Association of signal transduction for TLR-4 with oral cancer

我们研究了口腔癌的发生是否与口腔细菌诱导的炎症有关，而口腔细菌诱导的炎症主要是通过TLR-4介导的。在TLR-4信号转导分子中，我们重点研究了MyD88和MAP激酶磷酸酶-巨噬细胞（MAP kinase phosphatase-macrophage, MKP-M）， mkp是JNK1/2的磷酸酶。我们制备了myd88基因组改变小鼠和mkpm基因组改变小鼠，其中MKP-M蛋白失活。用4-硝基喹啉1-氧化物（4-NQO）作为致癌物诱导小鼠口腔癌。组织学分析显示，4-NQO对对照小鼠的舌头有癌变作用。然而，在myd88基因组改变的小鼠中观察到致癌作用的减少，与对照小鼠相比，对细菌的反应减少了。此外，在MKP-M灭活小鼠中观察到更大的致癌作用。接下来，我们从这些舌中分离总RNA，然后用反转录PCR和实时PCR检测c-met的表达，c-met是一个原基因，已知是肝细胞生长因子（HGF）的受体。4-NQO处理后，c-met在对照组小鼠的舌头中表达增加，而在myd88基因组改变小鼠的舌头中，c-met在4-NQO处理后的表达水平与未处理的对照组小鼠相同。另一方面，与4-NQO处理的对照组小鼠相比，4-NQO处理的MKP-M灭活小鼠的舌头中c-met的表达增加。这些结果与4-NQO处理后舌的组织学分析一致。接下来，我们从牙龈中培养成纤维细胞，并用IL-1和/或LPS刺激培养的牙龈成纤维细胞。然后采用反转录PCR和实时PCR检测c-met和HGF的表达。因此，这些刺激不会增加myd88（-/-）成纤维细胞中HGF和c-met的表达。另一方面，通过IL-1和/或LPS刺激，MKP-m基因组改变小鼠的牙龈成纤维细胞中HGF的表达增加，这些结果与我们之前报道的正常牙龈成纤维细胞中HGF表达的变化一致。这些结果表明，IL-1和TLR-4信号通过MyD88途径介导HGF表达。细菌和IL-1刺激与口腔癌细胞生长相关的HGF表达增强综上所述，口腔细菌感染激活TLR-4信号转导通路（如MyD88和JNK1/2介导的通路），通过增加c-met和HGF的表达，在口腔癌变中起重要作用。少

We examined if the generation of oral cancer was associated with the oral bacteria induced inflammation, that was mainly mediated though TLR-4. Among TLR-4 signal transducing molecules, we focused on MyD88 and MAP kinase phosphatase-macrophage (MKP-M), which was phosphatase for JNK1/2. We prepared myd88 genome altered mice and mkpm genome altered mice, in which MKP-M protein was inactivated. These mice were treated with 4-nitroquinoline 1-oxide (4-NQO) as a carcinogen material to induce oral cancer. Histological analysis showed carcinogenesis in tongues from control mice treated with 4-NQO. However a decrease in carcinogenesis was observed in myd88 genome altered mice in which the response against bacteria was reduced compared with control mice. Furthermore, a greater carcinogenesis was observed in MKP-M inactivated mice. Next we isolated total RNA from these tongues, and then we performed reverse transcription PCR and real time PCR to measure the expression of c-met which was found as … More a protoncogene and known to be a receptor of hepatocyte growth factor (HGF). The expression of c-met increased in tongues from control mice treated with 4-NQO, whereas its expression in tongue from myd88 genome altered mice treated with 4-NQO was the same level as that from non-treated control mice. While on the other hand, the expression of c-met increased in tongues from MKP-M inactivated mice treated with 4-NQO compared with 4-NQO treated control mice. These results of PCR consistent with histological analysis of 4-NQO treated tongues.Next, we established fibroblasts by outgrowth form gingiva, and stimulated cultured gingival fibroblasts with IL-1 and/or LPS. And then we performed reverse transcription PCR and real time PCR to measure the expression of c-met and HGF. Consequently, these stimulations did not increase HGF and c-met expressions in myd88(-/-) fibroblasts. On the other hand, the expression of HGF increase in gingival fibroblasts from MKP-m genome altered mice by stimulations with IL-1 and/or LPS, these results were consists with our previous reports which described changes of expression of HGF in normal gingival fibroblasts. These observations suggested that IL-1 and TLR-4 signal induced HGF expression mediated through MyD88 pathway. Enhancement of HGF expression by stimulation with bacteria and IL-1 associated with growth of oral cancer cells.In summary, activation of TLR-4 signal transduction pathway (such as MyD88 and JNK1/2 mediated pathway) by infections of oral bacteria plays an important role in oral carcinogenesis mediated through increase the expression of c-met and HGF. Less