Effects of rG-CSF on Proliferation of Urological Cancer Cells

OK-432培养的PBMCs可抑制PBMCs产生rg-CSF。这种抑制作用可能是OK-432细胞因子介导的抗肿瘤作用机制之一。KK-47细胞和T24细胞等肿瘤细胞和PBMC在分离的琼脂中相互接种。培养上清液中未检测到IL-6、EGF、碱性成纤维细胞生长因子、IL-2等细胞因子。KK-47细胞在培养上清液中结构性地产生碱性成纤维细胞生长因子和IL-6，而T24细胞则产生IL-6。当rg-CSF与KK-47细胞共同培养96h时，KK-47细胞产生的碱性成纤维细胞生长因子呈剂量依赖性增加，直至浓度达到10 ng/ml，且在PBMC和rg-CSF同时存在的情况下，碱性成纤维细胞生长因子的产生增加。加入碱性成纤维细胞生长因子后，四甲基偶氮唑蓝比色法测定的KK-47细胞数增加，且呈剂量依赖关系。Rg-CSF和PBMC联合培养的KK-47细胞数明显高于单独使用Rg-CSF的KK-47细胞。流式细胞仪测定KK-47细胞在rg-CSF作用下的S期细胞数增加，而T24细胞在相同条件下无明显变化。逆转录聚合酶链式反应显示KK-47细胞表达IL-6受体、FGF1和Rg-CSF受体的mRNAs，而未见FGF2mRNA的表达。T24细胞表达IL-6mRNA，而不表达rg-CSFmRNA。PBMC表达rg-CSF、FGF1和FGF2受体的mRNAs。提示IL-6和jbasicFGF2可能是KK-47细胞的自分泌因子。Rg-CSF激活的KK-47细胞产生碱性成纤维细胞生长因子可能参与了rg-CSF对KK-47细胞增殖的影响。

The cultivation of PBMCs with OK-432 inhibited the production of rG-CSF by PBMCs. The inhibition may play a role in the mechanism of the cytokine-mediated antitumor effect of OK-432.Tumor cells such as KK-47 cells and T24 cells, and PBMCs were seeded in separated agar layr each other in the following experiments. Any cytokines such as IL-6, EGF,basic-FGF,IL-2 were not detected in the supernatant of cultured PBMCs. KK-47 cells constitutively produced basic-FGF and IL-6 in the supernatant and T24 cells produced IL-6. When KK-47 cells were cultured with rG-CSF for 96 hours, productio of basic-FGF by KK-47 cells increased in a dose-dependent manner until rG-CSF concentraion of 10 ng/ml. Furthermore, the production of basic-FGF increased under the presence of both of PBMCs and rG-CSF in the culture medium. When basic-FGF was added into culture medium, cell number estimated by MTT assay of KK-47 cells increased in a dose-dependent manner on basic-FGF concentrations. Cell number of KK-47 cells cultured with both of rG-CSF and PBMCs significantly increased than those cultured with rG-CSF only. Cell number of S-phase fraction estimated by FCM of KK-47 cells increased in cultivation with rG-CSF,however, that of T24 cells did not in the same condition. In RT-PCR,KK-47 cells expressed mRNAs of IL-6 receptor, FGF receptor 1 and rG-CSF receptor, however, no expression of mRNA of FGF receptor 2 was observed. T24 cells expressed mRNA of IL-6, however, no expression of mRNA of rG-CSF was observed. PBMCs expressed mRNAs of rG-CSF,FGF receptor 1 and 2. It was suggested that IL-6 and jbasic FGF might be autocrine factors of KK-47 cells. Production of basic FGF by KK-47 cells activated by rG-CSF might take part in effect of rG-CSF on proliferation of KK-47 cells.