Expression of heat shock proteins (Hsps) 27 and 70 in kidney in cases of fatal hemorrhage.

The expression levels of intracellular heat shock proteins (Hsps), specialized chaperone proteins, increase in cases of cellular stress with protein misfolding and aggregation. In a previous study, we demonstrated that there is an extensive increase in intracellular Hsp27 and 70 expression levels in renal tissues in fire fatality cases. Hsp expression can be induced by not only heat, but also by tissue hypoxia. In cases of fatal hemorrhage, the individual suffers hypoxemia and consequently tissue hypoxia. Here, we examined 43 cases of fatal hemorrhage and a control group of 85 deaths not related to blood loss or temperature exposure. We evaluated Hsp27 and 70 protein expression levels in renal tissue using immunohistochemistry. The results revealed that no extensive Hsp27 or 70 expression is induced in the fatal hemorrhage cases. The renal Hsp levels were similar to those of the control group. Fatal blood loss does not cause relevant cell stress.

Cisplatin-based Electrochemotherapy Significantly Downregulates Key Heat Shock Proteins in MDA-MB-231-Human Triple-Negative Breast Cancer Cells.

Heat shock proteins (HSPs) are available and/or induced for the survival of all organisms, including eukaryotic, prokaryotic, and plants, from higher temperature stresses. They are the chaperone proteins that protect all cells against heat, as the name implies. In addition to thermal stress, they also protect them from chemical, physical, and other stresses, including exposure to oxidative stress, nutritional deficiencies, ultraviolet radiation, ethanol, viral infection, ischemia-reperfusion injury, and cancer-related stresses. They are classified based on their molecular weights in kDa, such as HSP90 and HSP70. In our label-free, high-throughput, quantitative LC-MS/MS-based proteomic studies of MDA-MB-231, human, triple-negative breast cancer cells, treated with electrical pulses (EP) and cisplatin (CsP), we identified a number of HSPs, such as HSP90AA1, and others to be significantly downregulated in EP + CsP, compared to CsP alone. This indicates that cells will undergo apoptotic cell death and hence could cause effective cancer cure/treatment. Considering that over 2 million new cases and over 600,000 deaths in 2020, of which ~ 15% are TNBC, heat shock proteins could be the untapped resources, available for the next biomarkers and/or inhibitors for new/additional therapies.

The Concentration of Selected Inflammatory Cytokines (IL-6, IL-8, CXCL5, IL-33) and Damage-Associated Molecular Patterns (HMGB-1, HSP-70) Released in an Early Response to Distal Forearm Fracture and the Performed Closed Reduction With Kirschner Wire Fixation in Children.

The evaluation of trauma after surgery through objective analysis of biochemical markers can help in selecting the most appropriate therapy. Thus the aim of the study was the evaluation of the concentration of selected inflammatory cytokines (IL-6, IL-8, CXCL5, IL-33), C-reactive protein (CRP), and damaged-associated molecular patterns (DAMPs): HMGB-1, HSP-70 in the plasma of children in response to bone fracture and 12-14 hours after subsequent surgery performed by closed reduction with percutaneous Kirschner wire fixation (CRKF). The study will answer the question if the CRFK procedure leads to excessive production of inflammatory and damage markers. Blood samples from 29 children with distal forearm fractures were collected 30 min. before CRKF procedure and 12-14 hours after performance of the procedure. The control group was composed of 17 healthy children. IL-6 and CRP concentrations were analyzed using routinely performed in vitro diagnostics tests; the remaining proteins were analyzed with the use of the ELISA method. Increased values of IL-6, CRP, and HSP-70 represented an early inflammatory response to distal forearm fractures classified as SH-II type according to the Salter-Harris classification system. However, the median CRP concentration was within the reference values not indicative of inflammation. The CRKF procedure may be a good solution for the treatment of bone fractures, as damaged associated molecular patterns - HMGB-1 and HSP-70 - did not significantly differ 12-14 hours after the approach was applied as compared to the control group. Moreover, the increase in IL-6 concentration after the CRKF procedure was 1.5-fold to the level before CRKF, while the increase of this marker in response to the distal forearm fracture was 4.3-fold compared to the control group. Based on this data, it appears reasonable to suggest that the CRKF approach caused less damage and inflammatory response in comparison to the response to the fracture itself.

Attenuation Effect of Radiofrequency Irradiation on UV-B-Induced Skin Pigmentation by Decreasing Melanin Synthesis and through Upregulation of Heat Shock Protein 70.

Excess melanin deposition in the skin causes cosmetic problems. HSP70 upregulation decreases microphthalmia-associated transcription factor (MITF) expression, which eventually decreases tyrosinase activity and melanogenesis. Ultraviolet (UV) radiation upregulates p53, which increases the melanocortin receptor (MC1R) and MITF. Furthermore, HSP70 decreases p53 and radiofrequency irradiation (RF) increases HSP70. We evaluated whether RF increased HSP70 and decreased p53, consequently decreasing the MITF/tyrosinase pathway and melanogenesis in UV-B radiated animal skin. Various RF combinations with 50, 100, and 150 ms and 5, 10, and 15 W were performed on the UV-B radiated mouse skin every 2 d for 28 d. When RF was performed with 100 ms/10 W, melanin deposition, evaluated by Fontana-Masson staining, decreased without skin crust formation in the UV-B radiated skin. Thus, we evaluated the effect of RF on decreasing melanogenesis in the HEMn and UV-B radiated skin at a setting of 100 ms/10 W. HSP70 expression was decreased in the UV-B radiated skin but was increased by RF. The expression of p53, MC1R, and MITF increased in the UV-B radiated skin but was decreased by RF. The expression of p53, MC1R, and MITF increased in the α-MSH treated HEMn but was decreased by RF. The decreasing effects of RF on p53, MC1R, CREB and MITF were higher than those of HSP70-overexpressed HEMn. The decreasing effect of RF on p53, MC1R, CREB, and MITF disappeared in the HSP70-silenced HEMn. MC1R, CREB, and MITF were not significantly decreased by the p53 inhibitor in α-MSH treated HEMn. RF induced a greater decrease in MC1R, CREB, and MITF than the p53 inhibitor. Therefore, RF may have decreased melanin synthesis by increasing HSP70 and decreasing p53, thus decreasing MC1R/CREB/MITF and tyrosinase activity.

Immunohistochemical Expression of AQP2 and HSP70 in Broiler Kidney Tissue Treated with Salix tetrasperma Roxb. Extract under Heat Exposure.

The administration of plant extracts to broilers may be a way to mitigate the effects of heat stress. The importance of AQP2 and HSP70 compounds in maintaining the homeostasis of the chicken body when it is subjected to heat stress is well established. This study aims to determine the effect of giving the ethanolic extract of the leaves of Salix tetrasperma Roxb. on the immunohistochemical expression of AQP2 and HSP70 in exposed and unexposed broiler kidney tissue. This study used 36 samples of 28-day-old chicken kidneys. Chickens were kept in individual cages, provided with feed and drinking water ad libitum. The design used was a completely randomized design with 6 treatments and 6 replications: (a) chickens were reared in conditions exposed to heat (HS + 0); (b) chickens were reared in conditions exposed to heat and given Salix extract at a dose of 50 mg/L drinking water (HS + 50); (c) chickens were reared under heat-exposed conditions and given Salix extract at a dose of 100 mg/L drinking water (HS + 100); (d) chickens were reared in conditions without exposure to heat (n-HS + 0); (e) chickens were reared in conditions without exposure to heat and given Salix extract at a dose of 50 mg/L drinking water (nHS + 50); and (f) chickens were reared in conditions exposed without exposure to heat and given 100 mg/L drinking water (nHS + 100) of Salix extract. Salix extract was given for 24 hours and was renewed every 6 hours. The results showed that giving Salix extract 100 mg/L in drinking water to chickens exposed to heat (HS + 100) reduced the value of the H/L ratio. Giving Salix extract 50-100 mg/L in drinking water caused an upregulated AQP2 expression; on the other hand, it downregulated HSP-70 expression, in chicken kidney tubules both exposed to heat stress and nonexposed to heat stress. In conclusion, exposure to heat stress in broiler chickens and giving Salix extract can increase the formation of aquaporin 2 compounds and suppress the formation of HSP70.

Inhibition of the Heat Shock Protein A (HSPA) Family Potentiates the Anticancer Effects of Manumycin A.

Manumycin A (MA) is a well-tolerated natural antibiotic showing pleiotropic anticancer effects in various preclinical in vitro and in vivo models. Anticancer drugs may themselves act as stressors to induce the cellular adaptive mechanism that can minimize their cytotoxicity. Heat shock proteins (HSPs) as cytoprotective factors can counteract the deleterious effects of various stressful stimuli. In this study, we examined whether the anticancer effects of MA can be counteracted by the mechanism related to HSPs belonging to the HSPA (HSP70) family. We found that MA caused cell type-specific alterations in the levels of HSPAs. These changes included concomitant upregulation of the stress-inducible (HSPA1 and HSPA6) and downregulation of the non-stress-inducible (HSPA2) paralogs. However, neither HSPA1 nor HSPA2 were necessary to provide protection against MA in lung cancer cells. Conversely, the simultaneous repression of several HSPA paralogs using pan-HSPA inhibitors (VER-155008 or JG-98) sensitized cancer cells to MA. We also observed that genetic ablation of the heat shock factor 1 (HSF1) transcription factor, a main transactivator of HSPAs expression, sensitized MCF7 cells to MA treatment. Our study reveals that inhibition of HSF1-mediated heat shock response (HSR) can improve the anticancer effect of MA. These observations suggest that targeting the HSR- or HSPA-mediated adaptive mechanisms may be a promising strategy for further preclinical developments.

Withaferin A Induces Heat Shock Response and Ameliorates Disease Progression in a Mouse Model of Huntington's Disease.

Impairment of proteostasis network is one of the characteristic features of many age-related neurodegenerative disorders including autosomal dominantly inherited Huntington's disease (HD). In HD, N-terminal portion of mutant huntingtin protein containing expanded polyglutamine repeats accumulates as inclusion bodies and leads to progressive deterioration of various cellular functioning including proteostasis network. Here we report that Withaferin A (a small bioactive molecule derived from Indian medicinal plant, Withania somnifera) partially rescues defective proteostasis by activating heat shock response (HSR) and delays the disease progression in a HD mouse model. Exposure of Withaferin A activates HSF1 and induces the expression of HSP70 chaperones in an in vitro cell culture system and also suppresses mutant huntingtin aggregation in a cellular model of HD. Withaferin A treatment to HD mice considerably increased their lifespan as well as restored progressive motor behavioral deficits and declined body weight. Biochemical studies confirmed the activation of HSR and global decrease in mutant huntingtin aggregates load accompanied with improvement of striatal function in Withaferin A-treated HD mouse brain. Withaferin A-treated HD mice also exhibit significant decrease in inflammatory processes as evident from the decreased microglial activation. These results indicate immense potential of Withaferin A for the treatment of HD and related neurodegenerative disorders involving protein misfolding and aggregation.

Expression, Rapid Purification and Functional Analysis of DnaK from Rhodococcus ruber.

BACKGROUND: Heat shock proteins (HSPs) represent a group of important proteins which are produced by all kinds of organisms especially under stressful conditions. DnaK, an Hsp70 homolog in prokaryotes, has indispensable roles when microbes was confronted with stress conditions. However, few data on DnaK from Rhodococcus sp. were available in the literature. In a previous study, we reported that toluene and phenol stress gave rise to a 29.87-fold and 3.93-fold increase for the expression of DnaK from R. ruber SD3, respectively. Thus, we deduced DnaK was in correlation with the organic solvent tolerance of R. ruber SD3. OBJECTIVE: To elucidate the role of DnaK in the organic solvent tolerance of R. ruber SD3, expression, purification and functional analysis of Dnak from R. ruber SD3 were performed in the present paper. METHODS: In this article, DnaK from R. ruber SD3 was heterologously expressed in E. coli BL21(DE3) and purified by affinity chromatography. Functional analysis of DnaK was performed using determination of kinetics, docking, assay of chaperone activity and microbial growth. RESULTS: The recombinant DnaK was rapidly purified by affinity chromatography with the purification fold of 1.9 and the recovery rate of 57.9%. Km, Vmax and Kcat for Dnak from R. ruber SD3 were 80.8 µM, 58.1 nmol/min and 374.3 S-1, respectively. The recombinant protein formed trimer in vitro, with the calculated molecular weight of 214 kDa. According to in-silico analysis, DnaK interacted with other molecular chaperones and some important proteins in the metabolism. The specific activity of catalase in the presence of recombinant DnaK was 1.85 times or 2.00 times that in the presence of BSA or Tris-HCl buffer after exposure to 54 °C for 1h. E. coli transformant with pET28-dnak showed higher growth than E. coli transformant with pET28 at 43°C and in the presence of phenol, respectively. CONCLUSION: The biochemical properties and the interaction analysis of DnaK from R. ruber SD3 deepened our understanding of DnaK function. DnaK played an important role in microbial growth when R. ruber was subjected to various stress such as heating and organic solvent.

Knockdown of miR-384-3p Protects Against Myocardial Ischemia-Reperfusion Injury in Rats Through Targeting HSP70.

BACKGROUND: Myocardial infarction (MI) and heart failure remain critical states of heart disease with high mortality. Previous studies have indicated that miRNA has cardioprotective effects and can resist myocardial ischemia-reperfusion (I/R) injury. However, the role of mir-384-3p in MI has not been reported, and whether this miRNA can regulate the apoptosis of cardiomyocytes needs to be verified. METHODS: The effect of hypoxia-reperfusion (H/R) on cardiomyocyte activity was detected using MTT assay. MiR-384-3p was knocked down or overexpressed in cardiomyocytes H/R models by pretreatment with miR-384-3p mimic or inhibitor to verify the function of miR-384-3p in H/R. Circulating levels of miR-384-3p was detected by quantitative realtime PCR, and protein expression was detected by western blotting. TUNEL staining and flow cytometry demonstrated a high degree of myocardium apoptosis after H/R induction. Dual-Luciferase Reporter Assay detected dynamic expression of miR-384-3p and HSP70. The infarction size of I/R rats was detected by 2,3,5-triphenyltetrazolium chloride (TTC) staining. RESULTS: MiR-384-3p was closely related to cardiomyocyte activity in H/R progression. Increased expression of mir-384-3p can promote the production of cleaved caspase-3 and cleaved PARP, thereby regulating cardiomyocyte apoptosis. HSP70 was a target of miR-384-3p and HSP70 silencing aggravated H/R-induced cardiomyocyte dysfunction. In an animal model, the expression level of HSP70 is regulated by miR-384-3p, and miR-384-3p inhibition remarkably reduced I/R-induced MI in rats. CONCLUSION: In conclusion, the present report identified that HSP70 was a potential target of miR-384-3p, and miR-384-3p inhibition remarkably reduced I/R-induced MI in rats. Therefore, this study provides a novel therapeutic approach for the treatment of MI from bench to clinic.

Genome-wide identification, classification and expression analysis of the Hsf and Hsp70 gene families in maize.

Heat shock factors (Hsfs) and heat shock proteins (Hsps) play a critical role in the molecular mechanisms such as plant development and defense against abiotic. As an important food crop, maize is vulnerable to adverse environment such as heat stress and water logging, which leads to a decline in yield and quality. To date, very little is known regarding the structure and function of Hsf and Hsp genes in maize. Although some Hsf and Hsp genes have been characterized in maize, analysis of the entire Hsf and Hsp70 gene families were not completed following Maize (B73) Genome Sequencing Project. Therefore, studying their molecular mechanism and revealing their biological function in plant stress resistance process will contribute to reveal important theoretical significance and application value for improving corn yield and quality. In this study, we have identified 25 ZmHsf and 22 ZmHsp70 genes in maize. The structural characteristics and phylogenetic relationships of the Hsf and Hsp70 gene families of Arabidopsis thaliana, rice and maize were compared. The final 25 ZmHsf proteins and 22 ZmHsp70 proteins were divided into three and four subfamilies, respectively. In addition, chromosomal localization indicated that the ZmHsf and ZmHsp70 genes were unevenly distributed on the chromosome, and the gene structure map revealed the characteristics of their structures. Finally, transcriptome analysis indicated that most of the ZmHsf and ZmHsp70 genes showed different expression patterns at different developmental stages of maize. Further, by semi-quantitative RT-PCR and quantitative real-time PCR analysis, all 25 ZmHsf and 22 ZmHsp70 genes were confirmed to respond to heat stress treatment, indicating that they have potential effects in heat stress response. The analyses performed by combining co-expression network with protein-protein interaction network among the members of the Hsf and Hsp70 gene families in maize further enabled us to recognize components involved in the regulatory network associated with hsfs and hsp70s complex. The predicted subcellular location revealed that maize Hsp70 proteins exhibited a various subcellular distribution, which may be associated with functional diversification in heat stress response. Taken together, our study provides comprehensive information on the members of Hsf and Hsp70 gene families and will help in elucidating their exact function in maize.

Heat stress induced arginylation of HuR promotes alternative polyadenylation of Hsp70.3 by regulating HuR stability and RNA binding.

Arginylation was previously found to promote stabilization of heat shock protein 70.3 (Hsp70.3) mRNA and cell survival in mouse embryonic fibroblasts (MEFs) on exposure to heat stress (HS). In search of a factor responsible for these phenomena, the current study identified human antigen R (HuR) as a direct target of arginylation. HS induced arginylation of HuR affected its stability and RNA binding activity. Arginylated HuR failed to bind Hsp70.3 3' UTR, allowing the recruitment of cleavage stimulating factor 64 (CstF64) in the proximal poly-A-site (PAS), generating transcripts with short 3'UTR. However, HuR from Ate1 knock out (KO) MEFs bound to proximal PAS region with higher affinity, thus excluded CstF64 recruitment. This inhibited the alternative polyadenylation (APA) of Hsp70.3 mRNA and generated the unstable transcripts with long 3'UTR. The inhibition of RNA binding activity of HuR was traced to arginylation-coupled phosphorylation of HuR, by check point kinase 2 (Chk2). Arginylation of HuR occurred at the residue D15 and the arginylation was needed for the phosphorylation. Accumulation of HuR also decreased cell viability upon HS. In conclusion, arginylation dependent modifications of HuR maintained its cellular homeostasis, and promoted APA of Hsp70.3 pre-mRNA, during early HS response.

Intravenous Immunoglobulin G Suppresses Heat Shock Protein (HSP)-70 Expression and Enhances the Activity of HSP90 and Proteasome Inhibitors.

Intravenous immunoglobulin G (IVIgG) is approved for primary immunodeficiency syndromes but may induce anti-cancer effects, and while this has been attributed to its anti-inflammatory properties, IgG against specific tumor targets may play a role. We evaluated IVIgG alone, and with a Heat shock protein (HSP)-90 or proteasome inhibitor, using multiple myeloma and mantle cell lymphoma (MCL) cells in vitro, and with the proteasome inhibitor bortezomib in vivo. IVIgG inhibited the growth of all cell lines tested, induced G1 cell cycle arrest, and suppressed pro-tumor cytokines including Interleukin (IL)-6, IL-8, and IL-10. Genomic and proteomic studies showed that IVIgG reduced tumor cell HSP70-1 levels by suppressing the ability of extracellular HSP70-1 to stimulate endogenous HSP70-1 promoter activity, and reduced extracellular vesicle uptake. Preparations of IVIgG were found to contain high titers of anti-HSP70-1 IgG, and recombinant HSP70-1 reduced the efficacy of IVIgG to suppress HSP70-1 levels. Combining IVIgG with the HSP90 inhibitor AUY922 produced superior cell growth inhibition and correlated with HSP70-1 suppression. Also, IVIgG with bortezomib or carfilzomib was superior to each single agent, and enhanced bortezomib's activity in bortezomib-resistant myeloma cells. Moreover, IVIgG reduced transfer of extracellular vesicles (EVs) to cells, and blocked transfer of bortezomib resistance through EVs. Finally, IVIgG with bortezomib were superior to the single agents in an in vivo myeloma model. These studies support the possibility that anti-HSP70-1 IgG contained in IVIgG can inhibit myeloma and MCL growth by interfering with a novel mechanism involving uptake of exogenous HSP70-1 which then induces its own promoter.

Novel role of mortalin in attenuating HIV-1 Tat-mediated astrogliosis.

BACKGROUND: In human immunodeficiency virus-1 (HIV-1) infection, activation of astrocytes induces imbalance in physiological functions due to perturbed astrocytic functions that unleashes toxicity on neurons. This leads to inflammatory response finally culminating into neurocognitive dysfunction. In neuroAIDS, HIV-1 protein, transactivator of transcription (Tat) is detected in the cerebrospinal fluid of infected patients. Mortalin, a multifunctional protein, has anti-inflammatory role following its activation in various stress conditions. Recent studies demonstrate downregulation of mortalin in neurodegenerative diseases. Here, we explored the mechanisms of mortalin in modulating HIV-1 Tat-mediated neuroinflammation. METHODS: Expression of mortalin in autopsy section in normal and diseased individuals were examined using immunohistochemistry. To decipher the role of mortalin in HIV-1 Tat-induced activation, human fetal brain-derived astrocytes were transiently transfected with Tat and mortalin using expression vectors. HIV-1 Tat-mediated damage was analyzed using RT-PCR and western blotting. Modulatory role of mortalin was examined by coexpressing it with Tat, followed by examination of mitochondrial morphodynamics using biochemical assay and confocal and electron microscopy. Extracellular ATP release was monitored using luciferase assay. Neuroinflammation in astrocytes was examined using flow cytometry, dye based study, immunocytochemistry, immunoprecipitation, and western blotting. Indirect neuronal damage was also analyzed. RESULTS: HIV-1 Tat downregulates the expression of mortalin in astrocytes, and this is corroborated with autopsy sections of HIV-1 patients. We found that overexpression of mortalin with Tat reduced inflammation and also rescued astrocytic-mediated neuronal death. Using bioinformatics, we discovered that binding of mortalin with Tat leads to Tat degradation and rescues the cell from neuroinflammation. Blocking of proteosomal pathway rescued the Tat degradation and revealed the ubiquitination of Tat. CONCLUSION: Overall, our data demonstrated the protective role of mortalin in combating HIV-1 Tat-mediated damage. We also showed that mortalin could degrade Tat through direct binding with HIV-1 Tat. Overexpression of mortalin in the presence of Tat could significantly reduce cytotoxic effects of Tat in astrocytes. Indirect neuronal death was also found to be rescued. Our in vitro findings were validated as we found attenuated expression of mortalin in the autopsy sections of HIV-1 patients.

DnaJs, the critical drivers of Hsp70s: genome-wide screening, characterization and expression of DnaJ family genes in Sorghum bicolor.

The DnaJ/Hsp40s, are important components in the chaperone machine, and play pivotal roles in plant growth, development and stress tolerance. Sorghum, the semi-arid crop, is the drought resilient, model C4 crop. However, no reports of DnaJs have been available. Genome-wide analysis of Sorghum bicolor revealed 113 DnaJ/Hsp40 genes, classified into four groups; 8 genes in SbDnaJ-A class, 10 in SbDnaJ-B, 82 in SbDnaJ-C and 13 in SbDnaJ-D distributed unevenly on all the 10 chromosomes. Chromosomes 1 and 3 were found hot spots with 22 and 20 genes respectively. All genes displayed large number of introns, with an exception of 11 of the SbDnaJ-C which is devoid of introns. Out of 36 paralogous duplications, 7 tandem and 29 segmental duplications were noticed, indicating the major role of segmental duplications in the expansion. Analysis of digital data revealed tissue and stage-specific expressions. Transcriptional profiling of 12 selected genes representing all 4 classes revealed highly significant expression in leaf followed by root tissues. No expression was noticed in stems with an exception of SbDnaJ-C76. The SbDnaJ-A1, D1, and C subgroup genes displayed upregulation in roots, stems and leaves under cold, inferring the involvement of Hsp40s for cellular protection during cold stress. The results demonstrate that C76 and D1 are the candidate genes associated with multiple abiotic stresses. Present research furnishes valuable information about the role of sorghum DnaJs in abiotic stress response and establishes a foundation for understanding the molecular mechanisms associated with plant development and stress tolerance.

The transcriptional factor GATA-4 negatively regulates Hsp70 transcription in Crassostrea hongkongensis.

To better explore the application potential of heat shock protein Hsp70s in diverse areas including biomonitoring, a further investigation of the details of the regulatory mechanism governing Hsp70 transcription is required. A transcriptional factor ChGATA-4 that displayed affinity to the ChHsp70 promoter of Crassostrea hongkongensis was isolated and identified by DNA affinity purification as well as mass spectrometry analysis. The ChGATA-4 cDNA is 2162 bp in length and the open reading frame encodes a polypeptide containing 482 amino acids with a conserved zinc finger domain. The over-expression of ChGATA-4 significantly inhibited the expression of ChHsp70 promoter in heterologous HEK293T cells. However, the depletion of ChGATA-4 mRNA by RNAi technique resulted in significant increase of ChHsp70 transcription in oyster hemocytes. The RT-PCR results demonstrated that the transcription of both ChHsp70 and ChGATA-4 were induced by heat, Cd, or NP (Nonyl phenol) stress. This suggested a potential correlation between ChHsp70 and ChGATA-4 in the stress-mediated genetic regulatory cascade. This study demonstrated that ChGATA-4 acts in a negative manner in controlling ChHsp70 transcription in C. hongkongensis and promotes to further understand the mechanisms leading Hsp70 transcription.

Overexpression of heat shock protein 70 enhanced mesenchymal stem cell treatment efficacy in phosgene-induced acute lung injury.

In our previous study, we have confirmed that in phosgene-induced acute lung injury (ALI) rats, mesenchymal stem cells (MSCs) can treat the disease. Moreover, heat shock protein 70 (Hsp70) can be used as a protective protein, and Hsp70 upregulated drastically when exposed to stressful conditions. We aimed to assess that MSCs overexpressed Hsp70 could enhance the capacity of MSCs and have a good therapeutic effect on phosgene-induced ALI. We transduced MSCs with Hsp70 and then we tested the function of the transduced MSCs. Sprague Dawley rats inhaled phosgene in a closed container for 5 minutes. The transduced MSCs and MSCs were administered via the trachea immediately. Rats in each group were killed at 6, 24, and 48 hours after exposure. Compared to MSCs, MSCs overexpressed Hsp70 enhanced MSCs viability, antiapoptotic ability, and migration ability, and these effects disappeared when using the phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT) pathway inhibitor. Furthermore, the results of pathological alterations improved. The lung wet-to-dry ratio declined. The lung injury index total protein content and total cells in bronchoalveolar lavage fluid (BALF) also declined. The level of tumor necrosis factor α declined and the level of interleukin-10 improved in BALF and serum. MSCs overexpressed Hsp70 can enhance the capacity and efficacy of MSCs in the treatment of phosgene-induced ALI and may be mediated through the PI3k/AKT signaling pathway. This article introduces a new approach to stem cell therapy for improving the efficacy of phosgene-induced ALI.

Cadmium-induced developmental alteration and upregulation of serine-type endopeptidase transcripts in wild freshwater populations of Chironomus plumosus.

Cadmium, a toxic heavy metal, is a persistent environmental contaminant with irreversible toxicity to aquatic organisms. Chironomus plumosus, a natural species, is the largest sediment-burrowing aquatic midge in freshwater environments. In this study, we evaluated developmental defects in C. plumosus resulting from Cd exposure. In C. plumosus larvae, Cd exposure induced decreased survival and growth rates, reduction of emergence rate and sex ratio, and delayed emergence, as well as elevating the incidence of split tooth deformities. To identify potential biomarker genes to assess environmental pollutants such as Cd, we identified differentially expressed genes (DEGs) in C. plumosus exposed to various Cd concentrations. Among fourteen characterized DEGs, serine-type endopeptidase (SP) and heat shock protein 70 (HSP70) genes exhibited significant upregulation in C. plumosus larvae after Cd exposure. Therefore, we evaluated SP and HSP70 responses in natural C. plumosus populations collected from three sites of a Korean river and analyzed their correlations with eighteen environmental quality characteristics using principal component analysis. The highest expression of SP and HSP70 transcripts was observed in C. plumosus populations from Yeosu in Korea, which has high concentrations of polluting heavy metals. SP transcript expression was positively correlated with concentrations of Cd, Pb, Al, Fe, NO2, and NO3. These results suggested that environmental pollutants such as Cd can impair proteolytic activity in the digestive system of C. plumosus and may ultimately induce developmental alterations. We therefore suggest SP as a potential biomarker to assess the effects of environmental pollutants in aquatic ecosystems.

Enquiring beneath the surface: can a gene expression assay shed light into the heterogeneity among newborns with neonatal encephalopathy?

BACKGROUND: We aimed to assess whether a gene expression assay provided insights for understanding the heterogeneity among newborns affected by neonatal encephalopathy (NE). METHODS: Analysis by RT-qPCR of the mRNA expression of candidate genes in whole blood from controls (n = 34) and NE (n = 24) patients at <6, 12, 24, 48, 72 and 96 h of life, followed by determination of differences in gene expression between conditions and correlation with clinical variables. RESULTS: During the first 4 days of life, MMP9, PPARG, IL8, HSPA1A and TLR8 were more expressed and CCR5 less expressed in NE patients compared to controls. MMP9 and PPARG increased and CCR5 decreased in moderate/severe NE patients compared to mild. At 6-12 h of life, increased IL8 correlated with severe NE and death, decreased CCR5 correlated with chorioamnionitis and increased HSPA1A correlated with expanded multiorgan dysfunction, severe NE and female sex. CONCLUSIONS: MMP9, PPARG and CCR5 mRNA expression within first days of life correlates with the severity of NE. At 6-12 h, IL8 and HSPA1A are good reporters of clinical variables in NE patients. HSPA1A may have a role in the sexual dimorphism observed in NE. CCR5 is potentially involved in the link between severe NE and chorioamnionitis.

Role of Apg-1 in HSF1 activation and bortezomib sensitivity in myeloma cells.

Exposure of cells to bortezomib (BTZ) could activate heat shock protein (HSP) expression, which is regulated mainly by heat shock factor 1 (HSF1). We determined the role of Apg-1 (HSPA4L, a member of the HSP110 family) in HSF1 activation and bortezomib sensitivity by silencing HSF1 in multiple myeloma (MM) cells. We observed that the Apg-1 protein level was upregulated as BTZ concentration increased. To investigate the mechanism underlying Apg-1 induction, we evaluated the HSF1 translocation and found BTZ-inducible transposition to the nucleus of HSF1. In addition, cleaved caspase 3 and PARP might account for increased BTZ sensitivity on Apg-1 silencing. Furthermore, silencing HSF1 with shRNA or triptolide resulted in significant BTZ sensitivity. It had a more profound effect on cell death caused by BTZ when myeloma cells were adherent to bone marrow stromal cell lines (Hs-5). In summary, we found that Apg-1 knockdown sensitized myeloma cells to bortezomib treatment, which may provide a new approach in multiple myeloma treatment.