Moore, D. P. | Search bioRxiv

bioRxiv2025

Different kinetics of humoral response against individual antigens in the ovine model of Toxoplasma gondii infection and its pathological association

BackgroundToxoplasma gondii is a globally distributed protozoan parasite with major implications for human and animal health. In sheep, infection can cause reproductive failure and represents an important source of zoonotic transmission through contaminated meat. While cell-mediated immunity is the main protective mechanism, the contribution of humoral responses, particularly against individual recombinant antigens (rAGs), remains poorly characterized in ovine toxoplasmosis. MethodsWe analyzed the humoral response of lambs experimentally infected with two reference strains of T. gondii (RH and Me49) using different inoculation doses. Recombinant antigens rGra4Gra7, rGra8, rRop2, rCST9, and rBCLA were tested by IgG-ELISA to evaluate their kinetics during infection. Histopathological analyses of brain tissues were performed to assess lesion severity and cyst presence. Additionally, sera from naturally infected sheep were evaluated to test diagnostic performance. ResultsAn early IgG response against rGra4Gra7 and rGra8 was detected, consistent with markers of acute infections. Anti-rBCLA, rRop2, and rCST9 IgG responses were associated with the chronic phase and correlated with cyst burden. Notably, stronger anti-rGra4Gra7 IgG responses tended to associate with lower lesion scores, suggesting a potential protective role. In naturally infected sheep, IgG-ELISA against rGra4Gra7 and rGra8 demonstrated the highest diagnostic performance, outperforming total lysate antigen (TLA)-based assays. ConclusionOur findings highlight distinct kinetics of specific antibody responses against selected T. gondii antigens in sheep, suggesting their value as markers of infection stage and pathology. rGra4Gra7 and rGra8 appear particularly promising both as diagnostic candidates and for exploring mechanisms of humoral modulation in ovine toxoplasmosis.

pathology↗

bioRxiv2025

Advanced human iPSC-based modelling of LMNA-related congenital muscular dystrophy enables development of targeted genetic therapies for muscle laminopathies

LMNA-related congenital muscular dystrophy (L-CMD) is amongst the most severe forms of laminopathies, which are diseases caused by pathogenic variants in the LMNA gene. LMNA encodes the proteins Lamin A and C, which assemble with Lamin B1 and B2 to form the nuclear lamina: a meshwork providing structural stability to the nucleus that also regulates chromatin organisation and gene expression. Research into L-CMD mechanisms and therapies is hindered by lack of humanised, tissue-specific models that accurately recapitulate disease phenotypes. We previously reported that LMNA-mutant induced pluripotent stem cell (iPSC)-derived skeletal muscle cells have nuclear shape abnormalities and Lamin A/C protein mislocalisation. Here, we expand the selection of L-CMD patient- derived iPSCs and validate disease-associated readouts using a transgene-free based protocol which more accurately mimics skeletal myogenesis. Results showed no overt defects in developmental myogenesis, but recapitulation of pathological nuclear shape abnormalities in 2D and 3D cultures, nuclear envelope protein mislocalisation and transcriptomic alterations across multiple pathogenic LMNA variants. We then utilised this platform to assess LMNA gene editing strategies. CRISPR-based exon removal generated stable RNA and protein Lamin A/C species, without significant normalisation of nuclear morphological phenotypes or transcriptomic profile. Conversely, precise editing of the same mutation showed complete reversal of disease-associated nuclear morphometrics, alongside normalisation of the pro-inflammatory transcriptomic signature. Our data provide the foundation for a humanised in vitro disease and therapy modelling platform for this complex and severe muscle disorder. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=81 SRC="FIGDIR/small/660928v2_ufig1.gif" ALT="Figure 1"> View larger version (15K): org.highwire.dtl.DTLVardef@134276forg.highwire.dtl.DTLVardef@108f44forg.highwire.dtl.DTLVardef@dfa04eorg.highwire.dtl.DTLVardef@1956a90_HPS_FORMAT_FIGEXP M_FIG C_FIG HIGHLIGHTSO_LILMNA-mutant iPSCs undergo efficient skeletal myogenesis upon transgene-free, small molecule-based lineage-directed differentiation C_LIO_LIL-CMD iPSCs recapitulate hallmark disease-associated nuclear phenotypes and show a pro-inflammatory transcriptional profile C_LIO_LIDisease modelling platforms based on iPSC-derived skeletal muscle cells enable comparative testing of gene editing strategies C_LIO_LICRISPR-edited L-CMD iPSC-derived myogenic cells show amelioration of disease-associated readouts C_LI

cell biology↗