Deregulation of collagen phagocytosis in aging human fibroblasts: effects of integrin expression and cell cycle

Experimental Cell Research
W Lee, C A McCulloch

Abstract

Intracellular degradation of collagen by phagocytosis in fibroblasts is essential for physiological remodeling of the extracellular matrix in a wide variety of connective tissues but imbalances between degradation and synthesis can lead to loss of tissue collagen. As aging is associated with loss of dermal and periodontal collagen and with increased lysomomal enzyme content in fibroblasts, we examined the regulation of collagen phagocytosis by integrin expression and the cell cycle in an in vitro fibroblast aging model. Two different fibroblast lines (CL1; CL2) at the fourth subculture were passaged up to replicative senescence to model aging processes in vitro. Cells were incubated with collagen-coated or BSA-coated green fluorescent beads for 3 h to assess alpha 2 beta 1-integrin-mediated or nonspecific phagocytosis, respectively. Single-cell suspensions were stained with DAPI and sulforhodamine 101 to separate cycling G1 and noncycling G0 cells. Staining for alpha 2-integrin, bead internalization, and bivariate analyses of DNA/protein content were measured by three-color flow cytometry. Serum deprivation was used to induce increases in the proportion of G0 cells. For G1 cells, the proportion of collagen phagocytic cells was ...Continue Reading

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