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High-throughput 3D cell invasion chip enables accurate cancer metastatic assays.

Zhang Y, Zhou L, Qin L - J. Am. Chem. Soc. (2014)

Bottom Line: Chemotaxis is the phenomenon by which the migration and invasion of cells is directed in response to an extracellular chemical gradient.Chemotaxis of tumor cells and tumor-associated inflammatory and stromal cells is mediated by chemokines, chemokine receptors, growth factors, and growth factor receptors.Additionally, this microdevice generates opposing gradients for two types of cells on the same chip, which builds a controlled system with sequentially changing components to study environmental effects from basal and immune cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Nanomedicine, Houston Methodist Research Institute , Houston, Texas 77030, United States.

ABSTRACT
Chemotaxis is the phenomenon by which the migration and invasion of cells is directed in response to an extracellular chemical gradient. Chemotaxis of tumor cells and tumor-associated inflammatory and stromal cells is mediated by chemokines, chemokine receptors, growth factors, and growth factor receptors. Current techniques used to study chemotactic driven cell invasion and metastasis utilize two-dimensional migration assays involving imaging and analyzing tumor cells on glass slides or plastic surfaces, which requires large numbers of cells and often lacks real-time monitoring of vertical cell movement and systematically controlled chemotactic gradients, leading to contradictory results compared to those from clinical investigations and animal models. We addressed such challenges by developing a high-throughput microdevice with 4000 ultraminiaturized wells to monitor real-time, three-dimensional cell invasion over a wide range of cell densities and also screen drugs that inhibit cell invasion and potentially prevent metastatic malignancy. Additionally, this microdevice generates opposing gradients for two types of cells on the same chip, which builds a controlled system with sequentially changing components to study environmental effects from basal and immune cells.

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High-densitycell invasion assay. (a) GFP image of invasive cellson the top layer of the microwells. (b) Image of a partial MI-Chiptaken by a confocal microscope shows the cells located on the toplayer of the microwells. (c) 3D bright-field (BF) image of invasivecells on the top layer of the microwells. Scale bar: 200 μm.(d) Representative image taken by an optical microscope shows cellslocated on the top and bottom layers of the microwells (each circlerepresents a cell). Scale bar: 100 μm (e). The fractions ofinvasive SUM-159 cells at at a high-cell density (H) at differenttime points. (f) Comparison of fractions of invasive SUM-159 and MCF-7cells at a gradient cell density after 96 h.
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fig4: High-densitycell invasion assay. (a) GFP image of invasive cellson the top layer of the microwells. (b) Image of a partial MI-Chiptaken by a confocal microscope shows the cells located on the toplayer of the microwells. (c) 3D bright-field (BF) image of invasivecells on the top layer of the microwells. Scale bar: 200 μm.(d) Representative image taken by an optical microscope shows cellslocated on the top and bottom layers of the microwells (each circlerepresents a cell). Scale bar: 100 μm (e). The fractions ofinvasive SUM-159 cells at at a high-cell density (H) at differenttime points. (f) Comparison of fractions of invasive SUM-159 and MCF-7cells at a gradient cell density after 96 h.

Mentions: An invasion assay usinga wide range of cell densities of SUM-159and MCF-7 was performed using the gradient cell seeding method (Figures 2a and S10). After cellloading, bright-field and fluorescent images of cells located on thetop layer of the microwells were recorded daily by adjusting the focalplane of the microscope (Figure 4a–d).Thefraction of invasive cells in each microwell was calculated by countingcell numbers in the top layer of each microwell and dividing themby numbers of initial cells in the microwell; the average value atthe same cell density was used to evaluate the invasive capacity ofthe cancer cell. For metastatic cancer cell line SUM-159, the fractionof invasive cells at a high cell density was greater than the fractionof invasive cells calculated at single-cell or low-cell density atall time points (Figure 4e). For example, after96 h, 34% of single SUM-159 cells had migrated to the top layer ofthe microwells, and 46.5% of SUM-159 high-density cells had movedtoward the FBS. As shown in Figure 4f, increasedcell density enhanced the invasion capacity of the cancer cell. Forthe nonmetastatic MCF-7 cell line, increased cell density did notaffect the fraction of invasive cells significantly. We also studiedthe impact of well shape to cell invasion by using two very differentshapes shown in our original chip, round and square wells. For singleor high-density cell invasion, our observed results showed very littledifference between the two shapes, completely within the error range.Confocal microscopy was used to image the morphology of invading cancercells at higher cell densities. In addition to the many cells alreadylocated in the top layer, additional cells had migrated more than100 μm. Most invasive cancer cells displayed elongated cellmorphology. We suggest that the invading metastatic cells act in acooperative manner using a form of autocrine and paracrine signaling,and increased cell density may contribute to tumor metastasis.7,22


High-throughput 3D cell invasion chip enables accurate cancer metastatic assays.

Zhang Y, Zhou L, Qin L - J. Am. Chem. Soc. (2014)

High-densitycell invasion assay. (a) GFP image of invasive cellson the top layer of the microwells. (b) Image of a partial MI-Chiptaken by a confocal microscope shows the cells located on the toplayer of the microwells. (c) 3D bright-field (BF) image of invasivecells on the top layer of the microwells. Scale bar: 200 μm.(d) Representative image taken by an optical microscope shows cellslocated on the top and bottom layers of the microwells (each circlerepresents a cell). Scale bar: 100 μm (e). The fractions ofinvasive SUM-159 cells at at a high-cell density (H) at differenttime points. (f) Comparison of fractions of invasive SUM-159 and MCF-7cells at a gradient cell density after 96 h.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4227729&req=5

fig4: High-densitycell invasion assay. (a) GFP image of invasive cellson the top layer of the microwells. (b) Image of a partial MI-Chiptaken by a confocal microscope shows the cells located on the toplayer of the microwells. (c) 3D bright-field (BF) image of invasivecells on the top layer of the microwells. Scale bar: 200 μm.(d) Representative image taken by an optical microscope shows cellslocated on the top and bottom layers of the microwells (each circlerepresents a cell). Scale bar: 100 μm (e). The fractions ofinvasive SUM-159 cells at at a high-cell density (H) at differenttime points. (f) Comparison of fractions of invasive SUM-159 and MCF-7cells at a gradient cell density after 96 h.
Mentions: An invasion assay usinga wide range of cell densities of SUM-159and MCF-7 was performed using the gradient cell seeding method (Figures 2a and S10). After cellloading, bright-field and fluorescent images of cells located on thetop layer of the microwells were recorded daily by adjusting the focalplane of the microscope (Figure 4a–d).Thefraction of invasive cells in each microwell was calculated by countingcell numbers in the top layer of each microwell and dividing themby numbers of initial cells in the microwell; the average value atthe same cell density was used to evaluate the invasive capacity ofthe cancer cell. For metastatic cancer cell line SUM-159, the fractionof invasive cells at a high cell density was greater than the fractionof invasive cells calculated at single-cell or low-cell density atall time points (Figure 4e). For example, after96 h, 34% of single SUM-159 cells had migrated to the top layer ofthe microwells, and 46.5% of SUM-159 high-density cells had movedtoward the FBS. As shown in Figure 4f, increasedcell density enhanced the invasion capacity of the cancer cell. Forthe nonmetastatic MCF-7 cell line, increased cell density did notaffect the fraction of invasive cells significantly. We also studiedthe impact of well shape to cell invasion by using two very differentshapes shown in our original chip, round and square wells. For singleor high-density cell invasion, our observed results showed very littledifference between the two shapes, completely within the error range.Confocal microscopy was used to image the morphology of invading cancercells at higher cell densities. In addition to the many cells alreadylocated in the top layer, additional cells had migrated more than100 μm. Most invasive cancer cells displayed elongated cellmorphology. We suggest that the invading metastatic cells act in acooperative manner using a form of autocrine and paracrine signaling,and increased cell density may contribute to tumor metastasis.7,22

Bottom Line: Chemotaxis is the phenomenon by which the migration and invasion of cells is directed in response to an extracellular chemical gradient.Chemotaxis of tumor cells and tumor-associated inflammatory and stromal cells is mediated by chemokines, chemokine receptors, growth factors, and growth factor receptors.Additionally, this microdevice generates opposing gradients for two types of cells on the same chip, which builds a controlled system with sequentially changing components to study environmental effects from basal and immune cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Nanomedicine, Houston Methodist Research Institute , Houston, Texas 77030, United States.

ABSTRACT
Chemotaxis is the phenomenon by which the migration and invasion of cells is directed in response to an extracellular chemical gradient. Chemotaxis of tumor cells and tumor-associated inflammatory and stromal cells is mediated by chemokines, chemokine receptors, growth factors, and growth factor receptors. Current techniques used to study chemotactic driven cell invasion and metastasis utilize two-dimensional migration assays involving imaging and analyzing tumor cells on glass slides or plastic surfaces, which requires large numbers of cells and often lacks real-time monitoring of vertical cell movement and systematically controlled chemotactic gradients, leading to contradictory results compared to those from clinical investigations and animal models. We addressed such challenges by developing a high-throughput microdevice with 4000 ultraminiaturized wells to monitor real-time, three-dimensional cell invasion over a wide range of cell densities and also screen drugs that inhibit cell invasion and potentially prevent metastatic malignancy. Additionally, this microdevice generates opposing gradients for two types of cells on the same chip, which builds a controlled system with sequentially changing components to study environmental effects from basal and immune cells.

Show MeSH
Related in: MedlinePlus