Hsp70 mRNA Expression Research Articles

Effects of short-term exposure of paracetamol in the gonads of blue mussels Mytilus edulis.

A growing body of literature suggests that pharmaceutical contamination poses an increasing risk to marine ecosystems. Paracetamol or acetaminophen is the most widely used medicine in the world and has recently been detected in seawater. Here, we present the results of 7 days' exposure of blue mussel adults to 40 ng/L, 250 ng/L and 100 μg/L of paracetamol. Histopathology shows that haemocytic infiltration is the most observed condition in the exposed mussels. The mRNA expression of VTG, V9, ER2, HSP70, CASP8, BCL2 and FAS in mussel gonads present different patterns of downregulation. VTG and CASP8 mRNA expression show downregulation in all exposed mussels, irrespective of sex. The V9, HSP70, BCL2 and FAS transcripts follow a concentration-dependent variation in gene expression and may therefore be considered good biomarker candidates. ER2 mRNA expression shows a downregulated trend, with a clearer dose-response relationship in males. In conclusion, this study suggests that paracetamol has the potential to alter the expression of several genes related to processes occurring in the reproductive system and may therefore impair reproduction in blue mussels.

Polymorphism of Duck HSP70 Gene and mRNA Expression under Heat Stress Conditions

Polymorphism of Duck HSP70 Gene and mRNA Expression under Heat Stress Conditions

Oxidative stress, immunological response, and heat shock proteins induction in the Chinese Mitten Crab, Eriocheir sinensis following avermectin exposure.

In this study, the Chinese mitten crabs, Eriocheir sinensis were exposed to avermectin at 0.03, 0.06, 0.12, 0.24, and 0.48 mg/L respectively for 96 hours. The results showed that levels of superoxide dismutase, catalase, and glutathione peroxidase in hepatopancreas were slightly induced at concentration of 0.03 and 0.06 mg/L, but significantly (P < .05) decreased at higher concentrations, meanwhile similar trend of the activities of acid phosphatase, alkaline phosphatase and lysozyme were observed. Significant induction of HSP70 and HSP90 mRNA expression was detected at 24 hours whereas no significant change was found in HSP60. In addition, levels of reactive oxygen species in hepatocytes increased in dose- and time- dependent manners, and cell viabilities of hepatocytes and haemocytes decreased. These results indicated that sublethal concentration exposure of avermectin had a prominent oxidative stress effect on E. sinensis based on the antioxidative and immunological activity inhibition, and HSP60, 70, and 90 perform a protective response during the exposure.

Salinity and bisphenol A alter cellular homeostasis and immune defense by heat shock proteins in the intertidal crab Macrophthalmus japonicus

Heat shock proteins (HSPs) play crucial roles in cellular metabolic processes, as well as in protective immune responses. In this study, we determined the cellular and immune-related functions of HSPs in the intertidal mud crab Macrophthalmus japonicus under osmotic and bisphenol A (BPA) exposure stress. We performed transcriptional profiling of six HSP genes (HSP70, HSP60, HSP90, HSP83, HSP40, and HSP21) in M. japonicus gills and hepatopancreas. Tissue-specific responses were observed as differentially expressed HSP transcripts in M. japonicus. Under osmotic stress, expression levels of HSP70, HSP90 and HSP40 mRNA increased at high salinity (40 psμ), and decreased at low salinity (10 psμ) in M. japonicus gills. In contrast, HSP60 expression was significantly diminished under high salinity, and increased under low salinity. HSP21 was down-regulated under all salinity changes over 1, 4, and 7 days. In the hepatopancreas, under osmotic stress, most HSP transcripts were up-regulated at an early exposure time point (day 1) with subsequent down-regulation (day 7). On day 4, HSP70, HSP60, HSP90 and HSP40 transcript levels increased in M. japonicus, whereas HSP83 and HSP21 transcript levels decreased in the hepatopancreas following most salinity changes. In addition, BPA exposure generally induced HSP responses in the gills. In particular, HSP70 and HSP83 were significantly up-regulated at all concentrations of BPA on days 1 and 4. However, induced responses of HSP transcripts diminished by day 7, except for HSP83 and HSP40. In the hepatopancreas, HSP70 and HSP90 mRNA expression showed significant up-regulation, whereas transcription of HSP60, HSP83, and HSP21 was down-regulated on days 1 and 4. These results suggested that HSPs are differently involved in the molecular adaptation and defense responses of crabs to BPA and osmotic stress.

Relationship Between Heat Shock Protein Expression and Obesity With and Without Metabolic Syndrome.

Background: Obesity is considered a chronic inflammatory disease in which the physiological mechanism responsible for reducing inflammation is weakened, prompting low-grade inflammation throughout the body. One of the key stress response systems that is dysregulated in obesity is the heat shock response, which is a critical defense mechanism that is activated in stressful conditions. Obesity is primary to metabolic syndrome (MetS) as it appears to lead to the increase in other MetS risk factors. Aim of the Study: We aimed to investigate the different expression levels of intracellular heat shock protein (iHSP) 70 and iHSP27 in obese patients with and without MetS and compare these levels to those of a lean control group. Patients and Methods: One hundred ten lean subjects were compared with 44 obese subjects without MetS and 56 obese subjects with MetS. HSP70 and HSP27 mRNA expression levels were measured by quantitative real-time polymerase chain reaction. Results: iHSP70 mRNA expression was significantly higher in obese subjects without MetS than in lean subjects (p = 0.04), whereas iHSP70 mRNA expression was significantly lower in obese subjects with MetS than in those without MetS (p = 0.02) as well as in those in the lean group (p = 0.03). iHSP27 mRNA expression was significantly lower in obese subjects with MetS than in those without MetS and in lean subjects (p = 0.037 and 0.031, respectively). Conclusion: We conclude that the intracellular expression levels of HSP70 and HSP27 may play an important role in the pathogenesis of MetS.

Feather HSP70: a novel non-invasive molecular marker for monitoring stress induced by heat exposure in broilers.

Poultry well-being and economic burden due to heat stress (HS) are of great importance to the poultry industry. Efficient design and effective evaluation of any strategies to alleviate the adverse effects of HS on poultry production require an accurate measurement of stress. However, current methods for monitoring stress in poultry are less than ideal, as they are invasive or subjective, and therefore variable. Here, we demonstrated that HSP70, the well-established intracellular stress chaperone, is expressed in chicken feather and is responsive to HS, and could therefore be used as a stress marker. Growing feathers were collected from the same bird (Cobb500, n = 9) in the morning (barn temperature 24°C) and afternoon (barn temperature 29.5°C) in the summer (2016 June 21) in Arkansas, USA. In the afternoon, the birds were panting and their core body temperatures were significantly higher compared to the morning time, as illustrated by iButton data thermo-loggers. Concomitantly, blood HSP70 mRNA and feather HSP70 expression (mRNA and protein) were significantly increased in the afternoon compared to the morning time. Similarly, HSP70 protein expression in the duodenum was also significantly higher in the afternoon compared to the morning period. Together, these finding identify feather HSP70 as a novel non-invasive molecular signature that mirrors the intracellular and systemic stress, which can be useful to monitor well-being of chickens and other avian species under different challenges.

Supplementing an immunomodulatory feed ingredient to improve thermoregulation and performance of finishing beef cattle under heat stress conditions.

This experiment compared physiological and productive responses in finishing beef cattle managed under heat stress conditions, and supplemented (SUPP) or not (CON) with an immunomodulatory feed ingredient (Omnigen-AF; Phibro Animal Health, Teaneck, NJ). Crossbred yearling cattle (¾ Bos taurus × ¼ Bos indicus; 64 heifers and 64 steers) were ranked by initial body weight (BW) (440 ± 3 kg) and sex, and allocated to 1 of 16 unshaded drylot pens (8 heifers or steers/pen). Pens within sex were randomly assigned to receive SUPP or CON (n = 8/treatment). Cattle received a total-mixed ration (91% concentrate inclusion and 1.21 Mcal/kg of net energy for gain; dry matter [DM basis]) during the experiment (day 0 to 106). The immunomodulatory feed was offered as a top-dress to SUPP pens (56 g/d per animal; as-fed basis) beginning on day 7. Cattle BW were recorded on day 0, 14, 28, 42, 56, 70, 84, 98, and 106. Feed intake was evaluated from each pen by recording feed offer daily and refusals biweekly. Intravaginal temperature of heifers was recorded hourly from day 1 to 6, 29 to 41, and 85 to 97. Environmental temperature humidity index (THI) was also recorded hourly throughout the experiment, and averaged 79.8 ± 0.6. Concurrently with BW assessment, hair samples from the tail-switch were collected (3 animals/pen) for analysis of hair cortisol concentrations. Blood samples were collected on day 0, 28, 56, 84, and 106 from all animals for plasma extraction. Whole blood was collected on day 0, 56, and 106 (3 animals/pen) for analysis of heat shock protein (HSP) 70 and HSP72 mRNA expression. Cattle were slaughtered on day 107 at a commercial packing facility. Results obtained prior to day 7 served as independent covariate for each respective analysis. Heifers receiving SUPP had less (P ≤ 0.05) vaginal temperature from 1500 to 1900 h across sampling days (treatment × hour, P < 0.01; 39.05 vs. 39.19 °C, respectively; SEM = 0.04), when THI ranged from 85.3 to 90.1. Expression of HSP70 and HSP72 was less (P ≥ 0.03) for SUPP cattle on day 106 (22.6- vs. 51.5-fold effect for HSP70, SEM = 9.7, and 11.0- vs. 32.8-fold effect for HSP72; treatment × day, P ≤ 0.04). No treatment effects were detected (P ≥ 0.22) for performance, carcass traits, plasma concentrations of cortisol and haptoglobin, or hair cortisol concentrations. Results from this study suggest that SUPP ameliorated hyperthermia in finishing cattle exposed to heat stress conditions, but such benefit was not sufficient to improve productive responses.

Pretreatment with probiotic Enterococcus faecium NCIMB 11181 ameliorates necrotic enteritis-induced intestinal barrier injury in broiler chickens

The dysfunction of tight-junction integrity caused by necrotic enteritis (NE) is associated with decreased nutrient absorption and gut injury in broiler chickens. Although probiotic Enterococcus faecium (E. faecium) has been reported to possess immune-regulatory characteristics and can prevent diarrhea in pigs, very little information exists in relation to the specific regulatory impact of E. faecium NCIMB 11181 on NE-induced intestinal barrier injury of broiler chickens. This study was conducted to investigate the protective effects of probiotic E. faecium NCIMB 11181 on NE-induced intestinal barrier injury in broiler chickens. The study also aimed to elucidate the mechanisms that underpin these protective effects. One hundred and eighty Arbor Acres (AA) broiler chicks (one day old) were randomly assigned using a 2 × 2 factorial arrangement into two groups fed different levels of dietary E. faecium NCIMB 11181 (0 or 2 × 108 CFU/kg of diet) and two disease-challenge groups (control or NE challenged). The results showed that NE induced body weight loss, intestinal lesions, and histopathological inflammation, as well as intestinal-cell apoptosis. These symptoms were alleviated following the administration of probiotic E. faecium NCIMB 11181. Pretreatment with probiotic E. faecium NCIMB 11181 significantly upregulated the expression of the Claudin-1 gene encoding a tight-junction protein. Claudin-1 and HSP70 protein expression were also increased in the jejunum regardless of NE infection. Furthermore, NE-infected birds fed with E. faecium displayed notable increases in MyD88, NF-κB, iNOS, PI3K, GLP-2, IL-1β, IL-4, and HSP70 mRNA expression. E. faecium NCIMB 11181 administration also significantly improved the animals’ intestinal microbial composition regardless of NE treatment. These findings indicated that addition of E. faecium NCIMB 11181 to poultry feed is effective in mitigating NE-induced gut injury, possibly by strengthening intestinal mucosal barrier function, as well as modulating gut microflora and intestinal mucosal immune responses.

Alpha lipoic acid supplementation ameliorates the wrath of simulated tropical heat and humidity stress in male Murrah buffaloes.

A supplement which ameliorates temperature-humidity menace in food producing livestock is a prerequisite to develop climate smart agricultural packages. A study was conducted to investigate the heat stress ameliorative efficacy of alpha lipoic acid (ALA) in male Murrah water buffaloes (Bubalus bubalis). Eighteen animals (293.61 ± 4.66Kg Bwt) were randomly allocated into three groups (n = 6); NHSC (non-heat-stressed control), HS (heat-stressed) and HSLA (heat-stressed-supplemented with ALA@32mg/kg Bwt orally) based on the temperature humidity index (THI) and ALA supplementation. HS and HSLA were exposed to simulated heat challenge in a climatically controlled chamber (40°C) for 21 consecutive days, 6h daily. Physiological responses viz. Respiration rate (RR), Pulse rate (PR) and Rectal temperature (RT) wererecorded daily before and after heat exposure. Blood samples were collected at the end of heat exposure on days 1, 6, 11, 16, and 21 and on day 28 (7th day post exposure which is considered as recovery) for peripheral blood mononuclear cells (PBMCs) separation, followed by RNA and Protein extraction for Real time quantitative PCR and Western blot analysis respectively, of heat shock proteins (HSPs). Two-way repeated measure ANOVA was performed between groups at different experimental periods. RR (post exposure) in HS and HSLA was significantly higher (P < 0.05) than NHSC from day 1 onwards but HSLA varied significantly from the HS 8th day onwards. Post exposure RT and PR in both HS and HSLA varied (P < 0.05) from NHSC throughout the study; but between HS and HSLA, RT significantly varied on initial 2days and last 6days (from days 16 to 21). HSP70 mRNA expression significantly up regulated in high THI groups with respect to the low THI group throughout the experimental period. During chronic stress (days 16 and 21) HSP70 significantly (P < 0.05) increased in HS but not in HSLA (P > 0.05) with respect to NHSC. ALA supplementation up-regulates and sustains (P < 0.05) the expression of HSP90 in HSLA in comparison to the HS and NHSC. HSP105 expression was significantly up-regulated (P < 0.05) in HS on days 16 and 21 (during long-term exposure) but only on day 21 (P < 0.05) in HSLA. HSP70, HSP90, and HSP105 protein expression dynamics were akin to the mRNA transcript data between the study groups. In conclusion, supplementing ALA ameliorates the deleterious effect of heat stress as reflected by improved physiological and cellular responses. ALA supplementation improved cellular antioxidant status and sustained otherwise easily decaying heat shock responses which concertedly hasten the baton change from a limited window of thermo tolerance to long run acclimatization.

Analysis of the growth performances, muscle quality, blood biochemistry and antioxidant status of Micropterus salmoides farmed in in-pond raceway systems versus usual-pond systems

In-pond raceway system (IPRS) was introduced to China as a sustainable pond farming approach since 2012 that effectively reduces pollution and facilitates management. To assess how IPRS affect the growth and quality of the farmed fish Micropterus salmoides, various rearing parameters were compared to the usual-pond system (UPS). The growth performances, muscle nutritional quality, serum biochemical indexes, antioxidant enzymatic activities and genes expression were analyzed in Micropterus salmoides with body weight of (8.25 ± 0.51) g reared in IPRS versus usual-pond system(UPS) at stocking densities of 37,500 ind./ha with triplication for 240 days. The final body weight of M. salmoides reared in IPRS and UPS was 482.52 ± 22.53 g and 539.51 ± 34.91 g. Survival rates were higher, despite lower weight gain, in IPRS compared to UPS, with no significant differences in specific growth rate or the food conversion ratio. The hepatosomatic index, viscerosomatic index, and lipid content in liver was lower in IPTS compared to URS. Crude fat and crude protein content significantly differed in muscle. EAA/TAA and EAA/NEAA in the muscle was higher in fish reared in IPRS compared to UPS. Saturated fatty acids in the muscle was lower in fish reared in IPRS compared to UPS. IPRS fish had lower percentages of monounsaturated fatty acids and polyunsaturated fatty acids, including EPA and DHA, than in IPRS.Total protein content was higher in the serum of IPRS fish. Total cholesterol and triglyceride content were lower in IPRS fish. Total bilirubin content and alanine aminotransferase activity was lower in IPRS fish. Glutathione peroxidase activity was higher, while malondialdehyde content was lower, in the liver of IPRS fish. Superoxide dismutase and catalase activity did not differ between the two conditions. mRNA expression of gpx and hsp70 was significantly up-regulated in IPRS compared to UPS. The result show, compared to UPS, IPRS can improve the survival rates, nutritional quality of fish muscle, and liver biological functioning and antioxidant ability of the target fish.

Heat Shock Protein 70 Messenger RNA in Rat Leukocytes Elevates After Severe Intestinal Ischemia–Reperfusion

BackgroundHeat shock protein 70 (HSP70) confers protection against heat shock, oxidative stress, infection, and inflammation in many cell types. A recent study reported that the induction of HSP70 was associated with morphologic protection against ischemia–reperfusion injury (IRI) in the rat small intestine. This study investigated the dynamics of HSP70 in leukocytes during intestinal IRI in a rat model. Materials and methodsSerial blood samples were collected at 60-minute intervals up to 240 min from male Wistar rats (n = 15). The rats were divided into three groups of five each: the control group, the nonlethal IRI group, and the lethal IRI group. Rats belonging to the control group underwent a sham operation, and laparotomy was performed on rats in the lethal and nonlethal IRI groups. The nonlethal group experienced a 30-minute clamping of the superior mesenteric artery, and the lethal group experienced a 75-minute clamping of the superior mesenteric artery. The expression of HSP70 messenger RNA (mRNA) in leukocytes was measured by real-time quantitative polymerase chain reaction. Mixed-effects modeling of repeated measures was used to carry out the statistical analysis. The Bonferroni correction was applied to multiple comparisons. A P value < 0.0167 was considered to indicate statistical significance. ResultsThe expression of HSP70 mRNA in leukocytes increased 60 min after reperfusion in both IRI groups, and it was 12.8 times higher in the lethal group and 3.6 times higher in the nonlethal group compared with the control group. The expression of mRNA in the lethal group was significantly increased compared with the nonlethal group and the control group at 120 and 180 min after reperfusion. At 120 min after reperfusion, the expression of HSP70 mRNA was 6.1 times higher in the lethal group than in the nonlethal group (P = 0.0075) and 17.7 times higher than in the control group (P = 0.0011). At 180 min after reperfusion, the expression of HSP70 mRNA was 6.8 times higher in the lethal group than in the nonlethal group (P = 0.0007) and 4.3 times higher than in the control group (P = 0.0032). Although the expression of HSP70 mRNA in the nonlethal group was elevated in the early stages of reperfusion, there was no difference between the nonlethal group and the control group (P = 0.0212 at 60 min). ConclusionsThe expression of HSP70 mRNA in leukocytes may be a clinically useful indicator for evaluating pathologic conditions in intestinal IRI.

Whole flour and protein hydrolysate from common beans reduce the inflammation in BALB/c mice fed with high fat high cholesterol diet

Common bean (Phaseolus vulgaris L.) is a source of bioactive peptides, but little is known about its effects on hypercholesterolemia, oxidative stress, and the inflammatory process. Therefore, the aim of this study was to evaluate the effect of whole flour and bean protein hydrolysate of common bean variety Carioca on inflammation and oxidative stress in BALB/c mice. Four experimental groups were included in the study: standard diet (SD), high fat high cholesterol diet (HFC), high fat high cholesterol diet and whole bean flour (HFC-F); and high fat high cholesterol diet and bean protein hydrolysate (HFC-PH). Animals fed with bean protein hydrolysate showed lower weight gain and food intake. Animals fed with whole bean flour showed lower alanine aminotransferase and low-density lipoprotein cholesterol levels than animals fed with bean protein hydrolysate. SOD mRNA was lower in HFC, HFC-F and HFC-PH groups whereas SOD concentration was higher in HFC-F and HFC-PH groups. HSP72 mRNA expression was lower in the HFC-F group in relation to HFC-PH. IL-10 and PPARα mRNA expression was lower in HFC-F and HFC-PH groups in comparison with SD. The whole bean flour and bean protein hydrolysate reduced inflammation and the risk factors for cardiovascular diseases in BALB/c mice.

Effects of extracellular Hsp70 and cigarette smoke on differentiated THP-1 cells and human monocyte-derived macrophages

Extracellular Hsp70 (eHsp70) can act as pro-inflammatory mediator and is elevated in blood of chronic obstructive pulmonary disease (COPD) patients. Most of those patients are smokers, and it was suggested previously that cigarette smoke might induce Hsp70 secretion from the circulating cells. Therefore, we aimed to explore inflammation-associated effects of cigarette smoke extract (CSE) and its combinations with eHsp70 in monocyte-derived macrophages (MDMs) and THP-1 cell line, used as systemic component models of COPD. We hypothesized that eHsp70 induces inflammation, but that it can also modulate cigarette smoke extract (CSE)-stimulated inflammatory responses.We assessed IL-8 secretion, TLR2, TLR4 and Hsp70 expressions, MAPKs and NF-κB activation, and cytotoxicity after treating the cells with CSE (2.5 and 5%) and its combinations with low-endotoxin recombinant human (rh) Hsp70, used to mimic eHsp70 effects.CSE induced IL-8 secretion from both cell types, but its combinations with rhHsp70 increased IL-8 release compared to CSE alone only from MDMs. In THP-1, combinations of rhHsp70 with 2.5% CSE induced TLR2 and TLR4 mRNA, while 5% CSE decreased TLR2 expression. In MDMs, CSE alone attenuated TLR2, while rhHsp70 increased TLR2 and lowered TLR4 gene expression. Hsp70 mRNA expression was suppressed in THP-1 with rhHsp70 and CSE; however, the same treatments increased its level in MDMs. CSE had cytotoxic effect only on MDMs, but cytotoxicity was reduced in co-treatments with rhHsp70, which also triggered apoptosis. CSE and rhHsp70 activated p38 and JNK, while ERK was activated only by rhHsp70 in MDMs. In THP-1, 2.5% CSE activated ERK, and 5% CSE activated p38. Inhibition of NF-κB and JNK in MDMs, and ERK and JNK in THP-1 cells, attenuated IL-8 release after rhHsp70 treatment.In conclusion, rhHsp70 provoked pro-inflammatory effects and could also modulate inflammatory response to CSE on protein and gene expression levels in THP-1 cells and MDMs, which suggests that eHsp70 might be implicated in systemic inflammation induced by cigarette smoke.

Distinct contribution of Rac1 expression in cardiomyocytes to anthracycline-induced cardiac injury

Cardiotoxicity is the dose limiting adverse effect of anthracycline-based anticancer therapy. Inhibitor studies point to Rac1 as therapeutic target to prevent anthracycline-induced cardiotoxicity. Yet, supporting genetic evidence is still missing and the pathophysiological relevance of different cardiac cell types is unclear. Here, we employed a tamoxifen-inducible cardiomyocyte-specific rac1 knock-out mouse model (Rac1flox/flox/MHC-MerCreMer) to investigate the impact of Rac1 expression in cardiomyocytes on cardiac injury following doxorubicin treatment. Distinctive stress responses resulting from doxorubicin treatment were observed, including upregulation of systemic markers of inflammation (IL-6, IL-1α, MCP-1), cardiac damage (ANP, BNP), DNA damage (i.e. DNA double-strand breaks (DSB)), DNA damage response (DDR) and cell death. Measuring the acute doxorubicin response, the serum level of MCP-1 was elevated, cardiac mRNA expression of Hsp70 was reduced and cardiac DDR was specifically enhanced in Rac1 deficient mice. The frequency of apoptotic heart cells remained unaffected by Rac1. Employing a subactue model, the number of doxorubicin-induced DSB was significantly reduced if Rac1 is absent. Yet, the doxorubicin-triggered increase in serum ANP and BNP levels remained unaffected by Rac1. Overall, knock-out of rac1 in cardiomyocytes confers partial protection against doxorubicin-induced cardiac injury. Hence, the data provide first genetic evidence supporting the view that pharmacological targeting of Rac1 is useful to widen the therapeutic window of anthracycline-based anticancer therapy by alleviating acute/subacute cardiomyocyte damage. Furthermore, considering published data obtained from the use of pharmacological Rac1 inhibitors, the results of our study indicate that Rac1-regulated functions of cardiac cell types others than cardiomyocytes additionally influence the adverse outcomes of anthracycline treatment on the heart.

The effect of exogenous HSP70 on expression kinetics of HSP70, TLR2, and TLR4 in peripheral blood mononuclear cells and possible crosstalk between them in Black Bengal goat

ABSTRACT The present study aims to investigate the modulatory effect of eHSP70 on major heat stressed genes in in-vitro cell culture system. HSP70 (eHSP70) on mRNA expression profile of heat-shock protein 70 (HSP70), toll-like receptor 2 (TLR2) and TLR4 in peripheral blood mononuclear cells (PBMCs) in cell culture model of Black Bengal goat. After thermal challenge of cultured PBMCs were treated with a specific dose of eHSP70 (50 ng/ml) at 37°C for 0, 1, 2, 4, 8, 12, 16, 20, and 24 h. The mRNA expressions of the above-mentioned genes were determined by quantitative real-time PCR (qRT-PCR). The study showed that on administration of eHSP70 to the PBMCs cell culture system, the relative mRNA expression of iHSP70 and HSP90 increased significantly (p < 0.05) at 4, 20 and 24 h compared with control, and and TLR2 and TLR4 showed significantly higher expression (p < 0.05) at 2, 4, 8 and 24 h compared with control. Immunoreactivity revealed that all genes under study were localized in cytoplasm as well as in nucleus. The present findings specify that eHSP70 is probably involved in heat stress adaptive response in Black Bengal goat and the biphasic expression pattern may be providing a second window of protection during heat stress. Abbreviations: ANOVA: analysis of variance; GAPDH: gleceraldehyde 3-Phosphate dehydrogenase; bp: base pairs; cDNA: complementary DNA; Ct value: cycle threshold values; DEPC: diethyl pyrocarbonate; dNTP’s: deoxynucleoside triphosphate; DW: distilled water; DPBS: dulbecco’s phosphate buffer saline; FBS: fetal bovine serum; HSP: heat-shock protein; HSF: Heat shock factors; IG: immunoglobulin; KDa: kilo dalton; LN2: liquid nitrogen; NF-κB: nuclear factor kappa beta; TLR: toll-like receptor; PBMC: peripheral blood mononuclear cell; eHSP70: exogenous heat-shock protein 70.

Basal and dynamics mRNA expression of muscular HSP108, HSP90, HSF-1 and HSF-2 in thermally manipulated broilers during embryogenesis

BackgroundLimited data are available about the kinetics of mRNA expressions of Heat shock proteins (HSPs) and heat shock factors (HSFs) in the thermally manipulated (TM) broiler chicks during acute heat stress. Therefore, this study aimed to investigate effects of thermal manipulation (TM) of broiler chicken during embryonic days (ED) 12–18 on the basal and dynamics mRNA expression of heat shock proteins (HSP108 and HSP90) and heat shock factors (HSF-1 and HSF-2) in the muscle tissue during late embryogenesis, first week of life and during heat stress (HS) on post-hatch days 14 and 28. One thousand and five hundred fertile Ross 315 broiler eggs were randomly allocated to five groups: control group (37.8 °C), TM1 (38.5 °C for 18 h), TM2 (39 °C for 18 h), TM3 (39.5 °C for 18 h) and TM4 (40 °C for 18 h). Chicks from each treatment group were then randomly sub-divided into two further treatment groups, naïve and thermal challenged (TC). On post-hatch days 14 and 28, thirty chicks from each TC group were subjected to heat stress (41 °C for 6 h), while naïve chicks of each group (n = 30) were kept under regular conditions. The response of chicks to heat stress was investigated by evaluating the change in mRNA expressions of HSP108, HSP90, HSF-1 and HSF-2 in muscle tissue after 1, 3 and 5 h of heat stress.ResultsWhen compared to the control group, TM resulted in significant increases in the basal mRNA expression of HSPs and HSFs during embryogenesis and altered their dynamic expressions in the muscle tissue after heat stress on post-hatch days 14 and 28.Conclusionthe current study indicated short- and long-term enhancement of HSPs and HSFs gene expression which was associated with acquisition of improved thermotolerance in thermally manipulated chicks.

INFLUENCE OF DIETARY VITAMIN C SUPPLEMENTATION ON GROWTH PERFORMANCE, BLOOD BIOCHEMICAL PARAMETERS AND TRANSCRIPT LEVELS OF HEAT SHOCK PROTEINS IN HIGH STOCKING DENSITY REARED BROILER CHICKENS

This study was conducted to evaluate the effects of dietary supplementation of different levels of vitamin C (VC) in broiler chicken reared under high stocking density on the growth performance, blood biochemical parameters and the expression of heat shock protein genes. A total of 150, one day old mixed sex broiler chicks (Cobb 500) were randomly distributed in five equal groups. Group1 was reared in normal stocking density (10.6 birds/m2) and fed on the basal diet (BD) without VC (control). While, chicks in other groups (2, 3, 4 and 5) reared in high stocking density (15.6 birds/m2) and fed on BD supplemented with VC at 0, 200, 400 and 600 mg / kg diet, respectively. Birds reared in high stocking density showed a reduction of the final body weight and total feed intake, with high mortality (6.6%). Moreover, they revealed a significant up-regulation of HSP70 mRNA and elevated HSP90 and HSF1 mRNA expression in heart and liver tissue. Graded dietary levels of VC provided variable protection against the hazard of high density through improved final body weight and total feed intake, decreased the mortality % and downregulated liver HSP70 expression level. However, the best performance was observed in birds supplemented with 200 mg/kg VC (group 3).Key words: chicks' growth performance; gene expression; stocking density; vitamin C

Multigenerational heat acclimation increases thermal tolerance and expression levels of Hsp70 and Hsp90 in the rice leaf folder larvae.

Physiological response and acclimation to thermal stress is a key strategy of insects to cope with changing climate. The underlying mechanism of heat acclimation in insects is still unclear. Here, the heat selection and transcript level response in the larvae of the rice leaf folder Cnaphalocrocis medinalis Güenée, a serious pest of rice in summer, were studied. The survival and fecundity of larvae during multigenerational heat selection at 39 °C were examined, and heat tolerance and mRNA expression of heat shock protein 70 (Hsp70) and 90 (Hsp90) were examined under heat stress. The results showed that survival and fecundity of larvae increased notably and then kept constant after two or three generations of heat selection. Heat selection improved thermal tolerance of larvae. The Hsp70 mRNA expression of the 3rd-instar larvae increased in all five generations of heat selection, but Hsp90 increased only in the first two generations. The response of Hsp70 to 39 °C heat treatment in the larvae kept at 27 °C was different from the larvae exposed to the conditioning heat treatments, but the response of Hsp 90 was similar. Moreover, the Hsp70 and Hsp90 mRNA expression levels were significantly higher in the heat-acclimated larvae than that in the unacclimated larvae at a comparable duration of exposure to 37 and 41 °C. Selection at a high temperature across multiple generations led larvae to heat acclimation, and Hsp70 and Hsp90 were involved in this acclimation process.

Ammonia exposure induced abnormal expression of cytokines and heat shock proteins via glucose metabolism disorders in chicken neutrophils.

Ammonia (NH3) is a highly irritant, alkaline gas. Atmospheric emission of NH3 was recognized as an environmental challenge. As a global issue, the NH3 emission survey with spatially detailed information demonstrated that the sources of atmospheric NH3 include agriculture (livestock wastes, fertilizers) and some industrial activities. As an environmental pollution, excessive NH3 exposure can induce many bird dysfunction. Neutrophils respond to multiple invading pathogens through different mechanisms. In order to investigate the effect of NH3 exposure on broilers' neutrophil, 1-day-old broilers were treated with/without NH3 for 28days. We extracted neutrophils from peripheral blood of chicken with/without NH3 exposure and subsequently stimulated with PMA. Changes of cytokines and inflammatory bodies, heat shock proteins (HSPs), and glucose metabolism of neutrophil were examined in both cases. We not only explored that the index associated with inflammation changed due to NH3 exposure but also observed the status of neutrophils which was treated with PMA stimulation. After NH3 exposure, IL-1β and IL-6 were significantly increased on broilers neutrophil. Inflammatory-related factors (NLRP3, ASC, and caspase-1) were significantly elevated. The mRNA expression of HSP70 and HSP90 was increased significantly. All glucose metabolism indicators were reduced. In summary, we concluded that NH3 enhanced inflammation and disrupted glucose metabolism, and increased the expression of HSPs and inflammatory factors. In addition, the sensitivity of neutrophils to exogenous stimuli was diminished. This information can not only be used to evaluate the damage of NH3-spiked neutrophils to chickens, but also provide clues for human health pathophysiology caused by excess NH3, providing valuable information for NH3 risk management.

Innate-immune responses of tilapia (Oreochromis mossambicus) exposure to acute cold stress.

In this present study, Oreochromis mossambicus tilapia were transferred to cold water at 12°C for various time intervals (1, 4, 8, 24, and 48 hr) and its innate immune response was analyzed by studying cellular and humoral parameters. In vivo, alternative complement pathway activity in blood plasma was rapidly increased at 1 hr of cold water (12°C) exposure. Lysozyme activity and cortisol levels of plasma were increased at 4 and 1 hr, respectively. Surprisingly, only plasma cortisol levels remained unchanged through 24 hr of cold water transfer. Phagocytic ability, phagocytic capacity, and respiratory burst (RB) activity of head kidney (HK) leukocytes and splenocytes showed no any significant changes. In peripheral blood leukocytes, phagocytic capacity, and RB activity were increased at 24 hr of cold water exposure. The expressions of genes involved innate immunity in splenocytes and HK leukocytes of tilapia cold water exposure were analyzed, messenger RNA (mRNA) expressions of HSP70, HSP90, and immunoglobulin M failed to change upon exposure to cold stress. Major histocompatibility complex-I and II mRNAs were significantly increased in tilapia splenocytes at 1 hr of cold water transferred. Whereas myxovirus (Mx) expression was increased in splenocytes and HK leukocytes of tilapia after 1 hr of cold water exposed. Our result reveals that the exposure of tilapia to acute cold stress condition significantly enhances plasma acid phosphatase activity and both phagocytic capacity and RB activity. Furthermore, cold stress significantly stimulates Mx gene expression in splenocytes and HK leukocytes.