Browsing by Author "Dettleff, Phillip"

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    Dynamic Expression of Follicle-Stimulating Hormone and Estrogen mRNA Receptors Associated with microRNAs 34a and -let-7c in Canine Follicles during the Estrous Cycle
    (2024) De los Reyes, Monica; Dettleff, Phillip; Palomino, Jaime; Peralta, Oscar A.; Vergara, Ana
    The genes encoding for estrogen receptor (ESR2) and follicle-stimulating hormone receptor (FSHR) play crucial roles in ovarian follicular development. This study aimed to determine the expression levels of miRNAs predicted against FSHR and ESR2 mRNAs in follicular cells related to their target genes during the estrous cycle in canines. Antral follicles were dissected from 72 ovaries following ovariohysterectomies. MiRNAs regulating FSHR and ESR2 genes were selected from miRNA databases, and mature miRNA and mRNA expression profiling was performed using real-time polymerase chain reaction (PCR). The best miRNA for each target gene was selected considering the quantitative PCR (qPCR) performance and target prediction probability, selecting only miRNAs with a binding p-value of 1.0, and choosing cfa-miR-34a and cfa-let-7c for FSHR and ESR2, respectively. The expression levels comparing the different phases of the estrous cycle were evaluated using ANOVA. Pearson correlations between the expression pattern of each miRNA and their target genes were performed. Each miRNA and its target genes were expressed in the granulosa cells in all estrous phases. FSHR remained low in anestrus and proestrus, increased (p < 0.05) to the highest level in estrus, and decreased (p < 0.05) in diestrus. ESR2 showed the same trend as FSHR, with the highest (p < 0.05) expression in estrus and the lowest (p < 0.05) in anestrus and proestrus. A tendency for an inverse relationship was observed between the expression of miR-34a and FSHR only in the anestrus phase, while an inverse correlation (r = -0.8) was found between miRNA-7c and ESR2 (p < 0.01). The expression profile of miR-34a and miR-let-7c and their predicted target genes of dog ovarian follicles throughout the estrous cycle observed in this study suggest a role in the transcriptional regulation of FSHR and ESR2, which is the first evidence of the involvement of these miRNAs in the canine follicular function.
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    Embryo Buoyancy and Cell Death Gene Expression During Embryogenesis of Yellow-Tail Kingfish Seriola lalandi
    (2021) Palomino, Jaime; Gomez, Camila; Otarola, Maria Teresa; Dettleff, Phillip; Patino-Garcia, Daniel; Orellana, Renan; Moreno, Ricardo D.
    In pelagic fish, embryo buoyancy is a noteworthy aspect of the reproductive strategy, and is associated with overall quality, survival, and further developmental success. In captivity, the loss of buoyancy of early embryos correlates with high mortality that might be related to massive cell death. Therefore, the aim of this study was to evaluate under captivity conditions the expression of genes related to the apoptosis process during the early embryonic development of the pelagic fish Seriola lalandi, and its relationship to the buoyancy of embryos. The relative expression of bcl2, bax-like, casp9, casp8, and casp3 was evaluated by RT-qPCR and FasL/Fas protein levels by western blot in five development stages of embryos sorted as floating or low-floating. All genes examined were expressed in both floating and low-floating embryos up to 24 h of development. Expression of the pro-apoptotic factors bax, casp9, casp8, and casp3 was higher in low-floating as compared with floating embryos in a developmental stage-specific manner. In contrast, there was no difference in expression of bcl2 between floating and low-floating embryos. Fas protein was detected as a single band in floating embryos without changes in expression throughout development; however, in low-floating embryos, three higher intensity reactive bands were detected in the 24-h embryos. Interestingly, FasL was only detected at 24-h in floating embryos, whereas in low-floating samples this ligand was present at all stages, with a sharp increase as development progressed. Cell death, as evaluated by the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay, was highly increased in low-floating embryos as compared to floating embryos throughout all developmental stages, with the highest levels observed during the gastrula stage and at 24 h. The results of this study suggest that an increase in cell death, probably associated with the intrinsic and extrinsic apoptosis pathways, is present in low-floating embryos that might explain their lower developmental potential under captivity conditions.
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    Gills de novo assembly reveals oxidative stress, unfolded protein, and immune response on red cusk-eel ( Genypterus chilensis ) under thermal stress
    (2024) Dettleff, Phillip; Toloza, Carla; Fuentes, Marcia; Aedo, Jorge; Zuloaga, Rodrigo; Estrada, Juan Manuel; Molina, Alfredo; Valdes, Juan Antonio
    The heat waves on the South Pacific coast could lead to thermal stress in native fish. The red cusk-eel ( Genypterus chilensis ) is relevant for Chilean artisanal fisheries and aquaculture diversification. This study examined the effect of high-temperature stress in the gills of G. chilensis in control (14 degrees C) and high-temperature stress (19 degrees C) conditions. High-temperature stress induces a significant increase in gills cortisol levels. Additionally, oxidative damage was observed in gills (protein carbonylation and lipoperoxidation). RNA-seq data was used to build the first transcriptome assembly of gills in this species (23,656 annotated transcripts). A total of 1138 downregulated and 1531 up-regulated transcripts were observed in response to high-temperature stress in gills. The enrichment analysis showed immune response and replication enriched processes (on down-regulated transcripts), and processes related to the folding of proteins, endoplasmic reticulum, and transporter activity (on upregulated transcripts). The present study showed how gills could be affected by high-temperature stress.
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    High temperature induces oxidative damage, immune modulation, and atrophy in the gills and skeletal muscle of the teleost fish black cusk-eel (Genypterus maculatus)
    (2025) Becerra, Sofía; Arriagada-Solimano M.; Escobar Aguirre, Sebastián Gonzalo; Palomino, Jaime; Aedo, Jorge; Estrada, Juan Manuel; Barra Valdebenito, Verónica; Zuloaga, Rodrigo; Valdés, Juan Antonio; Dettleff, Phillip
    The high temperature associated with heat waves is a relevant abiotic factor that could impact the biology of teleost fish. The innate immune response, muscular growth, and oxidative stress status are relevant functions in fish tissues that could be affected by increased temperature. In this study, black cusk-eel (Genypterus maculatus) juveniles were subjected to increased temperature, to experimentally replicate heat waves registered from the South Pacific Ocean for five days. The results showed that thermal stress modulated the immune response in gills, with up-regulation of antibacterial peptides, pro-inflammatory cytokines, and Toll-like receptors genes, including hepcidin, gzma, tnfa, cxcl8, and tlr5, with no effect on complement system genes. In skeletal muscle, high temperature triggered atrophy-related gene expression, with up-regulation of foxo1, foxo3, fbxo32, murf1, and atg16l. Increased temperature also generated an up-regulation of transcripts encoding heat shock protein (hsp60 and hsp70) in gills and skeletal muscle, generating oxidative stress in both tissues, with increased expression of the antioxidant genes sod1 and gpx1 in gills and skeletal muscle, respectively, with oxidative damage observed at the DNA level (AP sites), protein (carbonyl content), and lipoperoxidation (HNE content) in both tissues. The present study shows that short-term increases in temperature like those observed in heat waves could affect the immune response in gills, induced atrophy in skeletal muscle, and generate oxidative stress in a teleost species important for Chilean aquaculture diversification, information relevant under the context of climate change scenario.
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    High-Temperature Stress Effect on the Red Cusk-Eel (Geypterus chilensis) Liver: Transcriptional Modulation and Oxidative Stress Damage
    (MDPI, 2022) Dettleff, Phillip; Zuloaga, Rodrigo; Fuentes, Marcia; Gonzalez, Pamela; Aedo, Jorge; Manuel Estrada, Juan; Molina, Alfredo; Antonio Valdes, Juan
    Simple Summary The red cusk-eel (Genypterus chilensis) is a native Chilean species important for aquaculture diversification in Chile. The effect of high-temperature stress on the liver, a key organ for fish metabolism, is unknown. In this study we determined for the first time the effects of high-temperature stress on the liver of red cusk-eel. The results showed that high-temperature stress increased hepatic enzyme activity in the plasma of stressed fish. Additionally, this stressor generated oxidative damage in liver, and generated a transcriptional response with 1239 down-regulated and 1339 up-regulated transcripts associated with several processes, including unfolded protein response, heat shock response and oxidative stress, among others. Together, these results indicate that high-temperature stress generates a relevant impact on liver, with should be considered for the aquaculture and fisheries industry of this species under a climate change scenario. Environmental stressors, such as temperature, are relevant factors that could generate a negative effect on several tissues in fish. A key fish species for Chilean aquaculture diversification is the red cusk-eel (Genypterus chilensis), a native fish for which knowledge on environmental stressors effects is limited. This study evaluated the effects of high-temperature stress on the liver of red cusk-eel in control (14 degrees C) and high-temperature (19 degrees C) groups using multiple approaches: determination of plasmatic hepatic enzymes (ALT, AST, and AP), oxidative damage evaluation (AP sites, lipid peroxidation, and carbonylated proteins), and RNA-seq analysis. High-temperature stress generated a significant increase in hepatic enzyme activity in plasma. In the liver, a transcriptional regulation was observed, with 1239 down-regulated and 1339 up-regulated transcripts. Additionally, high-temperature stress generated oxidative stress in the liver, with oxidative damage and transcriptional modulation of the antioxidant response. Furthermore, an unfolded protein response was observed, with several pathways enriched, as well as a heat shock response, with several heat shock proteins up regulated, suggesting candidate biomarkers (i.e., serpinh1) for thermal stress evaluation in this species. The present study shows that high-temperature stress generated a major effect on the liver of red cusk-eel, knowledge to consider for the aquaculture and fisheries of this species.
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    High-Temperature Stress Induces Autophagy in Rainbow Trout Skeletal Muscle
    (2023) Molina, Alfredo; Dettleff, Phillip; Valenzuela-Munoz, Valentina; Gallardo-Escarate, Cristian; Valdes, Juan Antonio
    Ectothermic animals, such as teleosts, have increasingly been exposed to stressful high-temperature events due to global warming. Currently, the effects of thermal stress on skeletal muscle, a key tissue for fish growth, are unknown. This study examined the impact of high-temperature stress on the skeletal muscle transcriptome of rainbow trout (Oncorhynchus mykiss) in control (15 & DEG;C) and high-temperature (20 & DEG;C) conditions. Additionally, we examined the plasmatic levels of cortisol, glucose, and creatine kinase activity, and examined oxidative damage and autophagy activation in skeletal muscle. High-temperature stress induced significant increases in cortisol and glucose plasmatic levels. Nevertheless, no changes were observed in creatine kinase activity in plasma and skeletal muscle oxidation. Skeletal muscle RNA was isolated and sequenced using the HiSeq Illumina platform. A total of 383,796,290 reads were mapped onto the reference rainbow trout genome. The transcriptomic analysis showed that 293 genes were upregulated in the high-temperature group, mainly associated with autophagosome assembly, amino acid transport, and the glutamine metabolic process. On the other hand, 119 genes were downregulated in the high-temperature group, mainly associated with digestion, proteolysis, and the muscle contraction process. In addition, RT-qPCR of differentially expressed representative genes and Western blot analysis of LC3-II/LC3-I levels confirmed skeletal muscle autophagy induced by high temperature. This study sheds light on intriguing facets of the adaptive response of rainbow trout skeletal muscle to high-temperature stress and provides significant insights into the physiology of autophagy in teleosts.
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    Thermal stress generate oxidative damage in black cusk-eel embryos (Genypterus maculatus)
    (WILEY, 2021) Dettleff, Phillip; Zuloaga, Rodrigo; Godoy, Camila; Palomino, Jaime; Molina, Alfredo; Antonio Valdes, Juan