ABSTRACT
Hematopoiesis generated from human embryonic stem cells (ES) and induced pluripotent stem cells (iPS) are unprecedented resources for cell therapy. We compared hematopoietic differentiation potentials from ES and iPS cell lines originated from various donors and derived them using integrative and non-integrative vectors. Significant differences in differentiation toward hematopoietic lineage were observed among ES and iPS. The ability of engraftment of iPS or ES-derived cells in NOG mice varied among the lines with low levels of chimerism. iPS generated from ES cell-derived mesenchymal stem cells (MSC) reproduce a similar hematopoietic outcome compared to their parental ES cell line. We were not able to identify any specific hematopoietic transcription factors that allow to distinguish between good versus poor hematopoiesis in undifferentiated ES or iPS cell lines. There is a relatively unpredictable variation in hematopoietic differentiation between ES and iPS cell lines that could not be predicted based on phenotype or gene expression of the undifferentiated cells. These results demonstrate the influence of genetic background in variation of hematopoietic potential rather than the reprogramming process.
Subject(s)
Cellular Reprogramming/genetics , Embryonic Stem Cells/cytology , Genetic Heterogeneity , Graft Survival , Hematopoiesis/genetics , Induced Pluripotent Stem Cells/cytology , Animals , Biomarkers/metabolism , Cell Differentiation , Cell Line , Cell Lineage/genetics , Chimerism , Embryonic Stem Cells/metabolism , Embryonic Stem Cells/transplantation , Gene Expression , Genetic Vectors , Humans , Induced Pluripotent Stem Cells/metabolism , Induced Pluripotent Stem Cells/transplantation , Lentivirus/genetics , Mice , Mice, Inbred NOD , Mice, SCID , Retroviridae/genetics , Tissue Donors , Transcription Factors/genetics , Transcription Factors/metabolism , Transplantation, HeterologousABSTRACT
BACKGROUND: Strategies on the basis of doxycycline-inducible lentiviruses in mouse cells allowed the examination of mechanisms governing somatic cell reprogramming. RESULTS: Using a doxycycline-inducible human reprogramming system, we identified unreported miRs enhancing reprogramming efficiency. CONCLUSION: We generated a drug-inducible human reprogramming reporter system as an invaluable tool for genetic or chemical screenings. SIGNIFICANCE: These cellular systems provide a tool to enable the advancement of reprogramming technologies in human cells. Reprogramming of somatic cells into induced pluripotent stem cells is achieved by the expression of defined transcription factors. In the last few years, reprogramming strategies on the basis of doxycycline-inducible lentiviruses in mouse cells became highly powerful for screening purposes when the expression of a GFP gene, driven by the reactivation of endogenous stem cell specific promoters, was used as a reprogramming reporter signal. However, similar reporter systems in human cells have not been generated. Here, we describe the derivation of drug-inducible human fibroblast-like cell lines that express different subsets of reprogramming factors containing a GFP gene under the expression of the endogenous OCT4 promoter. These cell lines can be used to screen functional substitutes for reprogramming factors or modifiers of reprogramming efficiency. As a proof of principle of this system, we performed a screening of a library of pluripotent-enriched microRNAs and identified hsa-miR-519a as a novel inducer of reprogramming efficiency.
Subject(s)
Cell Differentiation , Cytological Techniques/methods , Doxycycline/pharmacology , Genes, Reporter/drug effects , Stem Cells/cytology , Animals , Cell Differentiation/drug effects , Cell Line , Drug Evaluation, Preclinical/methods , Gene Expression/drug effects , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Humans , Lentivirus/genetics , Lentivirus/metabolism , Mice , MicroRNAs/genetics , MicroRNAs/metabolism , Stem Cells/metabolismABSTRACT
We studied the association between common haplotypes in six relevant lipid metabolism genes with plasma lipid levels. We selected single-nucleotide polymorphisms (SNPs) in the cholesterol ester transfer protein (CETP), lipoprotein lipase (LPL), hepatic triglyceride lipase (HL), low-density lipoprotein cholesterol receptor (LDLR), apolipoprotein E (ApoE) and lecithin-cholesterol acyltransferase (LCAT) genes, and studied 732 individuals from 184 German families. Total cholesterol (TC), low-density lipoprotein cholesterol (LDL) and high-density lipoprotein cholesterol (HDL) were similar to those reported in other European and American populations. Haplotypes derived from SNP combinations resulted in more significance and of a higher degree than did single SNPs in the genotype-phenotype association analysis. Reduction of the polygenic variance attributable to haplotypes was estimated using variance components analysis. Under the biometrical genetic model, allelic association of haplotypes was highly significant for HDL, LDL and the LDL/HDL ratio. The residual kinship correlation was reduced accordingly. The ApoE gene had a strong effect on trait variation; however, the other genes also contributed substantially. An epistatic interaction could not be demonstrated in this sample. The data are consistent with the notion that common genetic variants influence common traits.
