Mimicking Paracrine TGFβ1 Signals during Myofibroblast Differentiation in 3D Collagen Networks

Ansorge, Michael; Sapudom, Jiranuwat; Chkolnikov, Marina; Martin, W.D.; Anderegg, Ulf; Möller, Stephanie; Schnabelrauch, Matthias; Pompe, Tilo

doi:10.1038/s41598-017-05912-x

Cited by 21 publications

(23 citation statements)

References 32 publications

(56 reference statements)

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“…Interestingly, we found that a low concentration of TGF-β1 (300 pg/mL) in the co-culture is needed to trigger myofibroblast differentiation, when compared to the systematic addition of 10 ng/mL TGF-β1. This result is in line with the previous report where slow and sustained TGF-β1 release in a picogram range from polyethylene glycol (PEG) microbeads embedded in 3D collagen matrix could fully differentiate fibroblasts into myofibroblast [73]. We also demonstrated that fibril density could influence myofibroblast differentiation as shown by matrix dependent expression of aSMA and matrix protein secretion (Coll1a1 and EDA-FN) in mono-culture and in co-culture with M -IL-4/-IL-13 .…”

Section: Il-4/il-13 Triggers Fibroblast Differentiation Into Myofibsupporting

confidence: 93%

Collagen Fibril Density Modulates Macrophage Activation and Cellular Functions during Tissue Repair

Sapudom¹,

Mohamed

Garcia-Sabaté³

et al. 2020

Bioengineering

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Monocytes circulate in the bloodstream, extravasate into the tissue and differentiate into specific macrophage phenotypes to fulfill the immunological needs of tissues. During the tissue repair process, tissue density transits from loose to dense tissue. However, little is known on how changes in tissue density affects macrophage activation and their cellular functions. In this work, monocytic cell line THP-1 cells were embedded in three-dimensional (3D) collagen matrices with different fibril density and were then differentiated into uncommitted macrophages (MPMA) using phorbol-12-myristate-13-acetate (PMA). MPMA macrophages were subsequently activated into pro-inflammatory macrophages (MLPS/IFNγ) and anti-inflammatory macrophages (MIL-4/IL-13) using lipopolysaccharide and interferon-gamma (IFNγ), and interleukin 4 (IL-4) and IL-13, respectively. Although analysis of cell surface markers, on both gene and protein levels, was inconclusive, cytokine secretion profiles, however, demonstrated differences in macrophage phenotype. In the presence of differentiation activators, MLPS/IFNγ secreted high amounts of IL-1β and tumor necrosis factor alpha (TNFα), while M0PMA secreted similar cytokines to MIL-4/IL-13, but low IL-8. After removing the activators and further culture for 3 days in fresh cell culture media, the secretion of IL-6 was found in high concentrations by MIL-4/IL-13, followed by MLPS/IFNγ and MPMA. Interestingly, the secretion of cytokines is enhanced with an increase of fibril density. Through the investigation of macrophage-associated functions during tissue repair, we demonstrated that M1LPS/IFNγ has the potential to enhance monocyte infiltration into tissue, while MIL-4/IL-13 supported fibroblast differentiation into myofibroblasts via transforming growth factor beta 1 (TGF-β1) in dependence of fibril density, suggesting a M2a-like phenotype. Overall, our results suggest that collagen fibril density can modulate macrophage response to favor tissue functions. Understanding of immune response in such complex 3D microenvironments will contribute to the novel therapeutic strategies for improving tissue repair, as well as guidance of the design of immune-modulated materials.

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Section: Il-4/il-13 Triggers Fibroblast Differentiation Into Myofibsupporting

confidence: 93%

Collagen Fibril Density Modulates Macrophage Activation and Cellular Functions during Tissue Repair

Sapudom¹,

Mohamed

Garcia-Sabaté³

et al. 2020

Bioengineering

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“…Depending on the scientific question, various cell characteristics can be further analyzed manually or using an external analysis software from the obtained single cell trajectories including genealogies of dividing cells 36 , and cell behavior in complex bioengineered 3D systems e.g. biomimetic matrices with defined biophysical properties 8 or functionalized with other ECM components 39 , 40 , short-range cytokine gradients 38 , 41 and tissue boundaries 9 allowing for a better understanding of cell-cell-interactions and mimicking in vivo like microenvironments.…”

Section: Resultsmentioning

confidence: 99%

Quantitative label-free single cell tracking in 3D biomimetic matrices

Sapudom

Waschke

Franke

et al. 2017

Sci Rep

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Live cell imaging enables an observation of cell behavior over a period of time and is a growing field in modern cell biology. Quantitative analysis of the spatio-temporal dynamics of heterogeneous cell populations in three-dimensional (3D) microenvironments contributes a better understanding of cell-cell and cell-matrix interactions for many biomedical questions of physiological and pathological processes. However, current live cell imaging and analysis techniques are frequently limited by non-physiological 2D settings. Furthermore, they often rely on cell labelling by fluorescent dyes or expression of fluorescent proteins to enhance contrast of cells, which frequently affects cell viability and behavior of cells. In this work, we present a quantitative, label-free 3D single cell tracking technique using standard bright-field microscopy and affordable computational resources for data analysis. We demonstrate the efficacy of the automated method by studying migratory behavior of a large number of primary human macrophages over long time periods of several days in a biomimetic 3D microenvironment. The new technology provides a highly affordable platform for long-term studies of single cell behavior in 3D settings with minimal cell manipulation and can be implemented for various studies regarding cell-matrix interactions, cell-cell interactions as well as drug screening platform for primary and heterogeneous cell populations.

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“…Tumor cells invaded as groups of cells and were led by ASCs that stained positive for the myofibroblast marker α‐smooth muscle actin (α‐SMA). [ 15,31,35 ] Interestingly, co‐culture spheroids in which tumor cells were intermixed with ASCs isolated from human breast adipose tissue similarly promoted invasion (Figure 1f) suggesting that our data are of potential relevance to humans. Collectively, these data indicate that ASCs promote collective invasion of premalignant tumor cells via direct cell‐cell contact rather than secreted factors alone, and that myofibroblasts play a role in this process.…”

Section: Resultsmentioning

confidence: 76%

“…For example, cancer‐associated fibroblasts and thus, possibly obesity‐associated ASCs can promote tumor invasion via altered fibronectin deposition [ 12 ] and secretion of cytokines (including SDF‐1 and TGF‐ b 1). [ 31,34 ] Accordingly, our previous work suggests that obesity stimulates fibronectin deposition by ASCs [ 7 ] and that activated ASCs increase the secretion of factors contributing to elevated tumor malignancy. [ 40,43 ] Although secreted factors alone were not sufficient to promote extensive invasion in our experiments, cytokines can be sequestered within the ECM (e.g., due to differential binding to fibronectin).…”

Section: Discussionmentioning

confidence: 99%

Obesity‐Associated Adipose Stromal Cells Promote Breast Cancer Invasion through Direct Cell Contact and ECM Remodeling

Mulligan

Ouyang

et al. 2020

Adv Funct Materials

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Obesity increases the risk and worsens the prognosis for breast cancer due, in part, to altered adipose stromal cell (ASC) behavior. Whether ASCs from obese individuals increase migration of breast cancer cells relative to their lean counterparts, however, remains unclear. To test this connection, multicellular spheroids composed of MCF10A-derived tumor cell lines of varying malignant potential and lean or obese ASCs are embedded into collagen scaffolds mimicking the elastic moduli of interstitial breast adipose tissue. Confocal image analysis suggests that tumor cells alone migrate insignificantly under these conditions. However, direct cell-cell contact with either lean or obese ASCs enables them to migrate collectively, whereby obese ASCs activate tumor cell migration more effectively than their lean counterparts. Time-resolved optical coherence tomography imaging suggests that obese ASCs facilitate tumor cell migration by mediating contraction of local collagen fibers. Matrix metalloproteinase (MMP)-dependent proteolytic activity significantly contributes to ASC-mediated tumor cell invasion and collagen deformation. However, ASC contractility is also important, as co-inhibition of both MMPs and contractility is necessary to completely abrogate ASC-mediated tumor cell migration. These findings imply that obesity-mediated changes of ASC phenotype may impact tumor cell migration and invasion with potential implications for breast cancer malignancy in obese patients.

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Mimicking Paracrine TGFβ1 Signals during Myofibroblast Differentiation in 3D Collagen Networks

Cited by 21 publications

References 32 publications

Collagen Fibril Density Modulates Macrophage Activation and Cellular Functions during Tissue Repair

Collagen Fibril Density Modulates Macrophage Activation and Cellular Functions during Tissue Repair

Quantitative label-free single cell tracking in 3D biomimetic matrices

Obesity‐Associated Adipose Stromal Cells Promote Breast Cancer Invasion through Direct Cell Contact and ECM Remodeling

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