萎蔫酸铜离子络合物、镉离子络合物的抗结核活性及对结核分枝杆菌基因表达的影响

中国防痨杂志 ›› 2012, Vol. 34 ›› Issue (04): 224-228.

萎蔫酸铜离子络合物、镉离子络合物的抗结核活性及对结核分枝杆菌基因表达的影响

王娟黄宇虹陈伊李研彭毅潘嘉慧王军林永成赖小敏

510080 广州，中山大学中山医学院微生物学教研室、热带病防治研究教育部重点实验室、海洋微生物功能分子广东省高校重点实验室、广东省重大传染病预防和控制技术中心（王娟、黄宇虹、陈伊、李研、彭毅、赖小敏）；中山大学化学与化学工程学院有机化学研究所、海洋微生物功能分子广东省高校重点实验室（潘嘉慧、林永成）；中山大学药学院、海洋微生物功能分子广东省高校重点实验室（王军）

收稿日期:2011-09-17 出版日期:2012-04-10 发布日期:2012-06-14
通信作者: 赖小敏;林永成 E-mail:laixm@mail.sysu.edu.cn; ceslyc@mail.sysu.edu.cn
基金资助:
“十一五”国家高技术研究发展计划（863计划）（2007 AA092414）

The anti-tuberculosis activity of metal complexes of fusaric acid and their effect on gene expressions of M.tuberculosis

WANG Juan, HUANG Yu-hong, CHEN Yi, LI Yan, PENG Yi, PAN Jia-hui, WANG Jun, LIN Yong-cheng, LAI Xiao-min

Zhongshan School of Medicine, Ministry of Education Key Laboratory of Tropical Disease Control, Guangdong Provincial Department of Education Key Laboratory of Functional Molecules from Marine Microorganisms, Guangdong Provincial Research Center for Severe Infectious Disease Prevention and Control Technology, Sun Yat-sen University, Guangzhou 510080, China

Received:2011-09-17 Online:2012-04-10 Published:2012-06-14
Contact: LAI Xiao-min;LIN Yong-cheng E-mail:laixm@mail.sysu.edu.cn; ceslyc@mail.sysu.edu.cn

摘要/Abstract

摘要： 目的研究海洋微生物代谢产物萎蔫酸金属铜离子络合物与镉离子络合物体外抗结核活性，以及对Mtb H37Rv基因表达的影响。方法应用纸片扩散法（Kind-Bauer,K-B法）初步鉴定萎蔫酸铜离子络合物、萎蔫酸镉离子络合物的抗结核活性，重复实验3次；绝对浓度间接法测定萎蔫酸铜离子络合物、萎蔫酸镉离子络合物的最低抑菌浓度（minimum inhibitory concentration ，MIC），重复实验确定更准确的MIC；Mtb cDNA芯片检测萎蔫酸铜离子络合物、萎蔫酸镉离子络合物处理Mtb组与溶剂对照处理Mtb组表达差异的基因，重复实验一次。结果萎蔫酸铜离子络合物、萎蔫酸镉离子络合物在卡介苗（BCG）平板抑菌圈直径分别为（30.4±0.6）mm、（30.0±0.8）mm。萎蔫酸铜离子络合物抗Mtb H37Rv的MIC为10 μg/ml；对耐多药结核分枝杆菌，萎蔫酸镉离子络合物MIC为7.5 μg/ml，低于S（MIC=25 μg/ml）和RFP（MIC=25 μg/ml）；对Mtb临床分离株（0907961，耐S和EMB），萎蔫酸铜离子络合物的MIC（10 μg/ml）低于S（S的MIC为20 μg/ml），萎蔫酸镉离子络合物的MIC（5 μg/ml）均低于S（MIC=20 μg/ml）、EMB（MIC=6.4 μg/ml）。Mtb cDNA芯片检测发现萎蔫酸铜离子络合物处理组与溶剂对照组比较，有差异表达的基因总数为23条，其中已知功能基因有10条；萎蔫酸镉离子络合物处理组与溶剂对照组比较，有差异表达的基因总数为28条，其中已知功能基因有11条。这些存在差异的基因功能涉及核苷酸、脂质、能量、辅酶、氨基酸代谢，DNA的复制、转录、翻译、翻译后修饰以及细胞膜的生物合成等。结论萎蔫酸铜离子络合物与萎蔫酸镉离子络合物具有较好的体外抗结核活性，尤其对部分耐药菌株的抑制作用甚至优于抗结核一线用药，可作为抗结核药物的前导化合物；运用基因芯片所探究出的差异表达的基因为进一步深入研究其抗菌作用机制提供了一定的实验依据。

关键词: 镰刀菌酸, 配合物, 抗结核药, 寡核苷酸序列分析, 基因表达

Abstract: Objective To observe in vitro antituberculosis activity of copper ion complex and cadmium ion complex of fusaric acid, and their effects on the gene expressions of M.tuberculosis (Mtb) H37Rv. Methods The anti-tuberculosis activities of copper ion complex and cadmium ion complex of fusaric acid were determined for three times by Kind-Bauer method, and their minimum inhibitory concentrations (MICs) were detected twice by the absolute concentration method.The differences of the gene expressions of Mtb H37Rv between the compound group and non-compound control group were measured twice with DNA chips. Results The antibacterial diameters of cadmium complex and cupric complex of fusaric acid on the BCG plates were (30.0±0.8) mm and (30.4±0.6) mm, respectively. The MIC of cupric complex of fusaric acid on Mtb H37Rv was 10 μg/ml. The MIC of cadmium complex of fusaric acid on multi-drug-resistant Mtb was 7.5 μg/ml, which was lower than that of streptomycin (S, MIC=25 μg/ml) and rifampicin (RFP,MIC=25 μg/ml). The MIC of cupric complex of fusaric acid on Mtb clinical isolate（No.0907961，resistant to S and EMB) was 10 μg/ml, which was lower than that of S (MIC=20 μg/ml); and the MIC of cadmium complex of fusaric acid was 5 μg/ml, which was lower than that of S (MIC=20 μg/ml) and EMB (MIC=6.4 μg/ml). Of 3875 genes tested by the DNA array, 23 genes (10 known functional genes) were significantly different in copper ion complex group, and 28 genes (11 known functional genes) in cadmium ion complex group compared with the control groups. The functions of these differential genes were involved in nucleotide, lipid, energy, coenzyme, amino acid metabolism, DNA replication, transcription, translation, post-translational modification, biosynthesis of cell membranes etc. Conclusion Metal complexes of fusaric acid appeared considerable in vitro anti-tuberculosis activity, especially inhibitory activities of the compounds on some drug-resistant Mtb strains appeared higher than that of the first-line agents.Therefore, they could be used as anti-tuberculosis lead compounds. The genes with differential expressions showed by the DNA chip provide a valuable foundation for further study on their antibacterial mechanism.

Key words: Fusaric acid, Coordination complexes, Antitubercular agents, Oligonucleotide array sequence analysis, Gene expression

王娟黄宇虹陈伊李研彭毅潘嘉慧王军林永成赖小敏. 萎蔫酸铜离子络合物、镉离子络合物的抗结核活性及对结核分枝杆菌基因表达的影响[J]. 中国防痨杂志, 2012, 34(04): 224-228.

WANG Juan, HUANG Yu-hong, CHEN Yi, LI Yan, PENG Yi, PAN Jia-hui, WANG Jun, LIN Yong-cheng, LAI Xiao-min. The anti-tuberculosis activity of metal complexes of fusaric acid and their effect on gene expressions of M.tuberculosis[J]. Chinese Journal of Antituberculosis, 2012, 34(04): 224-228.

参考文献

［1］World Health Organization. Global tuberculosis controlsurveillance, planning, financing：report of WHO Scientific Group. Geneva：WHO，2008：393. http://www.who.int/tb/publications/global_report/2008/download_centre/en/index.html.

［2］Raman K，Rajagopalan P，Chandra N. Flux balance analysis of mycolic acid pathway: targets for anti-tubercular drugs. PLoS Comput Biol，2005，1(5)：349-358.

［3］Bugni TS，Ireland CM. Marine-derived fungi: a chemically and biologically diverse group of microorganisms. Nat Prod Rep，2004，21(1)：143-163.

［4］Mayer AM，Hamann MT. Marine pharmacology in 2000: marine compounds with antibacterial, anticoagulant, antifungal, anti-inflammatory, antimalarial, antiplatelet, antituberculosis, and antiviral activities; affecting the cardiovascular,immune, and nervous systems and other miscellaneous mechanisms of action. Mar Biotechnol（NY），2004，6 (1)：37-52.

［5］Wright AD，de Nys R，Angerhofer CK，et al. Biological activities and 3D QSAR studies of a series of Delisea pulchra (cf. fimbriata) derived natural products. J Nat Prod，2006，69(8)：1180-1187.

［6］Khalid A，El Sayed，Bartyzel P，et al. Marine natural products as antituberculosis agents. Tetrahedron，2000 (56)：949-953.

［7］Bacon CW，Porter JK，Norred WP，et al. Production of fusaric acid by Fusarium species. Appl Environ Microbiol，1996，62 (11)：4039-4043.

［8］Cameron DW，Deutscher KR，Feutrill GI. Synthesis of bostrycoidin and 8-0-methyl bostrycoidin. Tetrahedron Letters，1980，21(52)：5089-5090.

［9］黄宇虹，王娟，陈洪，等. Bostrycin 的抗结核菌活性及对细菌基因表达的影响.中国新药杂志，2011，20（6）：47-49.

［10］潘嘉慧. 南海真菌代谢物研究及萎蔫酸铜络合物生物合成机制初探. 广州：中山大学，2010.

［11］Medvedeva TM，Bezborodova AM，Zheludkova MD,et al. Synthesis and pharmacological activity of fusaric acid and its derivatives.Pharm Chem J，1978，12(5)：614-617.

［12］Wang H，Ng TB. Pharmacological activities of fusaric acid (5-butypicolinic acid). Life Sci，1999，65(9)：849-856.

［13］Hidaka H，Nagatsu T，Takeya K，et al. Fusaric acid, a hypotensive agent produced by fungi. J Antibiot(Tokyo)，1969，22(5)：228-230.

［14］Cole ST，Brosch R，Parkhill J，et al. Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence. Nature，1998，393(6685)：537-544.

[1]	梁小烟, 林玫, 梁大斌, 蓝如束, 覃慧芳, 叶婧, 黄莉雯. 广西壮族自治区耐多药结核分枝杆菌耐药情况与基因型特征分析[J]. 中国防痨杂志, 2020, 42(6): 578-582.
[2]	洪创跃, 杨婷婷, 李金莉, 李霜君, 吴利凯, 杨争, 谭卫国. 深圳市耐多药结核分枝杆菌耐药基因突变特征分析[J]. 中国防痨杂志, 2020, 42(6): 583-589.
[3]	张静, 陈曦, 王彬, 付雷, 陆宇, 陈效友. 改良单叠氮丙啶-荧光定量PCR法的建立及其检测抗结核药物活性的价值[J]. 中国防痨杂志, 2020, 42(5): 472-480.
[4]	高天慧,舒薇,高静韬,陆宇,李琦. 254例耐药肺结核患者克拉霉素耐药情况及影响因素分析[J]. 中国防痨杂志, 2020, 42(3): 259-265.
[5]	陈浩,武楠楠,胡文辉,杨忠金. 抗结核药物作用新靶点及其研究进展[J]. 中国防痨杂志, 2020, 42(3): 286-292.
[6]	王静,张蒙,贺建清. 抗结核药物引起的艰难梭状芽胞杆菌相关性腹泻一例并文献复习[J]. 中国防痨杂志, 2020, 42(3): 293-296.
[7]	邱磊,张少言,郭晓燕,付际游,田黎明,张惠勇,鹿振辉. 化疗方案加芩部丹片治疗复治涂阳肺结核患者的临床价值[J]. 中国防痨杂志, 2020, 42(2): 108-114.
[8]	王雪迪,江锋,代倩兰,王京,王冬梅. 中西医联合与单纯西医治疗结核病所致药物性肝损伤的对比分析(2000—2019年文献复习)[J]. 中国防痨杂志, 2020, 42(2): 126-132.
[9]	张蓝月,耿艺漫,贾红彦,肖婧,李自慧,潘丽萍,孙义成,张宗德. 耻垢分枝杆菌新型毒素-抗毒素系统MSMEG_3435-3436基因功能的初步研究[J]. 中国防痨杂志, 2020, 42(2): 133-142.
[10]	李蒙, 路丽苹, 江琦, 洪建军, 高谦, 杨崇广, 郭晓芹. 上海市松江区肺结核患者治疗药品的种类及费用分析[J]. 中国防痨杂志, 2020, 42(12): 1329-1332.
[11]	唐亮, 鲍玉成, 张文龙. 抗结核药品对肠道菌群的改变及其对机体的影响[J]. 中国防痨杂志, 2020, 42(12): 1333-1338.
[12]	陈曦,刘忠泉,王彬,朱慧,付雷,李媛媛,陆宇. 14种抗结核药物在巨噬细胞内的抗结核活性评价[J]. 中国防痨杂志, 2019, 41(9): 993-998.
[13]	刘佳文,吕红艳,丁北川,张治国. 129株非结核分枝杆菌采用两种分子检测技术行菌种鉴定的结果分析[J]. 中国防痨杂志, 2019, 41(9): 999-1003.
[14]	丁光贵,贺星,梁娟,刘亚亚,欧敏,陆坚,张国良. miR-20a-5p调控结核分枝杆菌诱导巨噬细胞凋亡相关基因表达的研究[J]. 中国防痨杂志, 2019, 41(7): 747-753.
[15]	《中国防痨杂志》编委会. 耐药结核病化疗过程中药品不良反应处理的专家共识[J]. 中国防痨杂志, 2019, 41(6): 591-603.