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Differential mechanical response of mesenchymal stem cells and fibroblasts to tumor-secreted soluble factors.

McGrail DJ, Ghosh D, Quach ND, Dawson MR - PLoS ONE (2012)

Bottom Line: In addition, unlike fibroblasts, MSCs exposed to tumor-secreted soluble factors display a functionally different phenotype characterized by morphological elongation, decreased actin stress fiber density, and decreased adhesion.Quantitative real-time PCR indicates these phenomena occur based on differential expression of small GTPases RhoA and Cdc42, but not Rac1.These findings demonstrate a fundamental difference in the recruitment of fibroblasts and MSCs.

View Article: PubMed Central - PubMed

Affiliation: School of Chemical & Biomolecular Engineering, Georgia Institute of Technology, Atlanta, Georgia, United States of America.

ABSTRACT
The progression of neoplastic malignancies is a complex process resulting not only from the accumulation of mutations within tumor cells, but also modulation of the tumor microenvironment. Recent advances have shown that the recruitment and subsequent heterotypic interactions of stromal cells--including fibroblasts and bone marrow-derived mesenchymal stem cells (MSCs)--are crucial for carcinogenesis. Though extensive work has been done analyzing the signals that recruit these cells, the governing mechanical properties have not been fully investigated. Here, we report that despite their initial similarities, MSCs respond not only faster but also more dramatically to pro-migratory tumor-secreted soluble factors. Utilizing multiple particle tracking microrheology to probe the cytoskeletal mechanical properties, we show that MSCs stiffen completely within one hour, three times faster than fibroblasts. In addition, unlike fibroblasts, MSCs exposed to tumor-secreted soluble factors display a functionally different phenotype characterized by morphological elongation, decreased actin stress fiber density, and decreased adhesion. Quantitative real-time PCR indicates these phenomena occur based on differential expression of small GTPases RhoA and Cdc42, but not Rac1. These findings demonstrate a fundamental difference in the recruitment of fibroblasts and MSCs.

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Tumor-secreted soluble factors alter MSC morphology.Brightfield images of murine MSCs (isolated from Balb/C mouse bone marrow; A–B), Swiss 3T3 fibroblasts (C–D), and primary fibroblasts (isolated from Balb/C mouse kidney; E–F) incubated for 24 hours in control media (CM, top) or tumor cell-conditioned media (TCM, bottom), then fixed in methanol, and stained with crystal violet. MSCs elongated dramatically in response to TCM (A–B); whereas, immortalized (C–D) and primary (E–F) fibroblasts did not respond to TCM treatment. A ‘sandwich’ co-culture model was used to study the migratory behavior of CM-DiL-labeled MSCs through a layer of polymerized collagen toward a monolayer of MSCs (G) or 4T1 tumor cells (H). Interestingly, MSCs elongated toward the tumor cells (H) but not toward the MSCs. (scale bar = 100 µm)
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pone-0033248-g002: Tumor-secreted soluble factors alter MSC morphology.Brightfield images of murine MSCs (isolated from Balb/C mouse bone marrow; A–B), Swiss 3T3 fibroblasts (C–D), and primary fibroblasts (isolated from Balb/C mouse kidney; E–F) incubated for 24 hours in control media (CM, top) or tumor cell-conditioned media (TCM, bottom), then fixed in methanol, and stained with crystal violet. MSCs elongated dramatically in response to TCM (A–B); whereas, immortalized (C–D) and primary (E–F) fibroblasts did not respond to TCM treatment. A ‘sandwich’ co-culture model was used to study the migratory behavior of CM-DiL-labeled MSCs through a layer of polymerized collagen toward a monolayer of MSCs (G) or 4T1 tumor cells (H). Interestingly, MSCs elongated toward the tumor cells (H) but not toward the MSCs. (scale bar = 100 µm)

Mentions: To investigate the response of MSCs to tumor cell-conditioned media (TCM), cells were incubated in serum-free TCM or control media (CM) for 24 hours. MSCs underwent a morphological change from a cobblestone appearance to an elongated phenotype (Fig. 2A–B). In comparison, Swiss 3T3 fibroblasts (Fig. 2C–D) and primary isolated kidney fibroblasts (Fig. 2E–F) had similar initial morphologies, but did not undergo a morphological change upon treatment with TCM. A 3D sandwich culture model was used to determine if tumor cells could stimulate elongation through a viscoelastic collagen gel. Moreover, MSCs elongated through the collagen gel toward a monolayer of 4T1 tumor cells (Fig. 2H) but did not elongate towards a monolayer of MSCs (Fig. 2G).


Differential mechanical response of mesenchymal stem cells and fibroblasts to tumor-secreted soluble factors.

McGrail DJ, Ghosh D, Quach ND, Dawson MR - PLoS ONE (2012)

Tumor-secreted soluble factors alter MSC morphology.Brightfield images of murine MSCs (isolated from Balb/C mouse bone marrow; A–B), Swiss 3T3 fibroblasts (C–D), and primary fibroblasts (isolated from Balb/C mouse kidney; E–F) incubated for 24 hours in control media (CM, top) or tumor cell-conditioned media (TCM, bottom), then fixed in methanol, and stained with crystal violet. MSCs elongated dramatically in response to TCM (A–B); whereas, immortalized (C–D) and primary (E–F) fibroblasts did not respond to TCM treatment. A ‘sandwich’ co-culture model was used to study the migratory behavior of CM-DiL-labeled MSCs through a layer of polymerized collagen toward a monolayer of MSCs (G) or 4T1 tumor cells (H). Interestingly, MSCs elongated toward the tumor cells (H) but not toward the MSCs. (scale bar = 100 µm)
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3306382&req=5

pone-0033248-g002: Tumor-secreted soluble factors alter MSC morphology.Brightfield images of murine MSCs (isolated from Balb/C mouse bone marrow; A–B), Swiss 3T3 fibroblasts (C–D), and primary fibroblasts (isolated from Balb/C mouse kidney; E–F) incubated for 24 hours in control media (CM, top) or tumor cell-conditioned media (TCM, bottom), then fixed in methanol, and stained with crystal violet. MSCs elongated dramatically in response to TCM (A–B); whereas, immortalized (C–D) and primary (E–F) fibroblasts did not respond to TCM treatment. A ‘sandwich’ co-culture model was used to study the migratory behavior of CM-DiL-labeled MSCs through a layer of polymerized collagen toward a monolayer of MSCs (G) or 4T1 tumor cells (H). Interestingly, MSCs elongated toward the tumor cells (H) but not toward the MSCs. (scale bar = 100 µm)
Mentions: To investigate the response of MSCs to tumor cell-conditioned media (TCM), cells were incubated in serum-free TCM or control media (CM) for 24 hours. MSCs underwent a morphological change from a cobblestone appearance to an elongated phenotype (Fig. 2A–B). In comparison, Swiss 3T3 fibroblasts (Fig. 2C–D) and primary isolated kidney fibroblasts (Fig. 2E–F) had similar initial morphologies, but did not undergo a morphological change upon treatment with TCM. A 3D sandwich culture model was used to determine if tumor cells could stimulate elongation through a viscoelastic collagen gel. Moreover, MSCs elongated through the collagen gel toward a monolayer of 4T1 tumor cells (Fig. 2H) but did not elongate towards a monolayer of MSCs (Fig. 2G).

Bottom Line: In addition, unlike fibroblasts, MSCs exposed to tumor-secreted soluble factors display a functionally different phenotype characterized by morphological elongation, decreased actin stress fiber density, and decreased adhesion.Quantitative real-time PCR indicates these phenomena occur based on differential expression of small GTPases RhoA and Cdc42, but not Rac1.These findings demonstrate a fundamental difference in the recruitment of fibroblasts and MSCs.

View Article: PubMed Central - PubMed

Affiliation: School of Chemical & Biomolecular Engineering, Georgia Institute of Technology, Atlanta, Georgia, United States of America.

ABSTRACT
The progression of neoplastic malignancies is a complex process resulting not only from the accumulation of mutations within tumor cells, but also modulation of the tumor microenvironment. Recent advances have shown that the recruitment and subsequent heterotypic interactions of stromal cells--including fibroblasts and bone marrow-derived mesenchymal stem cells (MSCs)--are crucial for carcinogenesis. Though extensive work has been done analyzing the signals that recruit these cells, the governing mechanical properties have not been fully investigated. Here, we report that despite their initial similarities, MSCs respond not only faster but also more dramatically to pro-migratory tumor-secreted soluble factors. Utilizing multiple particle tracking microrheology to probe the cytoskeletal mechanical properties, we show that MSCs stiffen completely within one hour, three times faster than fibroblasts. In addition, unlike fibroblasts, MSCs exposed to tumor-secreted soluble factors display a functionally different phenotype characterized by morphological elongation, decreased actin stress fiber density, and decreased adhesion. Quantitative real-time PCR indicates these phenomena occur based on differential expression of small GTPases RhoA and Cdc42, but not Rac1. These findings demonstrate a fundamental difference in the recruitment of fibroblasts and MSCs.

Show MeSH
Related in: MedlinePlus