Kent L W, Rahemtulla F, Hockett R D, Gilleland R C, Michalek S M
Department of Periodontics, School of Dentistry, The University of Alabama at Birmingham, 35294, USA.
Infect Immun. 1998 Feb;66(2):608-14. doi: 10.1128/IAI.66.2.608-614.1998.
Cytokines are hormone-like proteins which mediate and regulate inflammatory and immune responses. The purpose of this study was to investigate the effect of lipopolysaccharide (LPS) and inflammatory cytokines on regulation of interleukin-6 (IL-6) production by human gingival fibroblasts (HGF). The HGF cell lines used in this study, H-CL and F-CL, were established by the explant technique from healthy gingival tissue. Cultured cells were grown to confluency and incubated with various concentrations of LPS from Escherichia coli or Porphyromonas gingivalis or with the recombinant human cytokine tumor necrosis factor alpha (TNF-alpha), IL-1alpha, or IL-1beta. Culture supernatants were collected at various times and assessed for IL-6 production by enzyme-linked immunosorbent assay. Total RNA was isolated from the harvested cells and used to assess levels of IL-6 mRNA by the RNase protection assay. Both LPS preparations induced IL-6 production (1 to 4 ng of IL-6 per ml) by both HGF cell lines. Although TNF-alpha stimulated IL-6 production by HGF, > 10-fold-larger amounts were induced with IL-1alpha and IL-1beta. Furthermore, the addition of both IL-1alpha and TNF-alpha to cultured cells resulted in approximately 600- to 800-fold-higher levels of IL-6 than seen in control cultures, suggesting that these cytokines synergistically induced IL-6 production by HGF. IL-6 message in cultured cells was upregulated 20-fold by TNF-alpha, 1,000-fold by IL-1alpha and IL-1beta, and 1,400-fold by IL-1alpha plus TNF-alpha. IL-1alpha and TNF-alpha alone upregulate IL-6 production in a dose- and time-dependent fashion. The addition of IL-1alpha and TNF-alpha to cultured HGF cells resulted in a synergistic induction of IL-6 after 8 h of incubation and when greater than 10 pg of this combination per ml was used. Our studies show that inflammatory cytokines are hundreds of times more potent than LPS in stimulating IL-6 production by HGF.
细胞因子是一类类似激素的蛋白质，可介导和调节炎症及免疫反应。本研究的目的是调查脂多糖（LPS）和炎性细胞因子对人牙龈成纤维细胞（HGF）白细胞介素-6（IL-6）产生调节的影响。本研究中使用的HGF细胞系H-CL和F-CL，是通过外植体技术从健康牙龈组织建立的。将培养的细胞生长至汇合状态，并用来自大肠杆菌或牙龈卟啉单胞菌的不同浓度的LPS或用重组人细胞因子肿瘤坏死因子α（TNF-α）、IL-1α或IL-1β进行孵育。在不同时间收集培养上清液，并通过酶联免疫吸附测定评估IL-6的产生。从收获的细胞中分离总RNA，并用于通过核糖核酸酶保护测定评估IL-6 mRNA的水平。两种LPS制剂均诱导两种HGF细胞系产生IL-6（每毫升1至4纳克IL-6）。尽管TNF-α刺激HGF产生IL-6，但IL-1α和IL-1β诱导的量要大10倍以上。此外，向培养细胞中添加IL-1α和TNF-α导致IL-6水平比对照培养物中高约600至800倍，表明这些细胞因子协同诱导HGF产生IL-6。培养细胞中的IL-6信息被TNF-α上调20倍，被IL-1α和IL-1β上调1000倍，被IL-1α加TNF-α上调1400倍。单独的IL-1α和TNF-α以剂量和时间依赖的方式上调IL-6的产生。向培养的HGF细胞中添加IL-1α和TNF-α在孵育8小时后以及当每毫升使用大于10皮克的这种组合时导致IL-6的协同诱导。我们的研究表明，炎性细胞因子在刺激HGF产生IL-6方面比LPS强数百倍。
