Stem Cells and iPSCs

What are iPSCs

• iPSCs (induced pluripotent stem cells) are a type of pluripotent stem cell artificially derived from a non-pluripotent cell, typically an adult somatic cell, by inducing a "forced" expression of certain genes.
• First produced in 2006 from mouse cells and in 2007 from human cells.

Genes Responsible for Pluripotency

• Group 1: ES cell-specific transcription factors, essential for pluripotency in ES cells and early embryos, including Oct3/4, Sox2, and Nanog.
• Group 2: Proto-oncogenes, important for proliferation of ES cells, but not in early embryos, including TCLI, Stat3, c-Myc, and ERas.
• Group 3: Less famous genes, specifically expressed in ES cells, but with less defined functions, including ECATI, Esg1, and Fbx15.

Key Functions of Genes

• Oct3/4: involved in the maintenance of self-renewal of pluripotent cells, repression leads to formation of trophoectoderm, and overexpression leads to formation of various lineages, including primitive endoderm.
• Sox2: essential for embryonic development, downregulation leads to differentiation of cells in murine ES cells.
• KIf4: represses p53 directly, contributes to activation of Nanog and other ES cell-specific genes, acts as an inhibitor of c-Myc-induced apoptosis, and activates p21CIP, thereby suppressing cell proliferation.
• Nanog: necessary for promoting pluripotency in embryonic stem cells, along with Oct-3/4 and Sox2.
• LIN28: an mRNA binding protein expressed in embryonic stem cells and embryonic carcinoma cells, associated with differentiation and proliferation.

Production of iPSCs

• Typically derived by transfecting certain stem cell-associated genes into non-pluripotent cells, such as adult fibroblasts, using viral vectors, such as retroviruses.
• Small numbers of transfected cells become morphologically and biochemically similar to pluripotent stem cells, and are isolated through morphological selection, doubling time, or through a reporter gene and antibiotic selection.

First Generation of iPSCs

• First generated by Shinya Yamanaka's team at Kyoto University, Japan in 2006, using four key pluripotency genes: Oct-3/4, Sox2, c-Myc, and KIf4.
• Retroviruses were used to transfect mouse fibroblasts, and cells were isolated by antibiotic selection of Fbx15+ cells.
• Limitations: DNA methylation errors compared to original patterns in ESC lines and failed to produce viable chimeras if injected into developing embryos.

Second Generation of iPSCs in Mice

• Generated in June 2007, by the same group, using the same four endogenous pluripotent factors, but with Nanog instead of Fbx15.
• DNA methylation patterns and producing viable chimeras indicated that Nanog is a major determinant of cellular pluripotency.
• Limitations: one of the four genes used (c-Myc) is oncogenic, and 20% of the chimeric mice developed tumors.