Silva Tânia, Moreira Ana C, Nazmi Kamran, Moniz Tânia, Vale Nuno, Rangel Maria, Gomes Paula, Bolscher Jan G M, Rodrigues Pedro N, Bastos Margarida, Gomes Maria Salomé
i3S, Instituto de Investigação e Inovação em Saúde, Universidade do Porto, Porto, Portugal.
Instituto de Biologia Molecular e Celular (IBMC), Universidade do Porto, Porto, Portugal.
mSphere. 2017 Aug 30;2(4). doi: 10.1128/mSphere.00301-17. eCollection 2017 Jul-Aug.
Mycobacterial infections cause a significant burden of disease and death worldwide. Their treatment is long, toxic, costly, and increasingly prone to failure due to bacterial resistance to currently available antibiotics. New therapeutic options are thus clearly needed. Antimicrobial peptides represent an important source of new antimicrobial molecules, both for their direct activity and for their immunomodulatory potential. We have previously reported that a short version of the bovine antimicrobial peptide lactoferricin with amino acids 17 to 30 (LFcin17-30), along with its variants obtained by specific amino acid substitutions, killed in broth culture. In the present work, those peptides were tested against living inside its natural host cell, the macrophage. We found that the peptides increased the antimicrobial action of the conventional antibiotic ethambutol inside macrophages. Moreover, the d-enantiomer of the lactoferricin peptide (d-LFcin17-30) was more stable and induced significant killing of intracellular mycobacteria by itself. Interestingly, d-LFcin17-30 did not localize to -harboring phagosomes but induced the production of proinflammatory cytokines and increased the formation of lysosomes and autophagosome-like vesicles. These results lead us to conclude that d-LFcin17-30 primes macrophages for intracellular microbial digestion through phagosomal maturation and/or autophagy, culminating in mycobacterial killing. The genus comprises several pathogenic species, including , , , etc. Infections caused by these bacteria are particularly difficult to treat due to their intrinsic impermeability, low growth rate, and intracellular localization. Antimicrobial peptides are increasingly acknowledged as potential treatment tools, as they have a high spectrum of activity, low tendency to induce bacterial resistance, and immunomodulatory properties. In this study, we show that peptides derived from bovine lactoferricin (LFcin) improve the antimicrobial activity of ethambutol against growing inside macrophages. Moreover, the d-enantiomer of a short version of lactoferricin containing amino acids 17 to 30 (d-LFcin17-30) causes intramacrophagic death of by increasing the formation of lysosomes and autophagosomes. This work opens the way to the use of lactoferricin-derived peptides to treat infections caused by mycobacteria and highlights important modulatory effects of d-FLcin17-30 on macrophages, which may be useful under other conditions in which macrophage activation is needed.
分枝杆菌感染在全球范围内造成了重大的疾病负担和死亡。由于细菌对现有抗生素产生耐药性，其治疗疗程长、毒性大、成本高，且越来越容易失败。因此，显然需要新的治疗选择。抗菌肽作为新的抗菌分子的重要来源，不仅具有直接的抗菌活性，还具有免疫调节潜力。我们之前报道过，一种由17至30个氨基酸组成的牛抗菌肽乳铁蛋白素的短版本（LFcin17 - 30）及其通过特定氨基酸取代获得的变体，在肉汤培养中具有杀菌作用。在本研究中，我们测试了这些肽对寄生于其天然宿主细胞巨噬细胞内的结核分枝杆菌的作用。我们发现这些肽增强了传统抗生素乙胺丁醇在巨噬细胞内的抗菌作用。此外，乳铁蛋白素肽的d - 对映体（d - LFcin17 - 30）更稳定，并且自身就能显著杀死细胞内的分枝杆菌。有趣的是，d - LFcin17 - 30并不定位于含有结核分枝杆菌的吞噬体，而是诱导促炎细胞因子的产生，并增加溶酶体和自噬体样小泡的形成。这些结果使我们得出结论，d - LFcin17 - 30通过吞噬体成熟和/或自噬作用使巨噬细胞做好细胞内微生物消化的准备，最终导致分枝杆菌被杀死。结核分枝杆菌属包括几种致病菌种，如结核分枝杆菌、牛分枝杆菌、鸟分枝杆菌等。由这些细菌引起的感染特别难以治疗，因为它们具有内在的通透性差、生长速率低和细胞内定位的特点。抗菌肽作为潜在的治疗工具越来越受到认可，因为它们具有广泛的活性谱、低诱导细菌耐药性的倾向以及免疫调节特性。在本研究中，我们表明源自牛乳铁蛋白素（LFcin）的肽增强了乙胺丁醇对巨噬细胞内生长的结核分枝杆菌的抗菌活性。此外，一种由17至30个氨基酸组成的乳铁蛋白素短版本的d - 对映体（d - LFcin17 - 30）通过增加溶酶体和自噬体的形成导致巨噬细胞内的结核分枝杆菌死亡。这项工作为使用源自乳铁蛋白素的肽治疗分枝杆菌感染开辟了道路，并突出了d - FLcin17 - 30对巨噬细胞的重要调节作用，这在其他需要激活巨噬细胞的情况下可能是有用的。
