title: Hypocapnia leads to enhanced expression of pluripotency and meso-endodermal differentiation genes in mouse embryonic stem cells.
abstract: The efficient utilization of embryonic stem cells for applications like cell based therapy, transplantation and drug discovery largely depends upon the culturing conditions of these cells. In this report, we have analyzed gene, protein expression and morphological changes of embryonic stem cells when subjected to lowered CO2 levels i.e. hypocapnia. We studied the quantitative expression of pluripotent genes, Oct3/4, Nanog and Sox2 and genes involved in the differentiation to the three lineages, under varying CO2 levels. Enhanced expression of these genes was seen at cultures maintained at 1.5% CO2 as compared to those maintained at 5% CO2. The cells exposed to hypocapnic conditions when subjected to immunocytochemical analysis stained positive for Oct-3/4, Nanog and Sox2 transcription factors. Flow cytometry and western blot further showed that the pluripotent proteins in the 1.5% CO2 maintained cultures have higher levels of expression as compared to the ES cells at 5% CO2. In addition, there was enhanced differentiation particularly towards the mesodermal and endodermal lineages at cultures maintained and differentiated at 1.5% CO2 at all the time periods analyzed i.e. day 10 (5+5d), 12 (5+7d) and day 15 (5+10d). These results, which we feel are the first of their kind, indicate that lowered CO2 levels seem to be preferred for the maintenance of pluripotency and the subsequent differentiation.
authors.affiliation: Department of Biotechnology and Bioinformatics, Jaypee University of Information Technology, Waknaghat 173234, India.
authors.forename: Saras
authors.initials: S
authors.lastname: Jyoti
authors.affiliation: Department of Biotechnology and Bioinformatics, Jaypee University of Information Technology, Waknaghat 173234, India. Electronic address: simran.tandon@juit.ac.in.
authors.forename: Simran
authors.initials: S
authors.lastname: Tandon
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D015415:Biomarkers / Q000378:metabolism*; D015153:Blotting, Western; D002454:Cell Differentiation; D019070:Cell Lineage; D002478:Cells, Cultured; D053595:Embryonic Stem Cells / Q000166:cytology / Q000502:physiology*; D004707:Endoderm / Q000166:cytology* / Q000502:physiology; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D018507:Gene Expression Regulation, Developmental; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D016857:Hypocapnia / Q000503:physiopathology*; D008648:Mesoderm / Q000166:cytology* / Q000502:physiology; D051379:Mice; D008856:Microscopy, Fluorescence; D000071317:Nanog Homeobox Protein; D050814:Octamer Transcription Factor-3 / Q000235:genetics / Q000378:metabolism; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055748:SOXB1 Transcription Factors / Q000235:genetics / Q000378:metabolism
chemical_list: D015415:Biomarkers; D018398:Homeodomain Proteins; D000071317:Nanog Homeobox Protein; C475540:Nanog protein, mouse; D050814:Octamer Transcription Factor-3; D012333:RNA, Messenger; D055748:SOXB1 Transcription Factors; C527783:Sox2 protein, mouse; D000469:Alkaline Phosphatase
keywords: Differentiation; Embryonic stem cells; Hypocapnia; Pluripotency