期刊
JOURNAL OF INTEGRATIVE MEDICINE-JIM
卷 14, 期 6, 页码 456-464出版社
ELSEVIER SCIENCE BV
DOI: 10.1016/S2095-4964(16)60279-0
关键词
allylpyrocatechol; catalase; superoxide dismutase; alkyl hydroperoxide reductase; reactive oxygen species; Staphylococcus aureus; Piper betle
资金
- Faculty of Health Sciences, MARA University of Technology (UiTM) Selangor, Puncak Alam
- UiTM LESTARI grant [600-IRMI/MYRA 5/3/LESTARI (16/2016)]
OBJECTIVE: This study investigated the effects of allylpyrocatechol (APC), the major component in ethanolic extract of Piper betle, on key oxidative stress resistance enzymes important for the survival of Staphylococcus aureus, a major pathogen in the human host. METHODS: Effects of APC on expressions of genes encoding catalase (katA), superoxide dismutases (SODs), including sodA and sodM, and alkyl hydroperoxide reductase (ahpC) in S. aureus were quantitated by RT-qPCR in reference to gyrA and 16S rRNA. Corresponding activities of the enzymes were also investigated. The Livak analysis was performed for verification of gene-fold expression data. Effects of APC on intracellular and extracellular reactive oxygen species (ROS) levels were determined using the nitroblue tetrazolium (NBT) reduction assay. RESULTS: APC-treated S. aureus cells had higher sodA and sodM transcripts at 1.5-fold and 0.7-fold expressions respectively with corresponding increase in total SOD activity of 12.24 U/mL compared to untreated cells, 10.85 U/mL (P<0.05). Expression of ahpC was highest in APC-treated cells with 5.5-fold increased expression compared to untreated cells (P<0.05). Correspondingly, ahpC activity was higher in APC-treated cells at 0.672 (A(310nm)) compared to untreated cells which was 0.394 (A(310nm)). In contrast, katA expression was 1.48-fold and 0.33-fold lower respectively relative to gyrA and 16S rRNA. Further, APC-treated cells showed decreased catalase activity of 1.8 x 10(-4) (U/L or mu mol/(min.L)) compared to untreated cells, which was 4.8 x 10(-4) U/L (P<0.05). Absorbance readings (A(575nm)) for the NBT reduction assay were 0.709 and 0.695 respectively for untreated and treated cells, which indicated the presence of ROS. APC-treated S. aureus cells had lower ROS levels both extracellularly and intracellularly, but larger amounts remained intracellularly compared to extracellular levels with absorbances of 0.457 and 0.137 respectively (P<0.05). CONCLUSION: APC induced expressions of both sodA and sodM, resulting in increased total SOD activity in S. aureus. Higher sodA expression indicated stress induced intracellularly involving O-2(-), presumably leading to higher intracellular pools of H2O2. A concommittant decrease in katA expression and catalase activity possibly induced ahpC expression, which was increased the highest in APC-treated cells. Our findings suggest that in the absence of catalase, cells are propelled to seek an alternate pathway involving ahpC to reduce stress invoked by O-2(-) and H2O2. Although APC reduced levels of ROS, significant amounts eluded its antioxidative action and remained intracellularly, which adds to oxidative stress in treated cells.
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