{"id":{"repo_id":"vcu","oai_identifier":"oai:scholarscompass.vcu.edu:etd-1636"},"canonical_url":"https://search.dev.ndltd.org/etd/vcu/oai:scholarscompass.vcu.edu:etd-1636","repository":{"repo_id":"vcu","name":"Virginia Commonwealth University","base_url":"https://scholarscompass.vcu.edu/do/oai/"},"display":{"title":"Approaches to Reduce Selection of Genomic Variants in Human Pluripotent Stem Cell Culture","abstract":"Optimizing culture conditions that reduce genomic instability in human pluripotent stem cells (hPSCs) is an unmet challenge in the field. Results from our lab and numerous research groups demonstrate that hPSCs are prone to genomic aberrations and single-cell passaging increases the rate of genomic alterations. However, single-cell based passaging maintains advantages for scale-up and standardizing differentiation protocols. In this study, we investigated the problem of genomic instability in hPSC cultures with the goal towards identifying and characterizing candidate genes that could contribute to generation and survival of abnormal hPSCs. Based on microarray analysis, we identify ARHGDIA, located on 17q25, as a candidate gene conferring selective advantage to trisomy 17 hPSCs. Using lentiviral approaches to overexpress ARHGDIA in hPSCs, [hPSC (Arg)], we functionally validate that in enzymatically passaged co-cultures, hPSC (Arg) lines exhibit competitive advantage against wild type hPSCs, [hPSC (WT)]. Additionally, hPSC (Arg) lines exhibit increased single-cell survival at low density plating. In co-cultures with hPSC (WT), ROCKi exposure attenuated the competitive advantage of hPSC (Arg) subpopulations. For the first time, this work demonstrates that increased expression of a gene on 17q25 confers selective advantage to hPSCs. In parallel studies, using medium devoid of bFGF containing LIF plus two inhibitors, MEK inhibitor (PD0325901) and p38 inhibitor (SB203580), we demonstrate that hPSCs are LIF responsive and can be stably maintained in naive pluripotent culture conditions. Based on their clonal viability, we propose that naive hPSCs are a more genetically stable population than primed hPSCs, when passaged as single- cells. These studies will aid the long-term goal of hPSC scale-up while promoting stable propagation of genomically normal hPSCs.","abstract_html":"Optimizing culture conditions that reduce genomic instability in human pluripotent stem cells (hPSCs) is an unmet challenge in the field. Results from our lab and numerous research groups demonstrate that hPSCs are prone to genomic aberrations and single-cell passaging increases the rate of genomic alterations. However, single-cell based passaging maintains advantages for scale-up and standardizing differentiation protocols. In this study, we investigated the problem of genomic instability in hPSC cultures with the goal towards identifying and characterizing candidate genes that could contribute to generation and survival of abnormal hPSCs. Based on microarray analysis, we identify ARHGDIA, located on 17q25, as a candidate gene conferring selective advantage to trisomy 17 hPSCs. Using lentiviral approaches to overexpress ARHGDIA in hPSCs, [hPSC (Arg)], we functionally validate that in enzymatically passaged co-cultures, hPSC (Arg) lines exhibit competitive advantage against wild type hPSCs, [hPSC (WT)]. Additionally, hPSC (Arg) lines exhibit increased single-cell survival at low density plating. In co-cultures with hPSC (WT), ROCKi exposure attenuated the competitive advantage of hPSC (Arg) subpopulations. For the first time, this work demonstrates that increased expression of a gene on 17q25 confers selective advantage to hPSCs. In parallel studies, using medium devoid of bFGF containing LIF plus two inhibitors, MEK inhibitor (PD0325901) and p38 inhibitor (SB203580), we demonstrate that hPSCs are LIF responsive and can be stably maintained in naive pluripotent culture conditions. Based on their clonal viability, we propose that naive hPSCs are a more genetically stable population than primed hPSCs, when passaged as single- cells. These studies will aid the long-term goal of hPSC scale-up while promoting stable propagation of genomically normal hPSCs.","abstract_has_math":false,"creators":["Riggs, Marion"],"institution":null,"degree_name":"Doctor of Philosophy","degree_level":"Dissertation","degree_discipline":"Integrative Life Sciences","degree_department":null,"school":null,"contributors":["Raj Rao"],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2014,"date_issued":"2014-05-13T07:00:00Z","date_published":"2014-05-13T07:00:00Z","updated_at":"2026-07-24T05:54:11Z","subjects":["Human Embryonic Stem Cells","Cell Culture","Culture Adaptation","Pluripotent","Genomic Instability","Life Sciences"],"languages":[],"rights":["© The Author"],"rights_urls":[],"identifier_entries":[{"key":"dc:identifier","label":"Identifier","values":["https://scholarscompass.vcu.edu/etd/637"],"render_values":[{"text":"https://scholarscompass.vcu.edu/etd/637","href":"https://scholarscompass.vcu.edu/etd/637","code":true}]}]},"links":{"outbound_url":"https://doi.org/10.25772/JW0X-ZE37","outbound_label":"DOI","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Raj Rao"]},{"key":"dc:creator","label":"Author","values":["Riggs, Marion"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date.available","label":"Dc Date Available","values":["2019-05-16T07:00:00Z"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Integrative Life Sciences"]},{"key":"thesis:degree_level","label":"Degree Level","values":["Dissertation"]},{"key":"thesis:degree_name","label":"Degree Name","values":["Doctor of Philosophy"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["Human Embryonic Stem Cells","Cell Culture","Culture Adaptation","Pluripotent","Genomic Instability","Life Sciences"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:rights","label":"Dc Rights","values":["© The Author"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["https://doi.org/10.25772/JW0X-ZE37","https://scholarscompass.vcu.edu/etd/637"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["Optimizing culture conditions that reduce genomic instability in human pluripotent stem cells (hPSCs) is an unmet challenge in the field. Results from our lab and numerous research groups demonstrate that hPSCs are prone to genomic aberrations and single-cell passaging increases the rate of genomic alterations. However, single-cell based passaging maintains advantages for scale-up and standardizing differentiation protocols. In this study, we investigated the problem of genomic instability in hPSC cultures with the goal towards identifying and characterizing candidate genes that could contribute to generation and survival of abnormal hPSCs. Based on microarray analysis, we identify ARHGDIA, located on 17q25, as a candidate gene conferring selective advantage to trisomy 17 hPSCs. Using lentiviral approaches to overexpress ARHGDIA in hPSCs, [hPSC (Arg)], we functionally validate that in enzymatically passaged co-cultures, hPSC (Arg) lines exhibit competitive advantage against wild type hPSCs, [hPSC (WT)]. Additionally, hPSC (Arg) lines exhibit increased single-cell survival at low density plating. In co-cultures with hPSC (WT), ROCKi exposure attenuated the competitive advantage of hPSC (Arg) subpopulations. For the first time, this work demonstrates that increased expression of a gene on 17q25 confers selective advantage to hPSCs. In parallel studies, using medium devoid of bFGF containing LIF plus two inhibitors, MEK inhibitor (PD0325901) and p38 inhibitor (SB203580), we demonstrate that hPSCs are LIF responsive and can be stably maintained in naive pluripotent culture conditions. Based on their clonal viability, we propose that naive hPSCs are a more genetically stable population than primed hPSCs, when passaged as single- cells. These studies will aid the long-term goal of hPSC scale-up while promoting stable propagation of genomically normal hPSCs."]},{"key":"dc:title","label":"Title","values":["Approaches to Reduce Selection of Genomic Variants in Human Pluripotent Stem Cell Culture"]}]}],"canonical_facts":{"dc:contributor":["Raj Rao"],"dc:creator":["Riggs, Marion"],"dc:date.available":["2019-05-16T07:00:00Z"],"dc:description.abstract":["Optimizing culture conditions that reduce genomic instability in human pluripotent stem cells (hPSCs) is an unmet challenge in the field. Results from our lab and numerous research groups demonstrate that hPSCs are prone to genomic aberrations and single-cell passaging increases the rate of genomic alterations. However, single-cell based passaging maintains advantages for scale-up and standardizing differentiation protocols. In this study, we investigated the problem of genomic instability in hPSC cultures with the goal towards identifying and characterizing candidate genes that could contribute to generation and survival of abnormal hPSCs. Based on microarray analysis, we identify ARHGDIA, located on 17q25, as a candidate gene conferring selective advantage to trisomy 17 hPSCs. Using lentiviral approaches to overexpress ARHGDIA in hPSCs, [hPSC (Arg)], we functionally validate that in enzymatically passaged co-cultures, hPSC (Arg) lines exhibit competitive advantage against wild type hPSCs, [hPSC (WT)]. Additionally, hPSC (Arg) lines exhibit increased single-cell survival at low density plating. In co-cultures with hPSC (WT), ROCKi exposure attenuated the competitive advantage of hPSC (Arg) subpopulations. For the first time, this work demonstrates that increased expression of a gene on 17q25 confers selective advantage to hPSCs. In parallel studies, using medium devoid of bFGF containing LIF plus two inhibitors, MEK inhibitor (PD0325901) and p38 inhibitor (SB203580), we demonstrate that hPSCs are LIF responsive and can be stably maintained in naive pluripotent culture conditions. Based on their clonal viability, we propose that naive hPSCs are a more genetically stable population than primed hPSCs, when passaged as single- cells. These studies will aid the long-term goal of hPSC scale-up while promoting stable propagation of genomically normal hPSCs."],"dc:identifier":["https://doi.org/10.25772/JW0X-ZE37","https://scholarscompass.vcu.edu/etd/637"],"dc:rights":["© The Author"],"dc:subject":["Human Embryonic Stem Cells","Cell Culture","Culture Adaptation","Pluripotent","Genomic Instability","Life Sciences"],"dc:title":["Approaches to Reduce Selection of Genomic Variants in Human Pluripotent Stem Cell Culture"],"thesis:degree_discipline":["Integrative Life Sciences"],"thesis:degree_level":["Dissertation"],"thesis:degree_name":["Doctor of Philosophy"]},"updated_at":"2026-07-24T05:54:11Z"}