The very property that puts adult stem cells in high demand, their ability to differentiate into mature cells with specialized functions, is a barrier to their expansion in culture. In many cases, tissue cell differentiation programs conclude with terminal division arrest and, eventually, cell death. Therefore, propagation and expansion of many types of normal mature cells in sufficient quantities for therapeutic applications is impractical. A method to expand adult somatic stem cells (SSCs), which are programmed for indefinite division, would provide a solution to this problem. After
expanding SSCs to the quantities required, subsequent cell differentiation could yield large numbers of needed mature
functional cells. A general approach is described for clonal expansion of adult SSCs. The method is based on Suppression
of asymmetric cell kinetics (”SACK”), a strategy that limits stem cell differentiation. As a first proof of principle, the
SACK method was recently used to derive hepatic stem cells from adult rat liver. In addition to providing B potential
method for routine expansion of adult SSCs from diverse mammalian tissues, the success of these initial studies
elucidates the nature of “stem cell differentiation” in vitro and in vivo.
Currently, stem cell research is driven by interests in developing innovative cell replacement therapies based on
the potential for stem cells to yield differentiated cells with clinically desirable specialized functions. The cell functions
commonly cited for potential cell replacement treatment include pancreatic beta-cell function for diabetes,
hepatocyte function for liver disease, and neuron regeneration for Parkinson’s disease. Of course, some stem
cell-based therapies are already in routine clinical practice. These include bone marrow transplantation and skin
grafting. Moreover, all organ transplantation formally qualities as stem cell-based therapy, because the continued
function of stem cells in transplanted organs is required for long-term transplant engraftment.
Continued success in biological engineering of stem cells for therapeutic applications will depend on a better
understanding of the biology of stem cell differentiation: However, current ideas on fundamental aspects of this basic
tissue process are widely disparate from one stem cell related research discipline to the next (e.g., tissue engineering research versus cancer research). Here, I discuss a basic point of departure in current concepts regarding the nature of stem cell differentiation and relate new developments in stem cell expansion research that may help to unify thinking on this important topic in biomedical science.
A. Stem cell differentiation, transit cell differentiation, and stem cell transpotency
The biomedical focus on specialized cell function has led to confusion regarding the nature of “stem cell differentiation”. This phrase can have at least two different meanings when used, and often the meaning intended is not explicitly stated. The first meaning is semantically erroneous, as it indicates changes in the stem cell itself. As far as semantics go, this is certainly an incorrect embodiment; because a change in a stem cell would be antithetical to sternness. The essential quality of sternness is that the stem cell produces cells that differentiate, but it itself remains constant and undifferentiated [I]. This meaning for stem cell differentiation may not only be incorrect semantically. It may also he an incorrect characterization of SSCs per se.
The concept that a stem cell can itself differentiate has two origins. The first is the stochastic model of stem cell renewal [1,2]. In this model, stem cells in tissue units are postulated to exist in stable pools from which some differentiate to form specialized tissue cells, whereas others remain as stem cells. The second origin is the recent emergence of reports of stem cell “trans-differentiation.”
Many laboratories have reported that when stem cells are transferred to a heterologous tissue site, they can begin to produce differentiated cells that are unique to the recipient tissue [3,4]. Trans-differentiation has been presented from the perspective that the stem’ cell itself changes fundamentally in different tissue environments . Therefore, though plastic in some respects, stem cell differentiation might he restricted by local tissue factors to yield only the mature cell types of a stem cell’s current tissue of residence.
The second meaning for “stem cell differentiation” makes the phrase itself a misnomer. In this case, what is meant is that the stem cell does not differentiate, but instead it produces progeny cells that differentiate and mature into specialized tissue cells [1,4]. These progeny cells are called transit cells to indicate the fact that they, unlike their parent stem cell, are not long-lived in tissues. The cell kinetics program characterized by an undifferentiated stem cell that continuously divides to produce another undifferentiated stem cell and a transit cells is referred to as deterministic asymmetric cell kinetics [I,4, 6-81, When this meaning is intended, the phrase “transit cell differentiation” should be used. Transit cell differentiation could be responsible for some cases of trans-differentiation. All adult stem cells may share the property of producing transit cells that subsequently are instructed by local micro environment factors to adopt specific differentiation lineages. Thus, adult SSCs would have the property of “transpotency” , defined as the ability to produce differentiated cells of any mature tissue type by virtue of instruction of their transit cell progeny and not themselves.
B. Suppression of asymmefric cell kinetics (SACK) and evidence for adult stem cell transpotency
Asymmetric cell kinetics are required for adult stem cell function in vivo. Ex vivo, the same cell kinetics present a major barrier to the propagation of adult SSCs in culture [7-91. Conversion of SSCs to symmetric cell kinetics, which produce two daughter stem cells, is predicted to promote their exponential expansion in culture. Recently, SACK agents were identified that reversibly convert asymmetric cell kinetics to symmetric cell kinetics. The SACK nucleoside xanthosine (Xs) was used to clonally derive somatic stem cell lines from adult rat liver . The SACK-derived liver stem cells exhibit Xs-dependent asymmetric stem cell kinetics, express alpha-fetoprotein, and produce progeny that express the following indicators of mature hepatocyte differentiation: self-adhesion, hepatocyte morphology, albumin expression, adult hepatocyte H4 antigen expression, and expression of cytochrome p450 activity on the scale of primary hepatocytes [9,10]. The SACK-derived lines divide and produce progeny cells that can be instructed to differentiate into diverse cell types. In different culture environments, the SACK-derived stem cells divide to produce progeny that adopt hepatocyte, neuronal progenitor, or glial-specific phenotypes, representing cell types that develop from at least two different embryonic germ layers, endoderm and ectoderm [Ill. These findings are consistent with the idea that SSCs are transpotent, However, to date, the stable undifferentiated stem cells postulated to exist amidst differentiating transit cells in different culture environments have not been directly identified. Visualizing these predicted constant stem cells would support the concepts of adult SSC transpotency and transit cell-specific instruction and differentiation.
In contrast to cell therapy research, gene therapy research acknowledged a stable undifferentiated state for adult SSCs from its onset [12,13]. SSCs that divide according to a determined asymmetric cell kinetics program are the ideal vessels to carry therapeutic genes. Such cells could persist unchanged for the long periods in the body while continuously producing mature tissue cells that express the required gene product. Increased attention to the heterogeneous cell kinetics architecture of stem cell based tissue units in adult tissues should better inform current efforts to improve existing stem cell therapies (e.g., bone marrow transplant) and promote an acceleration of development of new cell-based treatments. It is also noteworthy that the ability to expand pure populations of stem cells in culture is a major step towards a solution to the other major problem in stem cell biology, development of unique markers for adult stem cells.
James L. Sherley
The Biological Engineering Division, Biotechnology Process Engineering Center
and the Center for Environmental Health Sciences
Massachusetts Institute of Technology, Cambridge, MA, USA