Generation and molecular characterization of human pluripotent stem cell-derived pharyngeal foregut endoderm.

Approaches to study human pharyngeal foregut endoderm-a developmental intermediate that is linked to various human syndromes involving pharynx development and organogenesis of tissues such as thymus, parathyroid, and thyroid-have been hampered by scarcity of tissue access and cellular models. We present an efficient stepwise differentiation method to generate human pharyngeal foregut endoderm from pluripotent stem cells. We determine dose and temporal requirements of signaling pathway engagement for optimized differentiation and characterize the differentiation products on cellular and integrated molecular level.

Using an Inducible CRISPR-dCas9-KRAB Effector System to Dissect Transcriptional Regulation in Human Embryonic Stem Cells.

CRISPR-Cas9 effector systems have wide applications for the stem cell and regenerative medicine field. The ability to dissect the functional gene regulatory networks in pluripotency and potentially in differentiation intermediates of all three germ layers makes this a valuable tool for the stem cell community. Catalytically inactive Cas9 fused to transcriptional/chromatin effector domains allows for silencing or activation of a genomic region of interest.

MiRNA Profiling in Human Induced Pluripotent Stem Cells.

Human iPS cells are capable of differentiation towards all three germ layer lineages. As well as being of use for the study of developmental biology, these cells also provide an excellent resource for disease modeling and cell replacement therapies. iPS cells derived from an individual with a disease type of interest can be differentiated towards the cell type afflicted in that particular disease.

MicroRNA expression analysis: techniques suitable for studies of intercellular and extracellular microRNAs.

MicroRNAs, the class of small ribo-regulators, have been implicated in the regulation of a range of different biological processes, including development and differentiation, proliferation, and cell death. Only for a small fraction of identified microRNAs has a function been elucidated; therefore, a great deal of research remains to be performed to fully understand the role and implications of microRNAs.This chapter discusses protocols for the isolation of microRNAs, reverse transcription, PCR, and large scale profiling using TaqMan low density miRNA arrays for analysis of microRNA expression levels.

Identification of microRNAs with a role in glucose stimulated insulin secretion by expression profiling of MIN6 cells.

MicroRNAs (miRNAs) are a family of endogenous small non-coding RNAs which regulate mRNAs at the post-transcriptional level. MiRNAs have been identified in both normal physiological and pathological conditions. To date, a limited number of miRNAs have been shown to be involved in the regulation of insulin secretion.

Molecular medicine of microRNAs: structure, function and implications for diabetes.

MicroRNAs (miRNAs) are a family of endogenous small noncoding RNA molecules, of 19-28 nucleotides in length. In humans, up to 3% of all genes are estimated to encode these evolutionarily conserved sequences. miRNAs are thought to control expression of thousands of target mRNAs.