Project description:Gene expression profiling of immortalized human mesenchymal stem cells with hTERT/E6/E7 transfected MSCs. hTERT may change gene expression in MSCs. Goal was to determine the gene expressions of immortalized MSCs.
Project description:Transcriptional profiling of human mesenchymal stem cells comparing normoxic MSCs cells with hypoxic MSCs cells. Hypoxia may inhibit senescence of MSCs during expansion. Goal was to determine the effects of hypoxia on global MSCs gene expression.
Project description:Gene expression profiling of immortalized human mesenchymal stem cells with hTERT/E6/E7 transfected MSCs. hTERT may change gene expression in MSCs. Goal was to determine the gene expressions of immortalized MSCs. One-condition experment, gene expression of 3A6
Project description:Transcriptional profiling of human mesenchymal stem cells comparing normoxic MSCs cells with hypoxic MSCs cells. Hypoxia may inhibit senescence of MSCs during expansion. Goal was to determine the effects of hypoxia on global MSCs gene expression. Two-condition experiment, Normoxic MSCs vs. Hypoxic MSCs.
Project description:Gene methylation profiling of immortalized human mesenchymal stem cells comparing HPV E6/E7-transfected MSCs cells with human telomerase reverse transcriptase (hTERT)- and HPV E6/E7-transfected MSCs. hTERT may increase gene methylation in MSCs. Goal was to determine the effects of different transfected genes on global gene methylation in MSCs.
Project description:MSCs comprise several percent of pluripotent-like cells named Multilineage-differentiating stress enduring (Muse) cells that express pluripotent markers at moderate levels, are collectable as cells positive for the pluripotent surface marker SSEA-3, are able to differentiate into triploblastic lineage cells, and self-renew at the single cell level (Kuroda et al, PNAS, 2010; Wakao et al, PNAS, 2011). Importantly, MSCs also comprise cells other than SSEA-3(+)-Muse cells, namely multipotent SSEA-3(-)-non-Muse MSCs that correspond to ~98% of the total MSC population. These SSEA-3(-)-non-Muse MSCs exhibit the same properties as conventional MSCs, although the non-Muse MSCs are multipotent, they are not pluripotent. In the present study, to clarify the key molecules that characterize pluripotent-like vs multipotent somatic stem cells, we separated human MSCs into SSEA-3(+)-Muse cells and SSEA-3(-)-non Muse MSCs, and analyzed both populations by scRNA-seq. SSEA-3(+) Muse cells and SSEA-3(-) non-Muse MSCs were sorted from human BM-MSCs. Sequencing libraries were prepared using Chromium single cell 3' Kit v3 (10x Genomics, Pleasanton, CA, USA) and sequenced on HiSeq2500 (Illumina, San Diego, CA, USA). Transcripts were mapped with CellRanger pipeline v3 (10x Genomics). Library construction, sequencing, and initial analysis were performed by GENEWIZ (South Plainfield, NJ, USA).
Project description:Gene methylation profiling of immortalized human mesenchymal stem cells comparing HPV E6/E7-transfected MSCs cells with human telomerase reverse transcriptase (hTERT)- and HPV E6/E7-transfected MSCs. hTERT may increase gene methylation in MSCs. Goal was to determine the effects of different transfected genes on global gene methylation in MSCs. Two-condition experiment, KP MSCs vs. 3A6 MSCs.
Project description:Microarray analysis of the gene expression profile of SSEA-3-positive human adipose-derived MSCs were performed. Liposuction aspirates were obtained from the abdomen and thighs. Stromal vascular fraction (SVF) was isolated from the aspirated fat. Muse cells that express human SSEA-3 were collected from SVF using anti SSEA-3 antibody conjugated with PE and anti-PE microbeads. SSEA-3-negative fraction was considered M-bM-^@M-^XMuse-poor populationM-bM-^@M-^Y.
Project description:Kynureninase is a member of a large family of catalytically diverse but structurally homologous pyridoxal 5'-phosphate (PLP) dependent enzymes known as the aspartate aminotransferase superfamily or alpha-family. The Homo sapiens and other eukaryotic constitutive kynureninases preferentially catalyze the hydrolytic cleavage of 3-hydroxy-l-kynurenine to produce 3-hydroxyanthranilate and l-alanine, while l-kynurenine is the substrate of many prokaryotic inducible kynureninases. The human enzyme was cloned with an N-terminal hexahistidine tag, expressed, and purified from a bacterial expression system using Ni metal ion affinity chromatography. Kinetic characterization of the recombinant enzyme reveals classic Michaelis-Menten behavior, with a Km of 28.3 +/- 1.9 microM and a specific activity of 1.75 micromol min-1 mg-1 for 3-hydroxy-dl-kynurenine. Crystals of recombinant kynureninase that diffracted to 2.0 A were obtained, and the atomic structure of the PLP-bound holoenzyme was determined by molecular replacement using the Pseudomonas fluorescens kynureninase structure (PDB entry 1qz9) as the phasing model. A structural superposition with the P. fluorescens kynureninase revealed that these two structures resemble the "open" and "closed" conformations of aspartate aminotransferase. The comparison illustrates the dynamic nature of these proteins' small domains and reveals a role for Arg-434 similar to its role in other AAT alpha-family members. Docking of 3-hydroxy-l-kynurenine into the human kynureninase active site suggests that Asn-333 and His-102 are involved in substrate binding and molecular discrimination between inducible and constitutive kynureninase substrates.