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Abstract 33: Transcriptomic Identification of Functionally Potent Umbilical Cord Blood Units
(Oxford University Press, 2024-08-21) Ropa, James; Gutch, Sarah; Beasley, Lindsay; Van't Hof, Wouter; Sun, Jessica; Capitano, Maegan; Kaplan, Mark; Medical and Molecular Genetics, School of Medicine
Introduction:
Umbilical cord blood (UCB) is an important donor source for standard of care cellular therapies as well as innovative new treatments. Universal potency criteria for cord blood unit (CBU) selection for different cellular therapy applications are still desired and efficient methods to elucidate these criteria remain elusive.
Objectives:
Our goal is to find molecular markers that identify potent CBUs for use in cellular therapies. Here, we utilized transcriptomics to reveal genes associated with hematopoietic stem and progenitor cell (HSC/HPC) potency in hematopoietic cell transplantation.
Methods:
We performed three separate transcriptomic analyses of human UCB used in mouse models of transplantation. This included bulk RNA-sequencing of HSCs/HPCs from CBUs with known engraftment capacities (n=9 CBUs), bulk RNA-sequencing of homed/early engrafted CD34+ cells (n=3 CBUs), and single cell RNA-sequencing of CD34+ cells expanded in varying oxygen tensions, which affects their transplantation potency (n=4 CBUs).
Results:
HSCs/HPCs enriched for dehydrogenase and cell cycle associated genes yield better repopulating cell frequency. Early homed CD34+ cells have enriched expression of immune activation and cell cycle genes compared to input transplanted cells. Distinct clusters of UCB cells marked by genes such as PRSS2 and AVP are enriched in oxygenation conditions that drive increased potency. Dehydrogenase and stress response genes are enriched in populations predicted to be more functional regardless of HSC/HPC subpopulation. Integration of all three studies reveals genes that may define highly potent CBUs, including DDIT4, a stress response gene. Indeed, DDIT4 independently predicts engraftment outcomes in mouse models of transplantation. Future work will examine a qPCR based gene panel potency assay to predict outcomes in patient transplantations.
Discussion:
We have identified genes associated with HSC/HPC potency using transcriptomic approaches. These findings have immediate translational implications for CBU selection for transplantation, but also provides a blueprint for finding CBUs best suited for use in developing off-the-shelf immune effector therapies or those that are best for treating non-hematologic central nervous system disorders such as cerebral palsy, among other applications. Importantly, this study highlights the importance for omics technology as a valuable tool to define potency criteria for UCB as a donor source for cellular therapies.
Abstract 27: Physiologic Oxygen Expansion Enhances Lymphocyte and Neutrophil Recovery Following Transplantation
(Oxford University Press, 2024-08-21) Gutch, Sarah; Ropa, Jim; Beasley, Lindsay; Whitacre, Grace; Van't Hof, Wouter; Capitano, Maegan; Medical and Molecular Genetics, School of Medicine
Introduction:
Expeditious recovery of lymphocytes after hematopoietic cell transplantation is a major determinant of patient outcome. There are few efficient clinical therapies to enhance lymphocyte recovery, indicating a clear unmet need. Ex vivo expansion of cord blood (CB) units is an approved therapy to increase numbers of hematopoietic stem and progenitor cells, but impact on lymphocyte recovery remains uncertain. Moreover, culture in physioxic (physiological oxygen) conditions results in increased lymphoid-biased RNA levels.
Objectives:
We hypothesize that ex vivo expansion in physioxic conditions will increase lymphoid-biased cells and increase lymphocyte counts post-transplantation (PT). The objective of this study is to increase lymphocyte numbers following transplantation without sacrificing reconstitution of potent hematopoietic cells.
Methods:
Three independent transplants were conducted. 1) Murine lineage- bone marrow (BM) was expanded for 7 days then transplanted into lethally irradiated mice with/without additional common lymphoid progenitors (CLPs). 2) Murine lineage- BM was expanded in 1%, 3%, 5%, 14%, and 21% O2 for 7 days and transplanted into lethally irradiated mice. 3) Human CD34+ CB cells were expanded for 7 days in 1%, 3%, 5%, 14%, and 21% O2 and transplanted into NSG mice. Unexpanded BM or CB cells were used as controls.
Results:
Additional CLPs did not contribute to lymphocyte recovery. Mice transplanted with expanded BM had increased lymphocyte counts compared to transplantations with unexpanded BM at weeks 2 and 5 PT via complete blood count (CBC) and increased B-cell reconstitution in the spleen, BM, and peripheral blood (PB) at weeks 2, 5, and 8 PT. LSK (Lin- SCA1+ cKIT+) and neutrophil frequencies were increased at 3% O2 in the BM and 5% O2 in the PB, respectively. Compared to 21% O2, CB cells expanded at 1%, 3%, 5%, 14% O2 had increased neutrophil and lymphocyte frequencies in the PB at weeks 2 and 10, respectively, and demonstrated greater recovery than unexpanded at week 2.
Discussion:
Expansion increases lymphocyte counts via CBC and immunophenotyping. Physioxic expansion increases numbers of potent hematopoietic cell subpopulations and frequencies of specific lymphocyte compartments in multiple organs. Thus, expansion under physioxia is a viable strategy to enhance recovery of lymphocytes PT.
Optimization of a human induced pluripotent stem cell-derived sensory neuron model for the in vitro evaluation of taxane-induced neurotoxicity
(Springer Nature, 2024-08-17) Cantor, Erica L.; Shen, Fei; Jiang, Guanglong; Philips, Santosh; Schneider, Bryan P.; Medicine, School of Medicine
Human induced pluripotent stem cell-derived sensory neuron (iPSC-dSN) models are a valuable resource for the study of neurotoxicity but are affected by poor replicability and reproducibility, often due to a lack of optimization. Here, we identify experimental factors related to culture conditions that substantially impact cellular drug response in vitro and determine optimal conditions for improved replicability and reproducibility. Treatment duration and cell seeding density were both found to be significant factors, while cell line differences also contributed to variation. A replicable dose-response in viability was demonstrated after 48-h exposure to docetaxel or paclitaxel. Additionally, a replicable dose-dependent reduction in neurite outgrowth was demonstrated, demonstrating the applicability of the model for the examination of additional phenotypes. Overall, we have established an optimized iPSC-dSN model for the study of taxane-induced neurotoxicity.
Succinate dehydrogenase-complex II regulates skeletal muscle cellular respiration and contractility but not muscle mass in genetically induced pulmonary emphysema
(American Association for the Advancement of Science, 2024) Balnis, Joseph; Tufts, Ankita; Jackson, Emily L.; Drake, Lisa A.; Singer, Diane V.; Lacomis, David; Lee, Chun Geun; Elias, Jack A.; Doles, Jason D.; Maher, L. James, III; Jen, Annie; Coon, Joshua J.; Jourd’heuil, David; Singer, Harold A.; Vincent, Catherine E.; Jaitovich, Ariel; Anatomy, Cell Biology and Physiology, School of Medicine
Reduced skeletal muscle mass and oxidative capacity coexist in patients with pulmonary emphysema and are independently associated with higher mortality. If reduced cellular respiration contributes to muscle atrophy in that setting remains unknown. Using a mouse with genetically induced pulmonary emphysema that recapitulates muscle dysfunction, we found that reduced activity of succinate dehydrogenase (SDH) is a hallmark of its myopathic changes. We generated an inducible, muscle-specific SDH knockout mouse that demonstrates lower mitochondrial oxygen consumption, myofiber contractility, and exercise endurance. Respirometry analyses show that in vitro complex I respiration is unaffected by loss of SDH subunit C in muscle mitochondria, which is consistent with the pulmonary emphysema animal data. SDH knockout initially causes succinate accumulation associated with a down-regulated transcriptome but modest proteome effects. Muscle mass, myofiber type composition, and overall body mass constituents remain unaltered in the transgenic mice. Thus, while SDH regulates myofiber respiration in experimental pulmonary emphysema, it does not control muscle mass or other body constituents.
Impact of Intramyocardial Hemorrhage on Clinical Outcomes in ST-Elevation Myocardial Infarction: A Systematic Review and Meta-analysis
(Elsevier, 2022-08-26) Vyas, Rohit; Changal, Khalid H.; Bhuta, Sapan; Pasadyn, Vanessa; Katterle, Konrad; Niedoba, Matthew J.; Vora, Keyur; Dharmakumar, Rohan; Gupta, Rajesh; Medicine, School of Medicine
Background: Intramyocardial hemorrhage (IMH) occurs after ST-elevation myocardial infarction (STEMI) and has been documented using cardiac magnetic resonance imaging. The prevalence and prognostic significance of IMH are not well described, and the small sample size has limited prior studies.
Methods: We performed a comprehensive literature search of multiple databases to identify studies that compared outcomes in STEMI patients with or without IMH. The outcomes studied were major adverse cardiovascular events (MACE), infarct size, thrombolysis in myocardial infarction (TIMI) flow after percutaneous coronary intervention (PCI), left ventricular end-diastolic volume (LVEDV), left ventricular ejection fraction (LVEF), and mortality. Odds ratios (ORs) and standardized mean differences with corresponding 95% CIs were calculated using a random effects model.
Results: Eighteen studies, including 2824 patients who experienced STEMI (1078 with IMH and 1746 without IMH), were included. The average prevalence of IMH was 39%. There is a significant association between IMH and subsequent MACE (OR, 2.63; 95% CI, 1.79-3.86; P < .00001), as well as IMH and TIMI grade <3 after PCI (OR, 1.75; 95% CI, 1.14-2.68; P = .05). We also found a significant association between IMH and the use of glycoprotein IIb/IIIa inhibitors (OR, 2.34; 95% CI, 1.42-3.85; P = .0008). IMH has a positive association with infarct size (standardized mean difference, 2.19; 95% CI, 1.53-2.86; P < .00001) and LVEDV (standardized mean difference, 0.7; 95% CI, 0.41-0.99; P < .00001) and a negative association with LVEF (standardized mean difference, -0.89; 95% CI, -1.15 to -0.63; P = .01). Predictors of IMH include male sex, smoking, and left anterior descending infarct.
Conclusions: Intramyocardial hemorrhage is prevalent in approximately 40% of patients who experience STEMI. IMH is a significant predictor of MACE and is associated with larger infarct size, higher LVEDV, and lower LVEF after STEMI.