Project description:We report here genome wide identification of p63 binding sites in cycling neonatal foreskin keratinocytes using high throughput sequencing of ChIP enriched DNA. Analysis of gene ontology, database mining with integration with publicly available data, reveals a role for p63 in transcriptional regulation of multiple genes genetically linked to cleft palate. In addition, we identify AP-2α, a transcription factor which, when mutated, also results in craniofacial clefting syndrome, as a co-regulator of p63 responsive genes. Examination of p63 binding sites in neonatal foreskin keratinocytes
Project description:Pericytes derived from skin dermis can substantially enhance the short-term tissue-regenerative capacity of human epidermal cells already committed to differentiation; they also display both phenotypic and functional properties of mesenchymal stem cells. In this microarray analysis, we compared the gene expression profile of dermal pericytes to that of the remaining dermal cells of neonatal human foreskin. Experiment Overall Design: Human neonatal foreskin was digested overnight in dispase II at 4°C to separate the epidermis from the dermis. Subsequently the dermis was digested for 1-2 hours at 37°C in a mixed dispase and collagenase solution and then fractionated into two populations, i.e. pericytes (HD-1bri) and the remaining dermal cells (HD-1dim), on the basis of differential VLA-1 expression using fluorescence-activated cell sorting. Total RNA from 15,000 cells of each population was extracted from 4 independent replicate sorts. mRNAs were amplified using a T7-primer-based-2-round linear RNA amplification protocol (GeneChip Two-Cycle cDNA synthesis kit). Fragmented and biotin-labelled cRNA from each individual sample was hybridised to Affymetrix HG-U133 plus 2.0 arrays and scanned on a Affymetrix GeneChip scanner. Probe intensities were RMA normalized and log2-transformed expression values were compared using moderated t statistics to quantify differences between individual samples.
Project description:We report here genome wide identification of p63 binding sites in cycling neonatal foreskin keratinocytes using high throughput sequencing of ChIP enriched DNA. Analysis of gene ontology, database mining with integration with publicly available data, reveals a role for p63 in transcriptional regulation of multiple genes genetically linked to cleft palate. In addition, we identify AP-2α, a transcription factor which, when mutated, also results in craniofacial clefting syndrome, as a co-regulator of p63 responsive genes.
Project description:In this study human neonatal foreskin fibroblasts (HFF1 and BJ) were lipofected with an equal mixture of human reprogramming factor-encoding mRNAs. The cells were lysed and total RNA was harvested 24 hours post-transfection. Gene regulation was evaluated with respect to corresponding mock-transfected, Lipofectamine-treated control fibroblasts. For comparison, wildtype (un-treated) HFF1 and BJ fibroblasts as well as undifferentiated HFF1- and BJ-derived induced pluripotent stem cells and human embryonic stem cells (generated and maintained in our laboratory (refer to GSE26575) were included in the analysis. Total RNA obtained from untreated human neonatal foreskin fibroblasts, reprogramming factor- and mock(Lipofectamine-treated)-transfected human neonatal fibroblasts, undifferentiated hESCs and iPSCs (derived from human neonatal foreskin fibroblasts).
Project description:Melanocyte-keratinocyte interactions are vital for regulating melanogenesis and maintaining epidermal homeostasis. However, most 3D human skin equivalents lack melanocytes, limiting their relevance for pigmentation studies. To address this, we utilized a pigmented human epidermal equivalent (PmtHEE) that incorporates melanocytes into the epidermis. Using single-cell RNA sequencing (scRNA-seq), we characterized PmtHEE and compared it with neonatal foreskin epidermis (FsEpi) and a fibroblast-containing human skin equivalent model (FibHSE).
Project description:We sought to understand the differences between the basal and suprabasal layers of normal human skin epidermis. Comparison of transcriptomes among basal epidermis, suprabasal epidermis, whole epidermis, and reticular dermis
