PDF(Pubmed)
Abstract：
Staphylococcus aureus is a major causative pathogen of osteomyelitis. Intracellular infections of resident bone cells including osteocytes can persist despite gold-standard clinical intervention. The mechanisms by which intracellular S. aureus evades antibiotic therapy are unknown. In this study, we utilised an in vitro S. aureus infection model of human osteocytes to investigate whether antibiotic-mediated dysregulation of autophagy contributes to this phenomenon. Infected or non-infected osteocyte-like cells were exposed to combinations of rifampicin, vancomycin, and modulators of autophagy. Intracellular bacterial growth characteristics were assessed using colony-forming unit (CFU) analysis, viable bacterial DNA abundance, and the rate of escape into antibiotic-free medium, together with measures of autophagic flux. Rifampicin, alone or in combination with vancomycin, caused a rapid decrease in the culturability of intracellular bacteria, concomitant with stable or increased absolute bacterial DNA levels. Both antibiotics significantly inhibited autophagic flux. However, modulation of autophagic flux did not affect viable bacterial DNA levels. In summary, autophagy was shown to be a factor in the host-pathogen relationship in this model, as its modulation affected the growth state of intracellular S. aureus with respect to both their culturability and propensity to escape the intracellular niche. While rifampicin and vancomycin treatments moderately suppressed autophagic flux acutely, this did not explain the paradoxical response of antibiotic treatment in decreasing S. aureus culturability whilst failing to clear bacterial DNA and hence intracellular bacterial load. Thus, off-target effects of rifampicin and vancomycin on autophagic flux in osteocyte-like cells could not explain the persistent S. aureus infection in these cells.
摘要：
金黄色葡萄球菌是骨髓炎的主要病原体。尽管采取了金标准的临床干预措施，但包括骨细胞在内的常驻骨细胞的细胞内感染仍可持续。细胞内金黄色葡萄球菌逃避抗生素治疗的机制尚不清楚。在这项研究中,我们利用人骨细胞的金黄色葡萄球菌体外感染模型来研究抗生素介导的自噬失调是否促成了这一现象.感染或未感染的骨细胞样细胞暴露于利福平的组合，万古霉素,和自噬的调节剂。使用菌落形成单位（CFU）分析评估细胞内细菌生长特征，活的细菌DNA丰度，以及逃逸到无抗生素培养基中的速率，以及自噬通量的测量。利福平,单独或与万古霉素联合使用，导致细胞内细菌的可培养性迅速下降，伴随着稳定或增加的绝对细菌DNA水平。两种抗生素均显着抑制自噬通量。然而,自噬通量的调节不会影响活细菌DNA水平。总之,在这个模型中，自噬被证明是宿主-病原体关系中的一个因素，因为它的调节影响细胞内金黄色葡萄球菌的生长状态，就其可培养性和逃避细胞内生态位的倾向而言。虽然利福平和万古霉素治疗适度抑制自噬通量，这并不能解释抗生素治疗在降低金黄色葡萄球菌可培养性,同时未能清除细菌DNA和细胞内细菌负荷的矛盾反应.因此，利福平和万古霉素对骨细胞样细胞自噬通量的脱靶效应不能解释这些细胞中持续的金黄色葡萄球菌感染.
