Characteristics:
Human MSCs (hMSCs) are typically isolated from
the mononuclear layer of the bone marrow after
separation by density gradient centrifugation. The
mononuclear cells are cultured in medium with 10%
fetal calf serum, and the MSCs adhere to the tissue
culture plastic. Some hematopoietic cells also
adhere, but over time in culture these are washed
away, leaving adherent, fibroblast-like cells. After an
initial lag phase, the cells divide rapidly, with
population doubling time depending on the donor
and the initial plating density.
Human MSCs (hMSCs) are typically isolated from
the mononuclear layer of the bone marrow after
separation by density gradient centrifugation. The
mononuclear cells are cultured in medium with 10%
fetal calf serum, and the MSCs adhere to the tissue
culture plastic. Some hematopoietic cells also
adhere, but over time in culture these are washed
away, leaving adherent, fibroblast-like cells. After an
initial lag phase, the cells divide rapidly, with
population doubling time depending on the donor
and the initial plating density.
MSC-like cells have been isolated from pathological tissues such as the
rheumatoid arthritic joint, and these cells express bone morphogenetic
protein receptors. Indeed, it has been suggested that cells with mesenchymal
stem characteristics reside in virtually all postnatal organs and tissues. MSCs
have been isolated and cultured from many other species including mice,
rats, cats, dogs, rabbits, pigs, and baboons, albeit with varying success, as it
can be difficult to remove contaminating hematopoietic cells from species
such as mice. Nevertheless, enrichment for some species' MSCs can be
achieved by expansion and passaging in deprivational medium to eliminate
contamination. The resulting cultures are still morphologically heterogeneous,
containing cells ranging from narrow spindle-shaped cells to large polygonal
cells and, in confluent cultures, some slightly cuboidal cells.
Phenotypically, MSCs express a number of markers, none of which,
unfortunately, are specific to MSCs. It is generally agreed that adult human
MSCs do not express the hematopoietic markers CD45, CD34, CD14, or
CD11. They also do not express the costimulatory molecules CD80, CD86, or
CD40 or the adhesion molecules CD31 (platelet/endothelial cell adhesion
molecule [PECAM]-1), CD18 (leukocyte function-associated antigen-1 [LFA-
1]), or CD56 (neuronal cell adhesion molecule-1), but they can express
CD105 (SH2), CD73 (SH3/4), CD44, CD90 (Thy-1), CD71, and Stro-1 as well
as the adhesion molecules CD106 (vascular cell adhesion molecule [VCAM]
-1), CD166 (activated leukocyte cell adhesion molecule [ALCAM]),
intercellular adhesion molecule (ICAM)-1, and CD29.
There are several reports that describe the isolation of both human and
rodent MSCs using antibody selection based on the phenotype of MSCs.
Some have used a method of negative selection to enrich for MSCs, whereby
cells from the hematopoietic lineage are removed; others have used
antibodies to positively select for MSCs.MSCs from other species do not
express all the same molecules as those on human cells; for example,
although human and rat MSCs have been shown to be CD34, some papers
report variable expression of CD34 on murine MSCs [22]. It is generally
accepted that all MSCs are devoid of the hematopoietic marker CD45 and the
endothelial cell marker CD31. However, it is important to note that
differences in cell surface expression of many markers may be influenced by
factors secreted by accessory cells in the initial passages, and the in vitro
expression of some markers by MSCs does not always correlate with their
expression patterns in vivo.
There is also variable expression of many of the markers mentioned due to
variation in tissue source, the method of isolation and culture, and species
differences. For example, human adipose tissue is a source of multipotent
stem cells called processed lipoaspirate (PLA) cells which, like bone marrow
MSCs, can differentiate down several mesenchymal lineages in vitro.
However, there are some differences in the expressions of particular
markers: CD49d is expressed on PLA cells but not MSCs, and CD106 is
expressed on MSCs but not PLA cells. CD106 on MSCs in bone marrow has
been functionally associated with hematopoiesis, so the lack of CD106
expression on PLA cells is consistent with localization of these cells to a
nonhematopoietic tissue.Blood-derived mesenchymal precursor cells (BMPCs)
have also been described in the blood of normal individuals, and these
express many of the same markers as bone marrow MSCs, as well as
differentiating down the osteoblastic and adipogenic lineages. However, these
appear to be a separate population from fibrocytes, which are mesenchymal
precursor cells that circulate in the blood and can migrate into tissues.
Fibrocytes express CD34 and CD45 and appear to differentiate into
myofibroblasts, whereas BMPCs are reported to be CD34 negative.
Mesenchymal stem cells have also been isolated from human first- and
second-trimester fetal blood, liver, spleen, and bone marrow. Although
phenotypically similar, these culture-expanded MSCs exhibited heterogeneity
in differentiation potential, which related to the tissue source. Taken together,
these examples illustrate that mesenchymal precursor cells are
phenotypically heterogeneous, and the relationship between traditional bone
marrow-derived MSCs and these other MSC-like populations remains to be
fully clarified. Adult human MSCs are reported to express intermediate levels
of major histocompatibility complex (MHC) class I but do not express human
leukocyte antigen (HLA) class II antigens on the cell surface. The expression
of HLA class I on fetal hMSCs is lower [28]. Le Blanc and colleagues did
detect HLA class II by Western blot on lysates of unstimulated adult hMSCs,
suggesting intracellular deposits of the antigen [18], and found that cell-
surface expression can be induced by treatment of the cells with interferon-
{gamma} for 1 or 2 days. Unlike adult hMSCs, human fetal liver-derived
hMSCs have no MHC class II intracellularly or on the cell surface, suggesting
that MHC antigen expression by hMSCs changes from fetal to adult life.
References:
More References...
rheumatoid arthritic joint, and these cells express bone morphogenetic
protein receptors. Indeed, it has been suggested that cells with mesenchymal
stem characteristics reside in virtually all postnatal organs and tissues. MSCs
have been isolated and cultured from many other species including mice,
rats, cats, dogs, rabbits, pigs, and baboons, albeit with varying success, as it
can be difficult to remove contaminating hematopoietic cells from species
such as mice. Nevertheless, enrichment for some species' MSCs can be
achieved by expansion and passaging in deprivational medium to eliminate
contamination. The resulting cultures are still morphologically heterogeneous,
containing cells ranging from narrow spindle-shaped cells to large polygonal
cells and, in confluent cultures, some slightly cuboidal cells.
Phenotypically, MSCs express a number of markers, none of which,
unfortunately, are specific to MSCs. It is generally agreed that adult human
MSCs do not express the hematopoietic markers CD45, CD34, CD14, or
CD11. They also do not express the costimulatory molecules CD80, CD86, or
CD40 or the adhesion molecules CD31 (platelet/endothelial cell adhesion
molecule [PECAM]-1), CD18 (leukocyte function-associated antigen-1 [LFA-
1]), or CD56 (neuronal cell adhesion molecule-1), but they can express
CD105 (SH2), CD73 (SH3/4), CD44, CD90 (Thy-1), CD71, and Stro-1 as well
as the adhesion molecules CD106 (vascular cell adhesion molecule [VCAM]
-1), CD166 (activated leukocyte cell adhesion molecule [ALCAM]),
intercellular adhesion molecule (ICAM)-1, and CD29.
There are several reports that describe the isolation of both human and
rodent MSCs using antibody selection based on the phenotype of MSCs.
Some have used a method of negative selection to enrich for MSCs, whereby
cells from the hematopoietic lineage are removed; others have used
antibodies to positively select for MSCs.MSCs from other species do not
express all the same molecules as those on human cells; for example,
although human and rat MSCs have been shown to be CD34, some papers
report variable expression of CD34 on murine MSCs [22]. It is generally
accepted that all MSCs are devoid of the hematopoietic marker CD45 and the
endothelial cell marker CD31. However, it is important to note that
differences in cell surface expression of many markers may be influenced by
factors secreted by accessory cells in the initial passages, and the in vitro
expression of some markers by MSCs does not always correlate with their
expression patterns in vivo.
There is also variable expression of many of the markers mentioned due to
variation in tissue source, the method of isolation and culture, and species
differences. For example, human adipose tissue is a source of multipotent
stem cells called processed lipoaspirate (PLA) cells which, like bone marrow
MSCs, can differentiate down several mesenchymal lineages in vitro.
However, there are some differences in the expressions of particular
markers: CD49d is expressed on PLA cells but not MSCs, and CD106 is
expressed on MSCs but not PLA cells. CD106 on MSCs in bone marrow has
been functionally associated with hematopoiesis, so the lack of CD106
expression on PLA cells is consistent with localization of these cells to a
nonhematopoietic tissue.Blood-derived mesenchymal precursor cells (BMPCs)
have also been described in the blood of normal individuals, and these
express many of the same markers as bone marrow MSCs, as well as
differentiating down the osteoblastic and adipogenic lineages. However, these
appear to be a separate population from fibrocytes, which are mesenchymal
precursor cells that circulate in the blood and can migrate into tissues.
Fibrocytes express CD34 and CD45 and appear to differentiate into
myofibroblasts, whereas BMPCs are reported to be CD34 negative.
Mesenchymal stem cells have also been isolated from human first- and
second-trimester fetal blood, liver, spleen, and bone marrow. Although
phenotypically similar, these culture-expanded MSCs exhibited heterogeneity
in differentiation potential, which related to the tissue source. Taken together,
these examples illustrate that mesenchymal precursor cells are
phenotypically heterogeneous, and the relationship between traditional bone
marrow-derived MSCs and these other MSC-like populations remains to be
fully clarified. Adult human MSCs are reported to express intermediate levels
of major histocompatibility complex (MHC) class I but do not express human
leukocyte antigen (HLA) class II antigens on the cell surface. The expression
of HLA class I on fetal hMSCs is lower [28]. Le Blanc and colleagues did
detect HLA class II by Western blot on lysates of unstimulated adult hMSCs,
suggesting intracellular deposits of the antigen [18], and found that cell-
surface expression can be induced by treatment of the cells with interferon-
{gamma} for 1 or 2 days. Unlike adult hMSCs, human fetal liver-derived
hMSCs have no MHC class II intracellularly or on the cell surface, suggesting
that MHC antigen expression by hMSCs changes from fetal to adult life.
References:
- Colter DC, Class R, DiGirolamo CM et al. Rapid expansion of recycling
stem cells in cultures of plastic-adherent cells from human bone
marrow. Proc Natl Acad Sci U S A 2000;97:3213–3218.
- Campagnoli C, Roberts IA, Kumar S et al. Identification of
mesenchymal stem/progenitor cells in human first-trimester fetal
blood, liver, and bone marrow. Blood 2001;98:2396–2402.
- In't Anker PS, Scherjon SA, Kleijburg-van der Keur C et al. Amniotic
fluid as a novel source of mesenchymal stem cells for therapeutic
transplantation. Blood 2003;102:1548–1549.
- Nakahara H, Dennis JE, Bruder SP et al. In vitro differentiation of bone
and hypertrophic cartilage from periosteal-derived cells. Exp Cell Res
1991;195:492–503.
More References...
Fluorescent murine mesenchymal stem cells in
culture transfected (modified). [Courtesy of
Neurosciences, University of Genoa, Italy.]
culture transfected (modified). [Courtesy of
Neurosciences, University of Genoa, Italy.]
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