Human embryonic stem cell cultivation: historical perspective and evolution of xeno-free culture systems

Abstract

Human embryonic stem cells (hESC) have emerged as attractive candidates for cell-based therapies that are capable of restoring lost cell and tissue function. These unique cells are able to self-renew indefinitely and have the capacity to differentiate in to all three germ layers (ectoderm, endoderm and mesoderm). Harnessing the power of these pluripotent stem cells could potentially offer new therapeutic treatment options for a variety of medical conditions. Since the initial derivation of hESC lines in 1998, tremendous headway has been made in better understanding stem cell biology and culture requirements for maintenance of pluripotency. The approval of the first clinical trials of hESC cells for treatment of spinal cord injury and macular degeneration in 2010 marked the beginning of a new era in regenerative medicine. Yet it was clearly recognized that the clinical utility of hESC transplantation was still limited by several challenges. One of the most immediate issues has been the exposure of stem cells to animal pathogens, during hESC derivation and during in vitro propagation. Initial culture protocols used co-culture with inactivated mouse fibroblast feeder (MEF) or human feeder layers with fetal bovine serum or alternatively serum replacement proteins to support stem cell proliferation. Most hESC lines currently in use have been exposed to animal products, thus carrying the risk of xeno-transmitted infections and immune reaction. This mini review provides a historic perspective on human embryonic stem cell culture and the evolution of new culture models. We highlight the challenges and advances being made towards the development of xeno-free culture systems suitable for therapeutic applications.

Distinct Metabolic States Can Support Self-Renewal and Lipogenesis in Human Pluripotent Stem Cells under Different Culture Conditions

“…Successful establishment of culture conditions able to maintain human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) in the undifferentiated state represented critical steps in advancing these technologies to practice (Thomson et al, 1998, Takahashi and Yamanaka, 2006). However, the large quantity of cells needed for screening and tissue engineering applications poses a challenge that must still be addressed (Desai et al, 2015). Initial protocols for hPSC self-renewal mimicked the in vivo microenvironment by using feeder cell co-culture or medium conditioned by feeder cells to support hPSC expansion (Villa-Diaz et al, 2013, Desai et al, 2015).…”

Stem Cell-Based Therapy for Diseases of the Retinal Pigment Epithelium: From Bench to Bedside

“…The functional definition of ESCs also includes the capability of maintaining a normal karyotype during growth. 17 In 1998, preceding work in murine models and primates ultimately led to the isolation and characterization of embryonic stem cells from human blastocysts (hESCs) and the demonstration that these cells could be propagated by in vitro co-culture with feeder mouse embryonic fibroblast cells. 18 These hESCs demonstrated high levels of telomerase (indicating their longevity), expressed markers of undifferentiated cells (e.g.…”

“…The study of hESC-derived oligodendrocytes in spinal cord injury was not completed due to high costs and difficulty producing required quantities of oligodendrocyte precursors. 17 Therefore, recently published data from two Ocata Therapeutics (formerly known as Advanced Cell Technology; Marlborough, Massachusetts) trials using hESCs to treat AMD represent the very first proof of the safety of hESC-derived cells to treat human disease. 37,38 In the first of two papers, RPE was generated from hESCs by directed differentiation, adhering to strict cGMP and cGTP guidelines, resulting in a population of RPE with 499% purity confirmed by morphologic and functional characterization.…”

The success of stem cell transplantations and the potential post-transplantation complications may be dependent, among other factors, on the capacity of the recipient and the transplanted cells to repair DNA damage

“…The data about the role of polymorphisms in genes coding for key genes in DNA repair and maintenance of genomic integrity is still very limited. However there is a report on donor cell leukaemia in a recipient that received a transplant from an HLA matched sibling carrying the repair-deficient alleles of the polymorphisms XPD Lys751Gln (rs1052559) and XRCC3 Thr241Met (rs861539) in their homozygous state (Diamond et al 2011). The XPD (ERCC2) gene codes for one of the two helicases that unwind DNA in the vicinity of the lesion site, allowing free access of the machinery for repair by nucleotide excision (NER), whereas XRCC3 codes for a key protein of repair by homologous recombination.…”

“…Thus, potential use of existing ESC lines may be associated with an increased risk for induction of tumorigenesis in the recipient, whereas the use of iPSC may also confer risk for rapid loss of proliferative and differentiation capacity and for shortened lifespan (and respectively, diminished functionality) of the differentiated cell products. For the past few years, many researchers worldwide have recommended that the currently used pluripotent lines may be unsuitable for clinical applications and that establishment of ESC and iPSC lines ought to start anew, in controlled and strictly xenofree conditions (Lãser et al 2009,Arabadjiev et al 2010, Abbasalizadeh and Baharvand 2013, Chakarov et al 2014c, Desai et al 2015. Other authors focus their attention upon the mutation rate of stem cell lines and advise that quantitative evaluation of the mutation rate ought to become a routine criterion in assessment of the properties of the pre-existing and the newly established lines and specifically, in the decision-making about its potential suitability for medical applications (Sverdlov andMineev 2013, Heslop et al 2015).…”