UNASSIGNED: Schistosomiasis 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.
UNASSIGNED: In 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.
UNASSIGNED: This 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.
UNASSIGNED: The 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.

BACKGROUND: The primary pathogenic mechanism of schistosomiasis-associated liver fibrosis involves the deposition of schistosome eggs, leading to the formation of liver egg granulomas and subsequent liver fibrosis. Hepatic stellate cells are abnormally activated, resulting in excessive collagen deposition and fibrosis development. While specific long non-coding RNAs (lncRNAs) have been associated with fibrotic processes, their roles in schistosomiasis-associated liver fibrosis remain unclear.
METHODS: Our previous research indicated that downregulating the ICOSL/ICOS could partially alleviate liver fibrosis. In this study, we established a schistosomiasis infection model in C57BL/6 and ICOSL knockout (KO) mice, and the liver pathology changes were observed at various weeks postinfection (wpi) using hematoxylin and eosin and Masson\'s trichrome staining. Within the first 4 wpi, no significant liver abnormalities were observed. However, mice exhibited evident egg granulomas and fibrosis in their livers at 7 wpi. Notably, ICOSL-KO mice had significantly smaller pathological variations compared with simultaneously infected C57BL/6 mice. To investigate the impact of lncRNAs on schistosomiasis-associated liver fibrosis, quantitative real-time polymerase chain reaction (RT-qPCR) was used to monitor the dynamic changes of lncRNAs in hepatic stellate cells of infected mice.
RESULTS: The results demonstrated that lncRNA-H19, -MALAT1, -PVT1, -P21 and -GAS5 all participated in liver fibrosis formation after schistosome infection. In addition, ICOSL-KO mice exhibited significantly inhibited expression of lncRNA-H19, -MALAT1 and -PVT1 after 7 wpi. In contrast, they showed enhanced expression of lncRNA-P21 and -GAS5 compared with C57BL/6 mice, influencing liver fibrosis development. Furthermore, small interfering RNA transfection (siRNA) in JS-1 cells in vitro confirmed that lncRNA-H19, -MALAT1, and -PVT1 promoted liver fibrosis, whereas lncRNA-P21 and -GAS5 had the opposite effect on key fibrotic molecules, including α- smooth muscle actin and collagen I expression.
CONCLUSIONS: This study uncovers that ICOSL/ICOS may play a role in activating hepatic stellate cells and promoting liver fibrosis in mice infected with Schistosoma japonicum by dynamically regulating the expression of specific lncRNAs. These findings offer potential therapeutic targets for schistosomiasis-associated liver fibrosis.

Schistosomiasis is a parasitic disease characterized by liver fibrosis, a process driven by the activation of hepatic stellate cells (HSCs) and subsequent collagen production. Previous studies from our laboratory have demonstrated the ability of Schistosoma japonicum protein P40 (SjP40) to inhibit HSCs activation and exert an antifibrotic effect. In this study, we aimed to elucidate the molecular mechanism underlying the inhibitory effect of recombinant SjP40 (rSjP40) on HSCs activation. Using a cell model in which rSjP40 inhibited LX-2 cell activation, we performed RNA-seq analyses and identified ATF3 as the most significantly altered gene. Further investigation revealed that rSjP40 inhibited HSCs activation partly by suppressing ATF3 activation. Knockdown of ATF3 in mouse liver significantly alleviated S. japonicum-induced liver fibrosis. Moreover, our results indicate that ATF3 is a direct target of microRNA-494-3p, a microRNA associated with anti-liver fibrosis effects. rSjP40 was found to downregulate ATF3 expression by upregulating microRNA-494-3p in LX-2 cells. This downregulation led to the inhibition of the expression of liver fibrosis proteins α-SMA and COL1A1, ultimately alleviating liver fibrosis caused by S. japonicum.

BACKGROUND: Oncomelania hupensis is the exclusive intermediate host of Schistosoma japonicum in China. Snail control is an essential component of schistosomiasis elimination programme. With 70 years of continuous efforts, the range of O. hupensis had reduced significantly, but slowed down in last decades. A large number of levees against flooding were constructed along Yangtze River and its affiliated lakes in the middle and lower reaches, which influenced the hydrology and ecology in the alluvial plains. The purpose of this study was to assess the impact of levees on the distribution of O. hupensis in the middle and lower reaches of the Yangtze River.
METHODS: The snail habitats were digitalised by hand-held GPS system. The years for discovery and elimination of snail habitats were extracted from historical records. The accumulated snail-infested range for each habitat was calculated on the basis of annual reports. The current distribution of O. hupensis was determined by systematic and environmental sampling. The geographical distribution of levees was obtained from satellite imagery. To assess the impact of levees, the data pertaining to O. hupensis were divided into two parts: inside and outside the Yangtze River. Joinpoint regression was utilised to divide the study time span and further characterise the regression in each period. The 5-year-period moving averages of eliminated area infested by snails were calculated for the habitats inside and outside Yangtze River. The moving routes of corresponding geographical median centres were simulated in ArcGIS. Hotspot analysis was used to determine the areas with statistical significance clustering of O. hupensis density.
RESULTS: Three periods were identified according to Joinpoint regression both inside and outside Yangtze River. The area infested by O. hupensis increased in the first two periods. It decreased rapidly outside Yangtze River year over year after 1970, while that inside the Yangtze River did not change significantly. Furthermore, the latter was significantly higher than the former. It was observed that the present density of O. hupensis inside Yangtze River was lower than outside the Yangtze River. The median centre for eliminated ranges inside Yangtze River wavered between the east (lower reach) and the west (middle reach). In contrast, the median centre for eliminated ranges continuously moved from the east to the west.
CONCLUSIONS: Our findings indicated that the levees had a considerable negative impact on the distribution of O. hupensis outside Yangtze River. Some hotspots observed in the irrigation areas need a sluice system at the inlet of branch for snail control. The major distribution of O. hupensis located in Hubei might be caused by severe waterlogging. The intensive surveillance should be implemented there. The biggest two freshwater lakes, the major endemic regions historically, were identified as cold spots. The long-term impact of Three Gorges Dam on the distribution of O. hupensis in the lakes should be monitored and evaluated.

BACKGROUND: Schistosomiasis japonica represents a significant public health concern in South Asia. There is an urgent need to optimize existing schistosomiasis diagnostic techniques. This study aims to develop models for the different stages of liver fibrosis caused by Schistosoma infection utilizing ultrasound radiomics and machine learning techniques.
METHODS: From 2018 to 2022, we retrospectively collected data on 1,531 patients and 5,671 B-mode ultrasound images from the Second People\'s Hospital of Duchang City, Jiangxi Province, China. The datasets were screened based on inclusion and exclusion criteria suitable for radiomics models. Liver fibrosis due to Schistosoma infection (LFSI) was categorized into four stages: grade 0, grade 1, grade 2, and grade 3. The data were divided into six binary classification problems, such as group 1 (grade 0 vs. grade 1) and group 2 (grade 0 vs. grade 2). Key radiomic features were extracted using Pyradiomics, the Mann-Whitney U test, and the Least Absolute Shrinkage and Selection Operator (LASSO). Machine learning models were constructed using Support Vector Machine (SVM), and the contribution of different features in the model was described by applying Shapley Additive Explanations (SHAP).
RESULTS: This study ultimately included 1,388 patients and their corresponding images. A total of 851 radiomics features were extracted for each binary classification problems. Following feature selection, 18 to 76 features were retained from each groups. The area under the receiver operating characteristic curve (AUC) for the validation cohorts was 0.834 (95% CI: 0.779-0.885) for the LFSI grade 0 vs. LFSI grade 1, 0.771 (95% CI: 0.713-0.835) for LFSI grade 1 vs. LFSI grade 2, and 0.830 (95% CI: 0.762-0.885) for LFSI grade 2 vs. LFSI grade 3.
CONCLUSIONS: Machine learning models based on ultrasound radiomics are feasible for classifying different stages of liver fibrosis caused by Schistosoma infection.

OBJECTIVE: To screen differentially expressed long non-coding RNAs (lncRNAs) in the liver of mice infected with Schistosoma japonicum during the chronic pathogenic stage and identify their functions, so as to provide insights into unravelling the role of lncRNAs in S. japonicum infection-induced liver disorders.
METHODS: Twenty 6-week-old C57BL/6 mice were randomly divided into two groups, of 10 animals each group. Each mouse in the experimental group was infected with (15 ± 2) S. japonicum cercariae via the abdomen for modeling chronic S. japonicum infection in mice, and distilled water served as controls. All mice were sacrificed 70 days post-infection, and mouse liver specimens were sampled for RNA extraction and library construction. All libraries were sequenced on the Illumina NovaSeq 6000 sequencing platform. Data cleaning was performed using the fastp software, and reference genome alignment and gene expression (FPKM) calculation were performed using the HISAT2 software. Potential lncRNA sequences were predicted using the software CNIC, CPC, Pfam, and PLEK, and potential lncRNAs were screened. Differentially expressed lncRNAs were screened with the DESeq2 software and subjected to Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses to identify biological processes and metabolic pathways involved in target genes of differentially expressed lncRNAs.
RESULTS: A total of 333 potential lncRNAs were screened, and 67 were identified as differentially expressed lncRNAs, including 49 up-regulated and 18 down-regulated lncRNAs. A total of 53 target genes were predicted for differentially expressed lncRNAs. GO enrichment analysis showed that these target genes were mainly enriched in biological process and molecular function, among which Sema7a, Arrb1, and Ccl21b genes may be hub target genes for positive regulation of extracellular regulated protein kinase 1 (ERK1) and ERK2 cascades and may participate in the regulation of collagen expression. KEGG enrichment analysis showed that the target genes of differentially expressed lncRNAs were mainly enriched in cytokine-cytokine receptor interaction, viral protein interactions with cytokines and cytokine receptors, chemokine signaling pathway, and nuclear factor kappa-B (NF-κB) signaling pathway.
CONCLUSIONS: This study identifies differentially expressed lncRNAs and functional enrichment of their target genes in the liver of mice during the chronic pathogenic stage of S. japonicum infection. Up-regulated lncRNAs may affect biological processes of ERK1/2 cascades and chemokine signaling pathways via target genes Sema7a, Arrb1, and Ccl21b, thereby affecting collagen expression and inflammatory signal pathways, ultimately affecting the development of liver disorders.
［摘要］ 目的 筛选日本血吸虫感染小鼠慢性致病阶段肝脏中差异表达长非编码 RNA (long non-coding RNA, lncRNA) 并进行功能分析, 为探索 lncRNA 在日本血吸虫感染所致肝脏病变中的作用提供参考。方法 20 只 6 周龄 C57BL/6 小鼠 随机分为 2 组, 每组 10 只。实验组每只小鼠采用腹部贴片法感染 (15 ± 2) 条日本血吸虫尾蚴建立日本血吸虫慢性感染小 鼠模型, 以蒸馏水作为对照组。两组小鼠均于感染 70 d 后剖杀, 获取小鼠肝脏组织样本, 进行RNA抽提及文库构建。通 过 Illumina NovaSeq 6000 测序平台对文库进行测序, 采用 fastp 软件进行数据清洗, 使用 HISAT2 软件进行参考基因组比 对及基因表达量 (FPKM) 计算。采用 CNIC、CPC、Pfam、PLEK 软件对潜在lncRNA序列进行编码潜能预测, 筛选出潜在 lncRNA。采用 DESeq2 软件进行基因差异表达分析, 筛选出差异表达 lncRNA。通过基因本体论 (Gene Ontology, GO) 和京 都基因和基因组百科全书 (Kyoto Encyclopedia of Genes and Genomes, KEGG) 富集分析, 挖掘差异表达 lncRNA 靶基因参 与的生物学过程和代谢途径。结果 共筛选出 333 个潜在 lncRNA, 67 个鉴定为差异表达 lncRNA, 其中 49 个表达上调、18 个表达下调。差异表达 lncRNA 预测靶基因共 53 个, GO 富集分析显示这些靶基因主要富集在生物学过程和分子功 能; 其中 Sema7a、Arrb1、Ccl21b 等基因可能是细胞外调节蛋白激酶 1 (extracellular regulated protein kinase, ERK1) 和 ERK2 级联正调控生物学过程的关键靶基因, 可能参与调控胶原蛋白表达。KEGG 富集分析显示, 差异表达 lncRNA 靶基因主 要参与细胞因子-细胞因子受体相互作用、病毒蛋白与细胞因子和细胞因子受体的相互作用、趋化因子信号通路和核因 子κB (nuclear factor kappa-B, NF-κB) 通路等信号通路。结论 本研究鉴定了日本血吸虫感染小鼠慢性致病阶段肝脏中 差异表达 lncRNA 及其靶基因的功能富集, 其中上调表达的 lncRNA 可能通过调控 Sema7a、Arrb1、Ccl21b 等靶基因影响 ERK1/2 级联等生物学过程以及趋化因子信号通路等, 影响胶原蛋白表达及炎症相关信号通路, 从而影响肝脏病变发展。.

Schistosoma species are the causative agent of schistosomiasis and shows worldwide distribution. There is a great need to develop a sensitive diagnostic approach for controlling the disease. Previously, we identified large numbers of Extracellular Vesicle (EV) proteins from Schistosoma japonicum (S. japonicum), but rarely these proteins have been evaluated for their diagnostic potential. In the present study, we performed bioinformatic analyses of S. japonicum identified EV-associated proteins from the previous study and then identified Schistosoma-specific proteins with potentially secreted capability. Among them, we selected SJCHGC02838 protein, SJCHGC05593 protein, SJCHGC05668 protein and a hypothetical protein (SJHYP) to evaluate their diagnostic potential for detecting S. japonicum infection. First, we determined the expression of these four proteins at the transcript levels using qRT-PCR and revealed that all these genes showed higher expression in adult stage. Then, we cloned the full-length cDNA for each protein into a prokaryotic expression vector and successfully generated the recombinant proteins. Upon the purification of recombinant proteins, we developed an indirect ELISA method to evaluate the diagnostic potential of these purified recombinant proteins. The results showed high sensitivity for detecting Schistosoma infection. Additionally, these proteins also displayed a good potential for detecting Schistosoma infection, especially SJCHGC05668 protein at an early stage. The diagnostic potentials of these recombinant proteins were further evaluated by Western blot and comparatively analyzed by our previously developed cfDNA methods.

BACKGROUND: Schistosoma japonicum eggs lodge in the liver and induce a fibrotic granulomatous immune response in the liver of host. Galectin 3 (Gal-3) is a protein implicated in fibrosis in multiple organs. However, the pathology and molecular mechanisms promoting hepatic granuloma formation remain poorly understood.
METHODS: To investigate the effect of blocking galectin-receptor interactions by α-lactose on liver immunopathology in mice with S. japonicum infection, C57BL/6 mice were infected with S. japonicum and alpha (α)-lactose was intraperitoneally injected to block the interactions of galectins and their receptors.
RESULTS: Compared with S. japonicum-infected mice, there were significantly decreased Gal-3 mRNA and protein expression levels, decreased intensity of Gal-3 fluorescence in the liver, decreased serum ALT and AST levels, decreased egg numbers of S. japonicum in the liver section, attenuated hepatic and spleen pathology, and alleviated liver fibrosis accompanied with decreased protein expression levels of fibrosis markers [α-smooth muscle actin (α-SMA), collagen I, and collagen IV] in the liver of S. japonicum-infected mice blocked galectin-receptor interactions with hematoxylin-eosin staining, Masson\'s trichrome staining, immunohistochemistry, or Western blot analysis. Compared with S. japonicum-infected mice, blocking galectin-receptor interactions led to increased eosinophil infiltration and higher eosinophil cationic protein (ECP) expression in the liver, accompanied by increased mRNA levels of eosinophil granule proteins [ECP and eosinophil peroxidase (EPO)], IL-5, CCL11, and CCR3 in the liver and decreased mRNA levels of Gal-3 and M2 macrophage cytokines (TGF-β, IL-10, and IL-4) in the liver and spleen by using quantitative real-time reverse transcription-polymerase chain reaction. In addition, there were increased Beclin1 protein expression and protein expression ratio of LC3B-II/LC3B-I and decreased p62 protein expression and protein expression ratios of phospho-mTOR/mTOR and phospho-AKT/AKT by Western blot; increased double-labeled F4/80+/LC3B+ cells by immunofluorescence staining; increased M1 macrophage polarization in the liver of S. japonicum-infected mice blocked galectin-receptor interactions by flow cytometric analysis and immunofluorescence staining.
CONCLUSIONS: Our data found that blockage of galectin-receptor interactions downregulated Gal-3, which in turn led to reduced liver functional damage, elevated liver eosinophil recruitment, promoted macrophage autophagy through the Akt/mTOR signaling pathway, and alleviated liver pathology and fibrosis. Therefore, Gal-3 plays a pivotal role during S. japonicum infection and could be a target of pharmacologic potential for liver fibrosis induced by S. japonicum infection.

This study intends to use the basic information and blood routine of schistosomiasis patients to establish a machine learning model for predicting liver fibrosis. We collected medical records of Schistosoma japonicum patients admitted to a hospital in China from June 2019 to June 2022. The method was to screen out the key variables and six different machine learning algorithms were used to establish prediction models. Finally, the optimal model was compared based on AUC, specificity, sensitivity and other indicators for further modeling. The interpretation of the model was shown by using the SHAP package. A total of 1049 patients\' medical records were collected, and 10 key variables were screened for modeling using lasso method, including red cell distribution width-standard deviation (RDW-SD), Mean corpuscular hemoglobin concentration (MCHC), Mean corpuscular volume (MCV), hematocrit (HCT), Red blood cells, Eosinophils, Monocytes, Lymphocytes, Neutrophils, Age. Among the 6 different machine learning algorithms, LightGBM performed the best, and its AUCs in the training set and validation set were 1 and 0.818, respectively. This study established a machine learning model for predicting liver fibrosis in patients with Schistosoma japonicum. The model could help improve the early diagnosis and provide early intervention for schistosomiasis patients with liver fibrosis.

Schistosoma infection is one of the major causes of liver fibrosis. Emerging roles of hepatic progenitor cells (HPCs) in the pathogenesis of liver fibrosis have been identified. Nevertheless, the precise mechanism underlying the role of HPCs in liver fibrosis in schistosomiasis remains unclear. This study examined how autophagy in HPCs affects schistosomiasis-induced liver fibrosis by modulating exosomal miRNAs. The activation of HPCs was verified by immunohistochemistry (IHC) and immunofluorescence (IF) staining in fibrotic liver from patients and mice with Schistosoma japonicum infection. By coculturing HPCs with hepatic stellate cells (HSCs) and assessing the autophagy level in HPCs by proteomic analysis and in vitro phenotypic assays, we found that impaired autophagy degradation in these activated HPCs was mediated by lysosomal dysfunction. Blocking autophagy by the autophagy inhibitor chloroquine (CQ) significantly diminished liver fibrosis and granuloma formation in S. japonicum-infected mice. HPC-secreted extracellular vehicles (EVs) were further isolated and studied by miRNA sequencing. miR-1306-3p, miR-493-3p, and miR-34a-5p were identified, and their distribution into EVs was inhibited due to impaired autophagy in HPCs, which contributed to suppressing HSC activation. In conclusion, we showed that the altered autophagy process upon HPC activation may prevent liver fibrosis by modulating exosomal miRNA release and inhibiting HSC activation in schistosomiasis. Targeting the autophagy degradation process may be a therapeutic strategy for liver fibrosis during Schistosoma infection.