The role of reactive oxygen intermediates in the intracellular fate of Leptospira interrogans in the macrophages of different hosts
Authors: Dingming Wang
Jie Yan
Peili Li
Guangpeng Tang
Ming Wang
Ying Liu
Lei Zhang
Bijun Zhou
Shijun Li
Weilin Hu
Journal: PLoS ONE (2017)
Discipline: Multidisciplinary
Keywords: Leptospira; leptospirosis; macrophages; reactive oxygen species; superoxide
Affiliations: Institute of Communicable Disease Control and Prevention, Guizhou Provincial Center for Disease Control and Prevention, Guiyang, Guizhou, P.R. China., College of Animal Science, Guizhou University, Huaxi District, Guiyang, Guizhou, P.R. China., Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, Zhejiang, P.R. China., Department of Medical Microbiology and Parasitology, College of Medicine, Zhejiang University, Hangzhou, P.R. China.

Abstract:  Background: Pathogenic species of Leptospira cause leptospirosis, a global zoonotic disease. Our previous work showed that leptospires survive and replicate in human macrophages but are killed in murine macrophages. However, the mechanism responsible for the different intracellular fates of leptospires within the macrophages of different hosts remains unclear. Results: The present study demonstrates that infection with Leptospira interrogans caused significant up-regulation of reactive oxygen species (ROS) and superoxide in J774A.1 cells but did so to a lesser extent in THP-1 cells. The up-regulation of ROS and superoxide was significantly inhibited by the NADPH oxidase inhibitor apocynin. The damaged leptospires and remnants of leptospires within membrane-bound vacuoles were significantly inhibited by apocynin in J774A.1 cells but were less inhibited in THP-1 cells. In addition, apocynin significantly prevented damage to leptospires and the co-localization of L. interrogans with lysosomes in J774A.1 cells but did so to a lesser extent in THP-1 cells. Furthermore, the relative fluorescence intensity levels of intracellular leptospires and the viability of the intracellular leptospires increased in apocynin pretreated J774A.1 and THP-1 cells after 2 h of infection. Conclusions: The present study, based on our previous findings, further demonstrated that ROS contributed substantially to the bactericidal ability of mouse macrophages to kill intracellular leptospires. However, ROS did not contribute as much in human macrophages, which partially explains the different intracellular fates of L. interrogans in human and mouse macrophages. 

Retracted: No

PubPeer comments:
 Figure 6A. * Yellow boxes: The  Apocynin pretrated muJ774A.1 cells at 12h panel looks remarkably similar to the  Apocynin pretreated huTHP-1 cells at 2h panel. The similarity is not as high as expected if the same panel was inserted twice by accident. Rather, the blue and red signals are a bit different between these two panels.  , 
 Dear Dr. Bik, Thank you for bringing this issue to our attention. I have carefully checked the data after I received your comments. We performed the experiments using muJ774A.1 cell and huTHP-1 cells at the same time. We captured several images for each field of a slide of J774A.1 cells and huTHP-1 cells using the laser confocal microscope. The captured images were saved in a same file of the computer. When we prepared Figure 6A for submission, the images from J774A.1 cells and huTHP-1 cells were accidently mixed. Therefore, we inadvertently used the similar images of slide from J774A.1 cells for both cell types. When we captured the images of one field of a slide, we captured two or more images, but the images from the same field can be slightly different, because the fluorescence is quenched during the time of the image acquisition (the fluorescence gradually fades). Therefore, the problem of u201cthe blue and red signals are a bit different between these two panelsu201d was not manipulated intentionally, because they are actually two images of a same field, which were captured at different times. I believe the mix-up of the images does not affect the conclusion of this experiment and the conclusion of the study. We will contact the journal PLOS ONE to ask for advice and we would gladly correct the figure. I really appreciate your kind help to identify the u201cproblemu201d for our work. I sincerely apologize that we were not more careful and regret that the problem was not discovered during submission nor by the referees during review. Best, Shijun ,