FIBROBLASTS OF HEALTHY AND GRANULATION gingiva are phenotypically heterogeneous with regard to binding C1q collagen-like (cC1qR) or C1q globular-heads (gC1qR) regions, respectively. Here, isolated fibroblast subsets, expressing either the cC1qR or the gC1qR phenotype, were stimulated with C1q, 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+]iwere 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 PKCα/βI-βII/γ. Subsets manifested different fluctuations in [Ca2+]ilevels 20 seconds after C1q-stimulation in the presence of millimolar concentrations of external calcium. Whereas cC1qR fibroblasts responded with a 38% over baseline [Ca2+]iincrease which was sustained for 20 to 30 minutes, gC1qR 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 cC1qR fibroblasts responded with an IP3 increase of 32 ± 3 pmol/104cells over baseline after 5 seconds stimulation, gC1qR fibroblasts responded after 15 to 20 seconds with a lower increase (13 ± 0.8 IP3 pmol/104cells over baseline). Subsets differed in cPKCs redistribution which peaked in gC1qR-membranes 30 seconds after stimulation and remained sustained between 10 and 30 minutes. No cPKC redistribution was detectable in stimulated cC1qR-cells. We conclude that fibroblasts are heterogeneous in phosphoinositide-Ca2+signaling and cPKC redistribution to C1q, and suggest that these differences may affect activities of normal and granulation gingiva.
Fibroblast Heterogeneity of Signal Transduction Mechanisms to Complement-C1q. Analyses of Calcium Mobilization, Inositol Phosphate Accumulation, and Protein Kinases-C Redistribution / Bordin, Sandra; Costa, Lucio G.; Tan, Xiaoxia. - In: JOURNAL OF PERIODONTOLOGY. - ISSN 0022-3492. - 69:6(1998), pp. 642-649. [10.1902/jop.1998.69.6.642]
Fibroblast Heterogeneity of Signal Transduction Mechanisms to Complement-C1q. Analyses of Calcium Mobilization, Inositol Phosphate Accumulation, and Protein Kinases-C Redistribution
Costa, Lucio G.;
1998-01-01
Abstract
FIBROBLASTS OF HEALTHY AND GRANULATION gingiva are phenotypically heterogeneous with regard to binding C1q collagen-like (cC1qR) or C1q globular-heads (gC1qR) regions, respectively. Here, isolated fibroblast subsets, expressing either the cC1qR or the gC1qR phenotype, were stimulated with C1q, 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+]iwere 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 PKCα/βI-βII/γ. Subsets manifested different fluctuations in [Ca2+]ilevels 20 seconds after C1q-stimulation in the presence of millimolar concentrations of external calcium. Whereas cC1qR fibroblasts responded with a 38% over baseline [Ca2+]iincrease which was sustained for 20 to 30 minutes, gC1qR 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 cC1qR fibroblasts responded with an IP3 increase of 32 ± 3 pmol/104cells over baseline after 5 seconds stimulation, gC1qR fibroblasts responded after 15 to 20 seconds with a lower increase (13 ± 0.8 IP3 pmol/104cells over baseline). Subsets differed in cPKCs redistribution which peaked in gC1qR-membranes 30 seconds after stimulation and remained sustained between 10 and 30 minutes. No cPKC redistribution was detectable in stimulated cC1qR-cells. We conclude that fibroblasts are heterogeneous in phosphoinositide-Ca2+signaling and cPKC redistribution to C1q, and suggest that these differences may affect activities of normal and granulation gingiva.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
