hsp70 PCR-RFLP as an alternative tool to identify Sauroleishmania species.

Sauroleishmania spp. comprises one of the four Leishmania subgenera, which has been historically considered a non-pathogenic protozoan of reptiles. However, some strains appear to be transiently infective to mammals, and recent findings have detected these parasites in dogs and humans in areas where leishmaniasis is endemic. Herein, the digestion pattern of PCR-RFLP of the 234 bp-hsp70 fragment was evaluated as a simpler and cheaper tool to distinguish the Sauroleishmania species from the other Leishmania subgenera. As a result, the digestion of the 234 bp-hsp70 fragments with HaeIII produced a banding pattern specific to the four Sauroleishmania strains assessed. This technique could contribute to the identification of Leishmania parasites isolated from sandflies, reptiles, or even mammals in fieldworks as an alternative to the use of laborious and expensive methodologies.

Characterizing the Linkage of Systemic Hypoxia and Angiogenesis in High-Grade Glioma to Define the Changes in Tumor Microenvironment for Predicting Prognosis.

High-grade gliomas (HGG) comprising WHO grades 3 and 4 have a poor overall survival (OS) that has not improved in the past decade. Herein, markers representing four components of the tumor microenvironment (TME) were identified to define their linked expression in TME and predict the prognosis in HGG, namely, interleukin6 (IL6, inflammation), inducible nitric oxide synthase(iNOS), heat shock protein-70 (HSP70, hypoxia), vascular endothelial growth receptor (VEGF), and endothelin1 (ET1) (angiogenesis) and matrix metalloprotease-14 (MMP14) and intercellular adhesion molecule1 (ICAM1, extracellular matrix). To establish a non-invasive panel of biomarkers for precise prognostication in HGG. Eighty-six therapy-naive HGG patients with 45 controls were analyzed for the defined panel. Systemic expression of extracellular/secretory biomarkers was screened dot-immune assay (DIA), quantified by ELISA, and validated by immunocytochemistry (ICC). Expression of iNOS, HSP70, IL-6, VEGF, ET1, MMP14, and ICAM1 was found to be positively associated with grade. Quantification of circulating levels of the markers by ELISA and ICC presented a similar result. The biomarkers were observed to negatively correlate with OS (p < 0.0001). Cox-regression analysis yielded all biomarkers as good prognostic indicators and independent of confounders. On applying combination statistics, the biomarker panel achieved higher sensitivity than single markers to define survival. The intra-association of all seven biomarkers was significant, hinting of a cross-talk between the TME components and a hypoxia driven systemic inflammation upregulating the expression of other components. This is a first ever experimental study of a marker panel that can distinguish between histopathological grades and also delineate differential survival using liquid biopsy, suggesting that markers of hypoxia can be a cornerstone for personalized therapy. The panel of biomarkers of iNOS, HSP70, IL-6, VEGF, ET1, MMP14, and ICAM1 holds promise for prognostication in HGG.

The Heat Shock Response as a Condensate Cascade.

The heat shock response (HSR) is a gene regulatory program controlling expression of molecular chaperones implicated in aging, cancer, and neurodegenerative disease. Long presumed to be activated by toxic protein aggregates, recent work suggests a new functional paradigm for the HSR in yeast. Rather than toxic aggregates, adaptive biomolecular condensates comprised of orphan ribosomal proteins (oRP) and stress granule components have been shown to be physiological chaperone clients. By titrating away the chaperones Sis1 and Hsp70 from the transcription factor Hsf1, these condensates activate the HSR. Upon release from Hsp70, Hsf1 forms spatially distinct transcriptional condensates that drive high expression of HSR genes. In this manner, the negative feedback loop controlling HSR activity - in which Hsf1 induces Hsp70 expression and Hsp70 represses Hsf1 activity - is embedded in the biophysics of the system. By analogy to phosphorylation cascades that transmit information via the dynamic activity of kinases, we propose that the HSR is organized as a condensate cascade that transmits information via the localized activity of molecular chaperones.

Altered autophagic flux in GNE mutant cells of Indian origin: Potential drug target for GNE myopathy.

Autophagy phenomenon in the cell maintains proteostasis balance by eliminating damaged organelles and protein aggregates. Imbalance in autophagic flux may cause accumulation of protein aggregates in various neurodegenerative disorders. Regulation of autophagy by either calcium or chaperone play a key role in the removal of protein aggregates from the cell. The neuromuscular rare genetic disorder, GNE Myopathy, is characterized by accumulation of rimmed vacuoles having protein aggregates of ß-amyloid and tau that may result from altered autophagic flux. In the present study, the autophagic flux was deciphered in HEK cell-based model for GNE Myopathy harbouring GNE mutations of Indian origin. The refolding activity of HSP70 chaperone was found to be reduced in GNE mutant cells compared to wild type controls. The autophagic markers LC3II/I ratio was altered with increased number of autophagosome formation in GNE mutant cells compared to wild type cells. The cytosolic calcium levels were also increased in GNE mutant cells of Indian origin. Interestingly, treatment of GNE mutant cells with HSP70 activator, BGP-15, restored the expression and refolding activity of HSP70 along with autophagosome formation. Treatment with calcium chelator, BAPTA-AM restored the cytoplasmic calcium levels and autophagosome formation but not LC3II/I ratio significantly. Our study provides insights towards GNE mutation specific response for autophagy regulation and opens up a therapeutic advancement area in calcium signalling and HSP70 function for GNE related Myopathy.

Dexmedetomidine mitigates lipopolysaccharide-induced acute lung injury by modulating heat shock protein A12B to inhibit the toll-like receptor 4/nuclear factor-kappa B signaling pathway.

Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS): Life-threatening medical conditions characterized by high morbidity and mortality rates, where the inflammatory process plays a crucial role in lung tissue damage, especially in models induced by lipopolysaccharide (LPS). Heat shock protein A12B (HSPA12B) has strong anti-infammatory properties However, it is unknown whether increased HSPA12B is protective against LPS-induced ALI. And Dexmedetomidine (DEX) is a potent α2-adrenergic receptor (α2-AR) agonist that has been shown to protect against sepsis-induced lung injury, however, the underlying mechanisms of this protection are not fully understood. This study utilized bioinformatics analysis and an LPS-induced ALI model to explore how DEX alleviates lung injury by modulating HSPA12B and inhibiting the Toll-like receptor 4/nuclear factor-kappa B (TLR4/NF-κB) signaling pathway. Results indicate that HSPA12B overexpression and DEX pre-treatment markedly mitigated LPS-induced lung injury, which was evaluated by the deterioration of histopathology, histologic scores, the W/D weight ratio, and total protein expression, tumor necrosis factor-alpha (TNF-α), and interleukin-1ß (IL-1ß) in the BALF, and the levels of NO, MDA,SOD and MPO in the lung. Moreover, HSPA12B overexpression and DEX pre-treatment significantly reduces lung injury and inflammation levels by upregulating HSPA12B and inhibiting the activation of the TLR4/NF-κB signaling pathway. On the contrary, when the expression of HSPA12B is inhibited, the protective effect of DEX pre-treatment on lung tissue is significantly weakened.In summary, our research demonstrated that the increased expression of AAV-mediated HSPA12B in the lungs of mice inhibits acute inflammation and suppresses the activation of TLR4/NF-κB pathway in a murine model of LPS-induced ALI. DEX could enhance HSPA12B and inhibit the initiation and development of inflammation through down-regulating TLR4/NF-κB pathway.These findings highlight the potential of DEX as a therapeutic agent for treating ALI and ARDS, offering new strategies for clinical intervention.

Force-regulated chaperone activity of BiP/ERdj3 is opposite to their homologs DnaK/DnaJ.

Polypeptide chains experience mechanical tension while translocating through cellular tunnels, which are subsequently folded by molecular chaperones. However, interactions between tunnel-associated chaperones and these emerging polypeptides under force is not completely understood. Our investigation focused on mechanical chaperone activity of two tunnel-associated chaperones, BiP and ERdj3 both with and without mechanical constraints and comparing them with their cytoplasmic homologs: DnaK and DnaJ. While BiP/ERdj3 have been observed to exhibit robust foldase activity under force, DnaK/DnaJ showed holdase function. Importantly, the tunnel-associated chaperones (BiP/ERdj3) transitioned to a holdase state in the absence of force, indicating a force-dependent chaperone behavior. This chaperone-driven folding event in the tunnel generated an additional mechanical energy of up to 54 zJ, potentially aiding protein translocation. Our findings align with strain theory, where chaperones with higher intrinsic deformability act as mechanical foldases (BiP, ERdj3), while those with lower deformability serve as holdases (DnaK and DnaJ). This study thus elucidates the differential mechanically regulated chaperoning activity and introduces a novel perspective on co-translocational protein folding.

Genome-Wide Identification, Molecular Characterization, and Expression Analysis of the HSP70 and HSP90 Gene Families in Thamnaconus septentrionalis.

Heat shock proteins (HSPs) are a class of highly conserved proteins that play an important role in biological responses to various environmental stresses. The mariculture of Thamnaconus septentrionalis, a burgeoning aquaculture species in China, frequently encounters stressors such as extreme temperatures, salinity variations, and elevated ammonia levels. However, systematic identification and analysis of the HSP70 and HSP90 gene families in T. septentrionalis remain unexplored. This study conducted the first genome-wide identification of 12 HSP70 and 4 HSP90 genes in T. septentrionalis, followed by a comprehensive analysis including phylogenetics, gene structure, conserved domains, chromosomal localization, and expression profiling. Expression analysis from RNA-seq data across various tissues and developmental stages revealed predominant expression in muscle, spleen, and liver, with the highest expression found during the tailbud stage, followed by the gastrula, neurula, and juvenile stages. Under abiotic stress, most HSP70 and HSP90 genes were upregulated in response to high temperature, high salinity, and low salinity, notably hspa5 during thermal stress, hspa14 in high salinity, and hsp90ab1 under low salinity conditions. Ammonia stress led to a predominance of downregulated HSP genes in the liver, particularly hspa2, while upregulation was observed in the gills, especially for hsp90b1. Quantitative real-time PCR analysis corroborated the expression levels under environmental stresses, validating their involvement in stress responses. This investigation provides insights into the molecular mechanisms of HSP70 and HSP90 in T. septentrionalis under stress, offering valuable information for future functional studies of HSPs in teleost evolution, optimizing aquaculture techniques, and developing stress-resistant strains.

Heat shock protein 70 is associated with duration of cell proliferation in early pod development of soybean.

Pod is an important organ for seed production in soybean. Pod size varies among soybean cultivars, but the mechanism is largely unknown. Here we reveal one of the factors for pod size regulation. We investigate pod size differences between two cultivars. The longer pod of 'Tachinagaha' is due to more cell number than in the short pod of 'Iyodaizu'. POD SIZE OF SOYBEAN 8 (GmPSS8), a member of the heat shock protein 70 (HSP70) family, is identified as a candidate gene for determining pod length in a major QTL for pod length. Expression of GmPSS8 in pods is higher in 'Tachinagaha' than 'Iyodaizu' and is highest in early pod development. The difference in expression is the result of an in/del polymorphism　which includes an enhancer motif. Treatment with an HSP70 inhibitor reduces pod length and cell number in the pod. Additionally, shorter pods in Arabidopsis hsp70-1/-4 double mutant are rescued by overexpression of GmPSS8. Our results identify GmPSS8 as a target gene for pod length, which regulates cell number during early pod development through regulation of transcription in soybean. Our findings provide the mechanisms of pod development and suggest possible strategies enhancing yield potential in soybean.

Augmented microglial endoplasmic reticulum-mitochondria contacts mediate depression-like behavior in mice induced by chronic social defeat stress.

Extracellular ATP (eATP) signaling through the P2X7 receptor pathway is widely believed to trigger NLRP3 inflammasome assembly in microglia, potentially contributing to depression. However, the cellular stress responses of microglia to both eATP and stress itself remain largely unexplored. Mitochondria-associated membranes (MAMs) is a platform facilitating calcium transport between the endoplasmic reticulum (ER) and mitochondria, regulating ER stress responses and mitochondrial homeostasis. This study aims to investigate how MAMs influence microglial reaction and their involvement in the development of depression-like symptoms in response to chronic social defeat stress (CSDS). CSDS induced ER stress, MAMs' modifications, mitochondrial damage, and the formation of the IP3R3-GRP75-VDAC1 complex at the ER-mitochondria interface in hippocampal microglia, all concomitant with depression-like behaviors. Additionally, exposing microglia to eATP to mimic CSDS conditions resulted in analogous outcomes. Furthermore, knocking down GRP75 in BV2 cells impeded ER-mitochondria contact, calcium transfer, ER stress, mitochondrial damage, mitochondrial superoxide production, and NLRP3 inflammasome aggregation induced by eATP. In addition, reduced GRP75 expression in microglia of Cx3cr1CreER/+Hspa9f/+ mice lead to reduce depressive behaviors, decreased NLRP3 inflammasome aggregation, and fewer ER-mitochondria contacts in hippocampal microglia during CSDS. Here, we show the role of MAMs, particularly the formation of a tripartite complex involving IP3R3, GRP75, and VDAC1 within MAMs, in facilitating communication between the ER and mitochondria in microglia, thereby contributing to the development of depression-like phenotypes in male mice.

Heat shock protein 70 kDa (HSP70) is involved in the maintenance of pig sperm function throughout liquid storage at 17 °C.

At present, liquid storage is the most efficient method for pig semen preservation. This approach relies upon reducing sperm metabolism, allowing for the maintenance of cell lifespan. In this context, the study of proteins that could protect sperm during liquid storage is of high relevance. The 70 kDa Heat Shock Protein (HSP70) is an anti-apoptotic protein that has been reported to be relevant to sperm survival. Thus, we explored the role of HSP70 during prolonged storage of pig semen at 17 °C. Six semen pools were incubated with YM-1 (0, 0.05, 0.1 and 0.2 µM), an HSP70 inhibitor, and stored at 17 °C for 21 days. On days 0, 4, 10, 14 and 21, sperm quality and function were evaluated through flow cytometry and Computer-Assisted Sperm Analysis (CASA), and HSP70 activity and chromatin condensation were also determined. While inhibition of HSP70 increased progressive motility, Ca2+ and Reactive Oxygen Species (ROS) levels, and mitochondrial activity during the first 10 days of storage, it had a detrimental effect on sperm motility after 14 and 21 days. In spite of this, sperm viability was not altered. We can conclude that HSP70 contributes to the liquid storage of pig semen because it keeps mitochondrial activity low, which is needed for the maintenance of sperm function.

Ultrashort wave diathermy inhibits pulmonary inflammation in mice with acute lung injury in a HSP70 independent way: a pilot study.

BACKGROUND: Acute lung injury (ALI) is a clinical syndrome characterized by pulmonary inflammation. Ultrashort wave diathermy (USWD) has been shown to be effective at in inhibiting ALI inflammation, although the underlying mechanism remains unclear. Previous studies have demonstrated that USWD generates a therapeutic thermal environment that aligns with the temperature required for heat shock protein 70 (HSP70), an endogenous protective substance. In this study, we examined the correlation between HSP70 and USWD in alleviating lung inflammation in ALI. METHODS: Forty-eight male C57BL/6 mice were randomly divided into control, model, USWD intervention (LU) 1, 2, and 3, and USWD preintervention (UL) 1, 2, and 3 groups (n = 6 in each group). The mice were pretreated with LPS to induce ALI. The UL1, 2, and 3 groups received USWD treatment before LPS infusion, while the LU1, 2, and 3 groups received USWD treatment after LPS infusion. Lung function and structure, inflammatory factor levels and HSP70 protein expression levels were detected. RESULTS: USWD effectively improved lung structure and function, and significantly reduced IL-1ß, IL-10, TGF-ß1, and TNF-α levels in both the USWD preintervention and intervention groups. However, HSP70 expression did not significantly differ across the experimental groups although the expression of TLR4 was significantly decreased, suggesting that USWD may have anti-inflammatory effects through multiple signaling pathways or that the experimental conditions should be restricted. CONCLUSIONS: Both USWD intervention and preintervention effectively reduced the inflammatory response, alleviated lung injury symptoms, and played a protective role in LPS-pretreated ALI mice. HSP70 was potentially regulated by USWD in this process, but further studies are urgently needed to elucidate the correlation and mechanism.

Effect of Heat Shock Treatment on the Virulence of Grass Carp Reovirus in Rare Minnow Gobiocypris rarus.

The mode and outcome of fish-virus interactions are influenced by many abiotic factors, among which water temperature is especially important in poikilothermic fish. Rare minnow Gobiocypris rarus is a eurythermal small cyprinid fish that is sensitive to infection with genotype II grass carp reovirus (GCRV). HSP70, a conservative and key player in heat shock response, is previously identified as an induced pro-viral factor during GCRV infection in vitro. Here, rare minnow was subjected to heat shock treatment (HST), 1 h treatment at 32 °C followed by reverting to a normal temperature of 24 °C, and subsequently challenged with GCRV-II at a dosage of 1 × LD50. The effect of HST on GCRV virulence in vivo was evaluated by calculating virus-associated mortality and viral load in both dead and survival fish. The results revealed that HST enhanced the mortality of rare minnow infected with GCRV; the fact that viral loads in the tissue samples of HST-treated fish were significantly higher than those in samples of the control group at 6, 8 d p.i. reflected a faster infection process due to HST. Quantitative gene expression analysis was further employed to show that the expression levels of Hsp70 in intestine and liver tissues from the HST group declined faster than muscle tissue after HST. HST W/O GCRV challenge upregulated proinflammatory cytokines such as MyD88 and Nf-κB, which was in consistence with the inflammation observed in histopathological analysis. This study shed light on the complexity of the interaction between fish abiotic and biotic stress response, which suggested that HST, an abiotic stress, could enhance the virulence of GCRV in Gobiocypris rarus that involved modulating the gene expression of host heat shock, as well as a pro-inflammatory response.

Enhancing Cartilage Metabolism in Rats through a Novel Thermal Stimulation Technique with Photosensitizers.

Although the moderate thermal stimulation of articular cartilage exerts chondroprotective effects, it is difficult to effectively heat deep articular cartilage with conventional methods. Photosensitizers increase the ambient temperature using near-infrared (NIR) radiation, which has high tissue permeability. We hypothesized that the intra-articular administration of photosensitizers and NIR irradiation would exert a greater heating effect on articular cartilage. We aimed to evaluate the heating effect of this method on cultured chondrocytes and rat knee cartilage. In vitro, we irradiated a photosensitizer-containing medium with NIR and measured changes in the medium temperature, cytotoxicity, and gene expression of heat shock protein (HSP) 70 and aggrecan (ACAN). In vivo, the knee joints of rats treated with photosensitizers were irradiated with NIR, and changes in intra-articular temperature and gene expression were measured, alongside histological analysis. The results showed that the medium and intra-articular temperature were raised to approximately 40 °C with no apparent disruption to articular cartilage or the immunohistochemically enhanced staining of HSP70 in chondrocytes. The gene expression of HSP70 and ACAN was increased in both cultured and articular cartilage. In summary, this method can safely heat joints and enhance cartilage metabolism by inducing HSP70 expression in articular cartilage. It presents a new hyperthermia therapy with effective cartilage protection.

Protective effects of L-theanine and dihydromyricetin on reproductive function in male mice under heat stress.

Heat stress can impair the male reproductive function. L-Theanine and dihydromyricetin have biological activities against heat stress; however, their effects on reproductive function in heat-stressed males are unclear. In this study, male mice were given L-theanine, dihydromyricetin, or a combination of both for 28 days, followed by 2 h of heat stress daily for 7 days. All interventions alleviated heat stress-induced testicular damage, improving the testicular organ index, sperm density, acrosome integrity, sperm deformity rate, and hormone levels. Treatment increased the antioxidant enzyme activity and decreased the markers of oxidative and inflammatory stress in the testes. A combination dose of 200 + 200 mg kg-1 d-1 showed the best protective effect. The potential mechanism involves the regulation of HSP27 and HSP70, which regulate the levels of reproductive hormones through the StAR/Cyp11a1/Hsd3b1/Cyp17a1/Hsd17b3 pathway, alleviate inflammation and oxidative stress through the P38/NF-κB/Nrf2/HO-1 pathway, and regulate the Bcl-2/Fas/Caspase3 apoptotic pathway. Overall, L-theanine and dihydromyricetin may play a protective role against heat stress-induced reproductive dysfunction, suggesting their potential use in heat stress-resistant foods.

Correlation of High-Grade Osteosarcoma Response to Chemotherapy with Enhanced Tissue Immunological Response: Analysis of CD95R, IFN-γ, Catalase, Hsp70, and VEGF.

High-grade osteosarcoma, a primary malignant bone tumour, is experiencing a global increase in reported incidence with varied prevalence. Despite advances in management, which include surgery and neoadjuvant chemotherapy often an unsatisfactory outcome is found due to poor or heterogeneous response to chemotherapy. Our study delved into chemotherapy responses in osteosarcoma patients and associated molecular expressions, focusing on CD95 receptor (CD95R), interferon (IFN)-γ, catalase, heat-shock protein (Hsp)70, and vascular endothelial growth factor (VEGF). Employing immunohistochemistry and Huvos grading of post-chemo specimens, we analysed formalin-fixed paraffin-embedded (FFPE) osteosarcoma tissue of resected post-chemotherapy specimens from Dr. Soetomo General Academic Hospital in Surabaya, Indonesia (DSGAH), spanning from 2016 to 2020. Results revealed varied responses (poor 40.38%, moderate 48.08%, good 11.54%) and distinct patterns in CD95R, IFN-γ, catalase, Hsp70, and VEGF expression. Significant differences among response groups were observed in CD95R and IFN-γ expression in tumour-infiltrating lymphocytes. The trend of diminishing CD95R expression from poor to good responses, accompanied by an increase in IFN-γ, implied a reduction in the count of viable osteosarcoma cells with the progression of Huvos grading. Catalase expression in osteosarcoma cells was consistently elevated in the poor response group, while Hsp70 expression was highest. VEGF expression in macrophages was significantly higher in the good response group. In conclusion, this study enhances our understanding of immune-chemotherapy interactions in osteosarcoma and identifies potential biomarkers for targeted interventions.

Capsaicin combined with stem cells improved mitochondrial dysfunction in PIG3V cells, an immortalized human vitiligo melanocyte cell line, by inhibiting the HSP70/TLR4/mTOR/FAK signaling axis.

BACKGROUND: Vitiligo is a common autoimmune skin disease. Capsaicin has been found to exert a positive effect on vitiligo treatment, and mesenchymal stem cells (MSCs) are also confirmed to be an ideal cell type. This study aimed to explore the influence of capsaicin combined with stem cells on the treatment of vitiligo and to confirm the molecular mechanism of capsaicin combined with stem cells in treating vitiligo. METHODS AND RESULTS: PIG3V cell proliferation and apoptosis were detected using CCK-8 and TUNEL assays, MitoSOX Red fluorescence staining was used to measure the mitochondrial ROS level, and JC-1 staining was used to detect the mitochondrial membrane potential. The expression of related genes and proteins was detected using RT‒qPCR and Western blotting. Coimmunoprecipitation was used to analyze the protein interactions between HSP70 and TLR4 or between TLR4 and mTOR. The results showed higher expression of HSP70 in PIG3V cells than in PIG1 cells. The overexpression of HSP70 reduced the proliferation of PIG3V cells, promoted apoptosis, and aggravated mitochondrial dysfunction and autophagy abnormalities. The expression of HSP70 could be inhibited by capsaicin combined with MSCs, which increased the levels of Tyr, Tyrp1 and DCT, promoted the proliferation of PIG3V cells, inhibited apoptosis, activated autophagy, and improved mitochondrial dysfunction. In addition, capsaicin combined with MSCs regulated the expression of TLR4 through HSP70 and subsequently affected the mTOR/FAK signaling pathway CONCLUSIONS: Capsaicin combined with MSCs inhibits TLR4 through HSP70, and the mTOR/FAK signaling pathway is inhibited to alleviate mitochondrial dysfunction and autophagy abnormalities in PIG3V cells.

The effects of the predictability of acclimatory temperature on the growth and thermal tolerance of juvenile Spinibarbus sinensis.

Heated effluent injection, cold hypolimnetic water inputs from dams, and extreme weather events can lead to unpredictable temperature fluctuations in natural waters, impacting fish performance and fitness. We hypothesized that fish exposed to such unpredictable fluctuations would exhibit weaker growth and enhanced thermal tolerance compared to predictable conditions. Qingbo (Spinibarbus sinensis) was selected as the experimental subject in this study. The qingbo were divided into a constant temperature group (C, 22 ± 0.5 °C), a predictable temperature fluctuation group (PF, 22 ± 4 °C, first warming, then cooling within a day) and an unpredictable temperature fluctuation group (UF, 22 ± 4 °C, the order of warming or cooling is random). After 40 days of temperature acclimation, the growth, metabolic rate, spontaneous activity, thermal tolerance, plasma cortisol concentration and liver hsp70 level of the fish were measured. Unexpectedly, neither the PF nor the UF group showed decreased growth compared to the C group. This could be attributed to the fact that temperature variation did not lead to a substantial increase in basic energy expenditure. Furthermore, feeding rates increased due to temperature fluctuations, although the difference was not significant. Both the PF and UF groups exhibited increased upper thermal tolerance, but only the UF group exhibited improved lower thermal tolerance and higher liver hsp70 levels compared to the C group. The qingbo that experienced unpredictable temperature fluctuations had the best thermal tolerance among the 3 groups, which might have occurred because they had the highest level of hsp70 expression. This may safeguard fish against the potential lethal consequences of extreme temperatures in the future. These findings suggested that qingbo exhibited excellent adaptability to both predictable and unpredictable temperature fluctuations, which may be associated with frequent temperature fluctuations in its natural habitat.

Toxicity of the model protein 3×GFP arises from degradation overload, not from aggregate formation.

Although protein aggregation can cause cytotoxicity, such aggregates can also form to mitigate cytotoxicity from misfolded proteins, although the nature of these contrasting aggregates remains unclear. We previously found that overproduction (op) of a three green fluorescent protein-linked protein (3×GFP) induces giant aggregates and is detrimental to growth. Here, we investigated the mechanism of growth inhibition by 3×GFP-op using non-aggregative 3×MOX-op as a control in Saccharomyces cerevisiae. The 3×GFP aggregates were induced by misfolding, and 3×GFP-op had higher cytotoxicity than 3×MOX-op because it perturbed the ubiquitin-proteasome system. Static aggregates formed by 3×GFP-op dynamically trapped Hsp70 family proteins (Ssa1 and Ssa2 in yeast), causing the heat-shock response. Systematic analysis of mutants deficient in the protein quality control suggested that 3×GFP-op did not cause a critical Hsp70 depletion and aggregation functioned in the direction of mitigating toxicity. Artificial trapping of essential cell cycle regulators into 3×GFP aggregates caused abnormalities in the cell cycle. In conclusion, the formation of the giant 3×GFP aggregates itself is not cytotoxic, as it does not entrap and deplete essential proteins. Rather, it is productive, inducing the heat-shock response while preventing an overload to the degradation system.

Heat acclimation improves exercise performance in hot conditions and increases heat shock protein 70 and 90 of skeletal muscles in Thoroughbred horses.

This study aimed to determine whether heat acclimation could induce adaptations in exercise performance, thermoregulation, and the expression of proteins associated with heat stress in the skeletal muscles of Thoroughbreds. Thirteen trained Thoroughbreds performed 3 weeks of training protocols, consisting of cantering at 90% maximal oxygen consumption (VO2max) for 2 min 2 days/week and cantering at 7 m/s for 3 min 1 day/week, followed by a 20-min walk in either a control group (CON; Wet Bulb Globe Temperature [WBGT] 12-13°C; n = 6) or a heat acclimation group (HA; WBGT 29-30°C; n = 7). Before and after heat acclimation, standardized exercise tests (SET) were conducted, cantering at 7 m/s for 90 s and at 115% VO2max until fatigue in hot conditions. Increases in run time (p = 0.0301), peak cardiac output (p = 0.0248), and peak stroke volume (p = 0.0113) were greater in HA than in CON. Pulmonary artery temperature at 7 m/s was lower in HA than in CON (p = 0.0332). The expression of heat shock protein 70 (p = 0.0201) and 90 (p = 0.0167) increased in HA, but not in CON. These results suggest that heat acclimation elicits improvements in exercise performance and thermoregulation under hot conditions, with a protective adaptation to heat stress in equine skeletal muscles.