Josiah Brown Poster Abstract

Archive

Thanh D. Tran
Robert L. Modlin
Rosane M. B. Teles and Robert L. Modlin
Detection of cathelicidin in different clinical forms of leprosy
STTP

Background: Leprosy exists on a spectrum where the clinical form correlates with the effectiveness of the host human response to the mycobacteria. Production of antimicrobial peptides and autophagy are important cell defense mechanisms against mycobacteria. These are correlated with the self-limiting tuberculoid leprosy (T-lep) which is characterized by a few skin lesions, granulomas, few bacilli, and type II IFN signature. On the other spectrum, there is the disseminated lepromatous leprosy (L-lep) characterized by widespread skin lesions, many bacilli, and type I IFN signature. The aims of this study are to investigate the expression of cathelicidin in the site of the disease and subsequently to evaluate the colocalization of cathelicidin with the autophagosomes in L-lep, T-lep, and reversal reaction (RR) leprosy lesions as these are key components of vitamin D dependent autophagy.

Methods: All skin samples were taken from patients at the time of diagnosis before treatment. The skin biopsy slides were stained using immunoperoxidase to detect cathelicidin (CATH), CD68 (macrophage), and IgG1 (isotype). Pictures were taken using Leica A4 software andImageJ was used to quantify expression of each marker in the immunoperoxidase staining with the program ImmunoRatio. Immunofluorescent labeling along with confocal microscopy were used to assess the specific expression of CATH, PGL-1 (M. leprae), and LC3 (autophagy marker).

Results: The CATH antibody was titrated in concentrations of 5ug/ml and 10ug/ml in human tonsil sections before use in leprosy lesions, with greater expression seen in the 10ug/ml concentration used. As shown by the CD68 counterstain, the cathelicidin and CD68 were seen in the same areas and often organized in granulomas in the T-lep and RR lesions, while in the L-lep lesions the CATH expression was weaker than RR and T-lep lesions. As determined by ImmunoRatio, the average level of CATH in the L-lep lesions was 26.24%. The average level of CATH in the T-lep lesions was 41.26%, and the average level of CATH in the RR lesions was 33.17%. The three-color immunofluorescence and confocal microscopy showed lower levels of CATH and LC3 in the L-lep lesions compared to the T-lep and RR lesions. There was colocalization of CATH and LC3 in the T-lep and RR lesions which was not seen in the L-lep lesions.

Conclusion: The induction of vitamin D dependent autophagy plays a key role in determining whether a patient develops widespread lepromatous leprosy or controlled tuberculoid leprosy. Autophagy involves the formation of autophagosomes that deliver intracellular pathogens to lysosomes for degradation, allowing for the clearance of mycobacterial infection. This suggests that activating autophagy may be a potential therapy against microbial infections such as leprosy.

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