Bordin S, Costa L G, Tan X
Department of Periodontics, University of Washington, Seattle 98195, USA.
J Periodontol. 1998 Jun;69(6):642-9. doi: 10.1902/jop.1998.69.6.642.
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+]i 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 PKCalpha/betaI-betaII/gamma. Subsets manifested different fluctuations in [Ca2+]i levels 20 seconds after C1q-stimulation in the presence of millimolar concentrations of external calcium. Whereas cC1qR fibroblasts responded with a 38% over baseline [Ca2+]i increase 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/10(4) cells over baseline after 5 seconds stimulation, gC1qR 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 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.
健康牙龈和肉芽组织牙龈的成纤维细胞在分别结合C1q胶原样（cC1qR）或C1q球形头部（gC1qR）区域方面表现出表型异质性。在此，用C1q刺激表达cC1qR或gC1qR表型的分离成纤维细胞亚群，并评估胞质游离钙[Ca2+]i的变化、肌醇三磷酸（IP3）的积累以及钙依赖性蛋白激酶-C（cPKCs）从胞质溶胶到膜的重新分布。使用Indo-1荧光结合贴壁细胞分析和分选（ACAS）流式细胞术测定[Ca2+]i的变化。使用竞争性放射受体结合测定法定量IP3的积累。通过用抗PKCalpha/betaI-betaII/gamma抗体进行免疫印迹来评估cPKCs的重新分布。在存在毫摩尔浓度的细胞外钙的情况下，C1q刺激后20秒，亚群在[Ca2+]i水平上表现出不同的波动。cC1qR成纤维细胞的反应是[Ca2+]i比基线增加38%，并持续20至30分钟，而gC
