Advances in Cell biology, vol. 32, 2005, issue 1

The Primitive Cell Lineages of The Mouse Embryo – Source of Embryonal Stem Cells

Summary: During implantation the mouse embryo called blastocyst is comprised of three cell types with differing developmental potential and function. These cell lineages are specified during two successive rounds of differentiation. They lead to setting apart a group of pluripotent cells, progenitors for the future body capable to give rise to embryonic stem cells in vitro, from the extraembryonic lineages (trophectoderm and primitive endoderm) that take part in formation of the placenta and foetal membranes and give rise to lineage-specific stem cells during in vitro culture. Here, we review classic and contemporary models of primary cell lineages arising during development and perform molecular and cellular regulation of these processes. We describe the role of transcription factors involved in blastocyst structures formation and maintenance of three distinct stem cell types, that can be isolated from them in vitro.

Key words: blastocyst, cell lineages, inner cell mass, trophectoderm, epiblast, hypoblast, stem cells, Oct4, Cdx2, Nanog, Gata4/6.

Human Embryonic Stem Cells – Regulation of Pluripotency and Differentiation

Summary: Embryonic stem (ES) cells derived from preimplantation embryos at the blastocyst stage are unique in their unlimited self-renewal ability and pluripotency allowing their differentiation into any cell type. For these reasons ES cells are considered as a perfect material for basic research on their differentiation capacities, and also for the studies devoted to the development of novel therapies. This article focuses at the mechanisms regulating ES cells pluripotency, self-renewal, and also influencing their differentiation. It also describes alternative methods of the derivation of pluripotent stem cell lines, which do not require embryo destruction and thus do not raise ethical issues. A special attention is paid to the development of ES cell culture.

Summary: In 2006 first paper describing reprogramming of fibroblast into pluripotent cell, i.e. one that is able to differentiate into all tissues was published. Before that spectacular achievement many projects devoted to study the mechanisms of cell de-differentiation led to the establishment of animal cloning techniques and also derivation of embryonic stem cell lines. Present review summarizes the history of these studies and also describes pioneer works leading to derivation of induced pluripotent stem cells, current methods of reprogramming, and possible applications of these cells.

Key words: ES cells, iPS cells, pluripotency, Oct3/4, Sox2, Klf4, c-Myc, differentiation.

Very Small Embryonic Like Stem Cells (VSELs) Isolated From Adult Tissues – An Update

Summary: Accumulating evidence demonstrates that adult tissues contain a population of very primitive pluripotent stem cells and recently, our group identified a population of very small stem cells in murine bone marrow and other adult organs that express several markers characteristic for epiblast/germ line-derived cells. We named these rare cells „very small embryonic/epiblast like stem cells (VSELs)”. We hypothesized that these cells, which are deposited during early gastrulation in developing tissues/organs, play an important role in the turnover of tissue-specific/committed stem cells. Based on this, we envision that germ line is not only the origin but also a „basis/skeleton” for the stem cell compartment in adult life forms. We noticed that VSELs could be mobilized into peripheral blood and the number of these cells circulating in peripheral blood increases during stress and tissue/organ injuries (e.g., heart infarct, stroke). Furthermore, our data indicate that VSELs are protected from uncontrolled proliferation and teratoma formation by a unique pattern of methylation of selected somatic imprinted genes. Finally, we envision that, in pathological situations, VSELs could be involved in development of some malignan-cies (e.g., teratomas, germinal tumors, pediatric sarcomas).

Key words: VSELs, Oct-4, pluripotent stem cells, regeneration, "plasticity”.

Summary: Mesenchymal stem cells (MSC) are considered to be a promising tool for cellular transplantations. This population of cells can find potential applications in multiple clinical disorders as a stimulant for regeneration and/or an immuno-modulator for the treated tissue. As adult stem cells, MSC allow performance of autologous transplantations. Therefore, it is crucial to understand the relationships between ageing and properties of this cell population. This review presents the current knowledge of morphological and functional changes in bone marrow-derived MSC during their ageing both in vivo and in vitro. This review discusses the mechanisms of cell ageing, such as telomere shortening or free radical effects on MSC, along with the susceptibility of cells to negative environmental conditions, depending on donor's age. The analyzed results often differ from each other, but the majority of in vitro studies indicate that the donor's age has an impact on mesenchymal cell properties. The data suggest that cells derived from aged individuals display a lower therapeutic potential then the cells isolated from young organisms. Very few clinical trials conducted in humans do not allow drawing conclusions about the relationship between the donor's age and the transplantation outcome.

Key words: mesenchymal stem cells, ageing, cellular transplantation, differentiation potential.

Summary: Stem cells are cells with a capacity to self-renew and to generate daughter cells that can differentiate into different cell lineages to form all the cell types that are found in the mature tissue. The activity of mammary stem cells and their mitotic progeny is fundamental to normal mammary growth, differentiation and regeneration in successive cycles of pregnancy, lactation and involution. The isolation and characterization of mammary stem cells is fundamental to understanding mammary gland development and tissue homeostasis as well as breast oncogenesis. Several complementary approaches have been employed to isolate, identify and enrich mammary epithelial cells that maintain stem/progenitor cell characteristics. In situ studies of mammary tissue identified potential mammary stem and progenitor cells at the ultrastructural and light microscopy levels at all stages of mammary development. Evidence for the existence of mammary stem and progenitor cells has also been provided by in vitro and in vivo studies. The most useful markers for isolating stem/progenitor cells are the combination of integrin receptors, stem cell antigen-1 (Sca-1), CD24 and lack of steroid hormone receptors, but neither of them are exclusive markers of mammary gland stem cells. The challenge now is to identify new markers, so that these cells can be purified in such a way that meaningful gene expression profiles can be obtained.

Summary: The mammary gland is an organ comprised of branched ductal network terminated by secretory alveoli and embedded in mesenchymal stroma. Mammary alveoli, built by myoepithelial and epithelial cells with apico-basal polarity, obtain their functional differentiation only during lactogenesis. For this reason they represent a good model for studies on differentiation process of epithelium. The emergence of three-dimensional (3D) cell culture systems, based on the use of reconstituted basement membrane (rBM), commercially available as Matrigel, enabled to recreate the in vivo conditions, and brought a great progress in this area of research. The present review summarizes the latest achievements obtained with the use of 3D culture system of mammary gland acini. The results of these studies clearly indicate the important role of extracellular matrix (ECM) components in the regulation of polarization and functional differentiation of mammary luminal epithelium.

Keywords: mammary gland, epithelium, extracellular matrix, integrins, differentiation.

The Role of Antioxidant Genes in Endothelial Progenitor Cells and Muscle Satellite Cells

Summary: The research of last years have proved that regulation of production and neutralization of reactive oxygen species (ROS) play a key role in physiology of progenitor cells, among others endothelial progenitor cells (EPC) and muscle satellite cells. These cells are especially exposed on oxidative stress and are characterised by high expression of antioxidant genes. The paper presents mechanisms of cells protection from harmful, cytotoxic action of ROS as well as points out the significance of low ROS concentration for regulation of migration, proliferation and differentiation of progenitor cells.

Key words: antioxidant genes, endothelial progenitor cells, muscle satellite cells, oxidative stress.

Summary: Increasing number of type 2 diabetes patients (T2DM) represents a major health problem both for physicians and scientists. Despite characterization of endothelial dysfunction and application of new therapeutic strategies, numerous vascular complications lead to earlier death of diabetic patients. It was demonstrated that increased cardiovascular risk in diabetes positively correlates with impaired functions of endothelial progenitor cells (EPC). Moreover, hyperglycemia also attenuates their activities in vitro and in vivo. EPC are able to promote vascular growth and repair vascular injuries, however the exact mechanism of their action is not fully elucidated. They can incorporate into damaged vessels, but growing body of evidence also suggests a paracrine mode of action. Using EPC in cell therapy it may be possible to improve and accelerate healing of chronic wounds and leg ulcers in diabetic patients. However, more studies are needed to fully understand the potential risk of enhanced angiogenesis due to EPC for example in diabetic retinopathy.

Summary: The discovery of endothelial progenitor cells (EPCs) and demonstration of their participation in postnatal new blood vessel formation accelerated their application for therapeutic angiogenesis in cardiovascular diseases and vascularity-dependent disorders treatment. First experiments done on EPCs suggested that usage of these cells in clinic may improve the quality of life of the patients not curable with standard revascularizing surgery. However, the clinical trials of cell therapy with EPCs were not as fruitful as expected. In addition, lack of specified antigens helpful in EPCs population identification and lack of data concerning molecular mechanisms of EPCs function hamper the research on proangiogenic cell therapy. But despite these difficulties, researchers keep hardworking on the strategy of therapeutic angiogenesis and the new discoveries seem to be promising.

Key words: proangogenic cell therapy, endothelial progenitor cells, angiogenesis, vasculogenesis.

Summary: Skeletal muscle tissue is characterized by ability to regenerate in response to injury resulting for example from mechanical trauma, toxin or disease. The key role in this process is played by satellite cells localized in skeletal muscle. However, both under physiological and experimental conditions different types of stem cells originating either from skeletal muscle or other tissues can be also involved in regeneration. This raises the hope for potential use of stem cells in the therapy for skeletal muscles diseases, such as muscular dystrophies or spinal muscle atrophy. These pathologies are characterized by impairment of regeneration linked with depletion or reduction of endogenous population of satellite cells. Thus, transplantation of stem cells could improve regeneration of such muscle and, as a result, support their reconstruction. However, stem cells based therapy will be possible only after precise analysis of their features, especially their myogenic potential, i.e. the ability to differentiate into muscle cells and fibers. This review summarizes current state of knowledge concerning myogenic potential of different types of stem cells and indicates those that are likely to be used in a future as a source of cells transplanted during skeletal muscle therapy.

Key words: skeletal muscles, regeneration, stem cells, transplantation, myogenic potential.

Summary: In this review it has been presented current classification of stem cells indicating novel sources (tissues) of stem cells subtypes that can be used in clinical and pre-clinical attempts of heart regeneration. In the article an analysis of delivery of stem cells to myocardium has been provided dividing stem cells administration into: a) intramyocardial, b) percutaneous, c) systemic ones and its influence on stem cell homing. Quoting the so far published meta-analysis it was outlined the updated view for therapy of particular heart disease by using cell engineering that is for: a) post-infarction heart, b) chronic ischaemia, c) congestive heart failure. Novel approaches were indicated that may optimize clinical protocols of stem cell application for heart regeneration.

In Situ Evaluation of Stem Cell Function Grafted to Organs with 'Low Turnover'

Summary: Every human organ has its own capacity for self renewal due to progenitor cells of tissue reservoir. We can distinguish organs with 'high and low turnover' and for regenerative medicine the latter ones constitute the main target. These are represented by heart, central nervous system and pancreas. In this review, we focused on determination in situ of stem cell function in described organs and tissues. However, even in 'low turnover' systems the level of stem cell homing can be different. Surprisingly it is in favour of central nervous system. The remaining organs present low capacity for cell homing (instead of pancreas, where the ectopic sites can be used for insulin secretion and production) although the attempts to improve this critical issue are undergoing. Recent techniques of imaging as MRI (magnetic resonance imaging), SPECT (single photon emission computed tomography) or PET (positron emission tomography) have ability to precisely locate the cells within target organ and evaluate their functional (metabolic) activities. Beside of their sophistication, these instruments are based on chemical compounds that have their own characteristics, shortcomings and pitfalls. All of these are exemplified in this review together with specific examples of pre-clinical studies upon which the present controversies are outlined.
Key words: in situ imaging, stem cells, organs with 'low turnover'.

Key words: mesenchymal stem cells, ageing, cellular transplantation, differentiation potential.

The Role of Stem Cells in the Process of Endogenic Regeration of Central Nervous System Based on Selected Vascular Diseases

Summary: Central nervous system (CNS) is seen as crucial in the organism and plays critical role in regulation and decision making processes on multisystem and behavioral level. The variety of diseases affecting CNS, and lack of effective therapy raises a question of chance of maintenance or recovery of the nervous function within the organism own forces. The presence of heterogenic population of stem (SCs) and progenitor cells in various regions of the retina and brain is well documented. It has been also proved an association between SC mobilization from bone marrow into peripheral blood with nervous tissue injury. The recent discoveries indicating presence of very small embryonic-like, pluripotential stem cells within retina and mobilized peripheral blood suggest theoretically unlimited potential for self-repair of the organism. Nevertheless, the mechanism of SC action in the process of self regeneration of damaged nervous tissue remains to be elucidated.

Key words: central nervous system, stem cells, regeneration, retina, bone marrow, VSEL SCs.

Summary: Progress of research on stem cells including their genetic modifications allowed to obtain very promising results. However, one of the main barriers that (ex vivo) therapy has been encountered was delivery of genetic construct to the target cell. As well plasmids (delivered with using physico-chemical methods) as viral vectors have its prominent place at preparations aimed to implant genetically modified stem cells (auto- or allogenic origin) to recipient organ. Genetic modifications involve therapeutic, reporter or regulatory genes which may help to implantation of stem cells itself, to limit inflammatory process or apoptosis, to enhance physiological function of damaged organs/tissues. This review presents a progress in invention of novel genetic constructs and indicates main trends for application of genetically modified stem cells as therapeutic agents for either congenital or acquired pathologies.