Embryonic heat conditioning in chicks induces transgenerational heat/immunological resilience via methylation on regulatory elements.

The question of whether behavioral traits are heritable is under debate. An obstacle in demonstrating transgenerational inheritance in mammals originates from the maternal environment's effect on offspring phenotype. Here, we used in ovo embryonic heat conditioning (EHC) of first-generation chicks, demonstrating heredity of both heat and immunological resilience, confirmed by a reduced fibril response in their untreated offspring to either heat or LPS challenge. Concordantly, transcriptome analysis confirmed that EHC induces changes in gene expression in the anterior preoptic hypothalamus (APH) that contribute to these phenotypes in the offspring. To study the association between epigenetic mechanisms and trait heritability, DNA-methylation patterns in the APH of offspring of control versus EHC fathers were evaluated. Genome-wide analysis revealed thousands of differentially methylated sites (DMSs), which were highly enriched in enhancers and CCCTC-binding factor (CTCF) sites. Overlap analysis revealed 110 differentially expressed genes that were associated with altered methylation, predominantly on enhancers. Gene-ontology analysis shows pathways associated with immune response, chaperone-mediated protein folding, and stress response. For the proof of concept, we focused on HSP25 and SOCS3, modulators of heat and immune responses, respectively. Chromosome conformational capture (3C) assay identified interactions between their promoters and methylated enhancers, with the strongest frequency on CTCF binding sites. Furthermore, gene expression corresponded with the differential methylation patterns, and presented increased CTCF binding in both hyper- and hypomethylated DMSs. Collectively, we demonstrate that EHC induces transgenerational thermal and immunological resilience traits. We propose that one of the mechanisms underlying inheritance depends on three-dimensional (3D) chromatin reorganization.

Swimming in warm water is ineffective in heat acclimation and is non-ergogenic for swimmers.

Heat acclimation (HA) in air confers adaptations that improve exercise capabilities in hot and possibly temperate air. Swimmers may benefit from HA, yet immersion may constrain adaptation. Therefore, we examined whether warm-water swimming constitutes effective HA. In a randomized-crossover study, eight male swimmers swam 60 min/day on 7 days in 33 °C (HA) or 28 °C (CON) water. They performed 20-min distance trials before and after each regime: in 33 °C water (Warm); 28 °C water (Temperate); and cycling in 29 °C air (Terrestrial) following standardized exercise. Rectal temperature (Tre ) rose ∼ 1 °C in HA sessions, and sweat loss averaged 1.4 L/h. After accounting for CON, HA did not confer any clear expansion of plasma volume [1.9% (95% CI: 7.7)], reduction in heart rate during standardized cycling exercise [1 b/min (9)], reduction in Tre during rest [+0.1 °C (0.1)] or exercise, or change in sudomotor function. Only perceived temperature and discomfort tended to improve. Performance was clearly not improved for Warm [+0.3% (1.8)] or Temperate [+0.3% (1.9)], was unclear for Terrestrial [+0.4% (17.7)], and was unrelated to changes in resting plasma volume (r < 0.3). In conclusion, short-term HA using swimming in 33 °C water confers little adaptation and is not ergogenic for warm or temperate conditions.

Curing and low-temperature combined post-harvest storage enhances anthocyanin biosynthesis in blood oranges.

Anthocyanins are pigments present in blood oranges which can be enriched by post-harvest cold storage. Additionally, citrus fruits contain appreciable levels of other flavonoids, whose content increases under post-harvest heat treatments. Here, we investigated the effects of curing (37 °C for 3 days) and storage at low-temperature (9 °C) during 15, 30 and 45 days on accumulation of anthocyanins and other flavonoids in Moro and Sanguinelli Polidori blood oranges (Citrus sinensis L. Osbeck). Cured fruits reached up to 191.4 ± 1.4 mg/L of anthocyanins in their juice after cold storage and a 3-fold enrichment of other flavonoids such as flavones and flavanones, compared to 85.7 ± 3.3 mg/L anthocyanins from fruits with cold storage alone. Concomitantly, qPCR analysis showed that curing enhanced upregulation of the main structural and transcription factor genes regulating the flavonoid pathway. GC-MS analysis showed that no unpleasant compounds were generated in the cured plus cold-stored juice volatilome.

Embryonic Heat Conditioning Induces TET-Dependent Cross-Tolerance to Hypothalamic Inflammation Later in Life.

Early life encounters with stress can lead to long-lasting beneficial alterations in the response to various stressors, known as cross-tolerance. Embryonic heat conditioning (EHC) of chicks was previously shown to mediate resilience to heat stress later in life. Here we demonstrate that EHC can induce cross-tolerance with the immune system, attenuating hypothalamic inflammation. Inflammation in EHC chicks was manifested, following lipopolysaccharide (LPS) challenge on day 10 post-hatch, by reduced febrile response and reduced expression of LITAF and NFκB compared to controls, as well as nuclear localization and activation of NFκB in the hypothalamus. Since the cross-tolerance effect was long-lasting, we assumed that epigenetic mechanisms are involved. We focused on the role of ten-eleven translocation (TET) family enzymes, which are the mediators of active CpG demethylation. Here, TET transcription during early life stress was found to be necessary for stress resilience later in life. The expression of the TET family enzymes in the midbrain during conditioning increased in parallel to an elevation in concentration of their cofactor α-ketoglutarate. In-ovo inhibition of TET activity during EHC, by the α-ketoglutarate inhibitor bis-2-(5-phenylacetamido-1,3,4-thiadiazol-2-yl) ethyl sulfide (BPTES), resulted in reduced total and locus specific CpG demethylation in 10-day-old chicks and reversed both thermal and inflammatory resilience. In addition, EHC attenuated the elevation in expression of the stress markers HSP70, CRHR1, and CRHR2, during heat challenge on day 10 post-hatch. This reduction in expression was reversed by BPTES. Similarly, the EHC-dependent reduction of inflammatory gene expression during LPS challenge was eliminated in BPTES-treated chicks. Thus, TET family enzymes and CpG demethylation are essential for the embryonic induction of stress cross-tolerance in the hypothalamus.

Vitamin E Supplementation and Early Age Heat Conditioning to Alleviate the Negative Effects of Heat Stress in Quail Chicks.

BACKGROUND AND OBJECTIVE: Inclusion of vitamin E in poultry diets and manipulation of temperature at early age has been known to help birds cope with heat stress at later age of their life. This study was conducted to investigate the effects of early age heat conditioning (EHC) and vitamin E addition on the performance and blood parameters to alleviate deleterious impact of heat stress in quail chicks. MATERIAL AND METHODS: Three hundred one-day-old quail chicks were randomly divided into 4 groups of 5 replicates with 15 birds of each. Treatments were: Control, vitamin E: Chicks were fed the basal diet supplemented with 200 IU kg-1 diet vitamin E, EHC: Chicks were exposed to 40±1°C for 2 h at days 7th and 13th of embryogenesis and EHC+vitamin E: Chicks were exposed to 40±1°C for 2 h at days 7th and 13th of embryogenesis and fed the basal diet supplemented with 200 IU kg-1 vitamin E. The experiment lasted from 1-40 days of age. RESULTS: The results indicated that using early age heat conditioning and/or supplementation of vitamin E significantly (p<0.05) improved body weight, feed intake and feed conversion ratio at 40 days of age. Significantly (p<0.05) improvement was observed in blood pH, H/L ratio and Hb concentration, thyroid hormones (T3 and T4), plasma protein fractions and antioxidant status (Total antioxidant capacity, catalase and superoxide dismutase). CONCLUSION: Exposed quail chicks to early age heat conditioning and addition of vitamin E at 200 IU kg-1 diet can effectively alleviate the adverse effects of heat stress.

Insights into the Molecular Events That Regulate Heat-Induced Chilling Tolerance in Citrus Fruits.

Low non-freezing temperature may cause chilling injury (CI), which is responsible for external quality deterioration in many chilling-sensitive horticultural crops. Exposure of chilling-sensitive citrus cultivars to non-lethal high-temperature conditioning may increase their chilling tolerance. Very little information is available about the molecular events involved in such tolerance. In this work, the molecular events associated with the low temperature tolerance induced by heating Fortune mandarin, which is very sensitive to chilling, for 3 days at 37°C prior to cold storage is presented. A transcriptomic analysis reveals that heat-conditioning has an important impact favoring the repression of genes in cold-stored fruit, and that long-term heat-induced chilling tolerance is an active process that requires activation of transcription factors involved in transcription initiation and of the WRKY family. The analysis also shows that chilling favors degradation processes, which affect lipids and proteins, and that the protective effect of the heat-conditioning treatment is more likely to be related to the repression of the genes involved in lipid degradation than to the modification of fatty acids unsaturation, which affects membrane permeability. Another major factor associated with the beneficial effect of the heat treatment on reducing CI is the regulation of stress-related proteins. Many of the genes that encoded such proteins are involved in secondary metabolism and in oxidative stress-related processes.