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Cell Injury-Induced Release of Fibroblast Growth Factor 2: Relevance to Intracerebral Mesenchymal Stromal Cell Transplantations.

Aizman I, Vinodkumar D, McGrogan M, Bates D - Stem Cells Dev. (2015)

Bottom Line: We found that fibroblast growth factor (FGF)2 and FGF1, but not vascular endothelial growth factor and monocyte chemoattractant protein 1 levels were high in extracts despite being low in conditioned media.However, suspensions of killed cells strongly potentiated the astrogenic effects of live MSC.We therefore propose that both dying and surviving grafted MSC contribute to tissue regeneration.

View Article: PubMed Central - PubMed

Affiliation: 1 Department of Research, SanBio, Inc. , Mountain View, California.

ABSTRACT
Beneficial effects of intracerebral transplantation of mesenchymal stromal cells (MSC) and their derivatives are believed to be mediated mostly by factors produced by engrafted cells. However, the mesenchymal cell engraftment rate is low, and the majority of grafted cells disappear within a short post-transplantation period. Here, we hypothesize that dying transplanted cells can affect surrounding tissues by releasing their active intracellular components. To elucidate the type, amounts, and potency of these putative intracellular factors, freeze/thaw extracts of MSC or their derivatives were tested in enzyme-linked immunosorbent assays and bioassays. We found that fibroblast growth factor (FGF)2 and FGF1, but not vascular endothelial growth factor and monocyte chemoattractant protein 1 levels were high in extracts despite being low in conditioned media. Extracts induced concentration-dependent proliferation of rat cortical neural progenitor cells and human umbilical vein endothelial cells; these proliferative responses were specifically blocked by FGF2-neutralizing antibody. In the neuropoiesis assay with rat cortical cells, both MSC extracts and killed cells induced expression of nestin, but not astrocyte differentiation. However, suspensions of killed cells strongly potentiated the astrogenic effects of live MSC. In transplantation-relevant MSC injury models (peripheral blood cell-mediated cytotoxicity and high cell density plating), MSC death coincided with the release of intracellular FGF2. The data showed that MSC contain a major depot of active FGF2 that is released upon cell injury and is capable of acutely stimulating neuropoiesis and angiogenesis. We therefore propose that both dying and surviving grafted MSC contribute to tissue regeneration.

No MeSH data available.


Related in: MedlinePlus

Proliferative response of cortical Nestin-positive cells to extracts and the role of FGF2. (A) Proliferation in response to various concentrations of extract (E0) and CM from MSC. (B) Effect of FGF2 neutralization on cell proliferation stimulated by SB623-derived E0 at either 15% or 5% (equivalent to 0.2 and 0.07 ng/mL FGF2, respectively). *P<0.05; **P<0.01. Neutralizing anti-FGF2 (clone bFM1) or isotype control (mouse IgG1) were added at 2 μg/mL. (C, D) Immunostaining of rat cortical cells stimulated or not with 20% SB623-derived extract. After 7 days, cells cultures were labeled with 5-bromo-2-deoxyuridine (BRDU) and stained for either BRDU and Doublecortin (Dcx) (C) or for BRDU and Nestin (Nes) (D). We concluded that cells predominantly labeled with BRDU and increased in numbers in extract-treated cultures were nestin-positive cells.
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f2: Proliferative response of cortical Nestin-positive cells to extracts and the role of FGF2. (A) Proliferation in response to various concentrations of extract (E0) and CM from MSC. (B) Effect of FGF2 neutralization on cell proliferation stimulated by SB623-derived E0 at either 15% or 5% (equivalent to 0.2 and 0.07 ng/mL FGF2, respectively). *P<0.05; **P<0.01. Neutralizing anti-FGF2 (clone bFM1) or isotype control (mouse IgG1) were added at 2 μg/mL. (C, D) Immunostaining of rat cortical cells stimulated or not with 20% SB623-derived extract. After 7 days, cells cultures were labeled with 5-bromo-2-deoxyuridine (BRDU) and stained for either BRDU and Doublecortin (Dcx) (C) or for BRDU and Nestin (Nes) (D). We concluded that cells predominantly labeled with BRDU and increased in numbers in extract-treated cultures were nestin-positive cells.

Mentions: Rat embryonic cortical cell populations contain a large proportion of NPC that proliferate in response to FGF2. Therefore, we stimulated rat cortical cells with dilutions of MSC-derived E0 and CM varying from 0% to 75% and conducted proliferation assays using BRDU incorporation (Fig. 2A). The proliferation was increased by E0, but not by CM in a concentration-dependent manner. The proliferative response to E0 was diminished in the presence of the neutralizing anti-FGF2 (bFM1) mAb, while the control antibody had no effect (Fig. 2B).


Cell Injury-Induced Release of Fibroblast Growth Factor 2: Relevance to Intracerebral Mesenchymal Stromal Cell Transplantations.

Aizman I, Vinodkumar D, McGrogan M, Bates D - Stem Cells Dev. (2015)

Proliferative response of cortical Nestin-positive cells to extracts and the role of FGF2. (A) Proliferation in response to various concentrations of extract (E0) and CM from MSC. (B) Effect of FGF2 neutralization on cell proliferation stimulated by SB623-derived E0 at either 15% or 5% (equivalent to 0.2 and 0.07 ng/mL FGF2, respectively). *P<0.05; **P<0.01. Neutralizing anti-FGF2 (clone bFM1) or isotype control (mouse IgG1) were added at 2 μg/mL. (C, D) Immunostaining of rat cortical cells stimulated or not with 20% SB623-derived extract. After 7 days, cells cultures were labeled with 5-bromo-2-deoxyuridine (BRDU) and stained for either BRDU and Doublecortin (Dcx) (C) or for BRDU and Nestin (Nes) (D). We concluded that cells predominantly labeled with BRDU and increased in numbers in extract-treated cultures were nestin-positive cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4499789&req=5

f2: Proliferative response of cortical Nestin-positive cells to extracts and the role of FGF2. (A) Proliferation in response to various concentrations of extract (E0) and CM from MSC. (B) Effect of FGF2 neutralization on cell proliferation stimulated by SB623-derived E0 at either 15% or 5% (equivalent to 0.2 and 0.07 ng/mL FGF2, respectively). *P<0.05; **P<0.01. Neutralizing anti-FGF2 (clone bFM1) or isotype control (mouse IgG1) were added at 2 μg/mL. (C, D) Immunostaining of rat cortical cells stimulated or not with 20% SB623-derived extract. After 7 days, cells cultures were labeled with 5-bromo-2-deoxyuridine (BRDU) and stained for either BRDU and Doublecortin (Dcx) (C) or for BRDU and Nestin (Nes) (D). We concluded that cells predominantly labeled with BRDU and increased in numbers in extract-treated cultures were nestin-positive cells.
Mentions: Rat embryonic cortical cell populations contain a large proportion of NPC that proliferate in response to FGF2. Therefore, we stimulated rat cortical cells with dilutions of MSC-derived E0 and CM varying from 0% to 75% and conducted proliferation assays using BRDU incorporation (Fig. 2A). The proliferation was increased by E0, but not by CM in a concentration-dependent manner. The proliferative response to E0 was diminished in the presence of the neutralizing anti-FGF2 (bFM1) mAb, while the control antibody had no effect (Fig. 2B).

Bottom Line: We found that fibroblast growth factor (FGF)2 and FGF1, but not vascular endothelial growth factor and monocyte chemoattractant protein 1 levels were high in extracts despite being low in conditioned media.However, suspensions of killed cells strongly potentiated the astrogenic effects of live MSC.We therefore propose that both dying and surviving grafted MSC contribute to tissue regeneration.

View Article: PubMed Central - PubMed

Affiliation: 1 Department of Research, SanBio, Inc. , Mountain View, California.

ABSTRACT
Beneficial effects of intracerebral transplantation of mesenchymal stromal cells (MSC) and their derivatives are believed to be mediated mostly by factors produced by engrafted cells. However, the mesenchymal cell engraftment rate is low, and the majority of grafted cells disappear within a short post-transplantation period. Here, we hypothesize that dying transplanted cells can affect surrounding tissues by releasing their active intracellular components. To elucidate the type, amounts, and potency of these putative intracellular factors, freeze/thaw extracts of MSC or their derivatives were tested in enzyme-linked immunosorbent assays and bioassays. We found that fibroblast growth factor (FGF)2 and FGF1, but not vascular endothelial growth factor and monocyte chemoattractant protein 1 levels were high in extracts despite being low in conditioned media. Extracts induced concentration-dependent proliferation of rat cortical neural progenitor cells and human umbilical vein endothelial cells; these proliferative responses were specifically blocked by FGF2-neutralizing antibody. In the neuropoiesis assay with rat cortical cells, both MSC extracts and killed cells induced expression of nestin, but not astrocyte differentiation. However, suspensions of killed cells strongly potentiated the astrogenic effects of live MSC. In transplantation-relevant MSC injury models (peripheral blood cell-mediated cytotoxicity and high cell density plating), MSC death coincided with the release of intracellular FGF2. The data showed that MSC contain a major depot of active FGF2 that is released upon cell injury and is capable of acutely stimulating neuropoiesis and angiogenesis. We therefore propose that both dying and surviving grafted MSC contribute to tissue regeneration.

No MeSH data available.


Related in: MedlinePlus