Additive manufacturing, otherwise known as three-dimensional (3D) printing, is driving major innovations in many areas, such as engineering, manufacturing, art, education and medicine. Recent advances have enabled 3D printing of biocompatible materials, cells and supporting components into complex 3D functional living tissues. 3D bioprinting is being applied to regenerative medicine to address the need for tissues and organs suitable for transplantation. Compared with non-biological printing, 3D bioprinting involves additional complexities, such as the choice of materials, cell types, growth and differentiation factors, and technical challenges related to the sensitivities of living cells and the construction of tissues. Addressing these complexities requires the integration of technologies from the fields of engineering, biomaterials science, cell biology, physics and medicine. 3D bioprinting has already been used for the generation and transplantation of several tissues, including multilayered skin, bone, vascular grafts, tracheal splints, heart tissue and cartilaginous structures. Other applications include developing high-throughput 3D-bioprinted tissue models for research, drug discovery and toxicology.

3D bioprinting of tissues and organs

https://www.ics.uci.edu/~xhx/courses/references/Young_ES_regulator_cell06.pdf

Core transcriptional regulatory circuitry in human embryonic stem cells.

Functional expression cloning of nanog, a pluripotency sustaining factor in embryonic stem cells

https://www.cell.com/cell/pdf/S0092-8674(03)00393-3.pdf

The Homeoprotein Nanog Is Required for Maintenance of Pluripotency in Mouse Epiblast and ES Cells

In the three decades since pluripotent mouse embryonic stem (ES) cells were first described they have been derived and maintained by using various empirical combinations of feeder cells, conditioned media, cytokines, growth factors, hormones, fetal calf serum, and serum extracts. Consequently ES-cell self-renewal is generally considered to be dependent on multifactorial stimulation of dedicated transcriptional circuitries, pre-eminent among which is the activation of STAT3 by cytokines (ref. 8). Here we show, however, that extrinsic stimuli are dispensable for the derivation, propagation and pluripotency of ES cells. Self-renewal is enabled by the elimination of differentiation-inducing signalling from mitogen-activated protein kinase. Additional inhibition of glycogen synthase kinase 3 consolidates biosynthetic capacity and suppresses residual differentiation. Complete bypass of cytokine signalling is confirmed by isolating ES cells genetically devoid of STAT3. These findings reveal that ES cells have an innate programme for self-replication that does not require extrinsic instruction. This property may account for their latent tumorigenicity. The delineation of minimal requirements for self-renewal now provides a defined platform for the precise description and dissection of the pluripotent state.

/pdf/the-ground-state-of-embryonic-stem-cell-self-renewal-41xqrmi7s8.pdf

The ground state of embryonic stem cell self-renewal

We describe the derivation of pluripotent embryonic stem (ES) cells from human blastocysts. Two diploid ES cell lines have been cultivated in vitro for extended periods while maintaining expression of markers characteristic of pluripotent primate cells. Human ES cells express the transcription factor Oct-4, essential for development of pluripotential cells in the mouse. When grafted into SCID mice, both lines give rise to teratomas containing derivatives of all three embryonic germ layers. Both cell lines differentiate in vitro into extraembryonic and somatic cell lineages. Neural progenitor cells may be isolated from differentiating ES cell cultures and induced to form mature neurons. Embryonic stem cells provide a model to study early human embryology, an investigational tool for discovery of novel growth factors and medicines, and a potential source of cells for use in transplantation therapy.

Embryonic stem cell lines from human blastocysts: somatic differentiation in vitro.

https://www.cell.com/cell-stem-cell/pdf/S1934-5909(15)00267-2.pdf

Stem Cell Therapies in Clinical Trials: Progress and Challenges

The widespread use of clomiphene citrate and exogenous gonadotrophins for in vitro fertilization (IVF) in human frequently results in the production of multiple embryos. Replacement of more than two embryos increases pregnancy rate but may result in multiple pregnancies with increased pre- and post-natal abnormality. Preservation of embryos for a limited time allows fewer embryos to be replaced on several different occasions and thus the problems of multiple pregnancy can be minimized, the effectiveness of a single IVF procedure increased and embryo replacement in adverse maternal conditions avoided. Preimplantation embryos have been successfully cryopreserved in many animal species. The sensitivity of embryos to cooling and freezing varies between species and stages of embryo development. We report here the cryopreservation procedures that allow a high survival rate of four- and eight-cell human embryos and the establishment of a pregnancy following the freezing and storage of an eight-cell embryo for 4 months in liquid nitrogen. The pregnancy terminated at 24 weeks' gestation due to development of a septic Streptomyces agalactiae chorion amnionitis after premature membrane rupture.

Human pregnancy following cryopreservation, thawing and transfer of an eight-cell embryo

Embryonic stem (ES) cells are cells derived from the early embryo that can be propagated indefinitely in the primitive undifferentiated state while remaining pluripotent; they share these properties with embryonic germ (EG) cells. Candidate ES and EG cell lines from the human blastocyst and embryonic gonad can differentiate into multiple types of somatic cell. The phenotype of the blastocyst-derived cell lines is very similar to that of monkey ES cells and pluripotent human embryonal carcinoma cells, but differs from that of mouse ES cells or the human germ-cell-derived stem cells. Although our understanding of the control of growth and differentiation of human ES cells is quite limited, it is clear that the development of these cell lines will have a widespread impact on biomedical research.

Human embryonic stem cells

Ovarian steroid replacement therapy in the ovariectomized ewe, given in the correct sequence to mimic endogenous steroid changes in the normal ovulatory cycle, allows the development of embryos transferred in utero. A similar type of sequential therapy was designed for steroid replacement in women with primary ovarian failure. This produces the histological changes in uterine endometrial morphology and plasma oestradiol and progesterone similar to those observed in the normal ovulatory cycle. We now report that in one of these women a donated oocyte, fertilized by her husband's spermatozoa and cultured to the two-cell stage in vitro, was transferred in utero, resulting in a normal pregnancy and the delivery of a healthy child. Oestrogen therapy was withdrawn at 12 weeks and progesterone at 19 weeks gestation. This technique allows the treatment of human infertility due to primary ovarian failure.

Alan O Trounson

Papers

Embryonic stem cell lines from human blastocysts: somatic differentiation in vitro.

Stem Cell Therapies in Clinical Trials: Progress and Challenges

Human pregnancy following cryopreservation, thawing and transfer of an eight-cell embryo

Human embryonic stem cells

The establishment and maintenance of pregnancy using in vitro fertilization and embryo donation in a patient with primary ovarian failure