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Transcriptome analysis on leukocytes in heat-stressed Holstein calves treated with dexamethasone

ABSTRACT: Heat stress phenotype is mediated by inflammation and dexamethasone is a synthetic glucocorticoid to reduce inflammation in dairy cattle. A study was conducted with 16 Holstein bull calves of 3.5 ± 0.5 weeks age and 68.5 ± 1.37 kg BW, to assess dexamethasone induced differentially expressed genes (DEGs) and pathways in heat-stressed dairy calves. Calves were individually housed in climate-controlled room with an ambient temperature (AT) of 19.8 ± 0.8 °C for a 5d acclimatization period. Based on their BW, ADG and DMI, calves were randomly assigned to one of the two treatments (n=8) for 5 d as (1) heat stress (HS; AT of 40 °C for 12 h followed by 27 °C) and (2) heat stress dexamethasone (HSD; AT of 40 °C for 12 h followed by 27 °C).The HSD group received dexamethasone at a dose of 0.05 mg/kg BW and HS group received saline at a dose of 1.2 mL via intramuscular injection on d 1 and 3 at 0700 h. Cyclic heat stress was imposed on HS and HSD groups from 0800-1900 h. Blood samples were collected on 0, 12, and 108 h of the study for RNA extraction, sequencing and transcriptomics analysis. DEGs were called using a linear mixed effects model using the dream method from the variancePartition R package with combinations of treatment and timepoint as a fixed effect, and animal as random effect. Significance was declared at the level of FDR adjusted P < 0.05. The comparison between HSD vs HS treatments showed a total of 4855 DEGs (adj-P < 0.05) from which 2533 were upregulated and 2322 were downregulated. In summary, transcriptomics results showed that the HSD treatment induced downregulation of genes associated with cholesterol biosynthesis, cell cycle arrest, neutrophils degranulation, and genes related to the process of transcription and translation which may ameliorate the negative effects of heat stress in dairy calves’ health and productivity.

ORGANISM(S): Bos taurus

PROVIDER: GSE326757 | GEO | 2026/08/31

REPOSITORIES: GEO

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Project description:BACKGROUND: Large-scale farms struggle to effectively address summer heat stress (HS) in dairy goats, which impairs spermatogenesis and reduces reproductive performance by compromising gut and reproductive organ health in male animals. Although Clostridium butyricum (CB) and Enterococcus faecium (EF) have been used to alleviate gut dysbiosis, their protective effects and mechanisms against HS-induced spermatogenic dysfunction in dairy goats remain unclear. This study aims to evaluate the efficacy of CB and EF in mitigating HS-induced spermatogenic disorders in male dairy goats and elucidate the underlying mechanisms.RESULTS: HS leads to a significant decline in sperm quality. CB and EF ameliorate HS-induced sperm quality deterioration by reducing pro-inflammatory factors and oxidative stress levels in blood, while modulating the disrupted reproductive hormone levels caused by heat stress. 16S rRNA-based microbiota analysis revealed that CB and EF treatment shifted the gut microbiota of HS-treated male dairy goats toward a composition resembling that of the NC. To investigate whether the protective effects of these probiotics on sperm quality are mediated through gut microbiota modulation, we transplanted fecal microbiota from HS-treated dairy goats (with or without probiotic intervention) into antibiotic-induced microbiome-depleted (AIMD) mice, followed by heat stress exposure. We observed severe testicular damage and spermatogenic impairment in mice receiving HS group microbiota, whereas these pathological manifestations were significantly alleviated in mice receiving microbiota from probiotic-treated HS goats, indicating that CB and EF mitigate HS-induced spermatogenic damage in a microbiota-dependent manner. Furthermore, non-targeted metabolomics sequencing identified significant reductions in arginine (Arg) and pantothenic acid (PaA) levels in both HS-treated goats and HS microbiota-transplanted mice, metabolites that showed correlation with sperm quality parameters. Subsequent studies demonstrated that supplementation with Arg and PaA effectively attenuated the elevation of pro-inflammatory factors and oxidative stress in mouse blood, restored intercellular junctions, and ultimately ameliorated spermatogenic disorders.CONCLUSION: This study demonstrates that CB and EF alleviate HS-induced sperm quality decline in dairy goats by modulating the gut microbiota to increase blood levels of Arg and PaA. These findings highlight novel therapeutic avenues for preventing HS-induced spermatogenic disorders in dairy goats and provide fresh insights into the application of probiotics to mitigate heat stress-associated impairments in sperm production.

Project description:Heat stress (HS) poses a major challenge to the dairy industry by reducing milk production, yet its cell type-specific effects in the bovine mammary gland remain incompletely defined. In this study, we recorded production traits and collected mammary biopsies from cows under thermoneutral (TN), HS, and pair-fed (PF) conditions.We applied single-nucleus RNA-seq (snRNA-seq) on mammary biopsies to generate cell-resolved HS responses. We identified 14 distinct cell clusters, including epithelial, immune, and stromal populations. Under the TN condition, casein genes (e.g., CSN1S1, CSN2) were broadly expressed across luminal cells but were attenuated under HS, whereas luminal alveolar cells showed upregulation of casein genes. Heat shock protein genes were strongly induced by HS, primarily in epithelial clusters. Gene-set enrichment analyses revealed increased ribosomal activities across HS-responsive clusters and enrichment of protein folding and metabolic pathways in luminal alveolar cells, suggesting elevated proteostasis demands under stress. Pseudotime analysis positioned luminal cells along a progenitor-to-secretory trajectory under TN, accompanied by increased casein gene expression, whereas under HS, mature luminal cells shifted toward a homeostasis regulatory state. Cell-cell communication analysis demonstrated HS-induced remodeling of interepithelial signaling, including altered ERBB4-mediated signaling from luminal hormone-sensing to alveolar lineages. Finally, transcription factor activity profiling highlighted cell-type-specific HS-activated regulators and their downstream target genes. Together, this cell type-resolved atlas delineates how HS alters bovine mammary epithelial function, developmental state, and intercellular crosstalk. These findings point to proteostasis pressure, disrupted signaling pathways, and rewired regulatory networks as mechanistic contributors to reduced lactational performance under HS, offering insights for improving heat resilience in dairy cattle.

Project description:Heat stress (HS) can damage the integrity of the intestinal mucosal barrier, leading to decreased poultry productivity. This study aimed to identify candidate genes related to acute HS in breeder hens and provide insight into the molecular mechanisms underlying acute HS in gut health. Fifty 28-week-old breeder hens were divided into two groups (25 hens each) raised under thermoneutral zone (23 °C) and acute HS (36 °C, 6 hours) conditions. The heart rate and cloacal temperature were measured in all hens, and jejunal mucosa tissues were randomly collected from 12 hens per treatment for RNA-sequencing analysis. The results indicated a significant increase in the heart rate and cloacal temperature in hens exposed to acute HS (P < 0.05). Transcriptome analysis identified 138 differentially expressed genes (DEGs) in heat-stressed breeder hens, including 75 upregulated DEGs containing heat shock protein (HSP), energy homeostasis metabolism-related gene (PDK4), and fat metabolism-related genes (PPARA and CD36), and 63 downregulated DEGs containing the bile acid transporter gene (SLC10A2). Gene ontology analysis revealed significant enrichment in biological processes related to heat response and cholesterol biosynthesis. Kyoto Encyclopedia of Genes and Genomes analysis highlighted several significant pathways, including steroid biosynthesis, steroid hormone biosynthesis, protein processing in endoplasmic reticulum, the peroxisome proliferator-activated receptor (PPAR) signaling pathway, and the adipocytokine signaling pathway. Protein-protein interaction network analysis involving two large networks: one containing several upregulated HSPs and genes related to energy homeostasis and fat metabolism (PDK4, PPARA, and CD36) and glucose transporter (SLC2A5), and the other containing downregulated DEGs related to cholesterol biosynthesis. Overall, acute HS might affect energy metabolism, fat metabolism, and glucose transport in the jejunal mucosa of breeder hens. Heat-stressed hens could restore the nutritional function of the jejunal mucosa by increasing the expression of HSPs. These findings provide a theoretical framework for further investigation into the molecular regulatory mechanisms responsible for HS-induced changes in the gut health of poultry.

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Transcriptome analysis on leukocytes in heat-stressed Holstein calves treated with dexamethasone

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