nearly all NPC tumor cells, it remains unclear why NPC tumors evade immune targeting despite high levels of immune infiltration. Here, we investigate the phenotypes of peripheral EBV-specific T cells with the purpose of identifying clinically relevant, cancer-associated EBV-specific T cell phenotypes. We developed a 34-marker mass cytometry and multiplexed MHC-I tetramer panel to detect and phenotype CD8 T cells of up to 55 antigen-specificities, including EBV and other known viral and cancer-antigen epitopes. We used this approach to profile peripheral blood mononuclear cells (PBMCs) from treatment-naïve NPC patients (n=51). Compared to other antigen-specific T cells detected in NPC patients and EBV-specific T cells in healthy individuals, EBV-specific T cells in NPC patients expressed high levels of activation markers (CD38, HLA-DR), co-inhibitory markers (CD39, TIGIT, PD-1), and migratory markers (ITB7, CD103). Using unsupervised clustering, we found that the frequency of EBV-specific T cells in a phenotypic cluster characterized by high CD103, CD39, HLA-DR, and CD38 expression positively correlated with plasma EBV-DNA titer and gross tumor volume. Remarkably, we observed that regardless of antigen specificity, NPC patients had a higher frequency of total CD8 T cells with this activated/exhausted cluster phenotype. CD8 frequencies of this cluster were also positively correlated with plasma EBV-DNA titer and stage (TNM). To investigate the gene expression profiles and TCR clonal diversity of this cancer-associated phenotype, we performed single-cell multi-omics analysis on peripheral CD8 T cells from NPC patients (n=17). We identified that cell subsets within this phenotype had distinct gene expression patterns indicative of potential associations with the tumor and we also assessed the clonality of these subpopulations. Overall, this study identified a unique cancer-associated peripheral EBV-specific T cell phenotype in the context of nasopharyngeal carcinoma that is positively correlated with advanced disease. Frequencies of peripheral T cells with this phenotype could also be useful as a blood-based prognostic marker for NPC. Citation Format: Nandita Kumar, Long Nguyen, Saumya Jani, Darren WT Lim, Joe PS Yeong, Melvin LK Chua, Amit Jain, Evan W Newell. Comprehensive EBV-specific T cell profiling and multi-omics analysis of nasopharyngeal carcinoma CD8 T cells reveals novel cancer-associated phenotype [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2023 Oct 1-4; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Cancer Immunol Res 2023;11(12 Suppl):Abstract nr A052.

Abstract B026: The senescent tumor cells evade the immune system by secreting SASPs in colorectal cancer

Abstract In colorectal cancer, the transition of cancer cells from an adenoid to a collective invasion morphology, ultimately leading to a partial epithelial-mesenchymal transition phenotype at the invasive front, has been observed. Through comprehensive gene expression analysis, we identified distinct gene expression patterns between cancer cells at the invasive front and those in the central region, with cellular senescence playing a pivotal role in this transition. Trajectory analysis revealed a gradual transformation of non-senescent tumor cells in the central region into senescent cells at the invasive front, accompanied by altered gene expression patterns. Furthermore, we observed a progressive increase in the expression of CXCL12 and CSF1 at the invasive front. Senescent tumor cells were found to impair the directional migration of CD8+ T cells by secreting high concentrations of CXCL12. Additionally, CSF1 secreted by senescent tumor cells promoted the differentiation of monocytes into M2 macrophages, which subsequently inhibited CD8+ T cell activation. Importantly, inhibiting CXCL12 secretion from senescent tumor cells enhanced T cell infiltration and resulted in reduced tumor number and size. These findings highlight the generation of a cytokine barrier by senescent tumor cells, which serves to protect non-senescent tumor cells from immune attack. Moreover, this study identifies senescent tumor cells as a potential target for overcoming immunotherapy resistance in colorectal cancer. The development of interventions that disrupt the cytokine barrier and enhance T cell infiltration holds promise for improving the efficacy of immunotherapeutic approaches in the treatment of colorectal cancer. Citation Format: Tae Jun Park, Soon Sang Park. The senescent tumor cells evade the immune system by secreting SASPs in colorectal cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2023 Oct 1-4; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Cancer Immunol Res 2023;11(12 Suppl):Abstract nr B026.