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Extracellular protonation modulates cell-cell interaction mechanics and tissue invasion in human melanoma cells

View Article: PubMed Central - PubMed

ABSTRACT

Detachment of cells from the primary tumour precedes metastatic progression by facilitating cell release into the tissue. Solid tumours exhibit altered pH homeostasis with extracellular acidification. In human melanoma, the Na+/H+ exchanger NHE1 is an important modifier of the tumour nanoenvironment. Here we tested the modulation of cell-cell-adhesion by extracellular pH and NHE1. MV3 tumour spheroids embedded in a collagen matrix unravelled the efficacy of cell-cell contact loosening and 3D emigration into an environment mimicking physiological confinement. Adhesive interaction strength between individual MV3 cells was quantified using atomic force microscopy and validated by multicellular aggregation assays. Extracellular acidification from pHe7.4 to 6.4 decreases cell migration and invasion but increases single cell detachment from the spheroids. Acidification and NHE1 overexpression both reduce cell-cell adhesion strength, indicated by reduced maximum pulling forces and adhesion energies. Multicellular aggregation and spheroid formation are strongly impaired under acidification or NHE1 overexpression. We show a clear dependence of melanoma cell-cell adhesion on pHe and NHE1 as a modulator. These effects are opposite to cell-matrix interactions that are strengthened by protons extruded via NHE1. We conclude that these opposite effects of NHE1 act synergistically during the metastatic cascade.

No MeSH data available.


Related in: MedlinePlus

Single cell force spectroscopy using MV3 cells with different NHE1 expression levels.AFM experiments reveal that the cell-cell adhesion force, represented by the (a) maximum pulling force and the (b) adhesion energy, is lower in NHE1-overexpressing cells (0.62 nN (0.48/0.85 nN), N = 9 cells attached to the cantilever probing n = 354 cells on the underlying matrix; 1.96 fJ (0.97/2.65 fJ), N = 9, n = 315) than in MV3 control cells (1.45 nN (0.99/1.95 nN), N = 8, n = 326; 4.35 fJ (2.67/6.8 fJ), N = 8, n = 281). Statistical significance of the differences was assessed by the Mann-Whitney U test.
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f3: Single cell force spectroscopy using MV3 cells with different NHE1 expression levels.AFM experiments reveal that the cell-cell adhesion force, represented by the (a) maximum pulling force and the (b) adhesion energy, is lower in NHE1-overexpressing cells (0.62 nN (0.48/0.85 nN), N = 9 cells attached to the cantilever probing n = 354 cells on the underlying matrix; 1.96 fJ (0.97/2.65 fJ), N = 9, n = 315) than in MV3 control cells (1.45 nN (0.99/1.95 nN), N = 8, n = 326; 4.35 fJ (2.67/6.8 fJ), N = 8, n = 281). Statistical significance of the differences was assessed by the Mann-Whitney U test.

Mentions: We next tested whether the acid extruding NHE1 not only affects cell migration1519 but also cell-cell adhesion. Therefore, we compared MV3 cells overexpressing NHE1 and NHE1-deficient cells. The maximum pulling force necessary to separate two individual melanoma vector control cells was 1.45 nN, the adhesion energy was 4.35 fJ. (Fig. 3). In contrast, the maximum pulling force was 57% lower in MV3 cells overexpressing NHE1 and the adhesion energy was reduced by 55%. Moreover, NHE1-deficient MV3 cells showed a significant 12% increase in the maximum pulling force compared to the respective control (supplementary Fig. S2). Thus, because increased NHE1 expression is known to lower pericellular pHem, these findings were in line with the above described AFM measurements with varying proton concentrations in the bathing solution.


Extracellular protonation modulates cell-cell interaction mechanics and tissue invasion in human melanoma cells
Single cell force spectroscopy using MV3 cells with different NHE1 expression levels.AFM experiments reveal that the cell-cell adhesion force, represented by the (a) maximum pulling force and the (b) adhesion energy, is lower in NHE1-overexpressing cells (0.62 nN (0.48/0.85 nN), N = 9 cells attached to the cantilever probing n = 354 cells on the underlying matrix; 1.96 fJ (0.97/2.65 fJ), N = 9, n = 315) than in MV3 control cells (1.45 nN (0.99/1.95 nN), N = 8, n = 326; 4.35 fJ (2.67/6.8 fJ), N = 8, n = 281). Statistical significance of the differences was assessed by the Mann-Whitney U test.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5304230&req=5

f3: Single cell force spectroscopy using MV3 cells with different NHE1 expression levels.AFM experiments reveal that the cell-cell adhesion force, represented by the (a) maximum pulling force and the (b) adhesion energy, is lower in NHE1-overexpressing cells (0.62 nN (0.48/0.85 nN), N = 9 cells attached to the cantilever probing n = 354 cells on the underlying matrix; 1.96 fJ (0.97/2.65 fJ), N = 9, n = 315) than in MV3 control cells (1.45 nN (0.99/1.95 nN), N = 8, n = 326; 4.35 fJ (2.67/6.8 fJ), N = 8, n = 281). Statistical significance of the differences was assessed by the Mann-Whitney U test.
Mentions: We next tested whether the acid extruding NHE1 not only affects cell migration1519 but also cell-cell adhesion. Therefore, we compared MV3 cells overexpressing NHE1 and NHE1-deficient cells. The maximum pulling force necessary to separate two individual melanoma vector control cells was 1.45 nN, the adhesion energy was 4.35 fJ. (Fig. 3). In contrast, the maximum pulling force was 57% lower in MV3 cells overexpressing NHE1 and the adhesion energy was reduced by 55%. Moreover, NHE1-deficient MV3 cells showed a significant 12% increase in the maximum pulling force compared to the respective control (supplementary Fig. S2). Thus, because increased NHE1 expression is known to lower pericellular pHem, these findings were in line with the above described AFM measurements with varying proton concentrations in the bathing solution.

View Article: PubMed Central - PubMed

ABSTRACT

Detachment of cells from the primary tumour precedes metastatic progression by facilitating cell release into the tissue. Solid tumours exhibit altered pH homeostasis with extracellular acidification. In human melanoma, the Na+/H+ exchanger NHE1 is an important modifier of the tumour nanoenvironment. Here we tested the modulation of cell-cell-adhesion by extracellular pH and NHE1. MV3 tumour spheroids embedded in a collagen matrix unravelled the efficacy of cell-cell contact loosening and 3D emigration into an environment mimicking physiological confinement. Adhesive interaction strength between individual MV3 cells was quantified using atomic force microscopy and validated by multicellular aggregation assays. Extracellular acidification from pHe7.4 to 6.4 decreases cell migration and invasion but increases single cell detachment from the spheroids. Acidification and NHE1 overexpression both reduce cell-cell adhesion strength, indicated by reduced maximum pulling forces and adhesion energies. Multicellular aggregation and spheroid formation are strongly impaired under acidification or NHE1 overexpression. We show a clear dependence of melanoma cell-cell adhesion on pHe and NHE1 as a modulator. These effects are opposite to cell-matrix interactions that are strengthened by protons extruded via NHE1. We conclude that these opposite effects of NHE1 act synergistically during the metastatic cascade.

No MeSH data available.


Related in: MedlinePlus