Inflammatory Signatures of Pathogenic and Non-Pathogenic Leptospira Infection in Susceptible C3H-HeJ Mice

Abstract

The exact global impact of leptospirosis is unknown due to inadequate surveillance systems in place in most low-income countries. In this study, we analyzed the differences in mouse inflammatory signatures involved in pathogenic versus non-pathogenic Leptospira recognition at 24h and 72h post infection. Injection of C3H-HeJ mice with non-pathogenic L. biflexa increased circulation of a few chemokines (5/21, 24%) without secretion of cytokines in blood that resulted in engagement of resident macrophages, dendritic cells, neutrophils and NK cells without engagement of T cells. In contrast, pathogenic L. interrogans induced circulation of a much higher panel of chemokines (18/21, 86%) and pro- and anti-inflammatory cytokines (11/19, 58%) in blood with a resulting signaling cascade leading to engagement of macrophages, dendritic cells, monocytes, NK cells and T cells without engagement of neutrophils. Although neutrophils do not appear to be engaged, a considerable number of chemokines that recruit other granulocytes such as eosinophils and basophils were also increased at 72h post infection with L. interrogans. Overall, the data suggest that prevention of dissemination of L. biflexa is associated with an early engagement of the innate immune response characterized by upregulation of a few chemokines that results in an efficacious phagocytic response without an overwhelming increase of pro-inflammatory cytokines. However, when macrophages fail to clear a pathogenic serovar such as L. interrogans, the adaptive response (T cells) is engaged to help out, but the resulting chemo-cytokine storm mediates a robust but non-resolving inflammatory response to pathogenic Leptospira that results in dissemination, kidney colonization, pathology and disease.

Necroptosis Contributes to Persistent Inflammation During Acute Leptospirosis

“…Our recent work supports that pathogenic Leptospira induce splenomegaly by recruiting large numbers of immune cells in this secondary lymphoid organ and activating the chemo-cytokine response ( 2 ). Understanding the fate of those immune cells in spleen can further lead to understanding persistent inflammation in the host during leptospirosis.…”

“…We evaluated the interactions of increased chemo-cytokines present in serum 72h post infection with L. interrogans from our previous study ( 2 ) with the major necroptotic markers MLKL, RIP3 and RIP1 elevated in the spleen at 72h post infection in the present study through STRING proteome analysis ( Figure 7 ). This analysis informs on known and predicted protein-protein interactions as well as direct physical and indirect functional interactions.…”

“…Spleens were collected from different groups of mice after euthanasia and processed immediately for flow cytometry staining. Briefly, spleens were chopped and teared with frosted slides to prepare single cell suspensions and RBC lysis was performed as described previously ( 2 ). Live/dead cell stain was performed, and cells were counted using a Luna counter (Logos Biosystems, South Korea).…”

“…A total of 17 query proteins out of which 14 were increased chemo-cytokines in serum from our previous study ( 2 ) and 3 are elevated key necroptotic proteins from the present study at 72 h post infection with L. interrogans were used to generate a STRING proteome map using the online database (string-db.org). In these networks, the nodes are the proteins and the edges indicate the interactions between proteins as they were present in STRING (level of confidence >0.400).…”

“…Leptospirosis, a neglected emerging disease with worldwide distribution that affects virtually all vertebrates is caused by pathogenic Leptospira spp and is associated with robust inflammation orchestrated by chemo-cytokine release, recruitment of different immune cells as well as cellular processes like phagocytosis and NET/MET formation ( 1 , 2 ). Human leptospirosis ranges in severity from a mild, self-limited febrile illness to a fulminant life-threatening disease ( 3 ).…”