Penttinen Piia, Tampio Marjo, Mäki-Paakkanen Jorma, Vähäkangas Kirsi, Pelkonen Jukka, Hirvonen Maija-Riitta
National Public Health Institute, Department of Environmental Health, PO Box 95, FI-70701 Kuopio, Finland.
Toxicology. 2007 Jun 3;235(1-2):92-102. doi: 10.1016/j.tox.2007.03.005. Epub 2007 Mar 12.
Our recent studies have revealed that the co-cultivation of environmental microbes, Streptomyces californicus and Stachybotrys chartarum, potentiates the immunotoxic properties of the spores. In the present study, the spore-induced genotoxic potential of these microbes was investigated. Dose related differences in genotoxic and cytotoxic effects and in p53 level in mouse RAW264.7 macrophages were studied after 24h exposure to the spores of separately cultivated Streptomyces californicus or Stachybotrys chartarum alone, a simple spore-mixture of these microbes as well as to the spores of co-cultivated microbes. The genotoxic effect of the exposures was determined by the Comet assay and p53 level was analyzed by immunoblotting. Cytotoxicity was assessed by using flow cytometric analysis and also by the MTT test. The results revealed that the spores of co-cultivated microbes evoked DNA damage, p53 accumulation and cytotoxicity at a lower dose than the other exposures, and at the highest dose there was a 2.5-fold increase in DNA damage compared to control. In addition, the spores of Streptomyces californicus alone induced a 1.5-fold increase in DNA damage compared to control, dose dependent p53 accumulation and also extensive cytotoxicity. In contrast, the mixture of separately cultivated spores or the spores of Stachybotrys chartarum alone did not induce DNA damage with any tested dose although they triggered significant cytotoxicity and a slightly increased p53 level. Our results suggest that the detected genotoxic responses are the result of DNA damage in RAW264.7 cells by some genotoxically active metabolite(s) and the production of this compound was stimulated in Streptomyces californicus when it was co-cultivated with Stachybotrys chartarum.
我们最近的研究表明，环境微生物加州链霉菌和黑葡萄穗霉的共培养会增强孢子的免疫毒性。在本研究中，对这些微生物孢子诱导的遗传毒性潜力进行了研究。在分别单独培养的加州链霉菌或黑葡萄穗霉的孢子、这些微生物的简单孢子混合物以及共培养微生物的孢子暴露24小时后，研究了小鼠RAW264.7巨噬细胞中遗传毒性和细胞毒性效应以及p53水平的剂量相关差异。通过彗星试验确定暴露的遗传毒性效应，通过免疫印迹分析p53水平。使用流式细胞术分析并通过MTT试验评估细胞毒性。结果显示，与其他暴露相比，共培养微生物的孢子在较低剂量下即可引起DNA损伤、p53积累和细胞毒性，在最高剂量下，与对照相比DNA损伤增加了2.5倍。此外，单独的加州链霉菌孢子与对照相比，DNA损伤增加了1.5倍，呈剂量依赖性p53积累，并且还具有广泛的细胞毒性。相比之下，单独培养的孢子混合物或单独的黑葡萄穗霉孢子在任何测试剂量下均未诱导DNA损伤，尽管它们引发了显著的细胞毒性和p53水平略有升高。我们的结果表明，检测到的遗传毒性反应是RAW264.7细胞中某些具有遗传毒性活性的代谢物导致DNA损伤的结果，并且当加州链霉菌与黑葡萄穗霉共培养时，该化合物的产生受到刺激。
