Hsp70 Gene Expression Research Articles

Prenatal auditory stimulation and impacts on physiological response to feed restriction in broiler chickens at market age

Feed restriction could induce physiological stress in broiler chickens, leading to welfare issues. Prenatal stimulation could improve stress-coping mechanisms in poultry. The present study aimed to elucidate the effects of subjecting developing embryos to auditory stimulation on physiological stress response to feed restriction in broiler chickens at market age. A total of 423 hatching eggs of Cobb 500 (Gallus domesticus) were subjected to the following auditory treatments: 1) no additional sound treatment other than the background sound of the incubator's compressors at 40 dB (CONTROL), 2) exposure to pre-recorded traffic noise at 90 dB (NOISE), and 3) exposure to Mozart's Sonata for Two Pianos in D Major, K 488 at 90 dB) (MUSIC). The NOISE and MUSIC treatments were for 20 min/h for 24 h (a total of 8 h/d), starting from embryonic days (ED) 12 to hatching. On d 42, an equal number of birds from each prenatal auditory stimulation (PAS) group were subjected to either ad libitum feeding (AL) or 30-h of feed restriction (FR) in a completely randomised design. The FR chickens exhibited significantly higher serum levels of corticosterone (CORT), and heat shock protein (HSP) 70 compared to those of AL. Prenatal auditory stimulation, particularly NOISE, led to lower serum levels of CORT and alpha-1-acid glycoprotein (AGP) levels compared to the CONTROL group. Additionally, NOISE significantly increased brain mRNA glucocorticoid receptor and HSP70 gene expression. The cecal population of E. coli and Lactobacillus spp. was not significantly affected by prenatal auditory stimulation. In conclusion, our findings suggest that prenatal auditory stimulation, particularly NOISE, positively impacts broiler chickens' ability to cope with feed restriction.

Impact of Eight Weeks of Supplementation with Vitamins C and E on the Expression of CP Protein and the HSP70 Gene in the Brain Tissue of Rats with Alzheimer’s Disease

Introduction: Impact of antioxidant supplements on the health of patients with Alzheimer’s disease has not yet been clearly defined. Given that no study has been conducted so far to evaluate the effectiveness of vitamins C and E on the expression of Carbonyl Protein (CP) and Heat Shock 70 (HSP70) genes in rats with Alzheimer’s, this study aims to investigate the effect of an eight-week supplementation of vitamins C and E on the expression of CP and HSP70 in the brain tissue of rats with Alzheimer’s disease. Methods: In this experimental study, 28 rats with Alzheimer disease were injected with 10 mg/kg of trimethyltin and divided into four groups: patient control, sham (soluble vitamin E), Vitamin C, and Vitamin E (VE). Seven healthy rats were included in the control group to investigate the effects of disease induction on the variables under study. They were administered a daily supplement of 4 mg/kg of vitamin C and 30 mg/kg of vitamin E for eight weeks. After supplementation period, the groups were compared in terms of CP values and HSP70 gene expression. Data were analyzed by SPSS16 statistical software. Shapiro-Wilk test was used to check normality of the finding distributions. Effect of vitamin E and vitamin C supplementation was evaluated using one-way analysis of variance. Tukey's post hoc test was used to check the type of variable effects. A significant level of 0.05 was considered for all analyses. Results: Levels of carbonyl protein in both the vitamin C and vitamin E groups were significantly lower than those in the patient control group (P < 0.05). HSP70 levels in the vitamin C and E groups were significantly higher than those in the patient control group (P < 0.05). No significant difference was observed between vitamin C and E supplement groups (P > 0.05). Conclusion: Findings of this study suggest that both vitamin C and E exhibit similar antioxidant and cellular protective effects in the brain tissue of mice with Alzheimer’s disease. Given the impact of vitamins C and E on reducing carbonyl protein levels and increasing HSP70 levels in the hippocampal tissue of rats, it is recommended to explore the effects of these vitamins on other neurodegenerative disorders in future studies.

Molecular mechanisms of induced thermotolerance in two strains of Brachionus koreanus (Rotifera: Monogononta)

Temperature plays an important role in the occurrence and performance of organisms in aquatic ecosystems and is also one of the main environmental factors affecting species' survival and growth in aquaculture. As an important species for aquaculture sustainability, the rotifer Brachionus sp. benefits from inducible phenotypic traits that allow the organisms to cope with environmental stress. The exposure to high temperature has shown to increase production of heat shock proteins (HSPs) and histone modifications in several organisms, resulting in induced thermotolerance. This study aimed to evaluate the potential of non-lethal heat shock (NLHS) to induce thermotolerance in two strains of B. koreanus and pinpoint some of the molecular mechanisms involved in the process. Exposure of organisms to 42 °C for 30 min, with a subsequent recovery at 25 °C for 8 h demonstrated to increase thermotolerance in up to three-fold when organisms were, in a posterior phase, subjected to a lethal temperature. This study also showed that one of the strains is tendentially more thermotolerant than the other. Indeed, NLHS exposure resulted in increased mRNA expression of different Hsp genes and production of HSP70 in general, but different patterns of expression were observed between strains. However, a single NLHS showed to have no effects in epigenetic mechanisms, suggesting that the induced capacity to tolerate heat stress was transient and some more cycles of NLHS may be needed to promote a persistent effect.

Autoimmunity to stromal-derived autoantigens in rheumatoid ectopic germinal centers exacerbates arthritis and affects clinical response.

Ectopic lymphoid structures (ELSs) in the rheumatoid synovial joints sustain autoreactivity against locally expressed autoantigens. We recently identified recombinant monoclonal antibodies (RA-rmAbs) derived from single, locally differentiated rheumatoid arthritis (RA) synovial B cells, which specifically recognize fibroblast-like synoviocytes (FLSs). Here, we aimed to identify the specificity of FLS-derived autoantigens fueling local autoimmunity and the functional role of anti-FLS antibodies in promoting chronic inflammation. A subset of anti-FLS RA-rmAbs reacting with a 60 kDa band from FLS extracts demonstrated specificity for HSP60 and partial cross-reactivity to other stromal autoantigens (i.e., calreticulin/vimentin) but not to citrullinated fibrinogen. Anti-FLS RA-rmAbs, but not anti-neutrophil extracellular traps rmAbs, exhibited pathogenic properties in a mouse model of collagen-induced arthritis. In patients, anti-HSP60 antibodies were preferentially detected in RA versus osteoarthritis (OA) synovial fluid. Synovial HSPD1 and CALR gene expression analyzed using bulk RNA-Seq and GeoMx-DSP closely correlated with the lympho-myeloid RA pathotype, and HSP60 protein expression was predominantly observed around ELS. Moreover, we observed a significant reduction in synovial HSP60 gene expression followed B cell depletion with rituximab that was strongly associated with the treatment response. Overall, we report that synovial stromal-derived autoantigens are targeted by pathogenic autoantibodies and are associated with specific RA pathotypes, with potential value for patient stratification and as predictors of the response to B cell-depleting therapies.

Temperature Effects on Expression Levels of hsp Genes in Eggs and Second-Stage Juveniles of Meloidogyne hapla Chitwood, 1949.

Meloidogyne hapla is one of the most important nematode pathogens. It is a sedentary, biotrophic parasite of plants that overwinters in the soil or in diseased roots. The development of M. hapla is temperature dependent. Numerous studies have been performed on the effect of temperature on the development of M. hapla, but only a few of them analyzed the heat shock protein (hsp) genes. The aim of the study was to perform expression profiling of eight hsp genes (Mh-hsp90, Mh-hsp1, Mh-hsp4, Mh-hsp6, Mh-hsp60, Mh-dnj19, Mh-hsp43, and Mh-hsp12.2) at two development stages of M. hapla, i.e., in eggs and second-stage juveniles (J2). The eggs and J2 were incubated under cold stress (5 °C), heat stress (35 °C, 40 °C), and non-stress (10 °C, 20 °C, and 30 °C) conditions. Expression profiling was performed by qPCR. It was demonstrated that only two genes, Mh-hsp60 and Mh-dnj19, have been upregulated by heat and cold stress at both development stages. Heat stress upregulated the expression of more hsp genes than cold stress did. The level of upregulation of most hsp genes was more marked in J2 than in eggs. The obtained results suggest that the Mh-hsp90 and Mh-hsp1 genes can be used as bioindicators of environmental impacts on nematodes of the Meloidogyne genus.

High-energy and high-amino acid diet enhances production performance and antioxidant capacity in yellow-feathered broilers under heat stress

This study investigated the ameliorating effects of high-energy and high-amino acid (HEHA) diets on heat stress (HS) in yellow-feathered broilers. Broilers aged 35 d were randomly assigned to 3 groups: control and HS groups fed the basic normal diet, and the HEHA group fed the HEHA diet (basal diet + 100 kcal/kg AME + 15 % DAAs). The HS and HEHA groups were exposed to cyclic HS (30 ± 1 to 34 ± 1 ℃) for 2 wk, while the control group was maintained at 26 ± 1 ℃. The results indicated that the HEHA diet significantly alleviated HS-induced feed intake and body weight loss. HEHA feeding mitigated the increase in body temperature during HS. Compared with observations in the HS group, the HEHA diet reduced the levels of ALT, Alb, and corticosterone in the serum and downregulated the gene expression of HSP27 and HSP60 in the liver. Moreover, the HEHA group showed higher GSH-px activity in the serum and SOD and GSH-Px activity in the jejunal mucosa than that of the HS group. HEHA supplementation also reduced MDA levels in the liver. In conclusion, the HEHA diet improved the production performance of broilers under HS by increasing their antioxidant capacities. These findings suggest an effective strategy to combat HS in poultry production.

Effects of a phytogenic diet on growth, haemato-immunological parameters, expression of immune- and stress-related genes, and resistance of Oncorhynchus mykiss to Lactococcus garvieae infection

Phytogenic additives have been implicated in aquafeed owing to their proven growth-stimulating, antimicrobial, antioxidant, and immune-enhancing efficacies. Herein, a short-term feeding experiment (21 days) was designed to explore the effects of a phytogenic additive, PASSIF MOOD® (PM, a mixture of Valeriana officinalis and Passiflora incarnata extracts), on growth, haemato-biochemical variables, immunity, and mRNA expression responses of rainbow trout fingerlings. After feeding, fish were experimentally infected with Lactococcus garvieae and monitored for an additional 20 days. Six groups were fed a commercial diet supplied with different PM levels as 0, 470, 940, 1410, 1880, and 2350 mg/kg and labeled as PM0 (control), PM1, PM2, PM3, PM4, and PM5 groups, respectively. Results revealed significant improvements in the growth of PM-supplied groups compared to the control. Dietary PM significantly increased RBC count, hemoglobin, and hematocrit values compared to those fed on a control diet. Stress indicators such as blood glucose and cortisol levels were significantly decreased in all PM groups than in controls. Moreover, there were significant increases in myeloperoxidase, respiratory burst, and lysozyme activities in PM groups than the control. The highest expression levels of the lysozyme II gene were found in the PM3, PM4, and PM5 groups. Moreover, the mRNA transcripts of il-10 and IgM heavy chain genes were significantly increased in all PM groups compared to the controls. Alternatively, the expression of tnf-α and hsp70 genes were significantly decreased in all PM groups compared to the controls. After challenge, the survival rates in PM0, PM1, PM2, PM3, PM4, and PM5 groups were 37.34%, 84.0%, 85.34%, 68.0%, 64.0%, and 36.0%, respectively, suggesting enhancement of the fish resistance in PM1, PM2, PM3, and PM4 groups compared to control. The results suggest the growth-promoting, antistress, and immunomodulatory roles of the tested phytogenic additive.

Mitigation of extreme winter stress in European seabass, Dicentrarchus labrax through dietary supplementation

Climate change is driving the dangerous confluence of extreme weather events. Among others, aquatic ecosystems are the most affected one. This study sheds light on how dietary supplements affect the physiological responses of European seabass during extreme winter exposure. Fish were provided with three dietary supplements: vitamins C and E (diet-1), propolis (diet-2), and phycocyanin (diet-3) for 45 days. They were then exposed to a simulated extreme winter cold (7 °C) for 20 days. The results showed that dietary supplements do not have a significant impact on weight, growth rate, and protein efficiency ratio before cold exposure. Levels of Na + and Cl − in the fish remained stable across all diets during both control (20 °C) and 7 °C exposure. However, on the 20th day of cold exposure, fish fed diet-3 exhibited notably higher levels of K+ compared to the other two diets, whereas blood glucose was significantly lower in fish fed diet-1. On day 20, significantly higher GPT activities were observed in fish fed diet-1. No significant effects of dietary supplementation were observed in fish serum cholesterol and protein levels. Before cold stress, fish fed the control diet exhibited significantly lower triglyceride levels (p < 0.05). However, during cold exposure, triglyceride levels increased in fish fed diet-1 and the control diet. On day 20, fish fed diet-3 exhibited a comparatively higher lactate content. Conversely, cortisol levels were significantly lower (p < 0.05) in fish fed diets 2 and 3 on the same day. No dietary effects were observed on fish muscle HSP70 gene expression. On days 10 and 20, SREBP1 was significantly upregulated in fish fed all diets. On day 20 of cold exposure, FADS2 and GLUT2 genes were downregulated in the fish fed control diet compared to those fed diets 2 and 3. Before cold stress, fish on the control diet showed higher TNF-1α expression in the spleen. SREBP1 showed a significant upregulation on day 10 in fish fed diet-2. FADS2 gene expression remained unchanged during cold stress. GLUT2 expression showed no noticeable effects throughout the study. Overall, during extreme cold exposure, we observed decreased levels of blood cortisol, transaminase, dehydrogenase, and metabolites, along with a comparatively higher upregulation of growth, metabolic, and immune genes in fish fed diets supplemented with propolis, phycocyanin, and vitamins C&E. These results suggest that dietary supplements can help European seabass cope better with extreme cold, at least for short periods of up to 20 days.

Evidence for Beneficial Physiological Responses of the Land Snail Cornu aspersum to Probiotics' (Lactobacillus plantarum) Dietary Intervention.

A large variety of microorganisms ingested with food constitute animals' intestinal microflora, enhancing and maintaining the homeostasis of the host. Rations enriched with probiotics are a method recommended to prevent undesirable conditions. To date, research has been limited to farmed animals and reared fish, creating a knowledge gap concerning the effect of probiotics on the growth rate, physiological responses, and energy metabolism of invertebrates such as the land snail Cornu aspersum. Herein, juvenile snails (26.23 ± 0.5 mm shell diameter and 8.23 ± 1.02 g body weight) were fed L. plantarum probiotic-enriched rations in two different proportions (1.25 mg and 2.5 mg), and their growth rate was monitored for three months. Additionally, the RNA/DNA and Bax/Bcl-2 ratios, HSP gene expression and protein levels, and ND2 expression, were measured in the hepatopancreas, digestive tract, and mantle. Although the snails' growth rate was not affected, the RNA/DNA ratio presented an increase in various tissues, indicating an intense physiological response. Also, probiotic administration demonstrated low levels of the Bax/Bcl-2 ratio. HSP levels were higher in the presence of probiotics, probably signaling an attempt by the animal to face potentially stressful situations. Finally, ND2 expression levels in the hepatopancreas indicate intense metabolic and antioxidant activity.

Identification of reference genes and analysis of heat shock protein gene expression (Hsp90) in arta (Calligonum comosum L.) leaf under heat stress

In southern Tunisia, plants are subjected to high temperatures and disturbances, causing the degradation of soils and plants. Studying wild plants and their response to heat stress can facilitate their conservation and help ecosystem rehabilitation. Understanding heat stress responses has gradually become a hotspot in desert plant research. The adaptive responses of Calligonum comosum species to high temperature were examined in this regard. For C.comosum gene expression studies, reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) is the best technique, in addition to the use of appropriate reference genes. So, in our research, we analyzed four different housekeeping genes to select the most stable one during heat stress conditions. To analyze the results, we applied various statistical programs, including GeNorm, Normfinder, Bestkeeper, and the comparative ΔCt method, to identify the most appropriate reference genes for normalizing gene expression under heat stress conditions. Ultimately, our analysis revealed that α-tubulin (α-tub) emerged as the most stable reference gene.Additionally, we subjected C. comosum plants to thermal stress to elucidate the expression patterns of heat shock protein (hsp90) genes. Fourteen-day-old plants were exposed to a temperature of 60 °C for a period ranging from 30 to 240 min. The highest level of hsp90 gene expression was realized upon exposure to heat for 180 min, underscoring the upregulation of this protein in response to thermal stress.This study is the first systematic investigation for selecting reference genes for qPCR in C. comosum under heat stress. These findings will substantially benefit gene expression studies conducted under similar challenging conditions, contributing to our understanding of desert plant responses to heat stress.

Molecular and behavioural responses of the mussel Mytilus edulis exposed to a marine heatwave

Marine heatwaves (MHW) threaten marine organisms and tend to increase in frequency and intensity. We exposed the blue mussel Mytilus edulis to a MHW lasting 23 days, including two 10-d periods of thermal intensity increase of +5 °C (20 °C–25 °C) interspersed by 1 day back to 20 °C, followed by a 4-d recovery period. We investigated behaviour responses of mussels and gene expression changes relative to the circadian rhythm (Per), oxidative stress (SOD), cellular apoptosis (CASP3), energy production (ATPs), and general stress response (hsp70). Results showed that the MHW disturbed the valve activity of mussels. Particularly, mussels increased the number of valve micro-closures, showing a stressful state of organisms. Mussels also decreased Per, CASP3, ATPs, and Hsp70 gene expression. Some behavioural and molecular effects persisted after the MHW, suggesting a limited recovery capacity of individuals. This work highlighted the vulnerability of M. edulis to a realistic MHW.

Transcriptional modulation of heat shock proteins and adipogenic regulators in bovine adipocytes following heat exposure

This research endeavored to elucidate the transcriptional modulation of heat shock proteins and adipogenic regulators in bovine subcutaneous adipocytes following thermal exposure. Post-differentiation, mature adipocytes were subjected to three treatments of control (CON), moderate (MHS), and extreme (EHS) heat stress. These treatments consist of thermal conditions at temperatures of 38 °C (CON), 39.5 °C (MHS), or 41 °C (EHS) along with of 3 or 12 h. There was no statistically significant variations observed in the gene expressions of HSP27 and HSP70 when comparing CON with MHS across both exposures. Contrastingly, when comparing CON with EHS, an upregulation (P < 0.01) in HSP27 gene expression was evident for both 3 and 12 h of incubation, while HSP70 gene expression exhibited elevation (P < 0.01) at the 3-h mark, with no change observed at 12 h. Protein quantification, however, revealed an elevation (P < 0.01) in HSP27 and HSP70 for both CON vs. MHS and CON vs. EHS at the 12-h exposure. This trend in protein level mirrored (P < 0.05) that of proliferator-activated receptor-gamma (PPARγ). Elevated (P < 0.05) protein levels of fatty acid synthase (FAS) were exclusively discernible in the CON vs. MHS. Increased (P < 0.01) transcriptional activity of PPARγ, CCAAT/enhancer-binding protein alpha (C/EBPα), stearoyl-CoA desaturase (SCD), and FAS was evident in the CON vs. EHS comparison. Complementary to these molecular findings, an augmented lipid droplet accumulation was observed (P < 0.01) in EHS-exposed adipocytes progressively from day 6 through day 9. Our current study highlights how different levels and lengths of heat stress can impact the activity of important heat-related proteins and factors that play a role in fat development in beef cattle. These findings can help guide strategies to manage how beef cattle are exposed to heat, which can affect fat storage and ultimately the quality of the meat's marbling.

The Effect of high-intensity interval training with caloric restriction on gene expression of apoptosis indices HSP70 and HSP60 in the myocardium of male rats

Background and aims: The purpose of this study was to investigate the effect of high-intensity interval training combined with calorie restriction on the expression of HSP70 and HSP60 in the myocardium of male rats. Methods: A total of 27 male rats were randomly divided into three groups (9 rats in each group): control, caloric restriction, and caloric restriction with high-intensity interval training. During the study, the control group had free access to water and food, but the amount of food in the food restriction group was limited to 50% and the calorie restriction-exercise group was limited to 25% of the control group. The restriction-exercise group was subjected to intense interval training for eight weeks. The gene expression of apoptotic indicators HSP70 and HSP60 was assessed using the real-time PCR method. The data were analyzed using Shapiro-Wilk test and ANOVA test. Results: Body weight in both calorie restriction groups significantly decreased (P=0.001). The results of the post hoc test showed a significant increase in the expression of HSP70 and HSP60 genes in the myocardium of male rats in the calorie-restricted-exercise group compared to the calorie-restricted and control groups (P=0.001). Additionally, no significant difference was observed between the two restriction and control groups in terms of HSP70 and HSP60 (P=0.102). Conclusion: The combination of high-intensity interval training and mild food restriction significantly increased HSP60 and HSP70 expression.

Melatonin Application Induced Physiological and Molecular Changes in Carnation (Dianthus caryophyllus L.) under Heat Stress

Carnation is one of the most important ornamental plants worldwide; however, heat stress is a problem, which affects carnation cultivation. The harmful effects of heat stress include impaired vegetative development and reduced floral induction. In this study, to enhance carnation growth under conditions of heat stress, various concentrations of melatonin were added to in vitro culture media. The mechanism by which melatonin reduced heat stress damage was then studied by taking measurements of morphological parameters, levels of reactive oxygen species (ROS), antioxidant enzymes, and malondialdehyde (MDA), as well as differential gene expression, in carnation plants during in vitro culture. These data revealed that untreated carnation plants were more harmed by conditions of heat stress than plants treated with melatonin. Melatonin at concentrations of 5 and 10 mM increased chlorophyll content, fresh weight, and plant height to a greater extent than other concentrations. Melatonin may, thus, be used to alleviate damage to carnations caused by heat stress. The application of melatonin was also found to reduce oxidative damage and enhance antioxidant defense mechanisms. In addition, the expression of heat-related genes was found to be upregulated; in melatonin-treated plants, an upregulation was recorded in the expression of GAPDH, DcPOD1, DcPOD2, DcPOD3, Gols1, MBF1c, HSF30, HSP101, HSP70, and sHSP (MT) genes. In short, we found that melatonin treatment increased heat tolerance in carnation plants. The data presented here may serve as a reference for those seeking to enhance the growth of plants in conditions of heat stress.

Nucleotides supplementation (Nucleoforce fish™) in Caspian roach (Rutilus caspicus) diet: Growth performance, skin mucosal immune response, and resistance to salinity stress

In this study, the growth, epidermal mucosal immunity, expression of growth-related genes, cross-protection, and resistance to salinity stress of Caspian roach were scrutinized in response to dietary levels of nucleotides (NT). Accordingly, 1200 fish (0.51 ± 0.01 g) were fed ad libitum with a basal diet (38.88 % crude protein and 10.04 % crude lipid in dry basis) containing incremental levels of NT at 0 (NT-0; control), 0.3 g kg−1 (NT-0.3), 0.6 g kg−1 (NT-0.6), and 1.2 g kg−1 (NT-1.2) for 8 weeks in triplicates. The growth performance was significantly increased in the fish fed with NT-0.6 and NT-1.2 diets compared to the control group (p < 0.05). A significant elevation in the growth hormone and insulin-like growth factor-I gene expression was recorded in NT-added groups at 0.6 and 1.2 g kg−1 compared to the control group (p < 0.05). In contrast to the control group, feeding on NT-0.6 and NT-1.2 diets had a remarkable effect on the skin mucus soluble protein and immunoglobulin levels (p < 0.05). After the feeding trial, we examined how salinity stress (15 g/l salinity) lonely and salinity stress under non-lethal thermal shock (+10 °C) affected heat shock protein (HSP70). Then, the mRNA expression of HSP70 gene from the gill was analyzed at 0, 2, 8, and 24 h post-challenge tests. The HSP70 gene expression level was approximately up-regulated more than 2-fold in NT-6 and NT-1.2 treatments compared to the control group under the salinity stress. Altogether, this research represents that the addition of NT at 0.6 and 1.2 g kg−1 in Caspian roach diet can improve overall performance and resistance to salinity stress.

Effect of cryopreservation on motility, DNA integrity and gene expression in grey mullet, Mugil cephalus sperm

This study documents the effect of cryopreservation on motility, DNA integrity, and gene expression in Mugil cephalus sperm. Fresh sperm were cryopreserved using V2 extender (V2E) or 0.3 M glucose, each in combination with one of three cryoprotective agents (CPAs), i.e., 10 % of dimethylsulfoxide, ethylene glycol, or glycerol, all at once. After two different storage (7- vs 60- day) periods in liquid nitrogen, sperm samples were thawed. Single-cell gel electrophoresis was used to detect the DNA integrity. Heat shock proteins (HSPs), HSP70, HSP90 and glutathione peroxidase (GPx2) genes mRNA expression levels was documented using qRT-PCR. The results demonstrated that among 0.3 M glucose + CPAs combinations, EG recorded higher frozen-thawed motility 69 % (7- day) and 59 % (60- day). Similarly, in V2E + CPAs combinations, EG recorded higher frozen-thawed motility 31 % (7- day) and 26 % (60- day). The DNA integrity of all thawed sperm (both periods) did not differ from that of fresh sperm. The qRT-PCR results revealed that in the combination of 0.3 M glucose + CPAs, the level of HSP90 and GPx2 gene expression was found to be upregulated in frozen-thawed sperm on both periods. Whereas, the expression level of the HSP70 gene was down-regulated. On the contrary, in the combination of V2E + CPAs, the expression levels of HSP70, HSP90 and GPx2 genes could not be detected on both periods. Overall, the findings of this study demonstrate that the cryomedium (extender + cryoprotectant) has a more influential role in the motility and levels of gene expression in the frozen-thawed sperm of M. cephalus.

Potential synergistic regulation of hsp70 and antioxidant enzyme genes in Pyropia yezoensis under high temperature stress

High temperatures have been regarded as one of the main factors limiting Pyropia yezoensis cultivation in recent years. Both the heat shock proteins and antioxidant enzymes genes were activated timely before heat damage to the cell. Here, the activities of different antioxidant enzymes under different conditions, including HSF inhibitor, CaM inhibitor, and HSP70 protein inhibitor were determined. Then, RNA-seq was performed using samples that had been pretreated with corresponding chemicals under high temperature conditions. The principal component analysis inferred that the shift of gene transcription patterns under high temperature conditions was closely related to HSF since the addition of QR suppressed the changes in gene transcription pattern. The hsp70 genes and antioxidant enzyme genes were categorized into three groups by the method of WGCNA. And the HSE sites were identified using plantPAN and FIMO in most genes of the categories showed relationship to HSF. Quantitative RT-PCR analysis of designated genes (FMPK >10) indicated that the expression of hsp70 and antioxidant enzyme genes in Py. yezoensis showed various transcriptional patterns, including upregulation, downregulation, and no significant changes in response to high temperature stress. The classification between qRT-PCR and WGCNA results showed a certain consistency. Considered the results above together, it was speculated that there were different HSFs regulated the expression of different genes of hsp70 and antioxidant enzymes, and HSP70 might also regulate the expression of anti-oxidase genes under the conditions of long-term stress. Additionally, transcription factors other than HSF might be involved in the high temperature stress response. The results indicated the complexity of heat stress response mechanisms in Py. yezoensis.

The Physiological and Biochemical Response of Ribbed Mussels to Rising Temperatures: Benefits of Salt Marsh Cordgrass.

Salt marsh ecosystems are heavily reliant on ribbed mussel (Geukensia demissa) populations to aid in rapid recovery from droughts. The focus of this study was thus to document the effects of rising temperatures on ribbed mussel populations in a Georgia salt marsh. Seven lab and eight field experiments were used to assess the effects of current air temperatures on mussels at two high marsh (HM) sites with short and sparse cordgrass and one mid marsh (MM) site with tall and dense cordgrass. Field results in 2018 and 2019 indicate that ribbed mussels were experiencing extremely high temperatures for prolonged periods of time at the landlocked high marsh (LHM) site. In 2018, the highest temperature (54°C) and longest high temperature events, HTEs (58 days), that is, consecutive days with temperatures≥40°C, were recorded at this site. When laboratory temperatures were increased from 20 to 36°C, mean heart rates increased by an average of 19 bpm for mussels from both high and MM sites respectively. When field temperatures rose from 20°C in April to 40°C in September 2019, mean heart rates increased by an average of 10 bpm for HM mussels and by 26.3 bpm for MM mussels. Under identical laboratory and field conditions, mean heart rates for mussels from the LHM site with the highest temperatures, increased by<1 bpm and 3.7 bpm respectively. Evidence of the potential role of shade on mussel aggregates was provided by examining whether mussels from the edge of mussel aggregates with little to no cordgrass for shade were more stressed than those living at the center of mussel aggregates. In the absence of shade, mean body temperatures for mussels at the edge of mussel aggregates were up to 8°C higher than for those living in the center underneath a dense tuft of cordgrass. Despite high body temperatures, mean heart rates and Hsp70 gene expression were lower for mussels living at the edges. This agrees with the strategy that during prolong exposure to high temperatures, mussels may reduce their heart rate to conserve energy and enhance survival. Alternatively, heat-stressed mussels at the edges of aggregates may not have the resources to express high levels of Hsp70. Increase in the frequency, intensity, and duration of HTEs may stress the physiological and biochemical function of mussel populations to the limit, dictate mussel aggregate size, and threaten the functionality of SE salt marshes.

Assessing the impact of dietary polystyrene nanoplastics on growth performance, immunological parameters, and antioxidant defense in zebrafish (Danio rerio)

AbstractThis trial was performed in order to investigate the response of zebrafish (Danio rerio) to dietary exposure to polystyrene nanoplastics (PS‐NPs) under laboratory conditions on fish growth and health. Healthy zebrafish (n=240) were divided into 12 tanks and fed with diets including 0 (T0), 100 (T1), 500 (T2), and 1000 (T3) mg kg−1 synthesized polystyrenes nanoplastics (PS‐NPs) for 30 days. At the end of trial, fish fed the PS‐NPs supplementation showed weight gain percentages of 79.45%, 70.35%, and 61.88% for T1, T2, and T3 groups, respectively, compared with 87.39% in the control fish. The SOD, GPX and MDA activities and cortisol levels increased by 47%, 32% and 35% and 46%, respectively, especially at high‐dose administration (p &lt; 0.05). The expression of GPX (T2‐81% and T3‐82%) and SOD (T2‐101% and T3‐187%) were remarkably upregulated in T2 and T3 groups. Moreover, the relative gene expression of HSP70, interleukin‐1 (IL1), Interferon γ (IFN‐γ) and tumor necrosis factor‐α (TNF‐α) increased by 178%, 202%, 154% and 307%, respectively, especially at high‐dose administration (p &lt; 0.05). The results of the present study demonstrated that exposure to PS‐NPs especially at high concentrations (500 and 1000 mg kg−1 of diet) negatively influenced growth, health status‐, antioxidant‐, and immunity‐related gene expression responses of zebrafish.

Sperm HSP70: may not be an age-dependent gene but is associated with field fertility in Bali bulls (Bos sondaicus).

This study aimed to analyze the characteristics of the HSP70 gene and protein in spermatozoa of Bali bulls of different age groups and to examine its potential as a biomarker determining bull fertility. This study used frozen semen produced from six Bali bulls divided into two groups based on age (≤ 9 years and ≥ 12 years). Parameters of frozen semen quality analyzed included sperm motility and kinetics using computer-assisted semen analysis, sperm morphological defects using Diff-Quick staining, acrosome integrity using FITC-PNA staining, and DNA fragmentation using acridine orange staining. HSP70 gene expression characterization was analyzed using qRT-PCR, and HSP70 protein abundance was analyzed using enzyme immunoassays. Fertility field data were obtained by analyzing the percentage conception rate for each bull based on the artificial insemination service data contained in the Indonesian-integrated system of the National Animal Health Information System (iSIKHNAS). The results showed significant differences (P<0.05) in total and progressive motility, morphological defects of the neck and midpiece, and tail of sperm, and acrosome integrity between the age groups of Bali bulls. HSP70 gene expression and protein abundance showed no significant differences (P>0.05) in different age groups. HSP70 gene expression correlated with fertility rate (P<0.05). Age affected several semen quality parameters but did not affect HSP70 gene expression and protein abundance. The HSP70 gene molecule could be a biomarker that determines the fertility of Bali bulls.