Nitric oxide and cytokine production by glial cells exposed in vitro to neuropathogenic schistosome Trichobilharzia regenti

Abstract

BackgroundHelminth neuroinfections represent a serious health problem, but host immune mechanisms in the nervous tissue often remain undiscovered. This study aims at in vitro characterization of the response of murine astrocytes and microglia exposed to Trichobilharzia regenti which is a neuropathogenic schistosome migrating through the central nervous system of vertebrate hosts. Trichobilharzia regenti infects birds and mammals in which it may cause severe neuromotor impairment. This study was focused on astrocytes and microglia as these are immunocompetent cells of the nervous tissue and their activation was recently observed in T. regenti-infected mice.ResultsPrimary astrocytes and microglia were exposed to several stimulants of T. regenti origin. Living schistosomulum-like stages caused increased secretion of IL-6 in astrocyte cultures, but no changes in nitric oxide (NO) production were noticed. Nevertheless, elevated parasite mortality was observed in these cultures. Soluble fraction of the homogenate from schistosomulum-like stages stimulated NO production by both astrocytes and microglia, and IL-6 and TNF-α secretion in astrocyte cultures. Similarly, recombinant cathepsins B1.1 and B2 triggered IL-6 and TNF-α release in astrocyte and microglia cultures, and NO production in astrocyte cultures. Stimulants had no effect on production of anti-inflammatory cytokines IL-10 or TGF-β1.ConclusionsBoth astrocytes and microglia are capable of production of NO and proinflammatory cytokines IL-6 and TNF-α following in vitro exposure to various stimulants of T. regenti origin. Astrocytes might be involved in triggering the tissue inflammation in the early phase of T. regenti infection and are proposed to participate in destruction of migrating schistosomula. However, NO is not the major factor responsible for parasite damage. Both astrocytes and microglia can be responsible for the nervous tissue pathology and maintaining the ongoing inflammation since they are a source of NO and proinflammatory cytokines which are released after exposure to parasite antigens.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-016-1869-7) contains supplementary material, which is available to authorized users.

The peripheral immune response of mice infected with a neuropathogenic schistosome

“…The lyophilized rTrCB2 was dissolved in sterile PBS and concentrated using Amicon Ultra 0.5 mL centrifugal filters (10 000 MWCO, Merck Millipore). Endotoxin contamination after this preparation is <0.03 EU/μg 23 …”

“…They are gradually surrounded by massive clusters of neutrophils, macrophages and microglia 15,20 which are presumably responsible for killing the parasites. Microglia might also promote the neuroinflammation since they secrete a significant amount of IL‐6 and tumour necrosis factor (TNF)‐α, but no IL‐10 after in vitro stimulation by TrCB2 23 . Neuroinflammation peaks 14 dpi being steadily attenuated until 28 dpi (Macháček, unpublished), but no living schistosomula are found in the CNS after 21 dpi 20 .…”

Isoforms of Cathepsin B1 in Neurotropic Schistosomula of Trichobilharzia regenti Differ in Substrate Preferences and a Highly Expressed Catalytically Inactive Paralog Binds Cystatin

“…In a previous study, active TrCB1.1 and TrCB2 were shown to trigger nitric oxide production in murine astrocyte cultures, which could be harmful to both the parasite and the host (Macháček et al, 2016). Therefore, we wanted to test whether also the TrCB1.6 paralog has any effect on the immune response of the host.…”

“…In vitro Effect of Pro-TrCB1.6wt on Murine Astrocytes, Microglia and RAW 264.7 Macrophages Primary murine astrocyte and microglia cultures were obtained from mixed glial cultures as described elsewhere (Macháček et al, 2016). These cells were used because they naturally occur and are activated in the vicinity of T. regenti schistosomula migrating in murine spinal cord (Lichtenbergová et al, 2011).…”

“…Active cathepsin B1 of Schistosoma mansoni (SmCB1) diminished the proinflamatory reaction of activated macrophages (Donnelly et al, 2009), provoked mixed Th1/Th2/Th17 immune response in mice and induced a transient T-helper 17 response in acute schistosome infection (Soloviova et al, 2019), suggesting it has a considerable immunogenic capacity. As for T. regenti, an inflammationpromoting activity was noticed in case of active TrCB1.1 which triggered the production of nitric oxide and proinflammatory cytokines interleukin-6 and tumor necrosis factor α by astrocytes and/or microglia (Macháček et al, 2016), i.e., the cells activated in mice infected experimentally by T. regenti (Lichtenbergová et al, 2011). However, no specific immunomodulatory mechanisms were tested so far with inactive peptidase paralogs of helminths.…”