Reproduction

Inter-Tissue Gene Network Analyses of HPG, Muscle and Liver Transcriptome in IVP-Derived Bovine Calves Reveal Molecular Perturbations

---

Abstract

We have previously demonstrated that three-month-old male calves produced in vitro (IVP) exhibit molecular changes in hypothalamus, pituitary, gonadal, and adrenal organs of the HPG axis, as well as in liver and muscle, indicating disruptions in reproductive function and energy metabolism. However, the coordinated inter-tissue gene regulatory networks underlying these alterations remain undefined. This study employed multi-tissue transcriptomic profiling and weighted gene co-expression network analysis (WGCNA) on adrenal, hypothalamus, pituitary, testis, muscle, and liver samples from IVP and in vivo produced (MOET) Holstein calves (n=4 per group) to identify molecular signatures and regulatory mechanisms impacted by IVP culture conditions. We performed 100 bp paired-end RNA-Seq (~5 Gb/sample) on these six tissues collected postmortem from calves of the same breed and comparable genetic background raised under identical farm conditions. Initial alignment and gene counting detected 38,001 unique protein-coding genes across tissues. Raw counts were normalized using DESeq2 with batch effects corrected by ComBat-seq, followed by differential expression analysis. From the normalized dataset, the top 25% most variable transcripts per tissue (9,500 transcripts each) were selected for WGCNA. This identified 33 gene modules significantly associated with IVP calves (|r|≥0.60; FDR≤0.06). IVP calves displayed enhanced bioenergetic modules in muscle, liver, and adrenal enriched for oxidative phosphorylation, TCA cycle, and ribosomal functions, together with alterations in adhesion and morphogenesis modules in liver, muscle, and testis. Across all tissues, 1,456 hub genes were identified (MM >0.8; GS >0.5), of which 629 (43%) overlapped with differentially expressed genes. Inter-tissue analysis of 1,354 unique hubs revealed a 2.5-fold increase in IVP network modularity (10 vs. 4 modules), indicating fragmented cross-tissue coordination. IVP-specific transcription factor networks showed enrichment for pluripotency and immune pathways, whereas MOET networks maintained broader developmental signalling (Wnt, TGF-β, chromatin remodelling). These findings are to be considered as preliminary due to low sample size, but our initial results indicate that IVP procedures may reshape postnatal transcriptional landscapes by boosting metabolic-translation programs, reconfiguring developmental and adhesion networks, and reducing inter-tissue coordination, identifying hub genes as targets for optimizing IVP culture conditions to improve calf health and production efficiency.

Keywords: 2026

How to Cite:

Pathade, P., Rabaglino, M., Secher, J. & Kadarmideen, H., (2026) “Inter-Tissue Gene Network Analyses of HPG, Muscle and Liver Transcriptome in IVP-Derived Bovine Calves Reveal Molecular Perturbations”, World Congress on Genetics Applied to Livestock Production Digital Archive 2026(1): 2281978. doi: https://doi.org/10.31274/wcgalp.23466

Rights: 1