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[Preprint]. 2024 Jun 4:2024.06.04.597454. [Version 1] doi: 10.1101/2024.06.04.597454

Figure 1: Stiffening extracellular matrix lowers pHi in metastatic human lung carcinoma (H1299).

Figure 1:

a) Schematic of increased pHi and ECM stiffening (via increased protein secretion and increased protein crosslinking) associated with tumorigenesis. b) Schematic of synthetic ECM models with tunable-stiffness (~50 Pa-1,500 Pa). The Matrigel (or Geltrex) model mimics increased ECM protein secretion while hyaluronic acid (HA) gel system mimics increased ECM protein crosslinking. c) Representative images of H1299 cells stably expressing mCherry-pHluorin pH biosensor plated on varying HA gel stiffnesses. Images show ratiometric display of pHluorin/mCherry fluorescence. Scale bars: 50 μm. d) Quantification of single-cell pHi data collected as shown in (c). (n=3 biological replicates; n=91 0.5% PEGDA, n=90 1% PEGDA, n=102 2% PEGDA, n=89 4% PEGDA. Red lines show medians ± IQR). e) Representative images of H1299 cells stably expressing mCherry-pHluorin pH biosensor plated on varying Matrigel stiffnesses. Images show ratiometric display of pHluorin/mCherry fluorescence. Scale bars: 50 μm. f) Quantification of single-cell pHi data collected as shown in (e). (n=3 biological replicates; n=93 4mg/mL, n=92 6mg/mL, n=102 8mg/mL, n=97 12mg/mL. Red lines show medians ± IQR). For (d) and (f), significance was determined by a Kruskal-Wallis test (****P<0.0001).