| Summary: | 博士 === 國立臺灣大學 === 毒理學研究所 === 93 === The overdose and abuse of antibiotics have been a worldwide problem that it not only caused the antibiotics resistance in microbes but increased the risk in cancer development. However, there is no direct evidence about the relationship between carcinogenesis and antibiotics abuse. Several antibiotics, such as tetracyclines and chloramphenicol, can inhibit both bacterial and mitochondrial protein synthesis. The mitochondria are maternally inherited organelles and possess a circular extra-nuclear genome that encoded 13 polypeptides of mitochondrial oxidative phosphorylayion complexes. Hence, the damage in mitochondrial translation may cause ATP depletion and a series responses. In present study, we used the chloramphenicol to stress on mitochondria and to investigate the chloramphenicol-induced cellular responses.
We found that the chloramphenicol inhibited mitochondria-encoded protein synthesis specifically and decreased the cellular ATP level and the proliferative capacity. The chloramphenicol-treated cells also halted cell cycle at G0/G1 phase, elevated intracellular Ca2+ concentration, changed cell morphology to senescence-like shape and showed the appearance of senescence biomarker SA-β Gal. The chelation of intracellular Ca2+ will blocked the SA-β Gal activation suggested the involvement of Ca2+ in chloramphenicol-mediated senescence biogenesis. Moreover, the senescence-associated genes such as p21, galectin-3, matrix metalloproteinase-3 and -13 and the Ca2+ channel RyR were over-expressed in chloramphenicol-treated cells.
Subsequently, we attempted to investigate the physiological function p21 in chloramphenicol-treated cells. Pretreatment of HepG2 and H1299 cells with chloramphenicol rendered the cells resistant to mitomycin-induced apoptosis. Both mitomycin-induced caspase 3 activity and PARP activation were also inhibited. Both p21 antisense and siRNA could restore apoptotic-associated caspase 3 activity, PARP activation, and the sensitivity to mitomycin-induced apoptosis. Cellular levels of the p21 protein and mRNA were increased through a p53-independent pathway, possibly due to the stabilization of p21 mRNA in chloramphenicol-treated cells. The p21 was redistributed from the perinuclear region to the cytoplasm and co-localized with mitochondrial marker protein. These findings suggested that mitochondrial stress causes resistance to apoptosis through a p21-dependent pathway.
We also found the treatment of chloramphenicol could enhance MMP-3 and MMP-13 expression in JNK, PI-3K/Akt and AP-1 dependent manner. The chloramphenicol-mediated expression of MMP-13 could be inhibited by SP600125, LY294002 and curcumin. The JNK and c-Jun protein were phosphorylated in chloramphenicol-treated cells and the phospho-c-Jun protein will translocate into nucleus then activate MMP-13 promoter through AP-1 consensus site. The MMP-13 has been demonstrated in cancer invasion and metastasis. Chloramphenicol-treated cells showed strong invasive potential than untreated cells and the increase in invasive capacity could be completely prevented by MMP-13 inhibitor CL-82158. These findings suggested that chloramphenicol administration may accelerate cancer cell invasion through a MMP-13 dependent mechanism.
The similar effects also observed in minocycline, doxycycline and clindamycin. Our studies provided strong evidence for the antibiotics abuse in cancer development. The apoptotic resistance and MMP-13 dependent cell invasion may enhance cancer cell survival and accelerate cancer cell invasion, respectively. Finally, the clinical significant of present studies caution the clinical use of these antibiotics should be more carefully, especially during cancer chemotherapy.
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