Generating induced pluripotent stem cells from common marmoset (Callithrix jacchus) fetal liver cells using defined factors, including Lin28

Abstract

Although embryonic stem (ES) cell-like induced pluripotent stem (iPS) cells have potential therapeutic applications in humans, they are also useful for creating genetically modified human disease models in nonhuman primates. In this study, we generated common marmoset iPS cells from fetal liver cells via the retrovirus-mediated introduction of six human transcription factors: Oct-3/4, Sox2, Klf4, c-Myc, Nanog, and Lin28. Four to five weeks after introduction, several colonies resembling marmoset ES cells were observed and picked for further expansion in ES cell medium. Eight cell lines were established, and validation analyses of the marmoset iPS cells followed. We detected the expression of ES cell-specific surface markers. Reverse transcription-PCR showed that these iPS cells expressed endogenous Oct-3/4, Sox2, Klf4, c-Myc, Nanog and Lin28 genes, whereas all of the transgenes were silenced. Karyotype analysis showed that two of three iPS cell lines retained a normal karyotype after a 2-month culture. Both embryoid body and teratoma formation showed that marmoset iPS cells had the developmental potential to give rise to differentiated derivatives of all three primary germ layers. In summary, we generated marmoset iPS cells via the transduction of six transcription factors; this provides a powerful preclinical model for studies in regenerative medicine.

Naive-like Conversion Overcomes the Limited Differentiation Capacity of Induced Pluripotent Stem Cells

“…4B). Moreover, the increased expression of the candidate genes, KLF4 and KLF5, 16 ) and primed state iPS-S1 (PN 16 ) cells (left panels), and of the naivelike state iPS-L1 (N 34 ) and iPS-S1 (N 41 ) cell colonies (right panels), which all expressed EF1␣ and hOCT3/4-GFP (middle panels) and alkaline phosphatase activity (bottom panels). The passage number is denoted "PN number " or "N number."…”

“…C, increased expressions of rabbit KLF4 and KLF5 after naive-like conversion were confirmed by quantitative RT-PCR. D, EF1␣-OCT3/4-GFP expression in rabbit blastocysts 48 h after an injection into 8-cell embryos of primed state iPS-L1 (PN 16 ) cells and iPS-S1 (PN 16 ) cells (left eight panels) or naive-like iPS-L1 (N 39 ) and iPS-S1 (N 40 ) cells (right eight panels). When primed state iPS cells were injected, putatively differentiated iPS cells with almost no hOCT3/4-GFP expression could not contribute to the ICM of blastocysts (arrowheads).…”

Comparative Anatomy of Marmoset and Mouse Cortex from Genomic Expression

“…However, the disadvantages of using such monkeys for experimental manipulations include their large body size, inefficient fertility, and slow development to sexual maturity. Previously, the common marmoset (Callithrix jacchus), a small New World monkey species, has emerged as a promising model for scientific studies including the investigation of neurological disorders, basic and behavioral neuroscience, neuroimaging, stem cell research, drug toxicology, immunity and autoimmune diseases, and reproductive biology (Poswillo et al, 1972;Mansfield, 2003;Iwanami et al, 2005a,b;Sasaki et al, 2005;Leuner et al, 2007;Tomioka et al, 2010;Hikishima et al, 2011;Yamazaki et al, 2011). The marmoset has a number of advantageous characteristics as an experimental animal, including a small body size, high fertility, and early sexual maturity (they reach adulthood at 1.5-2 years).…”

iPS Cells: Born-Again Stem Cells for Biomedical Applications

“…iPSCs have several advantages in addition to overcoming ESC-specific obstacles. On a technique level, iPSCs are easier to derive than ESCs, and iPSC lines have already been derived from several species for which no ESC lines exist (Esteban et al, 2009;Li et al, 2009;Tomioka et al, 2010;Wu et al, 2009). This is because the optimal conditions for deriving ESCs vary across species [for example, human ESCs are maintained with basic fibroblast growth factor, while mouse ESCs are maintained with leukemia inhibitory factor], while induced pluripotency is conserved across mammals.…”