https://repository.kulib.kyoto-u.ac.jp/dspace/bitstream/2433/159777/1/j.cell.2006.07.024.pdf

Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors.

Somatic cell nuclear transfer allows trans-acting factors present in the mammalian oocyte to reprogram somatic cell nuclei to an undifferentiated state. We show that four factors (OCT4, SOX2, NANOG, and LIN28) are sufficient to reprogram human somatic cells to pluripotent stem cells that exhibit the essential characteristics of embryonic stem (ES) cells. These induced pluripotent human stem cells have normal karyotypes, express telomerase activity, express cell surface markers and genes that characterize human ES cells, and maintain the developmental potential to differentiate into advanced derivatives of all three primary germ layers. Such induced pluripotent human cell lines should be useful in the production of new disease models and in drug development, as well as for applications in transplantation medicine, once technical limitations (for example, mutation through viral integration) are eliminated.

https://science.sciencemag.org/content/sci/318/5858/1917.full.pdf

Induced Pluripotent Stem Cell Lines Derived from Human Somatic Cells

Cells of a multicellular organism are genetically homogeneous but structurally and functionally heterogeneous owing to the differential expression of genes. Many of these differences in gene expression arise during development and are subsequently retained through mitosis. Stable alterations of this kind are said to be 'epigenetic', because they are heritable in the short term but do not involve mutations of the DNA itself. Research over the past few years has focused on two molecular mechanisms that mediate epigenetic phenomena: DNA methylation and histone modifications. Here, we review advances in the understanding of the mechanism and role of DNA methylation in biological processes. Epigenetic effects by means of DNA methylation have an important role in development but can also arise stochastically as animals age. Identification of proteins that mediate these effects has provided insight into this complex process and diseases that occur when it is perturbed. External influences on epigenetic processes are seen in the effects of diet on long-term diseases such as cancer. Thus, epigenetic mechanisms seem to allow an organism to respond to the environment through changes in gene expression. The extent to which environmental effects can provoke epigenetic responses represents an exciting area of future research.

/pdf/epigenetic-regulation-of-gene-expression-how-the-genome-55fnpf16ec.pdf

Epigenetic regulation of gene expression: how the genome integrates intrinsic and environmental signals

We report here that cells co-purifying with mesenchymal stem cells--termed here multipotent adult progenitor cells or MAPCs--differentiate, at the single cell level, not only into mesenchymal cells, but also cells with visceral mesoderm, neuroectoderm and endoderm characteristics in vitro. When injected into an early blastocyst, single MAPCs contribute to most, if not all, somatic cell types. On transplantation into a non-irradiated host, MAPCs engraft and differentiate to the haematopoietic lineage, in addition to the epithelium of liver, lung and gut. Engraftment in the haematopoietic system as well as the gastrointestinal tract is increased when MAPCs are transplanted in a minimally irradiated host. As MAPCs proliferate extensively without obvious senescence or loss of differentiation potential, they may be an ideal cell source for therapy of inherited or degenerative diseases.

/pdf/pluripotency-of-mesenchymal-stem-cells-derived-from-adult-200s4qlizf.pdf

Pluripotency of mesenchymal stem cells derived from adult marrow

Fertilization of mammalian eggs is followed by successive cell divisions and progressive differentiation, first into the early embryo and subsequently into all of the cell types that make up the adult animal. Transfer of a single nucleus at a specific stage of development, to an enucleated unfertilized egg, provided an opportunity to investigate whether cellular differentiation to that stage involved irreversible genetic modification. The first offspring to develop from a differentiated cell were born after nuclear transfer from an embryo-derived cell line that had been induced to become quiescent. Using the same procedure, we now report the birth of live lambs from three new cell populations established from adult mammary gland, fetus and embryo. The fact that a lamb was derived from an adult cell confirms that differentiation of that cell did not involve the irreversible modification of genetic material required for development to term. The birth of lambs from differentiated fetal and adult cells also reinforces previous speculation that by inducing donor cells to become quiescent it will be possible to obtain normal development from a wide variety of differentiated cells.

Viable offspring derived from fetal and adult mammalian cells

Halothane positive and negative selection lines were established to estimate the effects of the halothane gene on performance in British Landrace pigs. Incidences of positive reaction diverged from 0·12 in the foundation generation to 0·93 in the positive and to 0·02 in the negative line in generation 4. Differences in litter productivity as a trait of the dam were estimated from a total of 399 positive × positive and negative × negative parity 1 and 2 matings in generations 1 to 4. Compared with negative contemporaries, positive females showed non-significant reductions in litter size at birth (−0·10, s.e. 0·26) and at 42 days (−0·28, s.e. 0·26), accompanied by significant reductions in average piglet weights at birth (−0·11, s.e. 0·02 kg) and 42 days (−0·4, s.e. 0·2 kg). There were no differences in conception rate or adult live weights. A subsample of 69 parity 2 and 3 sows slaughtered 30 days after mating showed no significant difference in ovulation rate or embryo survival, but for positive dams the length of embryos was significantly reduced (−3·1, s.e. 1·1 mm). The study suggests that the principal effect of the halothane gene was in reducing foetal growth rather than litter size. However, the phenotypic difference is expected to under-estimate the genetic difference between homozygotes, and it was not possible to distinguish the effects of dam and offspring genotypes. The present economic loss of roughly £4·70 per positive litter would be unlikely to justify elimination of the gene from a purebred maternal Landrace herd.

Performance of British Landrace pigs selected for high and low incidence of halothane sensitivity

In a previous study of mouse tetraploid[harr ]diploid chimaeric blastocysts, tetraploid cells were found to be more abundant in the trophectoderm than the inner cell mass (ICM) and more abundant in the mural trophectoderm than the polar trophectoderm. This non-random allocation of tetraploid cells to different regions of the chimaeric blastocyst may contribute to the restricted tissue distribution seen in postimplantation stage tetraploid[harr ]diploid chimaeras. However, the tetraploid and diploid embryos that were aggregated together differed in several respects: the tetraploid embryos had fewer cells and these cells were bigger and differed in ploidy. Each of these factors might underlie a non-random allocation of tetraploid cells to the chimaeric blastocyst. A combination of micromanipulation and electrofusion was used to produce two series of chimaeras that distinguished between the effects of cell size and ploidy on the allocation of cells to different tissues in chimaeric blastocysts. When aggregated cells differed in cell size but not ploidy, the derivatives of the larger cell contributed significantly more to the mural trophectoderm and polar trophectoderm than the ICM. When aggregated cells differed in ploidy but not cell size, the tetraploid cells contributed significantly more to the mural trophectoderm than the ICM. In both experiments the contributions to the polar trophectoderm tended to be intermediate between those of the mural trophectoderm and ICM. These experiments show that both the larger size and increased ploidy of tetraploid cells could have contributed to the non-random cell distribution that was observed in a previous study of tetraploid[harr ]diploid chimaeric blastocysts.

The effects of cell size and ploidy on cell allocation in mouse chimaeric blastocysts.

Summary. Corpora lutea induced in pigs on Day 6 regressed with the naturally formed corpora lutea towards the end of the cycle. Hormonal induction of corpora lutea on Days 8, 10 or 16 significantly prolonged the cycle despite involution of the original corpora lutea about 16 days after ovulation. The interval between the exogenous administration of hcg and the appearance of the next oestrus varied with the day on which the accessory corpora lutea were formed. The estimated functional life span of the corpora lutea induced on Days 6, 8, 10 and 16 was 12\m=.\5 14\m=.\5, 15\m=.\0and 19\m=.\3 days respectively. That these induced corpora lutea were fully functional was shown by the luteal progesterone levels estimated using the protein-binding and u.v. spectrophotometric techniques. Hysterectomy resulted in both the spontaneous corpora lutea and those induced on Day 6 being maintained for at least 60 days. It is concluded that Days 6 to 8 may represent a critical phase in the relationship between the uterus and the corpora lutea in non-pregnant pigs.

/pdf/the-relationship-between-day-of-formation-and-functional-2kftwya48s.pdf

The relationship between day of formation and functional life span of induced corpora lutea in the pig.

An argument for the benefits of cloning, co-written by a scientist whose team was responsible for a famous cloned sheep, presents the reasons for his opposition to the cloning of humans and explains that cloning technology can be ethically applied to free families from serious hereditary diseases.

Ian Wilmut

Papers

Performance of British Landrace pigs selected for high and low incidence of halothane sensitivity

The effects of cell size and ploidy on cell allocation in mouse chimaeric blastocysts.

The relationship between day of formation and functional life span of induced corpora lutea in the pig.

After Dolly: The Uses and Misuses of Human Cloning

Embryo cloning in sheep: work in progress.