Browsing by Author "McGuire, Patrick"

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    Cryopreservation Preserves Cell-Type Composition and Gene Expression Profiles in Bone Marrow Aspirates From Multiple Myeloma Patients
    (Frontiers Media, 2021-04-21) Chen, Duojiao; Abu Zaid, Mohammad I.; Reiter, Jill L.; Czader, Magdalena; Wang, Lin; McGuire, Patrick; Xuei, Xiaoling; Gao, Hongyu; Huang, Kun; Abonour, Rafat; Walker, Brian A.; Liu, Yunlong; Medical and Molecular Genetics, School of Medicine
    Single-cell RNA sequencing reveals gene expression differences between individual cells and also identifies different cell populations that are present in the bulk starting material. To obtain an accurate assessment of patient samples, single-cell suspensions need to be generated as soon as possible once the tissue or sample has been collected. However, this requirement poses logistical challenges for experimental designs involving multiple samples from the same subject since these samples would ideally be processed at the same time to minimize technical variation in data analysis. Although cryopreservation has been shown to largely preserve the transcriptome, it is unclear whether the freeze-thaw process might alter gene expression profiles in a cell-type specific manner or whether changes in cell-type proportions might also occur. To address these questions in the context of multiple myeloma clinical studies, we performed single-cell RNA sequencing (scRNA-seq) to compare fresh and frozen cells isolated from bone marrow aspirates of six multiple myeloma patients, analyzing both myeloma cells (CD138+) and cells constituting the microenvironment (CD138-). We found that cryopreservation using 90% fetal calf serum and 10% dimethyl sulfoxide resulted in highly consistent gene expression profiles when comparing fresh and frozen samples from the same patient for both CD138+ myeloma cells (R ≥ 0.96) and for CD138- cells (R ≥ 0.9). We also demonstrate that CD138- cell-type proportions showed minimal alterations, which were mainly related to small differences in immune cell subtype sensitivity to the freeze-thaw procedures. Therefore, when processing fresh multiple myeloma samples is not feasible, cryopreservation is a useful option in single-cell profiling studies.
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    CTSA 2 Community: www.ctsa2community.org
    (2011-08-31) Ackermann, Ronald; Hardwick, Emily; Comer, Karen; Hudson, Brenda; Odell, Jere D.; Arenson, Andrew; Barnett, Bill; McGuire, Patrick; Derr, Michelle; Reid, Tisha; Vandergraff, Donna; Marrero, David G.
    This poster describes the development an accessible, user-driven, and sustainable web resource for community and academic experts working together to identify, adopt, and implement a wide array of community engaged research infrastructures for enhancing community engagement in all forms of clinical and translational research. CTSA2Community aims to be a storage place for valuable resources referring to the set-up and running of a community engagement program. Resources are provided by experts in the field of community engagement.
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    Dynamic chromatin accessibility and transcriptome changes following PDGF-BB treatment of bone-marrow derived mesenchymal stem cells
    (Springer Nature, 2024-10-15) Liu, Sheng; Chu, Xiaona; Reiter, Jill L.; Yu, Xuhong; Fang, Fang; McGuire, Patrick; Gao, Hongyu; Liu, Yunlong; Wan, Jun; Wang, Yue; Medical and Molecular Genetics, School of Medicine
    Background: Mesenchymal stem cells (MSCs) are multipotent stem cells that are under investigation for use in clinical trials because they are capable of self-renewal and differentiating into different cell types under defined conditions. Nonetheless, the therapeutic effects of MSCs have been constrained by low engraftment rates, cell fusion, and cell survival. Various strategies have been explored to improve the therapeutic efficacy of MSCs, with platelet-derived growth factor (PDGF)-BB emerging as a promising candidate. To enhance our comprehension of the impact of PDGF-BB on the gene expression profile and chromosomal accessibility of MSCs, RNA-sequencing and analysis of chromatin accessibility profiles were conducted on three human primary MSCs in culture, both with and without stimulation by PDGF-BB. Results: Integrative analysis of gene expression and chromatin accessibility demonstrated that PDGF-BB treatment modified the chromatin accessibility landscape, marking regions for activation or repression through the AP-1 family transcription factors TEAD, CEBP, and RUNX2. These changes in AP-1 transcription factor expression, in turn, led to cell proliferation and differentiation potential towards osteoblasts, adipocytes, or chondrocytes. The degree of MSC differentiation varies among cells isolated from different donors. The presence of an enrichment of exosome-related genes is also noted among all the differentially expressed genes. Conclusions: In conclusion, the observed changes in AP-1 transcription factor expression not only induced cellular proliferation and differentiation, but also revealed variations in the degree of MSC differentiation based on donor-specific differences. Moreover, the enrichment of exosome-related genes among differentially expressed genes suggests a potential significant role for PDGF-BB in facilitating intercellular communication.
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    IndianaCTSI and INresearch
    (Office of the Vice Chancellor for Research, 2012-04-13) McGuire, Patrick
    The Indiana Clinical and Translational Sciences Institute (CTSI) is a statewide collaboration of Indiana University, Purdue University and the University of Notre Dame, as well as public and private partnerships, which facilitates the translation of scientific discoveries in the lab into clinical trials and new patient treatments in Indiana and beyond.
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    Integrated Single-Cell Multiomic Profiling of Caudate Nucleus Suggests Key Mechanisms in Alcohol Use Disorder
    (bioRxiv, 2024-08-06) Green, Nick; Gao, Hongyu; Chu, Xiaona; Yuan, Quiyue; McGuire, Patrick; Lai, Dongbing; Jiang, Guanglong; Xuei, Xiaoling; Reiter, Jill; Stevens, Julia; Sutherland, Greg; Goate, Alison; Pang, Zhiping; Slesinger, Paul; Hart, Ronald P.; Tischfield, Jay A.; Agrawal, Arpana; Wang, Yue; Duren, Zhana; Edenberg, Howard J.; Liu, Yunlong; Medical and Molecular Genetics, School of Medicine
    Alcohol use disorder (AUD) is likely associated with complex transcriptional alterations in addiction-relevant brain regions. We characterize AUD-associated differences in cell type-specific gene expression and chromatin accessibility in the caudate nucleus by conducting a single-nucleus RNA-seq assay and a single-nucleus RNA-seq + ATAC-seq (multiome) assay on caudate tissue from 143 human postmortem brains (74 with AUD). We identified 17 cell types. AUD was associated with a higher proportion of microglia in an activated state and more astrocytes in a reactive state. There was widespread evidence for differentially expressed genes across cell types with the most identified in oligodendrocytes and astrocytes, including genes involved in immune response and synaptic regulation, many of which appeared to be regulated in part by JUND and OLIG2. Microglia-astrocyte communication via interleukin-1 beta, and microglia-astrocyte-oligodendrocyte interaction via transforming growth factor beta 1 were increased in individuals with AUD. Expression quantitative trait loci analysis revealed potential driver genes of AUD, including ADAL, that may protect against AUD in medium spiny neurons and interneurons. This work provides a thorough profile of the effects of AUD in the human brain and identifies several promising genes for further study.
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    Kidney intercalated cells are phagocytic and acidify internalized uropathogenic Escherichia coli
    (Springer Nature, 2021-04-23) Saxena, Vijay; Gao, Hongyu; Arregui, Samuel; Zollman, Amy; Kamocka, Malgorzata Maria; Xuei, Xiaoling; McGuire, Patrick; Hutchens, Michael; Hato, Takashi; Hains, David S.; Schwaderer, Andrew L.; Pediatrics, School of Medicine
    Kidney intercalated cells are involved in acid-base homeostasis via vacuolar ATPase expression. Here we report six human intercalated cell subtypes, including hybrid principal-intercalated cells identified from single cell transcriptomics. Phagosome maturation is a biological process that increases in biological pathway analysis rank following exposure to uropathogenic Escherichia coli in two of the intercalated cell subtypes. Real time confocal microscopy visualization of murine renal tubules perfused with green fluorescent protein expressing Escherichia coli or pHrodo Green E. coli BioParticles demonstrates that intercalated cells actively phagocytose bacteria then acidify phagolysosomes. Additionally, intercalated cells have increased vacuolar ATPase expression following in vivo experimental UTI. Taken together, intercalated cells exhibit a transcriptional response conducive to the kidney’s defense, engulf bacteria and acidify the internalized bacteria. Intercalated cells represent an epithelial cell with characteristics of professional phagocytes like macrophages.
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    Osteogenic Differentiation Potential of Mesenchymal Stem Cells Using Single Cell Multiomic Analysis
    (MDPI, 2023-09-26) Chen, Duojiao; Liu, Sheng; Chu, Xiaona; Reiter, Jill; Gao, Hongyu; McGuire, Patrick; Yu, Xuhong; Xuei, Xiaoling; Liu, Yichen; Wan, Jun; Fang, Fang; Liu, Yunlong; Wang, Yue; Medical and Molecular Genetics, School of Medicine
    Mesenchymal stem cells (MSC) are multipotent stem cells that can differentiate into multiple cell types, including osteoblasts, chondrocytes, and adipocytes. Osteoblast differentiation is reduced during osteoporosis development, resulting in reduced bone formation. Further, MSC isolated from different donors possess distinct osteogenic capacity. In this study, we used single-cell multiomic analysis to profile the transcriptome and epigenome of MSC from four healthy donors. Data were obtained from ~1300 to 1600 cells for each donor. These cells were clustered into four groups, indicating that MSC from different donors have distinct chromatin accessible regulatory elements for regulating gene expression. To investigate the mechanism by which MSC undergo osteogenic differentiation, we used the chromatin accessibility data from the single-cell multiome data to identify individual-specific enhancer–promoter pairs and evaluated the expression levels and activities of the transcriptional regulators. The MSC from four donors showed distinct differentiation potential into osteoblasts. MSC of donor 1 showed the largest average motif activities, indicating that MSC from donor 1 was most likely to differentiate into osteoblasts. The results of our validation experiments were consistent with the bioinformatics prediction. We also tested the enrichment of genome-wide association study (GWAS) signals of several musculoskeletal disease traits in the patient-specific chromatin accessible regions identified in the single-cell multiome data, including osteoporosis, osteopenia, and osteoarthritis. We found that osteoarthritis-associated variants were only enriched in the regions identified from donor 4. In contrast, osteoporosis and osteopenia variants were enriched in regions from donor 1 and least enriched in donor 4. Since osteoporosis and osteopenia are related to the density of bone cells, the enrichment of variants from these traits should be correlated with the osteogenic potential of MSC. In summary, this study provides large-scale data to link regulatory elements with their target genes to study the regulatory relationships during the differentiation of mesenchymal stem cells and provide a deeper insight into the gene regulatory mechanism.
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    A Pilot Single Cell Analysis of the Zebrafish Embryo Cellular Responses to Uropathogenic Escherichia coli Infection
    (Case Western Reserve University, 2022-02-04) Rawson, Ashley; Saxena, Vijay; Gao, Hongyu; Hooks, Jenaya; Xuei, Xiaoling; McGuire, Patrick; Hato, Takashi; Hains, David S.; Anderson, Ryan M.; Schwaderer, Andrew L.; Pediatrics, School of Medicine
    Background: Uropathogenic Escherichia coli (UPEC) infections are common and when they disseminate can be of high morbidity. Methods: We studied the effects of UPEC infection using single cell RNA sequencing (scRNAseq) in zebrafish. Bulk RNA sequencing has historically been used to evaluate gene expression patterns, but scRNAseq allows gene expression to be evaluated at the single cell level and is optimal for evaluating heterogeneity within cell types and rare cell types. Zebrafish cohorts were injected with either saline or UPEC, and scRNAseq and canonical pathway analyses were performed. Results: Canonical pathway analysis of scRNAseq data provided key information regarding innate immune pathways in the cells determined to be thymus cells, ionocytes, macrophages/monocytes, and pronephros cells. Pathways activated in thymus cells included interleukin 6 (IL-6) signaling and production of reactive oxygen species. Fc receptor-mediated phagocytosis was a leading canonical pathway in the pronephros and macrophages. Genes that were downregulated in UPEC vs saline exposed embryos involved the cellular response to the Gram-negative endotoxin lipopolysaccharide (LPS) and included Forkhead Box O1a (Foxo1a), Tribbles Pseudokinase 3 (Trib3), Arginase 2 (Arg2) and Polo Like Kinase 3 (Plk3). Conclusions: Because 4-day post fertilization zebrafish embryos only have innate immune systems, the scRNAseq provides insights into pathways and genes that cell types utilize in the bacterial response. Based on our analysis, we have identified genes and pathways that might serve as genetic targets for treatment and further investigation in UPEC infections at the single cell level.