O. V. Payushina

Mesenchymal stem cells: Sources, phenotype, and differentiation potential

Mesenchymal stem cells present in the bone marrow and some other organs are primitive pluripotent precursors of osseous, cartilaginous, adipose, and other mesenchymal tissues. The recently revealed capacity of these cells for differentiation into nonmesenchymal derivatives is of considerable theoretical and practical interest. However, many aspects of the biology of these cells remain obscure despite active research. This review considers possible sources and methods for the isolation of mesenchymal stem cells, their potential for proliferation and differentiation in different directions, and outlooks of their therapeutic application. A model of parent-progeny relationships of stromal cells is proposed, and the problems of regulation of proliferation and differentiation of mesenchymal precursors as well as their role in the maintenance of regeneration and tissue functioning are discussed.

Comparative characterization of mesenchymal bone marrow stromal cells at early and late stages of culturing

The mesenchymal stromal cell is a multipotent precursor of osteoblasts, adipocytes, and some other cell types. In this study, a comparative analysis of cultured mesenchymal stromal cells from the rat bone marrow at the early and late stages of subculturing has been performed using molecular genetic and cytological methods. The culture has undergone 11 passages during 140 days. Upon long-term culturing, the mesenchymal stromal cells have proved to lose their potential for adipogenic differentiation but preserve the potential for osteogenesis. Morphological characters typical of osteogenic differentiation can be observed at the earlier stages of culturing (passages 1–4) but disappear at later stages (passages 9–11), despite mineralization of the extracellular matrix and the expression of osteogenic differentiation markers. A comparative analysis of the proliferation potential of stromal cells has shown that differences in the period of cell population doubling at the early and later stages of culturing are insignificant. An almost complete arrest of cell growth has been observed in the middle of the culture period (passages 5 and 6).

Comparative Investigation of Mesenchymal Stem Cells Isolated from the Bone Marrow and Fetal Liver of Mouse and Rat

We studied the properties of cells forming fibroblast colonies from the bone marrow and fetal liver of mouse and rat. Bone marrow and fetal liver cells formed colonies in vitro including fibroblasts as well as a considerable proportion of macrophages. The colonies formed from bone marrow and hepatic cells of rat differed from the murine ones by a higher proportion of fibroblasts. Most colonies derived from the bone marrow of both mouse and rat included a fraction of cells expressing alkaline phosphatase, and hence, capable of osteogenic differentiation; the colonies derived from the fetal liver included low proportions of such cells. The cell layers derived from the colony-forming fibroblasts of both bone marrow and fetal liver of mouse maintained hematopoiesis in the peritoneal cavity of irradiated mice, which indicated that these progenitor cells can form hematopoietic microenvironment.

Effects of growth factors on multipotent bone marrow mesenchymal stromal cells

Multipotent bone marrow mesenchymal stromal cells are progenitors of various cell types capable of long-term self-renewal. These cells are an adequate model for studying the most important problems in cell biology, such as self-renewal of stem cells and regulation of their differentiation. Moreover, these cells are a promising resource for regenerative medicine. In this context, isolation of the earliest multipotent mesenchymal stromal cells, their in vitro maintenance in an undifferentiated state, and stimulation of their differentiation in a desired direction appear to be most important. To successfully use the multipotent mesenchymal stromal cells both in fundamental studies and in therapy, it is necessary to modify and standardize the composition of culture medium, replacing blood serum with certain growth factors. These factors have influence on the proliferation and differentiation of most cell types, including multipotent mesenchymal stromal cells. This paper is a review of available data concerning the effects of some growth factors on the multipotent mesenchymal stromal cells of the bone marrow.

Analysis of sensitivity of stromal stem cells (CFU-f) from rat bone marrow and fetal liver to 5-fluorouracil

The sensitivity of stromal stem cells (CFU-f) from rat bone marrow and fetal liver to the cytotoxic effect of 5-fluorouracil (5-FU) was compared in vivo and in vitro. Cells from both tissues demonstrated a similar resistance to 5-FU in vitro; however, stromal stem cells from fetal liver proved notably more sensitive to 5-FU compared to marrow CFU-f in vivo. Cells forming colonies of different size were identified in stem cell populations from both tissues. Cells giving rise to small colonies had a higher resistance to 5-FU both in vivo and in vitro.

Clonal growth and differentiation of mesenchymal stromal cells from rat liver at different stages of embryogenesis

Fetal liver, during its hematopoietic activity, contains mesenchymal stromal cells (MSCs) generating its hematopoietic microenvironment. These cells are clonogenic and capable of multilineage differentiation; however, little is known about how their properties alter during embryogenesis. We compared the cloning efficiency of MSCs from rat fetal liver at 14, 16, and 20 days of development, as well as their capacity for osteo- and adipogenesis in vitro and chondrogenesis in vivo by ectopic transplantation of intact liver. The relative content of clonogenic MSCs in liver cell suspension was highest in 16-day fetuses and lowest in 20-day fetuses. Cells from 14-day fetuses exhibited high osteogenic and less apparent adipogenic and chondrogenic potential; cells from 20-day fetuses displayed weak adipogenic capacity and no osteo- or chondrogenic ability. These results show the correlation of MSC content and the cell differentiation potential with hematopoietic dynamics in developing rat liver. It may be thought that the changes we observed are related to the loss of hematopoietic activity and liver getting of definitive functions.

Spontaneous myogenesis in the primary culture of fetal rat liver

187 Fetal liver stroma is known as a source of mesenchymal stromal cells (MSCs), which exhibit osteogenic, adipogenic, and chondrogenic potencies [1]. It was reported that stromal cells of this organ can undergo myogenic differentiation in an induction medium [2], when cultured under specific conditions [3] and upon fusion with skeletal muscle cells [4]. We discovered that myosymplasts may spontaneously form in the primary culture of fetal rat liver.

Differentiation of Pluripotent Embryonic Stem Cells in the Peritoneal Cavity of Irradiated Mice

We studied the behavior and differentiation of pluripotent embryonic stem cells of R1 mice in vivo. Undifferentiated embryonic stem cells and differentiating embryoid bodies implanted in the abdominal cavity of irradiated mice were shown to form tumors containing the derivatives of all germ layers. Cells of the embryoid bodies form tumors two weeks after implantation, while undifferentiated embryonic stem cells form tumors only by week three.

Mesenchymal Stromal Cells of Rat Spleen during Pre-and Postnatal Ontogeny: Comparative Analysis of Clonal Growth, Phenotype and Differentiation Potencies

Comparison of mesenchymal stromal cells of embryonic and adult rat spleen showed that splenic cells from 20-day rat fetuses exhibit the capacity for clonal growth, express surface antigens CD73, CD90, and CD106, and have weak osteogenic and adipogenic potencies, while splenic cells from adult animals are characterized by lower cloning efficiency, rapid decrease of proliferative activity during passaging, the absence of CD73 and CD90 expression, and are incapable of osteogenesis The observed changes are probably related to extinction of myelopoiesis in the spleen during the postnatal ontogeny.

Cell Composition of the Primary Culture of Fetal Liver

Fetal liver is known as a source of multipotent mesenchymal stromal cells. These cells are routinely isolated by adhesion to plastic, but thus prepared culture is contaminated by other cells. For instance, primary cell culture of from rat fetal liver, apart from fibroblasts with phenotypic characteristics of mesenchymal stromal cells, contained skeletal muscle precursors, myofibroblasts, and epitheliocytes expressing cytokeratin-19 (the latter was also detected in some fibroblast-like cells probably undergoing epithelio-mesenchymal transition). During passaging, fibroblasts become practically the only cell type in the culture.