Fibroblast heterogeneity of signal transduction mechanisms to complement-C1q. Analyses of calcium mobilization, inositol phosphate accumulation, and protein kinases C redistribution

FIBROBLASTS OF HEALTHY AND GRANULATION gingiva are phenotypically heterogeneous with regard to binding Clq collagen-like (cClqR) or Clq globular-heads (gClqR) regions, respectively. Here, isolated fibroblast subsets, expressing either the cClqR or the gClqR phenotype, were stimulated with Clq, and assessed for changes in cytosolic free calcium [Ca2+](i), accumulation of inositol trisphosphate (IP3), and redistribution of Ca2+-dependent protein kinases-C (cPKCs) from cytosol to membranes. Changes in [Ca2+], were determined using Indo-1 fluorescence in combination with adhering cell analysis and sorting (ACAS) cytometry. Accumulation of IP3 was quantified using a competitive radioreceptor binding assay. Redistribution of cPKCs was evaluated by immunoblotting with antibodies to PKC alpha/beta I-beta II/gamma. Subsets manifested different fluctuations in [Ca2+](i) levels 20 seconds after Clq-stimulation in the presence of millimolar concentrations of external calcium. Whereas cClqR fibroblasts responded with a 38% over baseline [Ca2+](i) increase which was sustained for 20 to 30 minutes, gClqR fibroblasts responded with a higher (264% over baseline) and more rapid (2 to 3 minutes) transient. Likewise, subsets exhibited different kinetics of IP3 accumulation. Whereas cClqR fibroblasts responded with an IP3 increase of 32 +/- 3 pmol/10(4) cells over baseline after 5 seconds stimulation, gClqR fibroblasts responded after 15 to 20 seconds with a lower increase (13 +/- 0.8 IP3 pmol/10(4) cells over baseline). Subsets differed in cPKCs redistribution which peaked in gClqR-membranes 30 seconds after stimulation and remained sustained between 10 and 30 minutes. No cPKC redistribution was detectable in stimulated cClqR-cells. We conclude that fibroblasts are heterogeneous in phosphoinositide-Ca2+ signaling and cPKC redistribution to Clq, and suggest that these differences may affect activities of normal and granulation gingiva.