Department of Microbiology and Immunology, Louisiana State University Health Sciences Center-Shreveport, MSB 2-225, 1501 Kings Highway, Shreveport, LA 71130, USA.
Infect Immun. 2011 Jan;79(1):88-97. doi: 10.1128/IAI.00974-09. Epub 2010 Oct 25.
The human gastric pathogen Helicobacter pylori modifies host cholesterol via glycosylation and incorporates the glycosylated cholesterol into its membrane; however, the benefits of cholesterol to H. pylori are largely unknown. We speculated that cholesterol in the H. pylori membrane might alter the susceptibility of these organisms to membrane-disrupting antibacterial compounds. To test this hypothesis, H. pylori strains were cultured in Ham's F-12 chemically defined medium in the presence or absence of cholesterol. The two cultures were subjected to overnight incubations with serial 2-fold dilutions of 10 bile salts and four ceragenins, which are novel bile salt derivatives that mimic membrane-disrupting activity of antimicrobial peptides. H. pylori cultured with cholesterol was substantially more resistant to seven of the bile salts and three ceragenins than H. pylori cultured without cholesterol. In most cases, these cholesterol-dependent differences ranged from 2 to 7 orders of magnitude; this magnitude depended on concentration of the agent. Cholesterol is modified by glycosylation using Cgt, a cholesteryl glycosyltransferase. Surprisingly, a cgt knockout strain still maintained cholesterol-dependent resistance to bile salts and ceragenins, indicating that cholesterol modification was not involved in resistance. We then tested whether three putative, paralogous inner membrane efflux pumps, HefC, HefF, or HefI, played a role. While HefF and HefI appeared unimportant, HefC was shown to play a critical role in the resistance to bile salts and ceragenins by multiple methods in multiple strain backgrounds. Thus, both cholesterol and the putative bile salt efflux pump HefC play important roles in H. pylori resistance to bile salts and ceragenins.
人类胃病原体幽门螺杆菌通过糖基化修饰宿主胆固醇，并将糖基化胆固醇纳入其膜中；然而，胆固醇对幽门螺杆菌的益处在很大程度上尚不清楚。我们推测幽门螺杆菌膜中的胆固醇可能会改变这些生物体对破坏膜的抗菌化合物的敏感性。为了验证这一假设，在存在或不存在胆固醇的情况下，将幽门螺杆菌菌株在 Ham's F-12 化学定义培养基中培养。将这两种培养物分别用 10 种胆汁盐和 4 种半胱氨酸进行过夜孵育，这两种胆汁盐是新型的胆汁盐衍生物，可模拟抗菌肽的破坏膜活性。与不含胆固醇的培养物相比，用胆固醇培养的幽门螺杆菌对七种胆汁盐和三种半胱氨酸的抗性明显更高。在大多数情况下，这些胆固醇依赖性差异范围为 2 到 7 个数量级；这种幅度取决于试剂的浓度。胆固醇通过 Cgt （一种胆固醇糖苷转移酶）进行糖基化修饰。令人惊讶的是，cgt 敲除株仍然保持对胆汁盐和半胱氨酸的胆固醇依赖性抗性，表明胆固醇修饰不参与抗性。然后，我们测试了三个假定的、旁系同源的内膜外排泵 HefC、HefF 或 HefI 是否发挥作用。虽然 HefF 和 HefI 似乎不重要，但 HefC 通过多种方法在多种菌株背景下被证明在对胆汁盐和半胱氨酸的抗性中发挥关键作用。因此，胆固醇和假定的胆汁盐外排泵 HefC 都在幽门螺杆菌对胆汁盐和半胱氨酸的抗性中发挥重要作用。
