Protein Hsp Research Articles

Stress Pathways Induced by Volatile Anesthetics and Failure of Preconditioning in a Mitochondrial Complex I Mutant.

Carriers of mutations in the mitochondrial electron transport chain are at increased risk of anesthetic-induced neurotoxicity. To investigate the neurotoxicity mechanism and to test preconditioning as a protective strategy, this study used a Drosophila melanogaster model of Leigh syndrome. Model flies carried a mutation in ND23 (ND2360114) that encodes a mitochondrial electron transport chain complex I subunit. This study investigated why ND2360114 mutants become susceptible to lethal, oxygen-modulated neurotoxicity within 24 h of exposure to isoflurane but not sevoflurane. This study used transcriptomics and quantitative real-time reverse transcription polymerase chain reaction to identify genes that are differentially expressed in ND2360114 but not wild-type fly heads at 30 min after exposure to high- versus low-toxicity conditions. This study also subjected ND2360114 flies to diverse stressors before isoflurane exposure to test whether isoflurane toxicity could be diminished by preconditioning. The ND2360114 mutation had a greater effect on isoflurane- than sevoflurane-mediated changes in gene expression. Isoflurane and sevoflurane did not affect expression of heat shock protein (Hsp) genes (Hsp22, Hsp27, and Hsp68) in wild-type flies, but isoflurane substantially increased expression of these genes in ND2360114 mutant flies. Furthermore, isoflurane and sevoflurane induced expression of oxidative (GstD1 and GstD2) and xenobiotic (Cyp6a8 and Cyp6a14) stress genes to a similar extent in wild-type flies, but the effect of isoflurane was largely reduced in ND2360114 flies. In addition, activating stress response pathways by pre-exposure to anesthetics, heat shock, hyperoxia, hypoxia, or oxidative stress did not suppress isoflurane-induced toxicity in ND2360114 mutant flies. Mutation of a mitochondrial electron transport chain complex I subunit generates differential effects of isoflurane and sevoflurane on gene expression that may underlie their differential effects on neurotoxicity. Additionally, the mutation produces resistance to preconditioning by stresses that protect the brain in other contexts. Therefore, complex I activity modifies molecular and physiologic effects of anesthetics in an anesthetic-specific manner.

New insights into the structure and function of the complex between the Escherichia coli Hsp70, DnaK, and its nucleotide-exchange factor, GrpE

The 70 kDa heat shock proteins (Hsp70s) play a pivotal role in many cellular functions using allosteric communication between their nucleotide-binding domain (NBD) and substrate-binding domain, mediated by an interdomain linker, to modulate their affinity for protein clients. Critical to modulation of the Hsp70 allosteric cycle, nucleotide-exchange factors (NEFs) act by a conserved mechanism involving binding to the ADP-bound NBD and opening of the nucleotide-binding cleft to accelerate the release of ADP and binding of ATP. The crystal structure of the complex between the NBD of the Escherichia coli Hsp70, DnaK, and its NEF, GrpE, was reported previously, but the GrpE in the complex carried a point mutation (G122D). Both the functional impact of this mutation and its location on the NEF led us to revisit the DnaK NBD/GrpE complex structurally using AlphaFold modeling and validation by solution methods that report on protein conformation and mutagenesis. This work resulted in a new model for the DnaK NBD in complex with GrpE in which subdomain IIB of the NBD rotates more than in the crystal structure, resulting in an open conformation of the nucleotide-binding cleft, which now resembles more closely what is seen in other Hsp/NEF complexes. Moreover, the new model is consistent with the increased ADP off-rate accompanying GrpE binding. Excitingly, our findings point to an interdomain allosteric signal in DnaK triggered by GrpE binding.

Structural changes and features of heat shock protein HSP70 and HSP90 expression in the pyramidal neurons of the hippocampus of rat in the experimental discirculatory encephalopathy modeling

AbstractBackgroundThe heat shock protein (Hsp) system acts as intracellular chaperone molecules responsible for maintaining normal intracellular homeostasis by regulating the correct coagulation of newly synthesized peptides, as well as the transport and degradation of mature proteins. In addition to their role as chaperones, proteins of the Hsp family stabilize the cell membrane and inhibit the caspase‐dependent apoptosis cascadeMethodHypertensive encephalopathy was modeled on 12‐month‐old rats by the method of exposure to negative longitudinal overloads in the caudo‐cranial vector with a 9 G force for 5 minutes with 12 hours interval for 28 days. The behavioral responses, cognitive and mnestic functions, the relative number of pyramidal neurons with hyperchromic cytoplasm and shrunken perikariya, and the level of expression of heat shock proteins 70 kDa (Hsp70) and 90 kDa (Hsp90) were assessedResultAfter 28 days of exposure to 9 G negative longitudinal overloads, the rats showed functional and morphological manifestations characteristic of hypertensive encephalopathy. There was a decrease in the basic reflexes, disorders of the cognitive sphere, as well as decrease in memory and orientation‐exploratory activity. Morphological examination of the hippocampus against the background of pronounced edema revealed a significant increase in the relative number of pyramidal neurons with hyperchromatosis and perikaryon shrinkage in all zones of the pyramidal layer (p <0.01). These signs of reversible and irreversible damage were accompanied by a significant increase in Hsp70 expression, especially pronounced in the CA3 zone (p <0.01). A significant increase in Hsp90 expression was found in the CA1 zone (p <0.05)ConclusionDiscirculatory encephalopathy experimental modeling demonstrates signs of both reversible and irreversible alteration in all regions of the pyramidal layer of hippocampus. Differences in the ability of neurons of the pyramidal layer of different regions of the hippocampus to express Hsp70, Hsp90 were noted. In the CA1 and CA3 zones, compensatory reactions are more pronounced and are accompanied by a more pronounced expression of Hsp70, while in CA1, the expression of Hsp70 and Hsp90 is simultaneously enhanced

Small heat shock proteins as modulators of cell death in Plasmodium falciparum parasites and its human host

Plasmodium falciparum, the predominant cause of severe malaria, thrives within both poikilotherm mosquitoes and homeotherm humans, navigating challenging temperature shifts during its life cycle. Survival in such varying environments necessitate the development of robust mechanisms, including a sophisticated protein folding system to mitigate proteopathy. The parasite needs to control the survival of its host cells which affects its chances of development and propagation. Central to this system are heat shock proteins (Hsps), among which small Hsps (sHsps) play pivotal roles in maintaining proteostasis (protein homeostasis). In both humans and P. falciparum, numerous sHsps have been identified, making them attractive candidates as biomarkers for diagnostic and drug development strategies. Evidence is accumulating suggesting that these sHsps participate in cell death processes, potentially influencing disease pathogenesis. Despite their significance, the precise functions of sHsps in P. falciparum’s adaptation to stress conditions remains largely unknown. Comparative structural analysis of sHsps between human and P. falciparum reveals species-specific variations. Despite conserved tertiary structures, unique motifs are found in parasite sHsps which may modulate specialised chaperone functions. This review discusses the conserved and distinctive motifs of sHsps from the human host and the parasite, offering insights into shared and unique attributes. These findings illuminate the potential for species-specific targeting of sHsps, as players in cell death processes that may foster innovative biomarker identification approaches. As malaria continues to ravage Sub-Saharan Africa, understanding the molecular intricacies guiding parasite survival are essential in the development of interventions with heightened efficacy against this global health crisis.

Hspa5 Deficiency Blocks Intestinal Enterocyte Differentiation in Graft-Versus-Host Disease

Background: Allogeneic hematopoietic stem cell transplantation (allo-HSCT) has been established as an effective therapy for hematological malignancies. Graft-versus-host disease (GVHD) is a life-threatening complication, and the intestine is one of the major sites affected. Currently, immunosuppressive agents remain the preferred choice for GVHD treatment. However, treatment resistance still occurs in a large population, indicating that some tissue damage may be irreversible. Methods: The murine transplant and GVHD model were established as previously described. To characterize the intestinal damage mediated by radiation and GVHD, murine small intestine tissues were collected at day 14 after transplantation. Primary small intestinal crypts were isolated from each group and cultured for five consecutive days to evaluate their ability to proliferate and differentiate. Single-Cell analysis was performed to characterize the impaired intestinal repair after injury. Results: At day 14, the villus-to-crypt ratio was partially recovered in the BM group, but remained imbalanced in the GVHD group. We observed a significant decrease in the expression of Reg1, a gene expressed by enterocytes (EC), in the GVHD groups compared to the BM group. we further utilized a 3D organoid model to investigate intestinal growth and differentiation. The organoid size from the GVHD group was comparable to the BM and control groups. However, the number of buds per organoid was significantly reduced in the GVHD group compared to the BM and control groups. Immunofluorescence staining confirmed that cells in GVHD-derived organoids exhibited high proliferation (stained by Mki67) but impaired differentiation of enterocyte (stained by Aldob). To gain insights into the cellular mechanisms influencing enterocyte differentiation, we dissociated murine small intestinal tissues from each group into single cells and sequenced them using the 10X Genomics platform. Through pseudochronological analysis, each cell was ordered along the predicted lineages and assigned a pseudotime. We found that enterocytes in GVHD exhibited a long and discontinuous trajectory, suggesting explicit blockage of EC differentiation. Functional enrichment analysis showed that pathways related to cell growth and differentiation, such as MAPK, ERBB, and inositol phosphate metabolism, were highly enriched in M2 of the control and BM groups. In contrast, MAPK was dramatically downregulated at the early stage of ISC differentiation in the GVHD group. Next, we analyzed the differentially expressed genes in EP between the GVHD and BM groups. Endoplasmic reticulum (ER) chaperone-related genes from the heat shock protein (Hsp) 70 family (Hspa1a/1b/5), Hsp90 family (Hsp90aa1/ab1), and Hsp40 family (Dnajb1) were significantly decreased in GVHD compared to BM. Notably, Hspa5 (also known as GRP78 or Bip), which serves as a sensor of glucose levels and misfolded proteins, exhibited reduced protein levels, along with decreased IRE1ɑ phosphorylation (p-IRE1ɑ) and Xbp1s, as well as decreased PERK phosphorylation (p-PERK) and ATF6 expression. To validate our conjecture, we utilized small intestinal organoids cultured for 3 days for further experimentation. RT-qPCR demonstrated that energy restriction downregulated the mRNA expression of Hspa5 and other chaperones. Furthermore, we treated intestinal organoids with HA15, a BIP inhibitor, which resulted in significantly enlarged area but fewer buds. Consistent with our previous findings, Hspa5 was downregulated in the HA15-treated group, along with decreased expression of Hsp family and enterocyte markers. Since Hsp family plays key roles in restoring cellular homeostasis, we wondered if upregulating Hspa5 could protect intestinal cells from outside stimuli such as pro-inflammatory cytokines. When intestinal organoids were cocultured with IFN-g, we found that Bix, the selective Bip inducer, protected organoids from apoptosis by increasing Hsp70 expression. Conclusion: Given that immunosuppression partly resolves GVHD, strategies aimed at restoring intestinal homeostasis should be given sufficient attention. Our study reveals that intestinal enterocyte differentiation is blocked in GVHD, possibly due to starvation and energy restriction. Deficiency in Hspa5 results in impaired intestinal repair, and increasing Hsp levels may enhance cell resistance to cytokine storms.

Vitexin Regulates Heat Shock Protein Expression by Modulating ROS Levels Thereby Protecting against Heat-Stress-Induced Apoptosis.

Heat stress due to high temperatures can cause heat stroke, pyrexia, heat cramps, heart disease, and respiratory diseases, which seriously affect human health. Vitexin has been shown to alleviate heat stress; however, its mechanism of action remains unclear. Therefore, in this study, we used Caco-2 cells to establish a heat stress model and vitamin C as a positive control to investigate the regulatory effects of vitexin on heat-stress-induced apoptosis and the related mechanisms using Cell Counting Kit-8, flow cytometry, real-time quantitative polymerase chain reaction, and Western blot. The results showed that the mRNA expressions of Hsp27, Hsp70, and Hsp90 induced by heat stress could be effectively inhibited at vitexin concentrations as low as 30 μM. After heat stress prevention and heat stress amelioration in model cells based on this concentration, intracellular reactive oxygen species (ROS) levels and the mRNA level and the protein expression of heat shock proteins (Hsp70 and Hsp90) and apoptotic proteins were reduced. In addition, compared with the heat stress amelioration group, the expression of BCL2 mRNA and its protein (anti-apoptotic protein Bcl-2) increased in the heat stress prevention group, while the expression of BAX, CYCS, CASP3, and PARP1 mRNAs and their proteins (apoptotic proteins Bax, Cytochrome C, cle-Caspase-3, and cle-PARP1) were decreased. In summary, the heat-stress-preventive effect of vitexin was slightly better than its heat-stress-ameliorating effect, and its mechanism may be through the inhibition of intracellular ROS levels and thus the modulation of the expressions of Hsp70 and Hsp90, which in turn protects against heat-stress-induced apoptosis. This study provides a theoretical basis for the prevention and amelioration of heat stress using vitexin.

Exploration of the truncated cytosolic Hsp70 in plants - unveiling the diverse T1 lineage and the conserved T2 lineage.

The 70-kDa heat shock proteins (Hsp70s) are chaperone proteins involved in protein folding processes. Truncated Hsp70 (Hsp70T) refers to the variant lacking a conserved C-terminal motif, which is crucial for co-chaperone interactions or protein retention. Despite their significance, the characteristics of Hsp70Ts in plants remain largely unexplored. In this study, we performed a comprehensive genome-wide analysis of 192 sequenced plant and green algae genomes to investigate the distribution and features of Hsp70Ts. Our findings unveil the widespread occurrence of Hsp70Ts across all four Hsp70 forms, including cytosolic, endoplasmic reticulum, mitochondrial, and chloroplast Hsp70s, with cytosolic Hsp70T being the most prevalent and abundant subtype. Cytosolic Hsp70T is characterized by two distinct lineages, referred to as T1 and T2. Among the investigated plant and green algae species, T1 genes were identified in approximately 60% of cases, showcasing a variable gene count ranging from one to several dozens. In contrast, T2 genes were prevalent across the majority of plant genomes, usually occurring in fewer than five gene copies per species. Sequence analysis highlights that the putative T1 proteins exhibit higher similarity to full-length cytosolic Hsp70s in comparison to T2 proteins. Intriguingly, the T2 lineage demonstrates a higher level of conservation within their protein sequences, whereas the T1 lineage presents a diverse range in the C-terminal and SBDα region, leading to categorization into four distinct subtypes. Furthermore, we have observed that T1-rich species characterized by the possession of 15 or more T1 genes exhibit an expansion of T1 genes into tandem gene clusters. The T1 gene clusters identified within the Laurales order display synteny with clusters found in a species of the Chloranthales order and another species within basal angiosperms, suggesting a conserved evolutionary relationship of T1 gene clusters among these plants. Additionally, T2 genes demonstrate distinct expression patterns in seeds and under heat stress, implying their potential roles in seed development and stress response.

Characterization of genes encoding heat shock proteins reveals a differential response to temperature in two geographic populations of Liriomyza trifolii (Diptera: Agromyzidae)

Liriomyza trifolii is a significant, invasive pest that damages horticultural crops and vegetables. The distribution of L. trifolii is influenced by temperature, and prior research has demonstrated that variations in thermal adaptability differ among geographic populations of the insect. Heat shock proteins (Hsps) are involved in adaptation to temperatures; however, the underlying molecular mechanism for thermal adaption in different L. trifolii populations remains unclear. This study examines the temperature adaptability of two L. trifolii populations from Hainan (HN) and Jiangsu (JS) provinces. The results indicate that the HN population has a higher survival rate and a higher critical thermal maximum (CTmax) than the JS population under high temperature stress. Transcriptome data at 42 °C revealed that the JS population has more differentially expressed genes (DEGs) than the HN population, while the HN population has more upregulated DEGs. The two populations were similar in functional annotation of DEGs, and a large number of Hsps were upregulated. However, the HN population had larger numbers and higher expression levels of Hsps during heat stress as compared to the JS population. Additionally, the expression patterns of differentially expressed Hsps varied between the HN and JS populations in response to different elevated temperatures. Notably, the transcription levels of Hsp70s were higher in the HN population as compared to the JS population, while the expression level of genes encoding small heat shock proteins was higher in the JS population. These findings have significant scientific value in understanding the underlying mechanism of temperature adaption in L. trifolii and provide a fresh perspective on the distribution of this invasive pest.

Toxicological effects of copper on bioaccumulation and mRNA expression of antioxidant, immune, and apoptosis-related genes in Chinese striped-necked turtle (Mauremys sinensis).

Heavy metals are among the most ubiquitous environmental pollutants of recent decades. Copper is commonly used to control algal blooms or macrophyte and waste infestations, its ambient concentration has increased significantly, indicating possible environmental risk. To investigate the effects of copper exposure on bioaccumulation, antioxidant defense, immune response, and apoptosis in the Chinese Striped-necked Turtle Mauremys sinensis, three experimental groups, control (0.0mg/L), Cu2 (2mg/L) and Cu4 (4mg/L) were designed, and sampled at 14 and 28days. Results showed that copper accumulates in different organs depending on the concentration and exposure time, Liver > Kidney > Gut > Heart > Brain > Muscle and the time order was 28days > 14days. The liver enzymes AST, ALT, and ALP decreased when the turtles were exposed to copper stress, while the contents of bilirubin TBIL, DBIL, IBIL, and LDH showed a significant upward trend. Similarly, the mRNA expression level of acetylcholinesterase AChE in the brain was significantly downregulated upon copper exposure. An upward trend was noticed in the liver Metallothionein MT mRNA expression levels compared to the control group. The mRNA expression levels of antioxidant enzymes CAT, SOD, MnSOD, and GSH-PX1 in the liver increased initially and then significantly decreased. Furthermore, the relative mRNA expression levels of inflammatory cytokines IL-1β, IL-8, TNF-α, and IFN-γ involved in inflammatory response significantly upregulated. Copper significantly increased the hepatic mRNA transcription of heat shock proteins HSP70 and HSP90 at different exposure durations. In addition, the relative mRNA levels of caspase3, caspase8, and caspase9 related to the caspase-dependent apoptotic pathway significantly increased under copper stress. These results explain that copper toxicity causes bioaccumulation, promotes oxidative stress, obstructs immunity, and induces inflammation and apoptosis by altering their gene expression levels in M. sinensis.

A new look at Hsp70 activity in phosphatidylserine-enriched membranes: chaperone-induced quasi-interdigitated lipid phase

70 kDa heat shock protein Hsp70 (also termed HSP70A1A) is the major stress-inducible member of the HSP70 chaperone family, which is present on the plasma membranes of various tumor cells, but not on the membranes of the corresponding normal cells. The exact mechanisms of Hsp70 anchoring in the membrane and its membrane-related functions are still under debate, since the protein does not contain consensus signal sequence responsible for translocation from the cytosol to the lipid bilayer. The present study was focused on the analysis of the interaction of recombinant human Hsp70 with the model phospholipid membranes. We have confirmed that Hsp70 has strong specificity toward membranes composed of negatively charged phosphatidylserine (PS), compared to neutral phosphatidylcholine membranes. Using differential scanning calorimetry, we have shown for the first time that Hsp70 affects the thermotropic behavior of saturated PS and leads to the interdigitation that controls membrane thickness and rigidity. Hsp70-PS interaction depended on the lipid phase state; the protein stabilized ordered domains enriched with high-melting PS, increasing their area, probably due to formation of quasi-interdigitated phase. Moreover, the ability of Hsp70 to form ion-permeable pores in PS membranes may also be determined by the bilayer thickness. These observations contribute to a better understanding of Hsp70-PS interaction and biological functions of membrane-bound Hsp70 in cancer cells.

X-ray reflectivity study of the heat shock protein Hsp70 interaction with an artificial cell membrane model

Membrane-bound heat shock protein 70 (Hsp70) apart from its intracellular localization was shown to be specifically expressed on the plasma membrane surface of tumor but not normal cells. Although the association of Hsp70 with lipid membranes is well documented the exact mechanisms for chaperone membrane anchoring have not been fully elucidated. Herein, we addressed the question of how Hsp70 interacts with negatively charged phospholipids in artificial lipid compositions employing the X-ray reflectivity (XRR) studies. In a first step, the interactions between dioleoylphosphatidylcholine (DOPC) in the presence or absence of dioleoylphosphatidylserine (DOPS) and Hsp70 had been assessed using Quartz crystal microbalance measurements, suggesting that Hsp70 adsorbs to the surface of DOPC/DOPS bilayer. Atomic force microscopy (AFM) imaging demonstrated that the presence of DOPS is required for stabilization of the lipid bilayer. The interaction of Hsp70 with DOPC/DOPS lipid compositions was further quantitatively determined by high energy X-ray reflectivity. A systematic characterization of the chaperone-lipid membrane interactions by various techniques revealed that artificial membranes can be stabilized by the electrostatic interaction of anionic DOPS lipids with Hsp70.

Detection of Heat Shock Protein (Hsp70) Gene Expression, Some Physiological and Histological Alterations in the Nile Tilapia (Orechromis niloticus) Treated with Boric Acid and Cadmium

Detection of Heat Shock Protein (Hsp70) Gene Expression, Some Physiological and Histological Alterations in the Nile Tilapia (Orechromis niloticus) Treated with Boric Acid and Cadmium

Molecular docking technology and network pharmacology based on Rhapontici Radix-Cremastrae Pseudobulbus drug pair in treating breast cancer.

Network pharmacology is a bioinformatics-based research strategy for identifying the mechanisms of drugs and promoting drug development. This study used network pharmacology to investigate the mechanism of the Loulu-Cremastrae Pseudobulbus drug pair treating breast cancer (BC). The ingredients and potential targets of the drug pair were searched with Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCSMP). National Center for Biotechnology Information (NCBI) and gene cards were used to search the targets of BC. Networks of "drugs-components-targets" and protein-protein interaction were constructed through Cytoscape. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were carried out through common targets. Using AutoDock tool, molecular docking was performed to verify the binding between key targets and compounds. Finally, we selected 6 active compounds from the drug pair. A total of 61 targets were associated with the drug pair, and 15,295 targets were related to BC. 55 common targets were obtained after the intersection. The key targets included Transcription factor Jun (JUN), Heat shock protein HSP 90-alpha (HSP90AA1), and Caspase-3(CASP3). 327 terms were obtained by GO analysis. 78 pathways (p < 0.05) were identified through KEGG analysis. Molecular docking indicated that important compounds combined well with key targets. Various active compounds, including beta-sitosterol, 2-methoxy-9,10-dihydrophenanthrene-4,5-diol, and stigmasterol, can regulate multiple signaling pathways related to BC, such as the estrogen and prolactin signaling pathways, playing therapeutic roles in BC.

Small but powerful: The role of HSP in bacterial infectious disease pathogenesis- A mini-review of recent findings

Heat Shock Proteins (Hsp) are evolutionarily conserved ubiquitous molecular chaperones usually produced when cells are exposed to temperatures beyond their normal growth temperature. They are a group of ATP-independent chaperons pivotal for cell survival and proliferation. This group of proteins has been well studied over the years. However, recent investigations leveraging the available Whole Genome Sequencing (WGS) approach are beginning to shed more light on how this protein interacts with other intracellular proteins to modulate bacterial virulence and pathogenesis. Bacteria cells that manufacture HSPs can survive at higher temperatures. Certain heat-shock proteins have a refolding or degradation action that can save the bacterial cell even in extreme stress demands and even help some bacteria transition from being commensal to a pathogen. Certain HSPs are virulence factors, while others modulate bacterial pathogenesis increasing bacterial resistance to host immunological attack. Interestingly, others also influence certain virulence gene expressions. Several mechanisms govern heat shock response. Moreover, the employment of specific regulatory proteins that exert a positive or negative effect on the initiation of transcription by the RNA polymerase enzyme is successful and efficient in controlling heat-shock gene transcription. Interestingly, most pathogens use several approaches to reprogram gene transcription in response to diverse stress rapidly. In this mini-review, we tactfully discussed the recent update on HSP in bacteria virulence and how these proteins could serve as potential drug targets. Moreover, we provided an overview of how HSPs are regulated in bacterial pathogens favouring bacterial survival in diverse cellular niches.

Incubation in shaded hatcheries biases sex-determination but preserves Lepidochelys olivacea hatchling physiology

Some studies have associated ex situ conservation with cerebral and gonadal developmental delay, as well as decreased motor performance in Lepidochelys olivacea offspring. Ex situ management is also related to a more mature spleen and a differential leukocyte count in newly emerged Lepidochelys olivacea hatchlings. The physiological relevance of a more mature spleen is unknown in sea turtles, but studies in birds suggest an increased immune response. Because egg relocation to hatcheries is a common conservation practice, it is imperative to know its impact on hatchling physiology. Herein, plasma activity of superoxide dismutase, alkaline phosphatase and the alternative complement pathway, as well as total antioxidant capacity and hydrogen peroxide concentrations were quantified in hatchlings from in situ and ex situ nests under basal conditions at nest emergence. Toll-like receptor 4 (tlr4), heat shock proteins (hsp) 70 and hsp90 expression were quantified in the spleen and liver of the hatchlings. Hepatocyte density and nuclear area were quantified in histological sections of the liver and all turtles were sexed by histological sectioning of the gonads. Total antioxidant capacity and hydrogen peroxide concentrations in plasma were lower in turtles from ex situ nests, while tlr4 and hsp70 mRNA expression was higher in the spleen but not in the liver. Ex situ incubation produced 98% male hatchlings, whereas in situ incubation produced 100% females. There were no other differences in the attributes sampled between hatchlings emerging from ex situ and in situ treatments. The results suggest that ex situ relocated turtles may be less prone to oxidative stress than in situ incubated hatchlings and could have more mature splenic function. Together, the data suggest that ex situ relocation to shaded hatcheries biased sex determination but preserved the general physiological condition of sea turtle hatchlings.

Multi-omics reveals aging-related pathway in natural aging mouse liver

Aging is associated with gradual changes in liver structure, altered metabolites and other physiological/pathological functions in hepatic cells. However, its characterized phenotypes based on altered metabolites and the underlying biological mechanism are unclear. Advancements in high-throughput omics technology provide new opportunities to understand the pathological process of aging. Here, in our present study, both metabolomics and phosphoproteomics were applied to identify the altered metabolites and phosphorylated proteins in liver of young (the WTY group) and naturally aged (the WTA group) mice, to find novel biomarkers and pathways, and uncover the biological mechanism. Analysis showed that the body weights, alanine aminotransferase (ALT) and aspartate aminotransferase (AST) increased in the WTA group. The grips decreased with age, while the triglyceride (TG) and cholesterol (TC) did not change significantly. The increase of fibrosis, accumulation of inflammatory cells, hepatocytes degeneration, the deposition of lipid droplets and glycogen, the damaged mitochondria, and deduction of endoplasmic reticulum were observed in the aging liver under optical and electron microscopes. In addition, a network of metabolites and phosphorylated proteomes of the aging liver was established. Metabolomics detected 970 metabolites in the positive ion mode and 778 metabolites in the negative ion mode. A total of 150 pathways were pooled. Phosphoproteomics identified 2618 proteins which contained 16621 phosphosites. A total of 164 pathways were detected. 65 common pathways were detected in two omics. Phosphorylated protein heat shock protein HSP 90-alpha (HSP90A) and v-raf murine viral oncogene homolog B1(BRAF), related to cancer pathway, were significantly upregulated in aged mice liver. Western blot verified that protein expression of MEK and ERK, downstream of BRAF pathway were elevated in the liver of aging mice. However, the protein expression of BRAF was not a significant difference. Overall, these findings revealed a close link between aging and cancer and contributed to our understanding of the multi-omics changes in natural aging.

Hsp70 ameliorates sleep deprivation-induced anxiety-like behavior and cognitive impairment in mice

BackgroundMany neurobehavioral processes, including psychomotor, cognitive, and affection are negatively impacted by sleep deprivation (SD), which may be harmful to a person's physical and mental health. Heat shock proteins (Hsps) have been demonstrated to play a protective role in a number of neurodegenerative diseases and are essential for maintaining intracellular protein homeostasis, but their roles in SD remain elusive. MethodsA mouse SD model was constructed using a modified multi-platform water environment method. The cognitive function was tested by novel object recognition test and Y-maze test, and anxiety-like behaviors were assessed by open field test (OFT). Protein expression was determined by Western blotting assay and ELISA assay. ResultsWe found that SD could profoundly enhance anxiety levels and impair cognitive function in mice. SD also reduced the expression levels of p-cAMP-response element binding protein (CREB) and brain-derived neurotrophic factor (BDNF) and increased microglial activation and neuroinflammatory response in the hippocampus of mice. The intranasal injection of human recombinant Hsp70 protein could alleviate SD-induced anxiety and cognitive impairment, as well as restore pCREB and BDNF levels and reduce microglia-induced neuroinflammation in the hippocampus of SD mice. ConclusionsHsp70 treatment might serve as a potential treatment for mitigating SD-related unfavorable symptoms.

Temperature Elevation during Semen Delivery Deteriorates Boar Sperm Quality by Promoting Apoptosis.

Semen delivery practice is crucial to the efficiency of artificial insemination using high-quality boar sperm. The present study aimed to evaluate the effect of a common semen delivery method, a Styrofoam box, under elevated temperatures on boar sperm quality and functionality and to investigate the underlying molecular responses of sperm to the temperature rise. Three pooled semen samples from 10 Duroc boars (3 ejaculates per boar) were used in this study. Each pooled semen sample was divided into two aliquots. One aliquot was stored at a constant 17 °C as the control group. Another one was packaged in a well-sealed Styrofoam box and placed in an incubator at 37 °C for 24 h to simulate semen delivery on hot summer days and subsequently transferred to a refrigerator at 17 °C for 3 days. The semen temperature was continuously monitored. The semen temperature was 17 °C at 0 h of storage and reached 20 °C at 5 h, 30 °C at 14 h, and 37 °C at 24 h. For each time point, sperm quality and functionality, apoptotic changes, expression levels of phosphorylated AMPK, and heat shock proteins HSP70 and HSP90 were determined by CASA, flow cytometry, and Western blotting. The results showed that elevated temperature during delivery significantly deteriorated boar sperm quality and functionality after 14 h of delivery. Storage back to 17 °C did not recover sperm motility. An increased temperature during delivery apparently promoted the conversion of sperm early apoptosis to late apoptosis, showing a significant increase in the expression levels of Bax and Caspase 3. The levels of phosphorylated AMPK were greatly induced by the temperature rise to 20 °C during delivery but reduced thereafter. With the temperature elevation, expression levels of HSP70 and HSP90 were notably increased. Our results indicate that a temperature increase during semen delivery greatly damages sperm quality and functionality by promoting sperm apoptosis. HSP70 and HSP90 could participate in boar sperm resistance to temperature changes by being associated with AMPK activation and anti-apoptotic processes.

Effects of acute and chronic chromium stress on the expression of heat shock protein genes and activities of antioxidant enzymes in larvae of Orthetrum albistylum

The dragonfly species Orthetrum albistylum, can accumulate heavy metals from its aquatic environment and thus serves as a biological indicator for monitoring and evaluating water quality. Heat shock proteins (HSPs) play important biological roles in resistance to various types of environmental stress. The full-length cDNA sequences of the heat shock cognate (hsc) 70 and heat shock protein (hsp) 70 genes were cloned from O. albistylum larvae. Relative levels of expression of hsc70 and hsp70 in the head, epidermis, midgut, and adipose tissue were measured by qRT-PCR after chronic and acute contamination of 5–8 instar larvae with chromium (Cr) solution, and under control conditions. Activities of superoxide dismutase (SOD) and catalase (CAT) in chronically contaminated larvae were also measured. Phylogenetic analysis revealed that the cloned hsc70 and hsp70 genes were highly homologous to known HSP70 family members reported in other insects. The mRNA levels of hsc70 and hsp70 did not differ significantly in various larval tissues. Under chronic chromium stress, hsc70 and hsp70 expression were upregulated to a maximum and then downregulated; hsp70 mRNA levels were higher than those of hsc70 at all concentrations of chromium. Under acute chromium stress, hsc70 expression was inhibited at low chromium concentrations and upregulated at chromium concentrations higher than 125 mg/L. However, hsp70 expression was higher than that in the control group and markedly higher than that of hsc70. Changes in SOD and CAT activities displayed consistent trends for different chronic chromium concentrations, first increasing and then decreasing over time. Collectively, these findings demonstrated the response of the HSP family of genes and antioxidant enzymes following exposure to heavy metal stress, as well as their potential applicability as biomarkers for monitoring environmental pollutants.

Integrated metabolomic and transcriptomic responses to heat stress in a high-altitude fish, Triplophysa siluroides

Species in Triplophysa display strong adaptability to the extreme environment of the plateau, thus offering an ideal model to study the molecular mechanism of fish adaptation to environmental stress. In the present study, we conducted integrated analysis of the transcriptome and metabolism of liver tissue in Triplophysa siluroides under heat stress (28 °C) and control (10 °C) conditions to identify heat stress-induced genes, metabolites and pathways. RNA-Seq identified 2373 differentially expressed genes, which consisted of 1360 upregulated genes and 1013 downregulated genes, in the heat stress group vs. the control group. Genes in the heat shock protein (Hsp) family, including Hsp40, Hsp70, Hsp90 and other Hsps, were strongly upregulated by heat stress. Pathway enrichment analysis revealed that the PI3K/AKT/mTOR and protein processing in the endoplasmic reticulum (ER) pathways were significantly affected by heat stress. Metabolism sequencing identified a total of 155 differentially abundant metabolites, including 118 significantly upregulated metabolites and 37 downregulated metabolites. Combined analysis of the transcriptome and metabolism results showed that ubiquitin-dependent proteolysis and purine metabolism pathways were enhanced in response to acute heat stress to protect cells from damage under stress conditions. The results of this study may contribute to our understanding of the underlying molecular mechanism of the heat stress response in cold-water fish.