Melatonin administration during the first half of pregnancy improves physiological response and reproductive performance of rabbits under heat stress conditions.

Context Melatonin may have a heat-stress-alleviating role during pregnancy. Aims To investigate the effects of melatonin administration during the first half of pregnancy on heat-tolerance capacity and pregnancy outputs of naturally heat-stressed rabbits. Methods Forty female rabbits were stratified equally into two experimental groups and daily received 1mg melatonin/kg body weight or not (control) for 15 consecutive days post-insemination. Heat tolerance indices, hormone profile, ovarian structures, and fetal loss were determined. Key results Treatment with melatonin significantly decreased respiration rate and rectal temperature, improved concentrations of nitric oxide, and tended to decrease malondialdehyde concentrations (P =0.064) compared to control. Melatonin treatment significantly increased concentrations of high-density lipoprotein, oestradiol, and progesterone compared to control. No significant differences in the numbers of visible ovarian follicles, corpora lutea, and total implantation sites on day 18 of pregnancy were observed between experimental groups. However, melatonin treatment significantly reduced the number of absorbed implantation sites and significantly improved amniotic fluid volume and conception rate compared to control. Conclusions Melatonin administration during the first half of pregnancy can improve reproductive performance of heat-stressed female rabbits. Implications Melatonin can improve fetal survivability via improving heat-tolerance capacity of does and steroidogenesis.

The influence of occupational heat stress on serum inflammatory cytokines among traditional bakery workers in Iran.

Heat exposure exceeding the ISO7243:1989 standard limit can contribute to health problems among employees in a variety of workplaces. Ignoring heat standard requirements in hot working conditions such as bakeries results in physiologic and health problems, as well as an elevated risk of later illnesses. In this analytical case-control study, the serum levels of four inflammatory factors (interleukin-1 beta, interleukin-6, tumor necrosis factor-α, and C-reactive protein) were assessed using an enzyme-linked immunosorbent assay. 105 male artisan bakers (in four job classifications in bakeries and staff) were compared based on demographic characteristics and inflammatory factors. The findings of the study showed correlations between serum interleukin-1ß, interleukin-6, and C-reactive protein levels and thermal exposure in the occupational environment and employment type. Moreover, some differences in serum level of interleukin-1ß and job type were observed. Heat overexposure affected the increase of interleukin-1ß and C-reactive protein secretion. As a result of years of working in high-temperature conditions, inflammation can lead to subsequent diseases in workers. To protect their health from this occupational hazard, additional safeguards are needed. Our recommendations could also be applied to overly hot work environments that may cause heat stress in workers.

Impact of heat stress on health-related symptoms and physiological changes among workers at a palm oil mill in Mukah, Sarawak, Malaysia.

INTRODUCTION: The palm oil (PO) industry is one of the most important sectors in the Malaysian economy. Workers at PO mills are, however, at risk for a number of health and safety issues, including heat stress, as the PO is one of the industries with high heat exposure. Heat stress occurs when a person's body cannot get rid of excess heat. Heat stress can result in heat cramps, heat exhaustion, heat rash, and heat stroke. It also results in physiological and psychological changes that can have an impact on a worker's performance. Therefore, this study aimed to evaluate the impact of heat stress on health-related symptoms and physiological changes among workers in a PO mill. MATERIALS AND METHODS: This cross-sectional study was conducted in a PO mill located in Mukah, Sarawak, Malaysia. Thirty-one workers from the four workstations (sterilizer, boiler, oil, and engine rooms) were selected as the respondents in this study. Wet Bulb Globe Thermometer was used in this study to measure the environmental temperature (WBGTin). Body core temperature (BCT), blood pressure (BP), and heart rate (HR) were recorded both before and after working in order to assess the physiological effects of heat stress on workers. A set of questionnaires were used to determine sociodemographic characteristics of the respondents and their symptoms related to heat stress. Data were then analyzed using SPSS Ver28. RESULTS: The WBGTin was found to be above the ACGIH threshold limit value of heat stress exposure in the engine room, sterilizer, and boiler workstations (>28.0°C). Additionally, there was a significant difference in the worker's BCT in these three workstations before and after work (p<0.05). Only the systolic BP and HR of those working at the boiler workstation showed significant difference between before and after work (p<0.05). The most typical symptoms that workers experience as a result of being exposed to heat at work include headache and fatigue. However, statistical analysis using Spearman Rho's test showed that there is no correlation between heat stress level with physiological changes and health-related symptoms among study respondents (p>0.05). CONCLUSION: Results of the present study confirmed that workers in PO mill were exposed to high temperatures while at work. Although the evidence indicates the physiological parameters in general are not significantly affected while working, it also demonstrated that worker's body adapts and acclimates to the level of heat. Even so, precautions should still be taken to reduce future heat exposure. It is recommended that a physiological study be carried out that focuses on cognitive function impairment to support the evidence regarding the effects of heat stress on PO mill workers.

NINJ1 mediates inflammatory cell death, PANoptosis, and lethality during infection conditions and heat stress.

Innate immunity provides the first line of defense through multiple mechanisms, including pyrogen production and cell death. While elevated body temperature during infection is beneficial to clear pathogens, heat stress (HS) can lead to inflammation and pathology. Links between pathogen exposure, HS, cytokine release, and inflammation have been observed, but fundamental innate immune mechanisms driving pathology during pathogen exposure and HS remain unclear. Here, we use multiple genetic approaches to elucidate innate immune pathways in infection or LPS and HS models. Our results show that bacteria and LPS robustly increase inflammatory cell death during HS that is dependent on caspase-1, caspase-11, caspase-8, and RIPK3 through the PANoptosis pathway. Caspase-7 also contributes to PANoptosis in this context. Furthermore, NINJ1 is an important executioner of this cell death to release inflammatory molecules, independent of other pore-forming executioner proteins, gasdermin D, gasdermin E, and MLKL. In an in vivo HS model, mortality is reduced by deleting NINJ1 and fully rescued by deleting key PANoptosis molecules. Our findings suggest that therapeutic strategies blocking NINJ1 or its upstream regulators to prevent PANoptosis may reduce the release of inflammatory mediators and benefit patients.

Exertional Heat Illness-From Identifying Heat Rash to Treating Heat Stroke.

Heat-related illness commonly affects adolescent patients, especially as summer approaches and global temperature extremes worsen. Basic counseling on sunburn prevention can decrease the risk for future malignancies, and rapidly preventing, identifying, and treating heat stroke can prevent severe morbidity and mortality. This article will review the epidemiology of exertional heat-related illness and the variations in presentations and pathology, from heat rash and sunburn to heat exhaustion and heat stroke. By the end of this review clinicians should be able to identify and treat different heat-related illnesses in adolescents and potentially save a life. [Pediatr Ann. 2024;53(1):e17-e21.].

Effects of preconditioning with heat stress on acute exercise-induced intracellular signaling in male rat gastrocnemius muscle.

Heat stress (HS) induces Akt/mTOR phosphorylation and FoxO3a signaling; however, whether a prior increase in heat shock protein 72 (HSP72) expression affects intracellular signaling following eccentric exercise remains unclear. We analyzed the effects of HS pretreatment on intramuscular signaling in response to acute exercise in 10-week-old male Wistar rats (n = 24). One leg of each rat was exposed to HS and the other served as an internal control (CT). Post-HS, rats were either rested or subjected to downhill treadmill running. Intramuscular signaling responses in the red and white regions of the gastrocnemius muscle were analyzed before, immediately after, or 1 h after exercise (n = 8/group). HS significantly increased HSP72 levels in both deep red and superficial white regions. Although HS did not affect exercise-induced mTOR signaling (S6K1/ERK) responses in the red region, mTOR phosphorylation in the white region was significantly higher in CT legs than in HS legs after exercise. Thr308 phosphorylation of Akt showed region-specific alteration with a decrease in the red region and an increase in the white region immediately after downhill running. Overall, a prior increase in HSP72 expression elicits fiber type-specific changes in exercise-induced Akt and mTOR phosphorylation in rat gastrocnemius muscle.

Genistein alleviates chronic heat stress-induced lipid metabolism disorder and mitochondrial energetic dysfunction by activating the GPR30-AMPK-PGC-1α signaling pathways in the livers of broiler chickens.

The objective of this study was to investigate the preventive effects and mechanisms of genistein (GEN) on production performance and metabolic disorders in broilers under chronic heat stress (HS). A total of 120 male 3-wk-old Ross broilers were randomly assigned to 5 groups: a thermoneutral zone (TN) group maintained at normal temperature (21°C ± 1°C daily), an HS group subjected to cyclic high temperature (32°C ± 1°C for 8 h daily), and 3 groups exposed to HS with varying doses of GEN (50, 100, or 150 mg/kg diet). The experimental period lasted for 3 wk. Here, HS led to a decline in growth performance parameters and hormone secretion disorders (P < 0.05), which were improved by 100 and 150 mg/kg GEN treatment (P < 0.05). Moreover, the HS-induced increases in the liver index (P < 0.01) and abdominal fat rate (P < 0.05) were attenuated by 150 mg/kg GEN (P < 0.05). The HS-induced excessive lipid accumulation in the liver and serum (P < 0.01) was ameliorated after 100 and 150 mg/kg GEN treatment (P < 0.05). Furthermore, the HS-induced decreases in lipolysis-related mRNA levels and increases in lipid synthesis-related mRNA levels in the liver (P < 0.01) were effectively blunted after 100 and 150 mg/kg GEN treatment (P < 0.05). Importantly, the HS-stimulated hepatic mitochondrial energetic dysfunction and decreases in the mRNA or protein levels of peroxisome proliferator-activated receptor-gamma coactivator 1α (PGC-1α), nuclear respiratory factor 1, and mitochondrial transcription factor A in the liver were ameliorated by 150 mg/kg GEN (P < 0.05). Moreover, 50 to 150 mg/kg GEN treatment resulted in a significant increase in the mRNA or protein levels of G protein-coupled estrogen receptor (GPR30), AMP-activated protein kinase (AMPK) α1, phosphorylated AMPKα, and phosphorylated acetyl-CoA carboxylase α. Collectively, GEN alleviated metabolic disorders and hepatic mitochondrial energetic dysfunction under HS, possibly through the activation of GPR30-AMPM-PGC-1α pathways. These data provide a sufficient basis for GEN as an additive to alleviate HS in broilers.

Risk factors for heat-related illness resulting in death or hospitalization in the oil and gas extraction industry.

Many oil and gas extraction (OGE) activities occur in high-heat environments, resulting in a significant risk of heat-related illness among outdoor workers in this industry. This report highlights cases of occupational heat-related illness that resulted in death and identifies common risk factors for heat-related fatalities and hospitalizations among OGE workers. Two databases maintained by the National Institute for Occupational Safety and Health (NIOSH) and the Occupational Safety and Health Administration (OSHA) were reviewed to identify heat-related fatalities, hospitalizations, and associated risk factors among OGE workers. Nine fatalities and associated risk factors were identified during 2014-2019 from NIOSH's Fatalities in Oil and Gas Extraction (FOG) Database. Risk factors identified included those commonly associated with heat-related fatalities: new workers not acclimatized to heat, inadequate heat stress training, and underlying hypertension or cardiovascular disease. Of particular note, substance use was identified as a significant risk factor as more than half of the fatalities included a positive postmortem test for amphetamines or methamphetamines. Fifty heat-related hospitalizations were identified from OSHA's Severe Injury Report Database during January 2015-May 2021. Heat stress has been and will continue to be an important cause of fatality and adverse health effects in OGE as hot outdoor working conditions become more common and extreme. More emphasis on heat stress training, acclimatization regimens, medical screening, and implementation of workplace-supportive recovery programs may reduce heat-related fatalities and injuries in this industry.

Comparison of Transcriptomic Changes in Survivors of Exertional Heat Illness with Malignant Hyperthermia Susceptible Patients.

Exertional heat illness (EHI) is an occupational health hazard for athletes and military personnel-characterised by the inability to thermoregulate during exercise. The ability to thermoregulate can be studied using a standardised heat tolerance test (HTT) developed by The Institute of Naval Medicine. In this study, we investigated whole blood gene expression (at baseline, 2 h post-HTT and 24 h post-HTT) in male subjects with either a history of EHI or known susceptibility to malignant hyperthermia (MHS): a pharmacogenetic condition with similar clinical phenotype. Compared to healthy controls at baseline, 291 genes were differentially expressed in the EHI cohort, with functional enrichment in inflammatory response genes (up to a four-fold increase). In contrast, the MHS cohort featured 1019 differentially expressed genes with significant down-regulation of genes associated with oxidative phosphorylation (OXPHOS). A number of differentially expressed genes in the inflammation and OXPHOS pathways overlapped between the EHI and MHS subjects, indicating a common underlying pathophysiology. Transcriptome profiles between subjects who passed and failed the HTT (based on whether they achieved a plateau in core temperature or not, respectively) were not discernable at baseline, and HTT was shown to elevate inflammatory response gene expression across all clinical phenotypes.

Effects of dietary supplementation with L-arginine on the endogenous losses of amino acids in growing pigs exposed to heat stress.

Exposing pigs to heat stress (HS) provokes higher death of intestinal cells, resulting in elevated endogenous intestinal losses (EIL) of amino acids (AA) and damage to intestinal epithelia. Arginine (Arg) is precursor for the synthesis of polyamines, which are involved in proliferation of intestinal cells and restoration of the intestinal epithelia. Thus the effect of adding L-Arg to diets for HS pigs on the EIL of AA was analyzed. Twelve pigs (23.1 ± 1.1 kg body weight) implanted with T-type cannulas at the end of ileum were individually housed and allowed 15-days for surgery recovery under thermoneutral (TN) conditions (22 ± 2 °C). Following, the pigs were randomly assigned to one of three treatments: TN pigs fed a semi-purified, corn starch-3% casein basal diet (TN-B); HS pigs with the basal diet (HS-B); HS pigs consuming the basal diet supplemented with 0.20% L-Arg (HS-Arg). The experiment consisted of two 9-day periods; each period included 7-days of adaptation to their respective diet, followed by a 2-day ileal digesta collection period. Digesta was collected during 12 consecutive hours each day. The pigs were fed twice a-day. Ambient temperature (AT) inside the TN and HS rooms ranged from 18.6 to 27.6 °C and from 29.5 to 40.7 °C, respectively. Body temperature followed a pattern similar to that of AT. The daily EIL of indispensable AA increased (P < 0.01) in the HS-B pigs compared to both the TN-B and the HS-Arg pigs, however, there was no EIL difference between the TN-B and the HS-Arg pigs (P > 0.05). Likewise, with the exception of serine, daily losses of endogenous dispensable AA in the HS-B pigs were higher (P < 0.01) in comparison with those of TN-B and HS-Arg pigs. In summary, HS exposure compared to TN conditions increases the loss of endogenous AA, but dietary supplementation with L-Arg helped to counteract the negative HS effect.

Effect of heat stress on heat shock protein expression and hypertrophy-related signaling in the skeletal muscle of trained individuals.

Muscle mass is balanced between hypertrophy and atrophy by cellular processes, including activation of the protein kinase B-mechanistic target of rapamycin (Akt-mTOR) signaling cascade. Stressors apart from exercise and nutrition, such as heat stress, can stimulate the heat shock protein A (HSPA) and C (HSPC) families alongside hypertrophic signaling factors and muscle growth. The effects of heat stress on HSP expression and Akt-mTOR activation in human skeletal muscle and their magnitude of activation compared with known hypertrophic stimuli are unclear. Here, we show a single session of whole body heat stress following resistance exercise increases the expression of HSPA and activation of the Akt-mTOR cascade in skeletal muscle compared with resistance exercise in a healthy, resistance-trained population. Heat stress alone may also exert similar effects, though the responses are notably variable and require further investigation. In addition, acute heat stress in C2C12 muscle cells enhanced myotube growth and myogenic fusion, albeit to a lesser degree than growth factor-mediated hypertrophy. Though the mechanisms by which heat stress stimulates hypertrophy-related signaling and the potential mechanistic role of HSPs remain unclear, these findings provide additional evidence implicating heat stress as a novel growth stimulus when combined with resistance exercise in human skeletal muscle and alone in isolated murine muscle cells. We believe these findings will help drive further applied and mechanistic investigation into how heat stress influences muscular hypertrophy and atrophy.NEW & NOTEWORTHY We show that acute resistance exercise followed by whole body heat stress increases the expression of HSPA and increases activation of the Akt-mTOR cascade in a physically active and resistance-trained population.

The effects of betaine supplementation on fluid balance and heat tolerance during passive heat stress in men.

INTRODUCTION: Consuming intracellular osmolytes, like betaine (BET), may attenuate symptoms of heat stress. The purpose of this study was to examine the effects of BET supplementation on fluid balance and heat tolerance after a 7-day loading period and during passive heat exposure. METHODS: A double-blind, placebo controlled, crossover study compared BET or placebo consumption (50 mg·kg-1 , twice daily) for 7 days in young, recreationally active men (N = 11). RESULTS: During the loading period, no significant interactions were found for any marker of fluid balance between or within conditions. During heat exposure, significant time effects but no condition x time interactions, were found for plasma characteristics (i.e., volume, osmolality, sodium, albumin, and total protein). Plasma volume was significantly increased by min 30 in both conditions (PLA: +6.9. ± 5.0%, BET: +10.2 ± 7.4%) and remained elevated for the remainder of the experimental trial, but was not significantly different between conditions. After 60 min of passive heat exposure, both conditions experienced a similar increase in core temperature (PLA: +0.32 ± 0.22°C, BET: +0.31 ± 0.21°C; p = 0.912). CONCLUSIONS: Supplemental BET did not improve markers of fluid balance or heat tolerance during 7 days of loading or during passive heat exposure.

Alpha-lipoic acid attenuates heat stress-induced apoptosis via upregulating the heat shock response in porcine parthenotes.

Heat stress (HS) is a long-standing hurdle that animals face in the living environment. Alpha-lipoic acid (ALA) is a strong antioxidant synthesized by plants and animals. The present study evaluated the mechanism of ALA action in HS-induced early porcine parthenotes development. Parthenogenetically activated porcine oocytes were divided into three groups: control, high temperature (HT) (42 °C for 10 h), and HT + ALA (with 10 µM ALA). The results show that HT treatment significantly reduced the blastocyst formation rate compared to the control. The addition of ALA partially restored the development and improved the quality of blastocysts. Moreover, supplementation with ALA not only induced lower levels of reactive oxygen species and higher glutathione levels but also markedly reduced the expression of glucose regulatory protein 78. The protein levels of heat shock factor 1 and heat shock protein 40 were higher in the HT + ALA group, which suggests activation of the heat shock response. The addition of ALA reduced the expression of caspase 3 and increased the expression of B-cell lymphoma-extra-large protein. Collectively, this study revealed that ALA supplementation ameliorated HS-induced apoptosis by suppressing oxidative and endoplasmic reticulum stresses via activating the heat shock response, which improved the quality of HS-exposed porcine parthenotes.

Environment-induced heat stress causes structural and biochemical changes in the heart.

Prolonged exposure to heat can lead to environment-induced heat stress (EIHS), which may jeopardize human health, but the extent to which EIHS affects cardiac architecture and myocardial cell health are unknown. We hypothesized EIHS would alter cardiac structure and cause cellular dysfunction. To test this hypothesis, 3-mo old female pigs were exposed to thermoneutral (TN; 20.6 ± 0.2 °C; n = 8) or EIHS (37.4 ± 0.2 °C; n = 8) conditions for 24 h, hearts were removed and dimensions measured, and portions of the left ventricle (LV) and right ventricle (RV) were collected. Environment-induced heat stress increased rectal temperature 1.3 °C (P < 0.01), skin temperature 11 °C (P < 0.01) and respiratory rate 72 breaths per minute (P < 0.01). Heart weight and length (apex to base) were decreased by 7.6% (P = 0.04) and 8.5% (P = 0.01), respectively, by EIHS, but heart width was similar between groups. Left ventricle wall thickness was increased (22%; P = 0.02) and water content was decreased (8.6%; P < 0.01) whereas in RV, wall thickness was decreased (26%; P = 0.04) and water content was similar in EIHS compared to TN. We also discovered ventricle-specific biochemical changes such that in RV EIHS increased heat shock proteins, decreased AMPK and AKT signaling, decreased activation of mTOR (35%; P < 0.05), and increased expression of proteins that participate in autophagy. In LV, heat shock proteins, AMPK and AKT signaling, activation of mTOR, and autophagy-related proteins were largely similar between groups. Biomarkers suggest EIHS-mediated reductions in kidney function. These data demonstrate EIHS causes ventricular-dependent changes and may undermine cardiac health, energy homeostasis, and function.

Effects of dietary antioxidant supplementation on metabolism and inflammatory biomarkers in heat-stressed dairy cows.

Heat-stress-induced inflammation may be ameliorated by antioxidant supplementation due to the purported effects of increased production of reactive oxygen species or oxidative stress on the gastrointestinal tract barrier. Thus, study objectives were to evaluate whether antioxidant supplementation [AGRADO Plus 2.0 (AP); EW Nutrition] affects metabolism and inflammatory biomarkers in heat-stressed lactating dairy cows. Thirty-two mid-lactation multiparous Holstein cows were assigned to 1 of 4 dietary-environmental treatments: (1) thermoneutral (TN) conditions and fed a control diet (TN-CON; n = 8), (2) TN and fed a diet with AP (10 g antioxidant; n = 8), (3) heat stress (HS) and fed a control diet (HS-CON; n = 8), or (4) HS and fed a diet with AP (HS-AP; n = 8). The trial consisted of a 23-d prefeeding phase and 2 experimental periods (P). Respective dietary treatments were top-dressed starting on d 1 of the prefeeding period and continued daily throughout the duration of the experiment. During P1 (4 d), baseline data were collected. During P2 (7 d), HS was artificially induced using an electric heat blanket (Thermotex Therapy Systems Ltd.). During P2, the effects of treatment, day, and treatment-by-day interaction were assessed using PROC MIXED of SAS (SAS Institute Inc.). Heat stress (treatments 3 and 4) increased rectal, vaginal, and skin temperatures (1.2°C, 1.1°C, and 2.0°C, respectively) and respiration rate (33 breaths per minute) relative to TN cows. As expected, HS decreased dry matter intake, milk yield, and energy-corrected milk yield (32%, 28%, and 28% from d 4 to 7, respectively) relative to TN. There were no effects of AP on body temperature indices or production. Milk fat, protein, and lactose concentrations remained unaltered by HS or AP; however, milk urea nitrogen was increased during HS regardless of AP supplementation (26% relative to TN). Circulating glucose remained unchanged by HS, AP, or time. Additionally, HS decreased circulating glucagon (29% from d 3 to 7 relative to TN), but there was no additional effect of AP. There was a tendency for nonesterified fatty acid concentrations to be increased in HS-AP cows throughout P2 (60% relative to TN-CON), whereas it remained similar in all other treatments. Blood urea nitrogen increased for both HS treatments from d 1 to 3 before steadily decreasing from d 5 to 7, with the overall increase being most pronounced in HS-CON cows (27% relative to TN-CON). Further, supplementing AP decreased blood urea nitrogen in HS-AP on d 3 relative to HS-CON (15%). Circulating serum amyloid A tended to be and lipopolysaccharide binding protein was increased by HS, but neither acute-phase protein was affected by AP. Overall, AP supplementation appeared to marginally alter metabolism but did not meaningfully alter inflammation during HS.

Reliability of Biomarkers of Physiological Stress at Rest and Post-exertional Heat Stress.

The purpose of this study was to assess the reliability of blood biomarkers that can signify exercise-induced heat stress in hot conditions. Fourteen males completed two heat stress tests separated by 5-7 days. Venous blood was drawn pre- and post- heat stress for the concentration of normetanephrine, metanephrine, serum osmolality, copeptin, kidney-injury molecule 1, and neutrophil gelatinase-associated lipocalin. No biomarker, except copeptin, displayed systematic trial order bias (p≥0.05). Normetanephrine, copeptin and neutrophil gelatinase-associated lipocalin presented acceptable reliability (CV range: 0.9-14.3%), while greater variability was present in metanephrine, osmolality and kidney-injury molecule 1 (CV range: 28.6-43.2%). Normetanephrine exhibited the largest increase (p<0.001) in response to heat stress (trial 1=1048±461 pmol. L-1; trial 2=1067±408 pmol. L-1), whilst kidney-injury molecule 1 presented trivial changes (trial 1=-4±20 ng. L-1; trial 2=2 ± 16 ng. L-1, p>0.05). Normetanephrine, copeptin, and neutrophil gelatinase-associated lipocalin demonstrated good reliability and sensitivity to an acute bout of heat stress. These biomarkers may be suitable for application in laboratory and field research to understand the efficacy of interventions that can attenuate the risk of thermal injury whilst exercising in the heat.

Effects of betaine on viability, apoptosis, function protein expression and oxidative status in heat-stressed IEC-6 cells.

Intestinal epithelial dysfunction is one of the key factors in the pathogenesis of heat stress-induced disease. The purpose of this experiment was to investigate whether betaine protects IEC-6 cells from dysfunction induced by heat stress (HS) through antioxidative mechanism. The IEC-6 cells were divided into four groups: control group incubated at 37 °C, while those in heat treated groups (41 °C for 24 h) were pretreated with 0, 0.5 and 1 mmol/L betaine, respectively. Cell viability, apoptosis, barrier function protein and oxidative status were analyzed. Compared to control group, the rate of apoptosis and the Bax and caspase-3 expressions significantly increased in HS group (P < 0.05), however, cell activity, total antioxidative capacity (T-AOC), activities of glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and the expression of Bcl-2, claudin-1 and occludin decreased significantly (P < 0.05). Betaine (0.5 mmol/L) can significantly enhance IEC-6 cell viability, while significantly reduce the apoptosis rate of cell during HS (P < 0.05). Meanwhile, the expression of Bcl-2, claudin-1 and occludin proteins were also significantly upregulated (P < 0.05) when compared to HS group. HS had a negative impact on IEC-6 cells, while betaine protected from damage caused by HS via increasing the antioxidative capacity. This suggested that betaine might be an effective dietary additive to protect animals from detrimental intestinal reactions caused by HS.

Novel organic selenium source hydroxy-selenomethionine counteracts the blood-milk barrier disruption and inflammatory response of mice under heat stress.

Heat stress (HS) in summer has caused huge economic losses to animal husbandry production recently. When mammary gland is exposed to high temperatures, it will cause blood-milk barrier damage. Hydroxy-selenomethionine (HMSeBA) is a new selenium source with better guarantee of animals' production performance under stress, but whether it has protective effect on heat stress-induced blood-milk damage is still unclear. We established mammary epithelial cells and mice heat stress injury models to fill this research gap, and hope to provide theoretical basis for using HMSeBA to alleviate heat stress damage mammary gland. The results showed that (1) Heat stress significantly decreases in vitro transepithelial electrical resistance (TEER) and cell viability (P < 0.01), and significantly decreases clinical score, histological score, and total alveoli area of mice mammary gland tissue (P < 0.01). (2) HMSeBA significantly increases TEER and fluorescein sodium leakage of HS-induced monolayer BMECs (P < 0.01), significantly improves the milk production and total area of alveoli (P < 0.01), and reduces clinical score, histological score, mRNA expression of heat stress-related proteins, and inflammatory cytokines release of heat-stressed mice (P < 0.01). (3) HMSeBA significantly improves tight junction structure damage, and significantly up-regulated the expression of tight junction proteins (ZO-1, claudin 1, and occludin) as well as signal molecules PI3K, AKT, and mTOR (P < 0.01) in heat-stressed mammary tissue. (4) HMSeBA significantly increases glutathione peroxidase (GSH-Px), total antioxidant capacity (T-AOC), and superoxide dismutase release (SOD) (P < 0.01) and significantly reduce malondialdehyde (MDA) expression (P < 0.01) in heat-stressed mammary tissue. In conclusion, this study implemented heat-stressed cell and mice model and showed that HMSeBA significantly regulate antioxidant capacity, inhibited inflammation, and regulate tight junction proteins expression in blood-milk barrier via PI3K/AKT/mTOR signaling pathway, so as to alleviate mammary gland damage and ensure its structure and function integrity.

Effect of lowering blood cortisol level along with progesterone priming on intensifying the secondary symptoms of oestrus in Murrah buffaloes during the hot summer months.

The present study sought to intensify oestrus symptoms in heat-stressed pre-pubertal Murrah buffalo heifers. The first experiment aimed at lowering the blood cortisol level. Twenty pre-pubertal buffalo heifers approximately 36-40 months of age were randomly allocated to four groups of five buffaloes. Group Gly received 100 ml glycerol orally for 8 days (day 0 is the start of treatments), group Gly-E received intra-muscular injections of 500 mg tocopherol (vitamin E activity) and 150 mg sodium selenite on alternate days (four treatments) in addition to treatments received by group Gly, group Gly-E + B-Ch received intra-muscular injections of vitamin B-complex and choline daily for 8 days in addition to the treatments received by group Gly-E. The control group Co-1 did not received any treatments. Blood cortisol and glucose level was measured on day 7 post-treatment. The second experiment sought to test the effect of lowering the blood cortisol level and progesterone priming on the intensity of oestrus symptoms in heat-stressed pre-pubertal buffalo heifers. Eighty buffaloes were allocated into four groups of twenty buffaloes. P4 group received intra-vaginal progesterone CIDR for 5 days (day 0 is the day of start of treatment). Gly-E group received the same treatment as in the first experiment for 7 days (day 0-6) and P4 + Gly-E received the combination of P4 and Gly-E groups. The control group Co-2 did not receive any treatments; however, on day 6, all groups received intra-muscular injection of 2 mg estradiol benzoate. The buffaloes were graded for oestrus symptoms on the basis of mucus discharge (A-scanty and B-copious), vulvar redness (slightly pink and moist-A and deep red and moist-B), ferning (nil or poor-A and good with arborization-B) and cellularity of cervical mucus (Type A goblet cells with round nucleus and abundant cytoplasm and Type B goblet cells with cylindrical nucleus and scanty cytoplasm). The blood glucose level (mean ± SEM) of Gly group on day 7 of treatment in experiment 1 was 58.3 ± 0.4 mg/dl that was significantly greater than Co-1 (52.4 ± 0.4 mg/dl), Gly-E (53.5 ± 0.8 mg/dl) and Gly-E + B-Ch (52.7 ± 0.6 mg/dl) groups; however, the difference was found to be non-significant among Co-1, Gly-E and Gly-E + B-Ch groups. The cortisol level (mean ± SEM) on day 7 in groups Gly (11.4 ± 0.6 ng/ml), Gly-E (8.1 ± 0.5 ng/ml) and Gly-E + B-Ch (7.5 ± 0.4 ng/ml) was significantly lower than groups Co-1 (15.5 ± 0.3 ng/ml), though the difference between groups Gly-E and Gly-E + B-Ch was non-significant and the level in group Gly-E significantly lower than Gly. Thus, the combination of vitamin E and selenium along with glycerol had the best effect in the reduction of blood cortisol level in heat-stressed buffaloes. In the second experiment the blood estradiol level on day 7 in all groups was around 40 pg/ml. Cortisol reduction (group Gly-E) improved oestrus symptoms (mucus discharge and ferning) compared to control, but when compared to group P4 showed non-significant difference with respect to mucus discharge, vulvar redness and ferning of cervical mucus. However, when cortisol reduction treatments were combined with progesterone priming (group P4 + Gly-E) oestrus symptoms were improved with respect to mucus discharge and ferning thus, combination treatment had synergistic effect. This effect was also observed with cellularity of cervical mucus, where combination treatment group P4 + Gly-E produced cervical mucus with significantly lower percentage of Type B cells compared to group P4 and group Gly-E. Thus, administration glycerol and vitamin E selenium combination reduces blood cortisol level and along with progesterone priming can be effective to improve the oestrus symptoms of buffaloes during the hot summer months.

ROS/JNK-mediated lysosomal injury in rat intestinal epithelial-6 cells during heat stress.

Injury to the intestinal epithelial cells and loss of the intestinal barrier are critical to heatstroke. To reveal the mechanism through which heatstroke leads to intestinal epithelial injury, the relationship between reactive oxygen species (ROS), c-Jun NH2-terminal kinase (JNK), and lysosomes were studied in intestinal epithelial cells subjected to heat stress. Cells of heat stress groups were incubated at 43 °C for 1 h, then incubated at 37 °C as indicated. Control group cells were incubated at 37 °C. Cell-counting kit-8 assay was used to assess cell viability. Cells were labeled with 2'-7'dichlorofluorescin diacetate and acridine orange (AO) staining, respectively, the total ROS and AO were detected by confocal laser scanning microscopy and flow cytometry. Apoptosis was analyzed by flow cytometry using annexin V-fluorescein isothiocyanate/prodium iodide staining, the expressions of mitogen-activated protein kinases were detected by western blotting. Heat stress induced apoptosis and inhibited cell viability, the production of ROS, and lysosomal injury in IEC-6 cells. After pretreatment with the lysosomal cathepsin inhibitor E64, the JNK inhibitor SP600125, or the ROS scavenger NAC, the effect of heat stress on apoptosis or lysosomal injury was significantly attenuated. In conclusion, heat stress induced apoptosis, lysosomal injury, and the accumulation of ROS in IEC-6 cells; mechanistically, this occurred through the ROS-induced activation of JNK signaling, which mediated the lysosomal injury and ultimately apoptosis.