Biology of stem cells: an overview – PMC – National Center for …

Kidney Int Suppl (2011). 2011 Sep; 1(3): 6367.

1Department of Genetics, Universidade Federal do Rio Grande do Sul, Porto Alegre, Brazil

1Department of Genetics, Universidade Federal do Rio Grande do Sul, Porto Alegre, Brazil

2Postgraduation Program in Genetic and Molecular Diagnosis, Universidade Luterana do Brasil, Canoas, Brazil

1Department of Genetics, Universidade Federal do Rio Grande do Sul, Porto Alegre, Brazil

2Postgraduation Program in Genetic and Molecular Diagnosis, Universidade Luterana do Brasil, Canoas, Brazil

Stem cells are defined as precursor cells that have the capacity to self-renew and to generate multiple mature cell types. Only after collecting and culturing tissues is it possible to classify cells according to this operational concept. This difficulty in identifying stem cells in situ, without any manipulation, limits the understanding of their true nature. This review aims at presenting, to health professionals interested in this area, an overview on the biology of embryonic and adult stem cells, and their therapeutic potential.

Keywords: adult stem cells, biological characteristics, cell therapy, embryonic stem cells, human diseases

Although the initial concept of stem cells is more than 100 years old,1 and much of its biology and therapeutic potential has been explored in the past three decades, we still know little about their true nature. This review is intended to provide an overview on the biology of stem cells and their therapeutic potential to those interested in this field.

Stem cells are operationally defined as cells that have the potential for unlimited or prolonged self-renewal, as well as the ability to give rise to at least one type of mature, differentiated cells.2, 3 Although this basic definition of stemness' applies generally to stem cells, it is necessary to individually consider embryonic and adult stem cells as they do not share much more than the name and the basic definition above.

In humans, the embryo is defined as the organism from the time of implantation in the uterus until the end of the second month of gestation. Embryonic stem cells (ESCs), however, refer to a much more restricted period, resulting from the isolation and cultivation of cells from the blastocyst, which forms at approximately 5 days after fertilization.4

The zygote, which is the cell resulting from the fertilization of an oocyte by a spermatozoon, is totipotent. Several successive cell divisions generate the morula, with 3264 totipotent cells. After that stage, it develops into the blastocyst, which consists of a hollow ball of cells. Peripheral cells (the trophoblast) of the blastocyst generate the embryonic membranes and placenta, whereas the inner cell mass develops into the fetus. These are the cells that are used to establish stem cell cultures (). They are not totipotent, as they do not have the ability to support the formation of another embryo, and are considered to be pluripotent as they can produce all the cell types of the adult organism. Further development of the embryo leads to the formation of the gastrula, composed of the three germ layers (ectoderm, mesoderm, and endoderm), from which the complete organism develops.

Embryonic stem cell cultivation. The zygote undergoes successive mitotic divisions until a sphere of cellsthe blastocystis formed. In the blastocyst, the trophoblast at its periphery generates the embryonic membranes and placenta, whereas the inner cell mass develops into the fetus. Embryonic stem cells are immortal in culture, having been established from one pluripotent cell collected from the inner cell mass. These are capable of differentiating into any of the mature cell types present in the adult organism.

In 1981, two groups established the first ESC lines from mouse blastocysts, and in 1998 the first human ESC line was generated.5 Although seemingly simple, the procedure is technically demanding because of the need for strictly controlled conditions necessary for the maintenance of the cells in the undifferentiated state. This is particularly important for human ESCs.6 Once established, ESC lines may be maintained in permanent culture, frozen and thawed, and transported between laboratories. It is estimated that there are currently around 250 human ESC lines in the world, widely shared among different groups. The process of establishing an ESC line requires, however, the destruction of the blastocyst, raising ethical issues as scientific investigation alone is not capable of determining whether blastocysts constitute human beings. An alternative method involves the production of ESCs by collection of only one cell from the inner cell mass, allowing implantation of the remaining cells in the womb. However, ethical considerations still remain as it has to be tested whether the remaining cells can develop into a normal human being.

Cultured ESCs show defined characteristics: they are pluripotent, capable of differentiating into cells derived from all three germ layers; they are immortal in culture and may be maintained for several hundred passages in the undifferentiated state; and they maintain a normal chromosomal composition.

Molecular characterization of ESCs is well developed, and they are known to express surface markers such as CD9, CD24, and alkaline phosphatase, and several genes involved with pluripotency, including Oct-4, Rex-1, SOX-2, Nanog, LIN28, Thy-1, and SSEA-3 and -4.7 Expression of high levels of telomerase explains their immortality in culture.

ESC research focuses mainly on two issues, both of which have shown significant progress in the past few years.6 The first point explores how to better maintain the cells in long-term culture, without significant modifications of their genetic composition and, in the case of human ESCs, avoiding the need for animal products in the culture. Generally, the cells are maintained in culture on feeder cells such as mouse fibroblasts. The second point focuses on how to differentiate the cells into the many mature cell types that are necessary for the potential treatment of different diseases. ESCs can be induced to differentiate into various cell types in suspension culture, resulting in three-dimensional cell aggregates called embryoid bodies. This tendency of ESCs to differentiate spontaneously may not always be desirable. A technical challenge is to control the differentiation process: although the addition of growth factors directs the differentiation process, usually the cultures spontaneously differentiate into various cell types. It is thus necessary to use methods that allow removal of undifferentiated ESCs from cultures in which the differentiated cell types are the desired product.

Recently, methods for direct reprogramming of adult cells to induced pluripotent stem cells have been developed.8 In the process, mature cells from the patient are treated in vitro with genes that dedifferentiate' them to a pluripotent stage, similar to an ESC (). Induced pluripotent stem cells are believed to be identical to natural pluripotent ESCs in many respects, including the expression of specific genes and proteins, chromatin methylation patterns, culture kinetics, in vitro differentiation patterns, and teratoma formation. Besides avoiding the ethical issues associated with the destruction of human embryos, this approach allows the generation of patient-specific cells of any lineage. Problems related to the genetic modification of target cells, however, must still be resolved before induced pluripotent stem cells may be clinically tested.

Production of induced pluripotent stem (iPS) cells. iPS cells are produced by treating mature cells, such as fibroblasts, with genes that dedifferentiate' them to a pluripotent stage, similar to an embryonic stem cell. Viral vectors, such as retroviruses, are generally used for gene transfer. The transformed cells become morphologically and biochemically similar to pluripotent stem cells, with the advantage of representing autologous cells in therapeutic applications.

The principal advantage of ESCs over adult stem cells is related to their pluripotency and limitless expansion in culture, as they have the potential to give rise to all cell types composing the adult organism. This potential is exploited in vitro to develop specialized cells that are then used in therapy.

Owing mainly to safety issues, the clinical use of hESCs is much more restricted than that of adult stem cells. As proof of pluripotency, ESC lineages injected into immunodeficient mice must lead to teratoma formation, with derivatives of all three germ layers. Only differentiated cells derived from ESCs may be administered to patients, as any contaminating undifferentiated cells could give rise to cancer. The first clinical trial using human ESC-derived cells, which in this case are oligodendrocyte progenitor cells, was started in October 2010. Care must be taken, however, to not call this procedure human ESC therapy', as the cells to be used are no longer ESCs.

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