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Schistosome eggs stimulate reactive oxygen species production to enhance M2 macrophage differentiation and promote hepatic pathology in schistosomiasis.

ABSTRACT: Schistosomiasis is a neglected tropical disease of public health concern. The most devastating pathology in schistosomiasis japonica and mansoni is mainly attributed to the egg-induced granulomatous response and secondary fibrosis in host liver, which may lead to portal hypertension or even death of the host. Schistosome eggs induce M2 macrophages-rich granulomas and these M2 macrophages play critical roles in the maintenance of granuloma and subsequent fibrosis. Reactive oxygen species (ROS), which are highly produced by stimulated macrophages during infection and necessary for the differentiation of M2 macrophages, are massively distributed around deposited eggs in the liver. However, whether ROS are induced by schistosome eggs to subsequently promote M2 macrophage differentiation, and the possible underlying mechanisms as well, remain to be clarified during S. japonicum infection. Herein, we observed that extensive expression of ROS in the liver of S. japonicum-infected mice. Injection of ROS inhibitor in infected mice resulted in reduced hepatic granulomatous responses and fibrosis. Further investigations revealed that inhibition of ROS production in S. japonicum-infected mice reduces the differentiation of M2, accompanied by increased M1 macrophage differentiation. Finally, we proved that S. japonicum egg antigens (SEA) induce a high level of ROS production via both nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2 (NOX2) and mitochondria in macrophages. Our study may help to better understand the mechanism of schistosomiasis japonica-induced hepatic pathology and contribute to the development of potential therapeutic strategies by interfering with ROS production.

SUBMITTER: Yu Y

PROVIDER: S-EPMC8389433 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundHepatic granuloma formation and fibrosis as the consequence of tissue entrapped eggs produced by female schistosomes characterize the pathology of Schistosoma japonicum infection. It has been proposed that fucoidan, a sulfated polysaccharide existing naturally in brown seaweed Fucus vesiculosus, plays a diversified role to perform immunomodulatory activities. However, whether fucoidan functions in the host hepatic pathology is unknown and identifying the potential mechanism that is responsible for hepatic improvement is still necessary.MethodsWe evaluated the hepatic pathology from S. japonicum-infected mice after treatment with fucoidan. qRT-PCR and immunofluorescence were used to detect the pro- or anti-inflammatory factors and the phosphorylated p65 in the livers. In addition, flow cytometry was also performed to investigate the T cell subsets in the S. japonicum-infected mice after treatment with fucoidan, and functional molecules relatively specific to Treg cells were detected in vitro. Furthermore, macrophages were treated with fucoidan in vitro and to detect the inflammatory cytokines.ResultsTreatment with fucoidan significantly reduced the hepatic granuloma size and fibrosis response during S. japonicum infection. The attenuated phospho-p65 protein levels and the mRNA levels of pro-inflammatory cytokines (IL-6, IL-12 and TNF-α) were observed in the livers from fucoidan-treated S. japonicum-infected mice; however, the mRNA levels of anti-inflammatory cytokines (IL-4 and IL-13) were increased. In addition, the infiltration of Treg cells was significantly enhanced both in the livers and spleens from fucoidan-treated S. japonicum-infected mice. Consistent with this, the mRNA levels of IL-10 and TGF-β were dramatically increased in the livers from S. japonicum-infected mice after fucoidan treatment. Furthermore, in vitro stimulated splenocytes with fucoidan resulted in increasing Treg cells in splenocytes as well as the functional expression of CC chemokine receptor type 4 (CCR4) and CXC chemokine receptor type 5 (CXCR5) in Treg cells. Additionally, fucoidan promoted the mRNA levels of IL-4 and IL-13 in macrophages.ConclusionsThese findings suggest an important role of natural fucoidan in reducing hepatic pathology in the progress of S. japonicum infection with a stronger Treg response, which may reveal a new potential therapeutic strategy for hepatic disease caused by parasitic chronic infection.

Project description:BackgroundSchistosomiasis is a zoonotic parasitic disorder induced by the infestation of schistosomes, a genus of trematodes. MicroRNAs (miRNAs) in egg-derived exosomes are crucial for modulating the host's immune responses and orchestrating the pathophysiological mechanisms. Although the exosomes secreted by S. japonicum contain abundant miRNAs, the specific roles of these miRNAs in the pathogenesis of schistosomiasis-induced hepatic fibrosis are yet to be comprehensively elucidated. The egg exosomes of S. japonicum secrete miRNA-30, a novel miRNA.MethodsIn vitro, the effect of miRNA-30 was evaluated by transfecting HSCs with miRNA mimics. The target gene biosignature for miRNA-30 was predicted using the miRDB software. The effect of miRNA-30 in hepatic fibrosis was evaluated by either elevating its expression in healthy mice or by inhibiting its activity in infected mice by administration of recombinant adeno-associated virus serotype eight vectors expressing miRNA-30 or miRNA sponges.ResultsThis novel miRNA can activate hepatic stellate cells (HSCs), the prinary effector cells of hepatic fibrosis, in vitro, i.e., it significantly increases the fibrogenic factors Col1(α1), Col3(α1), and α-SMA at both mRNA and protein levels. In addition, miRNA-30 may activate HSCs by targeting the host RORA gene. In addition, in vivo experiments were conducted by administering a recombinant adeno-associated viral vector to modulate the expression levels of miRNA-30. The overexpression of miRNA-30 in healthy mice significantly elevated the expression of Col1(α1), Col3(α1), and α-SMA at both the transcriptomic and proteomic scales. This overexpression was coupled with a pronounced augmentation in the hepatic hydroxyproline content. Conversely, the in vivo silencing of miRNA-30 in infected mice induced a considerable reduction in the size of hepatic granulomas and areas of collagen deposition. Hence, in vivo, modulation of miRNA-30 expression may play a pivotal role in ameliorating the severity of hepatic fibrosis in mice afflicted with S. japonica.ConclusionsThe study results suggest that miRNA-30 may augment schistosomiasis-induced hepatic fibrosis through a probable interaction with the host RORA. Our study may improve the current theoretical framework regarding cross-species regulation by miRNAs of hepatic fibrosis in schistosomiasis.

Project description:BACKGROUND: Aedes aegypti Linnaeus is a peridomestic mosquito that lays desiccation-resistant eggs in water-filled human-made containers. Previous investigations connected egg hatching with declining dissolved oxygen (DO) that is associated with bacterial growth. However, past studies failed to uncouple DO from other potential stimulatory factors and they contained little quantitative information about the microbial community; consequently, a direct role for bacteria or compounds associated with bacteria in stimulating egg hatching cannot be dismissed. METHODOLOGY/PRINCIPAL FINDINGS: Environmental factors stimulating hatch of Ae. aegypti eggs were investigated using non-sterile and sterile white oak leaf (WOL) infusions and a bacterial culture composed of a mix of 14 species originally isolated from bamboo leaf infusion. In WOL infusion with active microbes, 92.4% of eggs hatched in 2-h at an average DO concentration of 2.4 ppm. A 24-h old bacterial culture with a DO concentration of 0.73 ppm also stimulated 95.2% of eggs hatch within 1-h. In contrast, only 4.0% of eggs hatched in sterile infusion, whose DO averaged 7.4 ppm. Effects of bacteria were uncoupled from DO by exposing eggs to bacterial cells suspended in NaCl solution. Over a 4-h exposure period, 93.8% of eggs hatched while DO concentration changed minimally from 7.62 to 7.50 ppm. Removal of bacteria by ultra-filtration and cell-free filtrate resulted in only 52.0% of eggs hatching after 4-h at an average DO concentration of 5.5 ppm. CONCLUSIONS/SIGNIFICANCE: Collectively, the results provide compelling evidence that bacteria or water-soluble compounds secreted by bacteria, not just low DO concentration, stimulate hatching of Ae. aegypti eggs. However, the specific cues involved remain to be identified. These research findings contribute new insight into an important aspect of the oviposition biology of Ae. aegypti, a virus vector of global importance, providing the basis for a new paradigm of environmental factors involved in egg hatching.

Project description:The schistosome egg represents an attractive developmental stage at which to target transgenes because of the high ratio of germ to somatic cells, because the transgene might be propagated and amplified by infecting snails with the miracidia hatched from treated eggs, and because eggs can be readily obtained from experimentally infected rodents.We investigated the utility of square wave electroporation to deliver transgenes and other macromolecules including fluorescent (Cy3) short interference (si) RNA molecules, messenger RNAs, and virions into eggs of Schistosoma mansoni. First, eggs were incubated in Cy3-labeled siRNA with and without square wave electroporation. Cy3-signals were detected by fluorescence microscopy in eggs and miracidia hatched from treated eggs. Second, electroporation was employed to introduce mRNA encoding firefly luciferase into eggs. Luciferase activity was detected three hours later, whereas luciferase was not evident in eggs soaked in the mRNA. Third, schistosome eggs were exposed to Moloney murine leukemia virus virions (MLV) pseudotyped with vesicular stomatitis virus glycoprotein (VSVG). Proviral transgenes were detected by PCR in genomic DNA from miracidia hatched from virion-exposed eggs, indicating the presence of transgenes in larval schistosomes that had been either soaked or electroporated. However, quantitative PCR (qPCR) analysis determined that electroporation of virions resulted in 2-3 times as many copies of provirus in these schistosomes compared to soaking alone. In addition, relative qPCR indicated a copy number for the proviral luciferase transgene of approximately 20 copies for 100 copies of a representative single copy endogenous gene (encoding cathepsin D).Square wave electroporation facilitates introduction of transgenes into the schistosome egg. Electroporation was more effective for the transduction of eggs with pseudotyped MLV than simply soaking the eggs in virions. These findings underscore the potential of targeting the schistosome egg for germ line transgenesis.

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Schistosome eggs stimulate reactive oxygen species production to enhance M2 macrophage differentiation and promote hepatic pathology in schistosomiasis.

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