Elevated Cholesterol in the Coxiella burnetii Intracellular Niche Is Bacteriolytic.
Elevated Cholesterol in the Coxiella burnetii Intracellular Niche Is Bacteriolytic.
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DOI：
10.1128/mbio.02313-16
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发表时间：
2017-02-28
期刊：
影响因子：
6.4
通讯作者：
Gilk SD
中科院分区：
文献类型：
--
作者：
Mulye M;Samanta D;Winfree S;Heinzen RA;Gilk SD
贝氏柯克斯体是一种细胞内细菌病原体，是美国培养阴性心内膜炎的重要原因。在感染后，新生的柯克斯体吞噬体与宿主的内吞途径融合，形成一个大的溶酶体样空泡，称为寄生虫空泡（PV）。PV膜富含固醇，干扰宿主细胞胆固醇稳态的药物抑制PV形成和细菌生长。使用无胆固醇细胞模型系统的胆固醇补充剂，我们发现随着细胞胆固醇浓度的增加，PV变小，Coxiella生长减少。此外，我们在含有胆固醇的细胞中观察到大量含有降解细菌的非融合PV，这是在不含胆固醇的细胞中未观察到的表型。胆固醇对无菌柯克斯体培养物没有影响，表明只有细胞内细菌对胆固醇敏感。活细胞显微镜显示，质膜衍生的胆固醇和外源性胆固醇载体蛋白低密度脂蛋白（LDL）交通到PV。为了测试增加PV胆固醇水平影响细菌存活的可能性，感染的细胞用U18666A处理，U18666A是一种将胆固醇捕获在溶酶体和PV中的药物。U18666A处理导致PV含有降解的细菌和细菌活力的显著损失。PV pH值显着更酸性的细胞与胆固醇或细胞与U18666A处理，和空泡ATP酶抑制剂巴弗洛霉素阻断胆固醇诱导的PV酸化和细菌死亡。此外，用几种FDA批准的改变胆固醇的药物治疗感染的HeLa细胞导致细菌活力的丧失，这也是巴弗洛霉素拯救的一种表型。总的来说，这些数据表明，增加PV胆固醇进一步酸化PV，导致柯克斯体死亡。细胞内革兰氏阴性细菌贝氏柯克斯体是培养阴性感染性心内膜炎的重要原因，如果不治疗，可能是致命的。现有的治疗策略需要长期的抗生素治疗，治疗患者的10年死亡率为19%。因此，需要新的临床治疗方法，可以通过更好地了解贝氏隐孢子虫的发病机制来实现。在感染宿主细胞后，贝氏隐孢子虫在一个专门的复制小生境中生长，即寄生虫空泡（PV）。最近的数据已经将胆固醇与胞内贝氏梭菌生长和PV形成联系起来，这使我们进一步破译胆固醇在贝氏梭菌-宿主相互作用期间的作用。我们观察到，增加PV胆固醇浓度导致PV的酸化增加和细菌死亡。此外，用FDA批准的改变宿主胆固醇稳态的药物治疗也通过PV酸化杀死了贝氏梭菌。我们的研究结果表明，靶向宿主胆固醇代谢可能被证明在临床上有效控制贝氏隐孢子虫感染。
Coxiella burnetii is an intracellular bacterial pathogen and a significant cause of culture-negative endocarditis in the United States. Upon infection, the nascent Coxiella phagosome fuses with the host endocytic pathway to form a large lysosome-like vacuole called the parasitophorous vacuole (PV). The PV membrane is rich in sterols, and drugs perturbing host cell cholesterol homeostasis inhibit PV formation and bacterial growth. Using cholesterol supplementation of a cholesterol-free cell model system, we found smaller PVs and reduced Coxiella growth as cellular cholesterol concentration increased. Further, we observed in cells with cholesterol a significant number of nonfusogenic PVs that contained degraded bacteria, a phenotype not observed in cholesterol-free cells. Cholesterol had no effect on axenic Coxiella cultures, indicating that only intracellular bacteria are sensitive to cholesterol. Live-cell microscopy revealed that both plasma membrane-derived cholesterol and the exogenous cholesterol carrier protein low-density lipoprotein (LDL) traffic to the PV. To test the possibility that increasing PV cholesterol levels affects bacterial survival, infected cells were treated with U18666A, a drug that traps cholesterol in lysosomes and PVs. U18666A treatment led to PVs containing degraded bacteria and a significant loss in bacterial viability. The PV pH was significantly more acidic in cells with cholesterol or cells treated with U18666A, and the vacuolar ATPase inhibitor bafilomycin blocked cholesterol-induced PV acidification and bacterial death. Additionally, treatment of infected HeLa cells with several FDA-approved cholesterol-altering drugs led to a loss of bacterial viability, a phenotype also rescued by bafilomycin. Collectively, these data suggest that increasing PV cholesterol further acidifies the PV, leading to Coxiella death. The intracellular Gram-negative bacterium Coxiella burnetii is a significant cause of culture-negative infectious endocarditis, which can be fatal if untreated. The existing treatment strategy requires prolonged antibiotic treatment, with a 10-year mortality rate of 19% in treated patients. Therefore, new clinical therapies are needed and can be achieved by better understanding C. burnetii pathogenesis. Upon infection of host cells, C. burnetii grows within a specialized replication niche, the parasitophorous vacuole (PV). Recent data have linked cholesterol to intracellular C. burnetii growth and PV formation, leading us to further decipher the role of cholesterol during C. burnetii-host interaction. We observed that increasing PV cholesterol concentration leads to increased acidification of the PV and bacterial death. Further, treatment with FDA-approved drugs that alter host cholesterol homeostasis also killed C. burnetii through PV acidification. Our findings suggest that targeting host cholesterol metabolism might prove clinically efficacious in controlling C. burnetii infection.