Biology subjects

Spiliotis, M.

Publications and source records attributed to Spiliotis, M..

3 recordsLinked to original sources

bioRxiv2026

Persistence of regenerative stem cells provides a cellular basis for the parasitostatic activity of albendazole against Echinococcus multilocularis

Albendazole (ABZ) is the mainstay of chemotherapy for alveolar echinococcosis caused by the larval stage of Echinococcus multilocularis, but its activity is predominantly parasitostatic rather than parasiticidal. The cellular basis of this remarkable parasite persistence remains poorly understood. Here, we investigated the effects of sustained ABZ exposure on the germinative stem cells that drive metacestode growth and regeneration and examined whether differential beta-tubulin isoform usage could contribute to their drug susceptibility. ABZ rapidly impaired microtubule-dependent tegumental transport and caused pronounced damage to metacestode vesicles. In contrast, proliferative germinative cells (GC) persisted during prolonged and continuous ABZ exposure. Importantly, cells surviving 14 to 28 days of treatment retained developmental competence, regenerated metacestode vesicles following drug withdrawal, and, after extended exposure to 10 micromolar ABZ, remained capable of generating parasite tissue following transplantation into a permissive host. Transcriptomic and immunohistochemical analyses identified the Y200 containing beta tubulin isoform Tub2 as a major metacestode beta tubulin that is almost exclusively associated with the proliferative GC lineage. Tub2 was present in virtually all S phase cells, colocalized with established GC markers, localized to mitotic spindles, and Tub2+ cells persisted during prolonged ABZ and ABZSO exposure. Structural analysis further revealed an E198-Y200 interaction in Tub2, providing a structural correlate of the resistance-associated Y200 configuration. Together, these findings provide a cellular explanation for the predominantly parasitostatic activity of benzimidazoles against E. multilocularis. ABZ profoundly damages parasite tissue while sparing a regenerative cell compartment capable of re-establishing metacestode growth after drug withdrawal. They further identify GC-associated Tub2 as a plausible molecular contributor to this persistence and as a potential target for curative anti-Echinococcus chemotherapy.

cell biology↗

bioRxiv2025

Canonical WNT signalling governs Echinococcus metacestode development

Alveolar echinococcosis (AE) is a lethal zoonosis caused by infiltrative growth of the metacestode larva of the tapeworm Echinococcus multilocularis in host organs. We previously showed that the Echinococcus metacestode is an evolutionarily unique, broadly posteriorized tissue, leading us to hypothesize that canonical WNT (cWNT) signalling--which patterns the body axis across metazoans--might be critical for metacestode formation. Here, we report effective RNAi-mediated knockdown of the E. multilocularis {beta}-catenin gene (bcat-1), the central effector of cWNT signalling, in a primary parasite cell culture system that produces metacestode vesicles. bcat-1(RNAi) cultures were markedly impaired in vesicle formation, exhibited stem-cell hyperproliferation, and displayed disrupted muscle-fibre organisation. Genome-wide transcriptomics revealed a general anteriorization of gene expression, and in situ hybridization showed an overproduction of cells expressing head-inducing factors such as sfrp upon bcat-1 knockdown. Conversely, metacestode-specific genes--including the tegumental factors muc-1, TNFR, and antigen B--as well as the posterior marker post2b were significantly downregulated, consistent with the observed vesicle-formation defects. In situ analyses further identified anterior markers--frizzled-10, nou-darake, notum, and follistatin--that were overexpressed in bcat-1(RNAi) cultures and localized to the future anterior pole at the earliest stages of protoscolex formation. Finally, pharmacological inhibition of WNT signalling with pyrvinium pamoate caused complete loss of posterior tissue in Echinococcus protoscoleces, killed metacestode vesicles, and reduced stem-cell proliferation at nanomolar concentrations. Together, these findings establish a central role for cWNT signalling in directing Echinococcus body-axis formation and the posteriorization events driving metacestode growth within the host, providing insight into asexual parasite proliferation mediated by this biologically unique larval stage and pointing to potential targets for chemotherapy against AE. Author SummaryAlveolar echinococcosis (AE) is a lethal disease caused by the cancer-like growth of the metacestode larva of the tapeworm Echinococcus multilocularis. From a developmental perspective, the Echinococcus metacestode is an unusual biological structure and even atypical among tapeworms. Previous work indicated that metacestode formation involves re-patterning of the body axis, eliminating head structures and producing broadly posteriorized tissue. How this is controlled at the molecular and cellular levels, however, was unknown. In this study, we perturbed expression of the {beta}-catenin gene (bcat-1), a central regulator of canonical WNT signalling, using RNA interference (RNAi). bcat-1(RNAi) parasite cultures failed to generate metacestode vesicles and instead showed stem-cell hyperproliferation and muscle-cell distortion. Genes required for posteriorized metacestode tissue were downregulated, whereas genes directing head formation in adult worms (follistatin, sfrp, fz10, ndk) were upregulated, indicating a general anteriorization of the culture system. Pharmacological inhibition of WNT signalling with pyrvinium pamoate caused complete loss of posterior structures in protoscoleces, reduced stem-cell proliferation, and killed metacestode tissue. These findings identify {beta}-catenin and the canonical WNT pathway as crucial regulators of the posteriorization that underlies metacestode formation. Given that WNT signalling is deregulated in many human cancers and that small-molecule inhibitors are available, our results suggest new avenues for anti-AE drug development.

developmental biology↗

bioRxiv2019

Regulation of Hepatic MicroRNAs in Response to Early Stage Echinococcus multilocularis Egg Infection in C57BL/6 mice

In this study, we present a comprehensive analysis of the hepatic miRNA transcriptome in mice suffering from experimental primary alveolar echinococcosis (AE), a parasitic infection caused upon ingestion of Echinococcus multilocularis (E. multilocularis) eggs. At one month post-infection, infected C57BL/6 mice, along with non-infected control mice, were euthanized. Subsequently livers were collected and used for small RNA library preparation and next-generation sequencing (NGS). The most significantly dysregulated hepatic miRNAs were validated by Stem-loop RT-qPCR. We identified 28 miRNAs with significantly altered expression levels upon infection with E. multilocularis. Of these, 9 were up-regulated (fold change (FC) [≥] 1.5) and 19 were down-regulated (FC [≤] 0.66) as compared to the non-infected controls. In infected liver tissues, mmu-miR-148a-3p and mmu-miR-101b-3p were 8- and 6-fold down-regulated, respectively, and the expression of mmu-miR-22-3p was reduced by 50%, compared to non-infected liver tissue. Conversely, significantly higher hepatic levels were noted for Mus musculus (mmu)-miR-21a-5p (FC = 2.3) and mmu-miR-122-5p (FC = 1.8). Down-regulated miRNAs were highly enriched in Reactome and KEGG pathways of angiogenesis and fatty acids biosynthesis. Moreover, relative mRNA expression levels of three pro-angiogenic (VEGFA, MTOR and HIF1-) and two lipogenic (FASN and ACSL1) genes were significantly higher in livers of E. multilocularis infected mice. Lastly, we studied the issue related to functionally mature arm selection preference (5p and/or 3p) from the miRNA precursor and showed that 9 pre-miRNAs exhibited different arm selection preferences in normal versus infected liver tissues. Our study provides first evidence of miRNA involvement in liver pathogenesis during AE. Our future research will focus on the characterization of miRNA transcriptome patterns in more advanced AE-stages towards the assessment of microRNA therapy for AE, and experimentally address functional characteristics of selected features presently found.\n\nAuthor SummaryVarious infectious diseases in humans have been associated with altered expression patterns of microRNAs (miRNAs), a class of small non-coding RNAs involved in negative regulation of gene expression. Herein, we revealed that significant alterations of miRNA expression occurred in murine liver subsequently to experimental infection with Echinococcus multilocularis (E. multilocularis) eggs when compared to non-infected controls. At the early stage of murine AE, hepatic miRNAs were mainly downregulated. Respective target genes of the most extensively downregulated miRNAs were involved in angiogenesis and fatty acid synthesis. Indeed, angiogenic and lipogenic genes were found to be significantly higher expressed in E. multilocularis infected livers relative to non-infected controls. These boosted cellular pathways are advantageous for development of the E. multilocularis metacestodes, since this larval stage is not able to undertake de novo fatty acid synthesis, and angiogenesis allows the larvae to be periparasitically supplied by oxygen and nutrients and to get rid of waste products. More research on the miRNA transcriptome at more advanced infection-stages, and on the role of angiogenesis in E. multilocularis larval growth and metastasis, is required to assess the usefulness of microRNA- and anti-angiogenic therapies against E. multilocularis infection.

microbiology↗