Caveolae and Rho Kinase: their implication of the senescent cell morphology and the secretion of the SASP in HeLa and A549 cancer cells

Purpose: Caveolae and Rho kinase signaling individually are both remodeled during cellular senescence and influence cell morphology and SASP secretion. However, how this interaction modulates senescent cancer-cell morphology and SASP remains unresolved. We evaluated the possible connection between caveolae and ROCK on the development of senescent cell morphology and the secretion of the SASP. Methods: Senescence was induced in HeLa and A549 cells with ouabain. Caveolin depletion was achieved with methyl-beta-cyclodextrin (M beta CD) or Caveolin-1 siRNA applications. Cell morphology was assessed by cell area, volume, and thickness parameters, while SASP secretion was assessed by measuring total protein, IL-6, and VEGF-A in senescent cell secretomes. We also evaluated the effect of caveolae depletion on ROCK expression, activation, and localization. Results: While the caveolae disruption with the preincubation of M beta CD did not alter the occurrence of ouabain-induced senescence, it significantly altered cellular morphological features, such as decreased cell area and volume. The amount of ROCK1 and ROCK2 in the membrane fraction of cells was decreased by M beta CD preincubation. These findings indicate that there may be a relationship between ROCK and the morphological changes observed in senescent cells as a result of the disruption of the caveolar structure. However, the incubation of M beta CD and caveolin-1 knocking down increased the secretory activity of senescent cells. Overall secretion increases with caveolar depletion, indicating that its secretion-enhancing effect is greater than any concomitant decrease that may occur with ROCK inhibition. Conclusion: These data suggest that maintaining the integrity of caveolar structures plays a critical role in mitigating the detrimental effects of SASP released from chemotherapy-induced senescent cells.