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Even though aberrant mechanistic target of rapamycin (mTOR) signaling is known to cause cardiomyopathy, its underlying mechanism remains poorly understood. Because augmentation of αB-crystallin and hspB2 was presented in the cortical tubers and lymphangioleiomyomatosis of tuberous sclerosis complex patients, we deciphered the role of αB-crystallin and its adjacent duplicate gene, hspB2, in hyperactive mTOR-induced cardiomyopathy. Cardiac Tsc1 deletion (T1-hKO) caused mouse mTOR activation and cardiomyopathy. Overexpression of αB-crystallin and hspB2 was presented in the hearts of these mice. Knockout of αB-crystallin/hspB2 reversed deficient Tsc1-mediated fetal gene expression, mTOR activation, mitochondrial damage, cardiomyocyte vacuolar degeneration, cardiomyocyte size, and fibrosis of T1-hKO mice. These cardiac-Tsc1; αB-crystallin; hspB2 triple knockout (tKO) mice had improved cardiac function, smaller heart weight to body weight ratio, and reduced lethality compared with T1-hKO mice. Even though activated mTOR suppressed autophagy in T1-hKO mice, ablation of αB-crystallin and hspB2 failed to restore autophagy in tKO mice. mTOR inhibitors suppressed αB-crystallin expression in T1-hKO mice and rat cardiomyocyte line H9C2. Starvation of H9C2 cells activated autophagy and suppressed αB-crystallin expression. Since inhibition of autophagy restored αB-crystallin expression in starved H9C2 cells, autophagy is a negative regulator of αB-crystallin expression. mTOR thus stimulates αB-crystallin expression through suppression of autophagy. In conclusion, αB-crystallin and hspB2 play a pivotal role in Tsc1 knockout-related cardiomyopathy and are therapeutic targets of hyperactive mTOR-associated cardiomyopathy., (© 2021 Wiley Periodicals LLC.)

Background: Phototherapy is a promising strategy for cancer therapy by reactive oxygen species (ROS) of photodynamic therapy (PDT) and hyperthermia of photothermal therapy (PTT). However, the therapeutic efficacy was restricted by tumor hypoxia and thermal resistance of increased expression of heat shock protein (Hsp). In this study, we developed albumin nanoparticles to combine hypoxia relief and heat shock protein inhibition to overcome these limitations for phototherapy enhancement., Results: Near-infrared photosensitizer (IR780) and gambogic acid (GA, Hsp90 inhibitor) were encapsulated into albumin nanoparticles via hydrophobic interaction, which was further deposited MnO 2 on the surface to form IGM nanoparticles. Both in vitro and in vivo studies demonstrated that IGM could catalyze overexpress of hydrogen peroxide to relive hypoxic tumor microenvironment. With near infrared irradiation, the ROS generation was significantly increase for PDT enhancement. In addition, the release of GA was promoted by irradiation to bind with Hsp90, which could reduce cell tolerance to heat for PTT enhancement. As a result, IGM could achieve better antitumor efficacy with enhanced PDT and PTT., Conclusion: This study develops a facile approach to co-deliver IR780 and GA with self-assembled albumin nanoparticles, which could relive hypoxia and suppress Hsp for clinical application of cancer phototherapy.

Hepatic ischemia-reperfusion injury (IRI) is a major unavoidable clinical problem often accompanying various liver surgery and transplantation. d-Pinitol, a cyclic polyol, exhibits hepatoprotective efficacy. The objective of this study is to determine the possible mechanism of action of pinitol against endoplasmic reticulum (ER) stress regulation-mediated hepatic IRI and compare its effects with thymoquinone (TQ) in experimental rats. Male Sprague Dawley rats were pre-treated orally with either vehicle (DMSO) or d-Pinitol (5, 10, and 20 mg/kg) or TQ (30 mg/kg) for 21 days and subjected to 60 min of partial hepatic ischemia followed by 24 h of reperfusion. Pre-treatment with pinitol (10 and 20 mg/kg) effectively ( P   <  0.05) protected against IRI-induced hepatic damage reflected by attenuation of elevated oxidative stress and pro-inflammatory cytokines. Additionally, western blot and ELISA analyses suggested that pinitol significantly ( P   <  0.05) down-regulated expression of endoplasmic reticulum stress apoptotic markers, namely glucose-regulated protein (GRP)-78, CCAAT/enhancer-binding protein homologous protein (CHOP), activating transcription factor (AFT)-4 and -6α, X-box binding protein-1, and caspase-3, 9, and 12. Additionally, pinitol pre-treatment effectively ( P   <  0.05) improved mitochondrial function and phosphorylation of Extracellular signal-regulated kinase (ERK)-1/2 and p38. Pinitol markedly ( P   <  0.05) protected hepatic apoptosis determined by flow cytometry. Further, pinitol provided effective ( P   <  0.05) protection against hepatic histological and ultrastructural aberrations induced by IRI. TQ showed more pronounced protective effect against attenuation of IRI-induced hepatic injury as compared to d-Pinitol. Pinitol offered protection against endoplasmic reticulum stress-mediated phosphorylation of ERK1/2 and p38, thereby inhibiting AFT4-CHOP/GRP78 signaling response and caspase-3 induced hepatocellular apoptosis during hepatic ischemia-reperfusion insults. Thus, Pinitol can be considered as a viable option for the management of hepatic IRI.

Background: Heat shock proteins (HSPs), a representative family of chaperone genes, play crucial roles in malignant progression and are pursued as attractive anti-cancer therapeutic targets. Despite tremendous efforts to develop anti-cancer drugs based on HSPs, no HSP inhibitors have thus far reached the milestone of FDA approval. There remains an unmet need to further understand the functional roles of HSPs in cancer., Methods: We constructed the network for HSPs across ~ 10,000 tumor samples from The Cancer Genome Atlas (TCGA) and ~ 10,000 normal samples from Genotype-Tissue Expression (GTEx), and compared the network disruption between tumor and normal samples. We then examined the associations between HSPs and cancer hallmarks and validated these associations from multiple independent high-throughput functional screens, including Project Achilles and DRIVE. Finally, we experimentally characterized the dual function effects of HSPs in tumor proliferation and metastasis., Results: We comprehensively analyzed the HSP expression landscape across multiple human cancers and revealed a global disruption of the co-expression network for HSPs. Through analyzing HSP expression alteration and its association with tumor proliferation and metastasis, we revealed dual functional effects of HSPs, in that they can simultaneously influence proliferation and metastasis in opposite directions. We experimentally characterized the dual function of two genes, DNAJC9 and HSPA14, in lung cancer cells. We further demonstrated the generalization of this dual direction of associations between HSPs and cancer hallmarks, suggesting the necessity to more carefully evaluate HSPs as therapeutic targets and develop highly specific HSP inhibitors for cancer intervention., Conclusions: Our study furnishes a holistic view of functional associations of HSPs with cancer hallmarks to aid the development of HSP inhibitors as well as other drugs in cancer therapy.

Rising antibiotic resistance urgently begs for novel targets and strategies for antibiotic discovery. Here, we report that over-activation of the periplasmic DegP protease, a member of the highly conserved HtrA family, can be a viable strategy for antibiotic development. We demonstrate that tripodal peptidyl compounds that mimic DegP-activating lipoprotein variants allosterically activate DegP and inhibit the growth of an Escherichia coli strain with a permeable outer membrane in a DegP-dependent fashion. Interestingly, these compounds inhibit bacterial growth at a temperature at which DegP is not essential for cell viability, mainly by over-proteolysis of newly synthesized proteins. Co-crystal structures show that the peptidyl arms of the compounds bind to the substrate-binding sites of DegP. Overall, our results represent an intriguing example of killing bacteria by activating a non-essential enzyme, and thus expand the scope of antibiotic targets beyond the traditional essential proteins or pathways.

Prostaglandin E2 (PGE2) is involved in numerous physiological and pathological processes of the kidney via its four receptors. A previous study has suggested that a defect in the PGE2 receptor 1 (EP1) gene markedly suppressed the transforming growth factor‑β1 (TGF‑β1)‑induced mesangial cell (MC) proliferation and extracellular matrix aggregation. Therefore, the present study aimed to adopt a pharmacological method of specifically suppressing or activating the EP1 receptor to further verify and demonstrate these results. The EP1 receptor antagonist SC‑19220 and EP1 receptor agonist 17‑phenyl‑trinor‑PGE2 ethyl amide (17‑pt‑PGE2) were selectively used to treat five‑sixths nephrectomy renal fibrosis model mice and TGF‑β1‑stimulated MCs. An Alpha screen PGE2 assay kit, flow cytometry, western blotting and immunohistochemical techniques were adopted to perform in vivo and in vitro experiments. The present results suggested that compared with the control group, the selective EP1 receptor antagonist SC‑19220 improved renal function, markedly reduced the plasma blood urea nitrogen and creatinine levels (P<0.05) and alleviated glomerulosclerosis (P<0.05). By contrast, the EP1 receptor agonist 17‑pt‑PGE2 aggravated renal dysfunction and glomerulosclerosis (P<0.05). To verify the renal protection mechanisms mediated by suppression of the EP1 receptor, the expression levels of endoplasmic reticulum stress (ERS)‑related proteins, including chaperone glucose‑regulated protein 78 (GRP78), transient receptor potential channel 1 (TRPC1) and protein kinase R‑like endoplasmic reticulum kinase (PERK), were further evaluated histologically. The expression of GRP78, TRPC1 and PERK in the antagonist treatment group were markedly downregulated (P<0.05), whereas those in the agonist treatment group were upregulated (P<0.05). The present in vitro experiments demonstrated that, compared with the control group, the EP1 receptor antagonist suppressed the expression of GRP78, TRPC1 and PERK (P<0.05), reduced the production of PGE2 (P<0.05) and decreased the MC apoptosis rate (P<0.05), thus alleviating TGF‑β1‑stimulated MC injury. Consequently, consistent with previous results, selectively antagonizing the EP1 receptor improved renal function and mitigated glomerulosclerosis, and its potential mechanism might be associated with the suppression of ERS.

An overwhelming endoplasmic reticulum stress (ERS) and the following unfolded protein response (UPR) can induce hepatic inflammation, fibrosis and hepatocellular carcinoma (HCC). Caudatin, one of the species of C‑21 steroidal glycosides mainly isolated from the roots of Cynanchum bungei Decne, exhibits potent anticancer activities in vivo. However, the effect of caudatin on HCC remains unclear. In the present study, a diethylnitrosamine (DEN)‑induced HCC model was established. Nodules and tumors in rat livers were monitored by T2‑/T1‑weighted‑magnetic resonance imaging (MRI) using a 1.5 T scanner. Caudatin reduced the number and size of nodules and alleviated the inflammatory foci in the liver. In addition, the hepatic pro‑inflammatory levels of interleukin (IL) 6, monocyte chemoattractant protein 1 and IL‑1β were decreased in caudatin‑treated rats. The DEN‑induced surge in malondialdehyde, aspartate aminotransferase, alanine transaminase and TBIL were alleviated following caudatin treatment. The expression of ERS chaperones glucose‑regulated protein, 94 kDa, glucose‑regulated protein, 78 kDa and protein disulfide‑isomerase A4 and the proliferation marker Ki‑67 in liver nodules were all downregulated by caudatin as demonstrated by immunohistochemistry, reverse transcription‑quantitative PCR and western blot analysis. Caudatin reduced the cytoprotective ERS sensor activating transcription factor 6‑mediated signal transduction and inhibited the PKR‑like endoplasmic reticulum kinase/eukaryotic initiation factor 2α/activating transcription factor 4 pathway. However, the effect of caudatin on inositol requiring enzyme 1 signaling was negligible. In conclusion, restoration of the dysregulated UPR program was involved in the antitumor efficacy of caudatin without inducing cumulative hepatotoxicity.

Several pathophysiological processes involve Hypoxia conditions, where the nervous system is affected as well. We postulate that the GABAergic system is especially sensitive. Furthermore, drugs improving the resistance to hypoxia have been investigated, such as the neurosteroid dehydroepiandrosterone sulfate (DHEAS) which has shown beneficial effects in hypoxic processes in mammals; however, at the cellular level, its exact mechanism of action has yet to be fully elucidated. Here, we used a chemical hypoxia model through sodium sulfite (SS) exposure in Caenorhabditis elegans (C. elegans), a nematode whose response to hypoxia involves pathways and cellular processes conserved in mammals, and that allows study the direct effect of DHEAS without its conversion to sex hormones. This work aimed to determine the effect of DHEAS on damage to the GABAergic system associated with SS exposure in C. elegans. Worms were subjected to nose touch response (Not Assay) and observed in epifluorescence microscopy. DHEAS decreased the shrinkage response of Not Assay and the level of damage in GABAergic neurons on SS-exposed worms. Also, the enhanced nuclear localization of DAF-16 and consequently the overexpression of chaperone HSP-16.2 by hypoxia were significantly reduced in SS + DHEAS exposed worms. As well, DHEAS increased the survival rate of worms exposed to hydrogen peroxide. These results suggest that hypoxia-caused damage over the GABAergic system was prevented at least partially by DHEAS, probably through non-genomic mechanisms that involve its antioxidant properties related to its chemical structure.

The 78-kDa glucose-regulated protein (GRP78), an endoplasmic reticulum (ER) chaperone, is a master regulator of the ER stress. A number of studies revealed that high levels of GRP78 protein in cancer cells confer multidrug resistance (MDR) to therapeutic treatment. Therefore, drug candidate that reduces GRP78 may represent a novel approach to eliminate MDR cancer cells. Our earlier studies showed that a set of 4 H -chromene derivatives induced selective cytotoxicity in MDR cancer cells. In the present study, we elucidated its selective mechanism in four MDR cancer cell lines with one lead candidate (CXL146). Cytotoxicity results confirmed the selective cytotoxicity of CXL146 toward the MDR cancer cell lines. We noted significant overexpression of GRP78 in all four MDR cell lines compared with the parental cell lines. Unexpectedly, CXL146 treatment rapidly and dose-dependently reduced GRP78 protein in MDR cancer cell lines. Using human leukemia (HL) 60/mitoxantrone (MX) 2 cell line as the model, we demonstrated that CXL146 treatment activated the unfolded protein response (UPR); as evidenced by the activation of inositol-requiring enzyme 1 α , protein kinase R-like ER kinase, and activating transcription factor 6. CXL146-induced UPR activation led to a series of downstream events, including extracellular signal-regulated kinase 1/2 and c-Jun N-terminal kinase activation, which contributed to CXL146-induced apoptosis. Targeted reduction in GRP78 resulted in reduced sensitivity of HL60/MX2 toward CXL146. Long-term sublethal CXL146 exposure also led to reduction in GRP78 in HL60/MX2. These data collectively support GRP78 as the target of CXL146 in MDR treatment. Interestingly, HL60/MX2 upon long-term sublethal CXL146 exposure regained sensitivity to mitoxantrone treatment. Therefore, further exploration of CXL146 as a novel therapy in treating MDR cancer cells is warranted. SIGNIFICANCE STATEMENT: Multidrug resistance is one major challenge to cancer treatment. This study provides evidence that cancer cells overexpress 78-kDa glucose-regulated protein (GRP78) as a mechanism to acquire resistance to standard cancer therapies. A chromene-based small molecule, CXL146, selectively eliminates cancer cells with GRP78 overexpression via activating unfolded protein response-mediated apoptosis. Further characterization indicates that CXL146 and standard therapies complementarily target different populations of cancer cells, supporting the potential of CXL146 to overcome multidrug resistance in cancer treatment., (Copyright © 2020 by The American Society for Pharmacology and Experimental Therapeutics.)

Background: Listeria monocytogenes (L. monocytogenes) is a global opportunistic intracellular pathogen that can cause many infections, including meningitis and abortion in humans and animals; thus, L. monocytogenes poses a great threat to public safety and the development of the aquaculture industry. The isolation rate of Listeria monocytogenes in fishery products has always been high. And the pore-forming toxin listeriolysin O (LLO) is one of the most important virulence factors of L. monocytogenes. LLO can promote cytosolic bacterial proliferation and help the pathogen evade attacks from the host immune system. In addition, L. monocytogenes infection can trigger a series of severe inflammatory reactions., Results: Here, we further confirmed that morin lacking anti-Listeria activity could inhibit LLO oligomerization. We also found that morin can effectively alleviate the inflammation induced by Listeria in vivo and in vitro and exerted an obvious protective effect on infected cells and mice., Conclusions: Morin does not possess anti-Listeria activity, neither does it interfere with secretion of LLO. However, morin inhibits oligomerisation of LLO and morin does reduce the inflammation caused during Listeria infection.

Antiretroviral therapy (ART) suppresses HIV-1 replication but fails to cure the infection. The presence of an extremely stable viral latent reservoir, primarily in resting memory CD4 + T cells, remains a major obstacle to viral eradication. The "shock and kill" strategy targets these latently infected cells and boosts immune recognition and clearance, and thus, it is a promising approach for an HIV-1 functional cure. Although some latency-reversing agents (LRAs) have been reported, no apparent clinical progress has been made, so it is still vital to seek novel and effective LRAs. Here, we report that thiostrepton (TSR), a proteasome inhibitor, reactivates latent HIV-1 effectively in cellular models and in primary CD4 + T cells from ART-suppressed individuals ex vivo TSR does not induce global T cell activation, severe cytotoxicity, or CD8 + T cell dysfunction, making it a prospective LRA candidate. We also observed a significant synergistic effect of reactivation when TSR was combined with JQ1, prostratin, or bryostatin-1. Interestingly, six TSR analogues also show reactivation abilities that are similar to or more effective than that of TSR. We further verified that TSR upregulated expression of heat shock proteins (HSPs) in CD4 + T cells, which subsequently activated positive transcriptional elongation factor b (p-TEFb) and NF-κB signals, leading to viral reactivation. In summary, we identify TSR as a novel LRA which could have important significance for applications to an HIV-1 functional cure in the future., (Copyright © 2020 American Society for Microbiology.)

Objectives: Xerostomia in SS patients has been associated with low quality and quantity of salivary mucins, which are fundamental for the hydration and protection of the oral mucosa. The aim of this study was to evaluate if cytokines induce aberrant mucin expression and whether tauroursodeoxycholic acid (TUDCA) is able to counteract such an anomaly., Methods: Labial salivary glands from 16 SS patients and 15 control subjects, as well as 3D acini or human submandibular gland cells stimulated with TNF-α or IFN-γ and co-incubated with TUDCA, were analysed. mRNA and protein levels of Mucin 1 (MUC1) and MUC7 were determined by RT-qPCR and western blot, respectively. Co-immunoprecipitation and immunofluorescence assays for mucins and GRP78 [an endoplasmic reticulum (ER)-resident protein] were also performed. mRNA levels of RelA/p65 (nuclear factor-κB subunit), TNF-α, IL-1β, IL-6, SEL1L and EDEM1 were determined by RT-qPCR, and RelA/p65 localization was evaluated by immunofluorescence., Results: MUC1 is overexpressed and accumulated in the ER of labial salivary gland from SS patients, while MUC7 accumulates throughout the cytoplasm of acinar cells; however, MUC1, but not MUC7, co-precipitated with GRP78. TUDCA diminished the overexpression and aberrant accumulation of MUC1 induced by TNF-α and IFN-γ, as well as the nuclear translocation of RelA/p65, together with the expression of inflammatory and ER stress markers in 3D acini., Conclusion: Chronic inflammation alters the secretory process of MUC1, inducing ER stress and affecting the quality of saliva in SS patients. TUDCA showed anti-inflammatory properties decreasing aberrant MUC1 accumulation. Further studies are necessary to evaluate the potential therapeutic effect of TUDCA in restoring glandular homeostasis in SS patients., (© The Author(s) 2019. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Purpose: Glutamine, as an essential part of enteral nutrition and parenteral nutrition agent, has been widely recognized to be a kind of important intestinal mucosa protectant in clinical practice and experimental research. However, the mechanisms of its protective effects are still not fully understand. Consequently, this study aimed to explore the potential mechanism of glutamine on ischemia-reperfusion (I/R) injury induced endoplasmic reticulum (ER) stress in intestine., Methods: An experimental model of intestinal I/R in rats was established by 1 hour occlusion of the superior mesenteric artery followed by 3 hours of reperfusion. Morphologic changes of intestinal mucosa, apoptosis of epithelial cells, and expression of intestinal Grp78, Gadd153, Caspase-12, ATF4, PERK phosphorylation (P-PERK) and elF2αphosphorylation(P-elF2α) were determined., Results: After I/R, the apoptotic index of intestinal mucosa epithelial cells observably increased with notable necrosis of intestinal mucosa, and the expressions of Grp78, Gadd153, Caspase-12, ATF4, P-PERK and P-elF2αall were increased. However, treatment with glutamine could significantly relieve intestinal I/R injury and apoptosis index. Moreover, glutamine could clearly up-regulate the expression of Grp78, restrain P-PERK and P-elF2α, and reduce ATF4, Gadd153 and Caspase-12 expressions., Conclusion: Glutamine may be involved in alleviating ER stress induced intestinal mucosa cells apoptosis.

Escherichia coli is a common pathogen of bacterial biofilm infections. Sub-minimum inhibitory concentration ceftazidime (sub-MIC CAZ) could inhibit the biofilm formation of E. coli . Deletion of the ibpAB genes could increase the extracellular indole concentration of E. coli and then inhibit biofilm formation. Therefore, we speculated that sub-MIC CAZ might inhibit biofilm formation via ibpAB . In this study, the results showed that sub-MIC CAZ could significantly inhibit biofilm formation, swimming motility and the expression of the ibpA gene, while it could increase the expression of tnaA gene and extracellular indole concentration. Knockout of the ibpA gene resulted in a decrease in biofilm formation and swimming motility and an increase in the indole concentration. When treated with sub-MIC CAZ, the tnaA gene expression, indole concentration, biofilm formation and swimming motility of MG1655 Δ ibpA were similar to those of the control group. The results indicated that sub-MIC CAZ might inhibit the biofilm formation of E. coli by increasing the extracellular indole concentration and downregulating the ibpA gene.

Our previous study had shown that chronic corticosterone (CORT) exposure causes excessive fat deposition in chicken liver, yet it remains unknown whether it is associated with inflammation and fibrosis. In general, heat shock proteins (HSPs) are activated in response to acute stress to play a cytoprotective role, and this activation is associated with m 6 A-mediated post-transcriptional regulation. However, changes of HSPs and the m 6 A methylation on their mRNAs in response to chronic CORT treatment in chicken liver have not been reported. In this study, chronic CORT exposure induced inflammation and fibrosis in chicken liver, associated with significantly modulated expression of HSPs that was significantly upregulated at mRNA level yet downregulated at protein level. Concurrently, m 6 A methyltransferases METTL3 content was upregulated together with the level of m 6 A methylation on HSPs transcripts. The m 6 A-seq analysis revealed 2-6 significantly (P < 0.05) hypermethylated m 6 A peaks in the mRNA of 4 different species of HSPs in CORT-treated chicken liver. HSP90B1 transcript had 6 differentially methylated m 6 A peaks among which peaks on exon 16 and exon 17 showed 3.14- and 4.72-fold of increase, respectively. Mutation of the 8 predicted m 6 A sites on exon 16 and exon 17 resulted in a significant (P < 0.05) increase in eGFP-fused content of HSP90B1 exon 16 and exon 17 fragment in 293 T cells, indicating a possible role of m 6 A in post-transcriptional regulation of HSPs. In conclusion, chronic CORT exposure induces inflammation and fibrosis in chicken liver along with an increase in the levels and m6A methylation of several HSPs mRNAs; HSPs levels were however reduced under the indicated conditions. Results presented suggest that the reduction in HSPs levels may be associated with m6A methylation in CORT-exposed chickens.

Upregulation of heat shock proteins (HSPs) is an approach to treatment of neurodegenerative disorders with impaired proteostasis. Many neurons, including motor neurons affected in amyotrophic lateral sclerosis (ALS), are relatively resistant to stress-induced upregulation of HSPs. This study demonstrated that histone deacetylase (HDAC) inhibitors enable the heat shock response in cultured spinal motor neurons, in a stress-dependent manner, and can improve the efficacy of HSP-inducing drugs in murine spinal cord cultures subjected to thermal or proteotoxic stress. The effect of particular HDAC inhibitors differed with the stress paradigm. The HDAC6 (class IIb) inhibitor, tubastatin A, acted as a co-inducer of Hsp70 (HSPA1A) expression with heat shock, but not with proteotoxic stress induced by expression of mutant SOD1 linked to familial ALS. Certain HDAC class I inhibitors (the pan inhibitor, SAHA, or the HDAC1/3 inhibitor, RGFP109) were HSP co-inducers comparable to the hydroxyamine arimoclomol in response to proteotoxic stress, but not thermal stress. Regardless, stress-induced Hsp70 expression could be enhanced by combining an HDAC inhibitor with either arimoclomol or with an HSP90 inhibitor that constitutively induced HSPs. HDAC inhibition failed to induce Hsp70 in motor neurons expressing ALS-linked mutant FUS, in which the heat shock response was suppressed; yet SAHA, RGFP109, and arimoclomol did reduce loss of nuclear FUS, a disease hallmark, and HDAC inhibition rescued the DNA repair response in iPSC-derived motor neurons carrying the FUS P525L mutation, pointing to multiple mechanisms of neuroprotection by both HDAC inhibiting drugs and arimoclomol.

Aim: Renal toxicity of adefovir disoproxil (ADV) and tenofovir disoproxil fumarate (TDF) is a significant concern in chronic hepatitis B (CHB) patients. Early observational clinical data suggested that telbivudine (LdT) might have renoprotective effects., Methods: In this prospective study, consecutive CHB patients on combined lamivudine (LAM) + ADV/TDF were switched to LdT + ADV/TDF at recruitment and were followed up for 24 months. Estimated glomerular filtration rate (eGFR) was calculated with the modification of diet in renal disease equation. The effects of LdT on cell viability and expression of kidney injury or apoptotic biomarkers were investigated in cultured renal tubular epithelial cell line HK-2., Results: Thirty-one patients (median age 55 years, 90.3% male) were recruited (54.8% TDF: 45.2% ADV). Serum HBV DNA was undetectable at all time points. Median eGFR was 70.2 (IQR 62.6-77.9) and 81.5 (IQR 63.6-99.1) mL/min/1.73 m 2 at baseline and 24 months, respectively (p < 0.001). Downstaging of chronic kidney disease was observed in eight (25.8%) patients and was more common in ADV-treated compared to TDF-treated patients (7/8 vs. 1/17, p = 0.011; OR 16, 95% CI 1.643-155.766, p = 0.017). In vitro data showed that adding LdT to ADV or TDF was associated with improved cell viability and lower expression of injury and apoptotic biomarkers compared with ADV or TDF alone. Treatment was prematurely discontinued in four(12.9%) patients due to myalgia., Conclusions: Clinical and in vitro data suggest that LdT has renoprotective effects in patients on long-term ADV/TDF treatment. LdT may be considered as an adjuvant therapy in this special group of patients with renal impairment (NCT03778567).

Heat shock proteins (HSPs) are evolutionary conserved proteins that work as molecular chaperones and perform broad and crucial roles in proteostasis, an important process to preserve the integrity of proteins in different cell types, in health and disease. Their function in cancer is an important aspect to be considered for a better understanding of disease development and progression. Glioblastoma (GBM) is the most frequent and lethal brain cancer, with no effective therapies. In recent years, HSPs have been considered as possible targets for GBM therapy due their importance in different mechanisms that govern GBM malignance. In this review, we address current evidence on the role of several HSPs in the biology of GBMs, and how these molecules have been considered in different treatments in the context of this disease, including their activities in glioblastoma stem-like cells (GSCs), a small subpopulation able to drive GBM growth. Additionally, we highlight recent works that approach other classes of chaperones, such as histone and mitochondrial chaperones, as important molecules for GBM aggressiveness. Herein, we provide new insights into how HSPs and their partners play pivotal roles in GBM biology and may open new therapeutic avenues for GBM based on proteostasis machinery.

Sirtuins have emerged as a promising novel class of anti-cancer drug targets. Inhibition of SIRT1 and SIRT2 induces apoptosis in cancer cells and they play multifaceted roles in regulating autophagy. In the present study, we found that salermide, a SIRT1/2-specific inhibitor or small interfering RNAs (siRNAs) to block SIRT1/2 expression could induce autophagy in human NSCLC cells. Moreover, SIRT1/2 inhibition increased the expression levels of ATF4 and DDIT4 and downregulated p-RPS6KB1 and p-EIF4EBP1, two downstream molecules of mTORC1. Moreover, ATF4 or DDIT4 knockdown attenuated salermide-induced autophagy, suggesting that SIRT1/2 inhibition induced autophagy through the ATF4-DDIT4-mTORC1 axis. Mechanistically, SIRT1/2 inhibition led to HSPA5 acetylation and dissociation from EIF2AK3, leading to ER stress response and followed by upregulation of ATF4 and DDIT4, triggering autophagy. Silencing of the autophagic gene ATG5 in lung cancer cells resulted in increased apoptotic cell death induced by SIRT1/2 inhibition. Our data show that inhibition of SIRT1/2 induces pro-survival autophagy via acetylation of HSPA5 and subsequent activation of ATF4 and DDIT4 to inhibit the mTOR signaling pathway in NSCLC cells. These findings suggest that combinatorial treatment with SIRT1/2 inhibitors and pharmacological autophagy inhibitors is an effective therapeutic strategy for cancer therapy.

Solar ultraviolet (UV) radiation is the main factor of photocarcinogenesis, photoaging, and photosensitivity; thus protection from biological damaging UV radiation is a concern. Sunscreens containing UV filters are the most preferred means of photoprotection but the safety and efficacy of UV filters are in question. Benzophenone (BP) and its derivatives, namely, benzophenone 1 (BP1), is commonly used in sunscreens as a UV blocker. The aim of this study was to assess the effects of BP and BP1 on the differential expression of proteins in human keratinocytes (HaCaT cells) under exposure to ultraviolet A radiation. Photosensitive proteins were screened from HaCaT cells by two-dimensional (2-D) gel electrophoresis, and identification of these differentially expressed proteins was performed by matrix-assisted laser desorption/ionization-time-of-flight (MALDI-TOF)/TOF mass spectrometry. Protein identification was performed using the search program MASCOT and a database made of SUMO and GhJMJ12 amino acid sequences. Our results showed that the proteins involved directly or indirectly in apoptosis are 70 kDa heat shock protein, long-chain specific acyl-CoA dehydrogenase, serine/threonine-protein kinase, and FAM78A protein, which were upregulated in comparison to control HaCaT cells. The expressions of binding immunoglobulin protein, podocalyxin-like protein, actin, cytoplasmic, and calreticulin precursors were downregulated. The altered protein expression indicated that cell growth arrest and apoptosis were potential mechanisms of cytotoxicity and genotoxicity of BPs. The results of 2-D gel electrophoresis followed by mass spectrometry showed expression of novel proteins involved in promoting or initiating apoptotic pathways. Hence, we conclude that BPs should be avoided as a UV blocker from sunscreens because of its potential to promote apoptotic proteins in human skin keratinocytes.

Green tea accounts for approximately 20% of the world's total tea yield. (-)-Epigallocatechin gallate (EGCG) is an active catechin in green tea, which suppresses tumor growth and enhances drug sensitivity in various cancers, but the molecular mechanism is still unclear. Chemotherapy drugs, such as 5-fluorouracil (5-FU), are a common strategy for clinical treatment of cancer patients; however, the lower response rate caused by prolonged use becomes the main reason for tumor recurrence. Therefore, discovering a safe and effective chemo-sensitizer is an urgent task required to be solved. Here, we report that EGCG reinforces the sensitivity of colon cancer cells to 5-FU, and the IC 50 values of 5-FU is decreased from 40 ± 4.2 μM to 5 ± 0.36 μM in one human colon carcinoma cell line-HCT-116, and from 150 ± 6.4 μM to 11 ± 0.96 μM in the other human colon carcinoma cell line-DLD1 when these cells are cotreated with 50 μM EGCG. Consistently, compared to 5-FU or EGCG treatment alone, the combination of both significantly promotes cancer cell apoptosis and DNA damage. Further mechanism research reveals that treatment of colorectal cancer (CRC) with 50 μM EGCG inhibits GRP78 expression, activates the NF-κB (2.55 ± 0.05-fold for HCT-116 and 2.27 ± 0.08-fold for DLD1) pathway, and enhances miR-155-5p (2.12 ± 0.02-fold for HCT-116 and 2.01 ± 0.01-fold for DLD1) level. The elevated miR-155-5p strongly suppresses target gene MDR1 expression, which blocks the efflux of 5-FU. The accumulation of 5-FU resulted in caspase-3 and PARP activation, Bcl-2 reduction, and Bad increase, which ultimately lead to cancer cell apoptosis. Overall, our data show that EGCG may be act as a novel chemo-sensitizer, and the GRP78/NF-κB/miR-155-5p/MDR1 pathway plays a vital role in EGCG enhancing the sensitivity of colorectal cancer to 5-FU.

Aim: Myocardial ischemia/reperfusion (I/R) injury is a severe trauma that cells undergo and is associated with cardiomyocyte apoptosis. Recently, miRNAs have been demonstrated to play an important role in cardiovascular biology and disease. However, whether the miR-133a and ER stress play a role in hydrogen sulfide (H2S) protection of cardiomyocytes against I/R-induced apoptosis remains unclear., Methods: The neonatal cardiomyocytes were prepared to be treated with H2S or transfected with miR-133a activator or miR-133a inhibitor, either separately or in combination. Non-treated cardiomyocytes served as control. The ER stress biomarker GRP78, CHOP, and eIF2α expression levels were measured by Western blot. Cell apoptosis was assessed by flow cytometry after staining with the Annexin V- FITC. Proliferation was monitored by BrdU labeling, while cell migration and invasion were determined by Transwell assays., Results: Pre-treatment of H2S and overexpression of miR-133a reversed I/R-induced ER stress and cardiomyocyte apoptosis in vitro and in vivo. The proliferation, migration, and invasion of cardiomyocytes were significantly increased by co-treatment with H2S and overexpression of miR-133a., Conclusion: These findings suggest the protective effect of miR-133a against I/R-induced ER stress and cardiomyocyte apoptosis and its enhancement of cell motility. Thus, cardioprotection by miR-133a overexpression provides a novel therapeutic approach to the treatment of ischemic heart diseases., (© 2018 S. Karger AG, Basel.)

In this study, we explored the presence and elevation of glucose-regulated protein 78 (GRP78) in aqueous humor of patients with diabetic macular edema (DME).After comparing DME patients with the controls, we analyzed GRP78 and vascular endothelial growth factor (VEGF) levels in DME patients. We examined factors associated with GRP78 levels in DME patients.GRP78 was detected in aqueous humor with elevated levels in DME patients. Stepwise backward regression analysis showed that GRP78 levels were associated with the VEGF levels and the duration of diabetes (P < .001 and P = .002, respectively). However, no statistical significance was observed between GRP78 levels and the decrease in CST following 3 monthly anti-VEGF treatments in univariate regression analysis (P = .695).We showed that GRP78 is elevated in DME patients. In addition, there is a correlation between GRP78 and VEGF levels in aqueous humor. However, GRP78 levels were not associated with the responsiveness of anti-VEGF in DME patients.

Background: Skeletal muscle atrophy is a striking example of the multiple changes in the physiological state of humans and animals induced by microgravity. Previous studies have shown that a blood circulation disorder may be a cause of this atrophy, and traditional Chinese medicine has been regarded as a potential countermeasure to reverse the atrophy in China. This study was carried out to test the effects of Xuefuzhuyu capsules (XFZY) on the skeletal muscle atrophy induced by simulated microgravity., Methods: The mass and cross-sectional area of the soleus muscle were compared in rats treated with XFZY that were hindlimb unloaded for 30 d (XFZY-TS group), untreated rats that were hindlimb unloaded for 30 d (TS group), and control rats (CON group). The expression and phosphorylation levels of key proteins of the sarcoplasmic reticulum stress system were also measured., Results: Treatment with XFZY attenuated the loss of muscle mass and cross-sectional area induced by hindlimb unloading. Western blot analysis showed that the phosphatidyl-inositol-3-kinase/phospho-Akt (PI3K/p-Akt) pathways were down-regulated after 30 d in the TS group compared with the CON group. This effect was partly reversed by XFZY. Hindlimb unloading increased the expression of glucose-regulated protein 78 (GRP78), cytosine-cytosine-adenosine-adenosine-thymidine/enhancer-binding protein homologous protein (CHOP), C-Jun N-terminal kinase (JNK), and Caspase 12. Treatment with XFZY alleviated this increased protein expression., Discussion: Our results suggest that XFZY could partially reverse the effects of hindlimb unloading on muscle atrophy and perhaps target the sarcoplasmic reticulum stress system, possibly through the GRP78-CHOP-JNK-Caspase 12 pathway.Zhang S, Yuan M, Cheng C, Xia D, Wu S. Chinese herbal medicine effects on muscle atrophy induced by simulated microgravity. Aerosp Med Hum Perform. 2018; 89(10):883-888.

ASS234 is a new multitarget molecule with multiple neuroprotective actions that significantly elevate mRNA levels of NRF2 and HSF1 transcriptional factors and of HSP105, HSP90AB1, HSPA1A, HSPA1B, HSPA5, HSPA8, HSPA9, HSP60, DNAJA1, DNAJB1, DNAJB6, DNAJC3, DNAJC5, DNAJC6, HSPB1, HSPB2, HSPB5, HSPB6, HSPB8, and HSP10 heat shock proteins (HSPs) family members in SH-SY5Y cells. This NRF2 and HSF1 overexpression may explain the upregulation of both the antioxidant enzymes previously described and the members of the HSPs family observed. These findings suggest that ASS234 is a potent HSPs inductor, which might be beneficial for preventing protein misfolding aggregation and cell death in Alzheimer's disease and other neurodegenerative diseases.

Small heat shock proteins (sHsps) are a group of proteins with molecular mass between 12 and 43 kDa. Currently, 11 members of this family have been classified, namely HspB1 to HspB11. HspB1, HspB2, HspB5, HspB6, HspB7, and HspB8, which are expressed in brain have been observed to be related to the pathology of neurodegenerative diseases, including Parkinson's, Alzheimer's, Alexander's disease, multiple sclerosis, and human immunodeficiency virus-associated dementia. Specifically, sHsps interact with misfolding and damaging protein aggregates, like Glial fibrillary acidic protein in AxD, β-amyloid peptides aggregates in Alzheimer's disease, Superoxide dismutase 1 in Amyotrophic lateral sclerosis and cytosine-adenine-guanine/polyglutamine (CAG/PolyQ) in Huntington's disease, Spinocerebellar ataxia type 3, Spinal-bulbar muscular atrophy, to reduce the toxicity or increase the clearance of these protein aggregates. The degree of HspB4 expression in brain is still debated. For neuroprotective mechanisms, sHsps attenuate mitochondrial dysfunctions, reduce accumulation of misfolded proteins, block oxidative/nitrosative stress, and minimize neuronal apoptosis and neuroinflammation, which are molecular mechanisms commonly accepted to mirror the progression and development of neurodegenerative diseases. The increasing incidence of the neurodegenerative diseases enhanced search for effective approaches to rescue neural tissue from degeneration with minimal side effects. sHsps have been found to exert neuroprotective functions. HspB5 has been emphasized to reduce the paralysis in a mouse model of experimental autoimmune encephalomyelitis, providing a therapeutic basis for the disease. In this review, we discuss the current understanding of the properties and the mechanisms of protection orchestrated by sHsps in the nervous system, highlighting the promising therapeutic role of sHsps in neurodegenerative diseases., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Enterotoxigenic Escherichia coli (ETEC) is a leading cause of diarrhea among children and travelers in developing countries, and heat-labile enterotoxin (LT) is one of the most important virulence factors. The pathogenesis of and virulence factors associated with ETEC have been well-characterized; however, the extent to which ETEC damages host cells remains unclear. In this study, we found that LT could induce decreases in intestinal epithelial cell viability and induce apoptosis in a dose- and time- dependent manner in both HCT-8 and Caco-2 cells. We analyzed the expression profiles of apoptosis-related proteins via protein array technology and found that Bax, p-p53(S46), cleaved caspase-3, and TNFRI/TNFRSF1A expression levels were significantly up-regulated in wild-type ETEC- but not in ΔLT ETEC-infected HCT-8 cells. Bax is essential for endoplasmic reticulum (ER) stress-triggered apoptosis, and our RNAi experiments showed that the PERK-eIF2-CHOP pathway and reactive oxygen species (ROS) are also main participants in this process. LT-induced ROS generation was decreased in CHOP-knockdown HCT-8 cells compared to that in control cells. Moreover, pretreatment with the ROS inhibitor NAC down-regulated GRP78, CHOP, Bim, and cleaved caspase-3 expression, resulting in a reduction in the apoptosis rate from 36.2 to 20.3% in LT-treated HCT-8 cells. Furthermore, ROS inhibition also attenuated LT-induced apoptosis in the small intestinal mucosa in the ETEC-inoculation mouse model.

Dengue virus (DENV) infection has become a public health issue of worldwide concern and is a serious health problem in Taiwan, yet there are no approved effective antiviral drugs to treat DENV. The replication of DENV requires both viral and cellular factors. Targeting host factors may provide a potential antiviral strategy. It has been known that up-regulation of PI3K/AKT signaling and GRP78 by DENV infection supports its replication. AR-12, a celecoxib derivative with no inhibiting activity on cyclooxygenase, shows potent inhibitory activities on both PI3K/AKT signaling and GRP78 expression levels, and recently has been found to block the replication of several hemorrhagic fever viruses. However the efficacy of AR-12 in treating DENV infection is still unclear. Here, we provide evidence to show that AR-12 is able to suppress DENV replication before or after virus infection in cell culture and mice. The antiviral activities of AR-12 are positive against infection of the four different DENV serotypes. AR-12 significantly down-regulates the PI3K/AKT activity and GRP78 expression in DENV infected cells whereas AKT and GRP78 rescue are able to attenuate anti-DENV effect of AR-12. Using a DENV-infected suckling mice model, we further demonstrate that treatment of AR-12 before or after DENV infection reduces virus replication and mice mortality. In conclusion, we uncover the potential efficacy of AR-12 as a novel drug for treating dengue., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Ionic liquids (ILs) are widely used as reaction media in various commercial applications. Many reports have indicated that most ILs are poorly decomposed by microorganisms and are toxic to aquatic organisms. In this study, differential gene expression profiling was conducted using a suppression subtraction hybridization cDNA library from hepatic tissue of the loach (Paramisgurnus dabryanus) after exposure to 1-hexadecyl-3-methylimidazolium chloride ([C 16 mim]Cl), a representative IL. Two hundred and fifty-nine differentially expressed candidate genes, whose expression was altered by >2.0-fold by the [C 16 mim]Cl treatment, were identified, including 127 upregulated genes and 132 downregulated genes. A gene ontology analysis of the known genes isolated in this study showed that [C 16 mim]Cl-responsive genes were involved in cell cycle, stimulus response, defense response, DNA damage response, oxidative stress responses, and other biological responses. To identify candidate genes that may be involved in [C 16 mim]Cl-induced toxicity, 259 clones were examined by Southern blot macroarray hybridization, and 20 genes were further characterized using quantitative real-time polymerase chain reaction. Finally, six candidate genes were selected, including three DNA damage response genes, two toxic substance metabolic genes, and one stress protein gene. Our results indicate that these changes in gene expression are associated with [C 16 mim]Cl-induced toxicity, and that these six candidate genes can be promising biomarkers for detecting [C 16 mim]Cl-induced toxicity. Therefore, this study demonstrates the use of a powerful assay to identify genes potentially involved in [C 16 mim]Cl toxicity, and it provides a foundation for the further study of related genes and the molecular mechanism of [C 16 mim]Cl toxicity. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 404-416, 2017., (© 2016 Wiley Periodicals, Inc.)

Glioblastoma (GBM) is the most common and aggressive brain tumour of adults. The metabolic phenotype of GBM cells is highly dependent on glycolysis; therefore, therapeutic strategies aimed at interfering with glycolytic pathways are under consideration. 3-Bromopyruvate (3BP) is a potent antiglycolytic agent, with a variety of targets and possible effects on global cell metabolism. Here we analyzed the changes in protein expression on a GBM cell line (GL15 cells) caused by 3BP treatment using a global proteomic approach. Validation of differential protein expression was performed with immunoblotting and enzyme activity assays in GL15 and U251 cell lines. The results show that treatment of GL15 cells with 3BP leads to extensive changes in the expression of glycolytic enzymes and stress related proteins. Importantly, other metabolisms were also affected, including pentose phosphate pathway, aminoacid synthesis, and glucose derivatives production. 3BP elicited the activation of stress response proteins, as shown by the phosphorylation of HSPB1 at serine 82, caused by the concomitant activation of the p38 pathway. Our results show that inhibition of glycolysis in GL15 cells by 3BP influences different but interconnected pathways. Proteome analysis may help in the molecular characterization of the glioblastoma response induced by pharmacological treatment with antiglycolytic agents., Significance: Alteration of the glycolytic pathway characterizes glioblastoma (GBM), one of the most common brain tumours. Metabolic reprogramming with agents able to inhibit carbohydrate metabolism might be a viable strategy to complement the treatment of these tumours. The antiglycolytic agent 3-bromopyruvate (3BP) is able to strongly inhibit glycolysis but it may affect also other cellular pathways and its precise cellular targets are currently unknown. To understand the protein expression changes induced by 3BP, we performed a global proteomic analysis of a GBM cell line (GL15) treated with 3BP. We found that 3BP affected not only the glycolytic pathway, but also pathways sharing metabolic intermediates with glycolysis, such as the pentose phosphate pathway and aminoacid metabolism. Furthermore, changes in the expression of proteins linked to resistance to cell death and stress response were found. Our work is the first analysis on a global scale of the proteome changes induced by 3BP in a GBM model and may contribute to clarifying the anticancer potential of this drug., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Background: To explore the role of the MAPK signaling pathway in the cardiomyocyte apoptosis of mice with post-infarction heart failure (HF)., Methods: Mice were divided into sham and myocardial infarction (MI) groups. Before surgery, the MI group was divided into SB203580 and PBS subgroups. A post-infarction HF model was established by ligating the left anterior descending coronary artery. Ventricular dilatation and cardiac function were observed by small animal echocardiography. The growth of primary cardiomyocytes was observed under an inverted phase contrast microscope. The mRNA and protein expressions of endoplasmic reticulum stress (ERS) markers, GRP78 and CHOP, were detected by qRT-PCR and immunofluorescence assay, respectively., Results: The MI group had enlarged left ventricle and decreased cardiac function. GRP78 and CHOP protein expressions in myocardial tissues, especially those of SB203580 subgroup, significantly increased (p < 0.05). The expressions of p-JNK and cleaved caspase 12 proteins, especially those of SB203580 subgroup, were significantly up-regulated. Cardiomyocytes of MI group were significantly more prone to apoptosis (p < 0.05), with SB203580 subgroup being more obvious., Conclusion: MI was accompanied by ERS, probably involving the MAPK signaling pathway. SB203580, a specific inhibitor of this pathway, can relieve cardiomyocyte apoptosis and protect the myocardium by suppressing such stress (Tab. 3, Fig. 7, Ref. 20).

Objective: To explore if improving the expression of TP53INP1 could enhance the sensitivity of A549 cells to arsenic., Methods: The eukaryotic express vector containing TP53INP1 gene was transferred into A549 cells by using lentivirus vector. Cell apoptosis and cell viability after arsenic treatment were assessed by flow cytometry and MTT assay, respectively., Results: The protein expression level of TP53INP1 was increased in A549 cells transferred with eukaryotic express vector containing TP53INP1 gene, which led to an increase in apoptosis and a decrease in cell viability. Compared with A549 cells, significant increase in apoptosis was found in A549-TP53INP1 cells when treated with As&#95;2O&#95;3( 5- 40 μmol/L). In addiation, the IC50 of As&#95;2O&#95;3 in A549-TP53INP1(( 44. 64 ± 6. 84) μmol/L) cells was significantly lower than that of the A549 group(( 54. 25 ± 6. 13) μmol/L)( P < 0. 05)., Conclusion: Enhancement of TP53INP1 can significantly improve apoptosis response and enhance sensitivity of A549 cells to arsenic. It is suggested that TP53INP1 could be used as a new target in arsenic-based cancer treatment.

We previously reported that endoplasmic reticulum (ER) stress is induced in the subfornical organ (SFO) and the hypothalamic paraventricular nucleus (PVN) of heart failure (HF) rats and is reduced by inhibition of mitogen-activated protein kinase (MAPK) signaling. The present study further examined the relationship between brain MAPK signaling, ER stress, and sympathetic excitation in HF. Sham-operated (Sham) and HF rats received a 4-wk intracerebroventricular (ICV) infusion of vehicle (Veh) or the ER stress inhibitor tauroursodeoxycholic acid (TUDCA, 10 μg/day). Lower mRNA levels of the ER stress biomarkers GRP78, ATF6, ATF4, and XBP-1s in the SFO and PVN of TUDCA-treated HF rats validated the efficacy of the TUDCA dose. The elevated levels of phosphorylated p44/42 and p38 MAPK in SFO and PVN of Veh-treated HF rats, compared with Sham rats, were significantly reduced in TUDCA-treated HF rats as shown by Western blot and immunofluorescent staining. Plasma norepinephrine levels were higher in Veh-treated HF rats, compared with Veh-treated Sham rats, and were significantly lower in the TUDCA-treated HF rats. TUDCA-treated HF rats also had lower mRNA levels for angiotensin converting enzyme, angiotensin II type 1 receptor, tumor necrosis factor-α, interleukin-1β, cyclooxygenase-2, and NF-κB p65, and a higher mRNA level of IκB-α, in the SFO and PVN than Veh-treated HF rats. These data suggest that ER stress contributes to the augmented sympathetic activity in HF by inducing MAPK signaling, thereby promoting inflammation and renin-angiotensin system activity in key cardiovascular regulatory regions of the brain.

Type 2 diabetes (T2D) is among the most common and costly disorders worldwide. The goal of current medical management for T2D is to transiently ameliorate hyperglycemia through daily dosing of one or more antidiabetic drugs. Hypoglycemia and weight gain are common side effects of therapy, and sustained disease remission is not obtainable with nonsurgical approaches. On the basis of the potent glucose-lowering response elicited by activation of brain fibroblast growth factor (FGF) receptors, we explored the antidiabetic efficacy of centrally administered FGF1, which, unlike other FGF peptides, activates all FGF receptor subtypes. We report that a single intracerebroventricular injection of FGF1 at a dose one-tenth of that needed for antidiabetic efficacy following peripheral injection induces sustained diabetes remission in both mouse and rat models of T2D. This antidiabetic effect is not secondary to weight loss, does not increase the risk of hypoglycemia, and involves a novel and incompletely understood mechanism for increasing glucose clearance from the bloodstream. We conclude that the brain has an inherent potential to induce diabetes remission and that brain FGF receptors are potential pharmacological targets for achieving this goal.

Protein aggregates are hallmarks of nearly all age-related neurodegenerative diseases, including Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, and several polyglutamine diseases such as Huntington's disease and different forms of spinocerebellar ataxias (SCA; SCA1-3, SCA6, and SCA7). The collapse of cellular protein homoeostasis can be both a cause and a consequence of this protein aggregation. Boosting components of the cellular protein quality control system has been widely investigated as a strategy to counteract protein aggregates or their toxic consequences. Heat shock proteins (HSPs) play a central part in regulating protein quality control and contribute to protein aggregation and disaggregation. Therefore, HSPs are viable targets for the development of drugs aimed at reducing pathogenic protein aggregates that are thought to contribute to the development of so many neurodegenerative disorders., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

We performed proteomic studies using the GRP78 chaperone-inhibitor drug AR-12 (OSU-03012) as bait. Multiple additional chaperone and chaperone-associated proteins were shown to interact with AR-12, including: GRP75, HSP75, BAG2; HSP27; ULK-1; and thioredoxin. AR-12 down-regulated in situ immuno-fluorescence detection of ATP binding chaperones using antibodies directed against the NH2-termini of the proteins but only weakly reduced detection using antibodies directed against the central and COOH portions of the proteins. Traditional SDS-PAGE and western blotting assessment methods did not exhibit any alterations in chaperone detection. AR-12 altered the sub-cellular distribution of chaperone proteins, abolishing their punctate speckled patterning concomitant with changes in protein co-localization. AR-12 inhibited chaperone ATPase activity, which was enhanced by sildenafil; inhibited chaperone - chaperone and chaperone - client interactions; and docked in silico with the ATPase domains of HSP90 and of HSP70. AR-12 combined with sildenafil in a GRP78 plus HSP27 -dependent fashion to profoundly activate an eIF2α/ATF4/CHOP/Beclin1 pathway in parallel with inactivating mTOR and increasing ATG13 phosphorylation, collectively resulting in formation of punctate toxic autophagosomes. Over-expression of [GRP78 and HSP27] prevented: AR-12 -induced activation of ER stress signaling and maintained mTOR activity; AR-12 -mediated down-regulation of thioredoxin, MCL-1 and c-FLIP-s; and preserved tumor cell viability. Thus the inhibition of chaperone protein functions by AR-12 and by multi-kinase inhibitors very likely explains why these agents have anti-tumor effects in multiple genetically diverse tumor cell types.

Aim: Heat shock proteins (HSPs) are synthesized in virtually all organisms in response to increases in temperature. They are associated with a relaxant effect on the human myometrium and are present in decreased concentration in the myometrium at the time of labor. Paeoniflorin is derived from Paeonia lactiflora and has been shown to induce the synthesis of HSPs in cultured mammalian cells. The purpose of the study was to evaluate the effect of paeoniflorin on human uterine contractility., Material and Methods: Samples of human myometrium were taken at lower segment cesarean section. Dissected muscle strips were suspended under isometric conditions and exposed to cumulative additions of paeoniflorin in concentrations ranging from 1 nmol/L to 10 mol/L. Control experiments were simultaneously performed., Results: Paeoniflorin was found to exert an inhibitory effect on spontaneous and agonist-induced contractions compared to control strips. The mean maximal inhibition values were: 42.21% ± 9.26 for spontaneous contractions (n = 6; P < 0.0001) and 47.84% ± 9.05 for oxytocin-induced contractions (n = 6; P < 0.0001)., Conclusion: The HSP inducing compound, paeoniflorin, had a relaxant effect on human uterine contractility in vitro. These results reinforce the fact that HSPs may play a physiological role in the onset of labor and may also provide future targets for novel tocolytic treatments., (© 2015 Japan Society of Obstetrics and Gynecology.)

Purpose: To investigate the anticancer activity of ellagic acid (EA) in U251 human glioblastoma cells and its possible molecular mechanism., Methods: The cells were treated with EA at various concentrations for different time periods. Cell viability and cell proliferation were detected by cell counting kit-8(CCK-8) assay and live/dead assay respectively. Cell apoptosis were measured with Annexin V-FITC/PI double staining method by flow cytometry and Mitochondrial membrane potential assay separately. Cell cycle was measured with PI staining method by flow cytometry. The expressions of Bcl-2, Survivin, XIAP, Caspase-3, Bax, JNK, p-JNK, ERK1/2, p-ERK1/2, p38, p-p38, DR4, DR5, CHOP and GRP78-related proteins were detected by western blot after EA treatment., Results: Cell viability and proliferation of glioblastoma cells treated with EA were significantly lower than the control group. EA caused robust apoptosis of the glioblastoma cells compared to the control group. EA significantly decreased the proportion at G0/G1 phases of cell cycling accompanied by increased populations at S phase in U251 cell lines. And the expressions of anti-apoptotic proteins were dramatically down-regulated., Conclusion: Ellagic acid potentially up-regulated DR4, DR5 and MAP kinases (JNK, ERK1/2 and p38). EA also caused significant increase in the expressions of CHOP and GRP78. Our findings suggest that EA would be beneficial for the treatment of glioblastoma.

We investigated the effects of exogenous glucagon-like peptide-2 (GLP-2) on mucosal atrophy and intestinal antioxidant capacity in a mouse model of total parenteral nutrition (TPN). Male mice (6-8 weeks old) were divided into three groups (n = 8 for each group): a control group fed a standard laboratory chow diet, and experimental TPN (received standard TPN solution) and TPN + GLP-2 groups (received TPN supplemented with 60 µg/day of GLP-2 for 5 days). Mice in the TPN group had lower body weight and reduced intestinal length, villus height, and crypt depth compared to the control group (all p < 0.05). GLP-2 supplementation increased all parameters compared to TPN only (all p < 0.05). Intestinal total superoxide dismutase activity and reduced-glutathione level in the TPN + GLP-2 group were also higher relative to the TPN group (all p < 0.05). GLP-2 administration significantly upregulated proliferating cell nuclear antigen expression and increased glucose-regulated protein (GRP78) abundance. Compared with the control and TPN + GLP-2 groups, intestinal cleaved caspase-3 was increased in the TPN group (all p < 0.05). This study shows GLP-2 reduces TPN-associated intestinal atrophy and improves tissue antioxidant capacity. This effect may be dependent on enhanced epithelial cell proliferation, reduced apoptosis, and upregulated GRP78 expression.

Five new chalcones, 4,2',4'-trihydroxy-3'-[(2E,5E)-7-methoxy-3,7-dimethyl-2,5-octadienyl]chalcone (1), (±)-4,2',4'-trihydroxy-3'-[(2E)-6-hydroxy-7-methoxy-3,7-dimethyl-2-octenyl]chalcone (2), 4,2',4'-trihydroxy-3'-[(2E)-3-methyl-5-(1,3-dioxolan-2-yl)-2-pentenyl]chalcone (3), 2',3'-furano-4-hydroxy-4'-methoxychalcone (4), and (±)-4-hydroxy-2',3'-(2,3-dihydro-2-methoxyfurano)-4'-methoxychalcone (5), were isolated from the aerial parts of Angelica keiskei Koidzumi together with eight known chalcones, 6-13, which were identified as (±)-4,2',4'-trihydroxy-3'-[(6E)-2-hydroxy-7-methyl-3-methylene-6-octenyl]chalcone (6), xanthoangelol (7), xanthoangelol F (8), xanthoangelol G (9), 4-hydroxyderricin (10), xanthoangelol D (11), xanthoangelol E (12), and xanthoangelol H (13), respectively. Chalcones 1-13 were evaluated for their promoter activity on heat shock protein 25 (hsp25, murine form of human hsp27). Compounds 1 and 6 activated the hsp25 promoter by 21.9- and 29.2-fold of untreated control at 10 μM, respectively. Further protein expression patterns of heat shock factor 1 (HSF1), HSP70, and HSP27 by 1 and 6 were examined. Compound 6 increased the expression of HSF1, HSP70, and HSP27 by 4.3-, 1.5-, and 4.6-fold of untreated control, respectively, without any significant cellular cytotoxicities, whereas 1 did not induce any expression of these proteins. As a result, 6 seems to be a prospective HSP inducer.

Acute brain ischemia can induce the activation of c-Jun N-terminal kinases (JNKs). Hypertension is a critical etiology for brain ischemia. We identified the effects of hypertension on the activation of JNK as well as its impact on SP600125, a JNK inhibitor, during endoplasmic reticulum stress (ERS) in the hippocampus using a rat model. Transient whole-brain ischemia was induced by 4-vessel occlusion (bilateral vertebral and bilateral common carotid arteries) in normal and spontaneous hypertensive rats. SP600125 (0.05 mg/kg, iv) was administered 30 min before ischemia. Morphological changes in hippocampal nerve cells were observed by cresyl violet staining. Phosphorylation of JNK, and expression levels of CHOP and GPR78, markers for ERS, were detected by western blot at 1, 6, 24, and 48 h, and neurological outcomes were measured using an eight-arm radial maze 48 h after ischemia. Hypertension apparently aggravated impairment of memory function, decreased the density of surviving neurons, increased phosphorylation of JNK, and enhanced CHOP expression, but reduced GPR78 levels in hippocampal tissues following brain ischemia. SP600125 alleviated neurological dysfunction, improved neuron survival, decreased phosphorylation of JNK and levels of CHOP, but increased expression of GPR78 in rats with hypertension during cerebral ischemia by inhibition of ERS.

The brainstem represents a major tissue area affected by sarin organophosphate poisoning due to its function in respiratory and cardiovascular control. While the acute toxic effects of sarin on brainstem-related responses are relatively unknown, other brain areas e.g., cortex or cerebellum, have been studied more extensively. The study objective was to analyze the guinea pig brainstem toxicology response following sarin (2×LD50) exposure by proteome pathway analysis to gain insight into the complex regulatory mechanisms that lead to impairment of respiratory and cardiovascular control. Guinea pig exposure to sarin resulted in the typical acute behavior/physiology outcomes with death between 15 and 25min. In addition, brain and blood acetylcholinesterase activity was significantly reduced in the presence of sarin to 95%, and 89%, respectively, of control values. Isobaric-tagged (iTRAQ) liquid chromatography tandem mass spectrometry (LC-MS/MS) identified 198 total proteins of which 23% were upregulated, and 18% were downregulated following sarin exposure. Direct gene ontology (GO) analysis revealed a sarin-specific broad-spectrum proteomic profile including glutamate-mediated excitotoxicity, calcium overload, energy depletion responses, and compensatory carbohydrate metabolism, increases in ROS defense, DNA damage and chromatin remodeling, HSP response, targeted protein degradation (ubiquitination) and cell death response. With regards to the sarin-dependent effect on respiration, our study supports the potential interference of sarin with CO2/H(+) sensitive chemoreceptor neurons of the brainstem retrotrapezoid nucleus (RTN) that send excitatory glutamergic projections to the respiratory centers. In conclusion, this study gives insight into the brainstem broad-spectrum proteome following acute sarin exposure and the gained information will assist in the development of novel countermeasures., (Published by Elsevier B.V.)

Cholera toxin (CT) intoxicates cells by trafficking from the cell surface to the endoplasmic reticulum (ER), where the catalytic CTA1 subunit hijacks components of the ER-associated degradation (ERAD) machinery to retrotranslocate to the cytosol and induce toxicity. In the ER, CT targets to the ERAD machinery composed of the E3 ubiquitin ligase Hrd1-Sel1L complex, in part via the activity of the Sel1L-binding partner ERdj5. This J protein stimulates BiP's ATPase activity, allowing BiP to capture the toxin. Presumably, toxin release from BiP must occur before retrotranslocation. Here, using loss-and gain-of-function approaches coupled with binding studies, we demonstrate that the ER-resident nucleotide exchange factors (NEFs) Grp170 and Sil1 induce CT release from BiP in order to promote toxin retrotranslocation. In addition, we find that after NEF-dependent release from BiP, the toxin is transferred to protein disulfide isomerase; this ER redox chaperone is known to unfold CTA1, which allows the toxin to cross the Hrd1-Sel1L complex. Our data thus identify two NEFs that trigger toxin release from BiP to enable successful retrotranslocation and clarify the fate of the toxin after it disengages from BiP., (© 2015 Williams et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).)

Background: The accumulation of advanced glycation end products (AGEs) appears to be the main factor responsible for modulating periodontal inflammation in diabetes. The aim of this study is to examine the effects of AGEs on inflammation in human periodontal ligament cells and to investigate the mechanism with a specific emphasis on the role of endoplasmic reticulum (ER) stress-induced nuclear factor-kappa B (NF-κB) pathway., Methods: Cytotoxicity was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The protein expressions of ER markers and NF-κB were examined by Western blot analysis. The translocation of NF-κB was observed by immunofluorescence assay. Proinflammatory chemokine production was determined by enzyme-linked immunosorbent assay., Results: Treatment with AGEs reduced cell viability in a concentration- and time-dependent manner. AGEs induced ER stress, as evidenced by survival molecules, such as glucose-regulated protein 78 (GRP78), double-stranded RNA-activated protein kinase-like ER kinase (PERK), and activating transcription factor 6 (ATF-6), and apoptotic molecules, such as CCAAT/enhancer binding protein homologous protein (CHOP) and caspase 12. AGEs upregulated the nucleoprotein expression of NF-κB, enhanced translocation of NF-κB from the cytoplasm to the nucleus, and increased the production of proinflammatory chemokines interleukin-6 and interleukin-8., Conclusion: AGEs mediate inflammation of human periodontal ligament cells via the ER stress-induced NF-κB pathway.

Dichloroacetic acid (DCA), a water disinfection by-product, has attained emphasis due to its prospect for clinical use against different diseases including cancer along with negative impact on organisms. However, these reports are based on the toxicological as well clinical data using comparatively higher concentrations of DCA without much of environmental relevance. Here, we evaluate cellular as well as organismal effects of DCA at environmentally and mild clinically relevant concentrations (0.02-20.0 μg/ml) using an established model organism, Drosophila melanogaster. Flies were fed on food mixed with test concentrations of DCA for 12-48 h to examine the induction of reactive oxygen species (ROS) generation, oxidative stress (OS), heat shock genes (hsps) and cell death along with organismal responses. We also examined locomotor performance, ROS generation, glutathione (GSH) depletion, expression of GSH-synthesizing genes (gclc and gclm), and hsps at different days (0, 10, 20, 30, 40, 50) of the age in flies after prolonged DCA exposure. We observed mild OS and induction of antioxidant defense system in 20.0 μg/ml DCA-exposed organism after 24 h. After prolonged exposure to DCA, exposed organism exhibited improved survival, elevated expression of hsp27, gclc, and gclm concomitant with lower ROS generation and GSH depletion and improved locomotor performance. Conversely, hsp27 knockdown flies exhibited reversal of the above end points. The study provides evidence for the attenuation of cellular and functional decline in aged Drosophila after prolonged DCA exposure and the effect of hsp27 modulation which further incites studies towards the therapeutic application of DCA.

Background: The metabolic syndrome is the most important risk factor for cardiovascular disease. The heat shock proteins (HSPs) are highly conserved families of proteins expressed by a number of cell types following exposure to stressful environmental conditions include several known risk factors for cardiovascular disease. Recent studies have shown the potential of constituents of saffron in the treatment of atherosclerosis. We aimed on investigating the effect of saffron on antibody titers to HSP in patients with metabolic syndrome., Methods: This was a randomized, placebo-controlled clinical trial. One-hundred and five subjects with metabolic syndrome were randomly allocated to one of the three groups: the case group received 100 mg/day saffron, the placebo control group received a capsule of placebo and a non-placebo control group received no capsule, for 12 weeks., Results: Antibodies against heat shock proteins 27, 60, 65 and 70 were determined in all patients before (week 0) and after (week 6 and 12) intervention. At 12 weeks, saffron produced a significantly decrease in AntiHSP27, 70 levels. Saffron can decrease AntiHSP27, 70 levels in patients with metabolic syndrome., Conclusions: The results of this study indicate the efficacy of saffron in the improvement of some markers of autoimmunity HSPs in patients with metabolic syndrome.

Arsenic trioxide (As2O3) has been used in the treatment of acute promyelocytic leukemia (APL) and many malignant solid tumors. Recently, endoplasmic reticulum (ER) stress plays an important role in As2O3-treated laryngeal squamous cell line Hep-2 cells. In the present work, the expression of ER stress-related proteins was investigated in As2O3-treated Hep-2 cells. The results showed that As2O3 increased the expression of GRP78, CHOP, phosphorylated eIF2α and ATF4, all of which are the molecule of ER stress. Therefore, As2O3 induced ER stress in Hep-2 cells., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

Aims: We hypothesized that oral l-glutamine supplementations could attenuate muscle damage and oxidative stress, mediated by glutathione (GSH) in high-intensity aerobic exercise by increasing the 70-kDa heat shock proteins (HSP70) and heat shock factor 1 (HSF1)., Main Methods: Adult male Wistar rats were 8-week trained (60-min/day, 5 days/week) on a treadmill. During the last 21 days, the animals were supplemented with either l-alanyl-l-glutamine dipeptide (1.5 g/kg, DIP) or a solution containing the amino acids l-glutamine (1g/kg) and l-alanine (0.67 g/kg) in their free form (GLN+ALA) or water (controls)., Key Findings: Plasma from both DIP- and GLN+ALA-treated animals showed higher l-glutamine concentrations and reduced ammonium, malondialdehyde, myoglobin and creatine kinase activity. In the soleus and gastrocnemius muscle of both supplemented groups, l-glutamine and GSH contents were increased and GSH disulfide (GSSG) to GSH ratio was attenuated (p<0.001). In the soleus muscle, cytosolic and nuclear HSP70 and HSF1 were increased by DIP supplementation. GLN+ALA group exhibited higher HSP70 (only in the nucleus) and HSF1 (cytosol and nucleus). In the gastrocnemius muscle, both supplementations were able to increase cytosolic HSP70 and cytosolic and nuclear HSF1., Significance: In trained rats, oral supplementation with DIP or GLN+ALA solution increased the expression of muscle HSP70, favored muscle l-glutamine/GSH status and improved redox defenses, which attenuate markers of muscle damage, thus improving the beneficial effects of high-intensity exercise training., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Arimoclomol is a hydroxylamine derivative, a group of compounds which have unique properties as co-inducers of heat shock protein expression, but only under conditions of cellular stress. Arimoclomol has been found to be neuroprotective in a number of neurodegenerative disease models, including Amyotrophic Lateral Sclerosis (ALS), and in mutant Superoxide Dismutase 1 (SOD1) mice that model ALS, Arimoclomol rescues motor neurons, improves neuromuscular function and extends lifespan. The therapeutic potential of Arimoclomol is currently under investigation in a Phase II clinical trial for ALS patients with SOD1 mutations. In this review we summarize the evidence for the neuroprotective effects of enhanced heat shock protein expression by Arimoclomol and other inducers of the Heat Shock Response. ALS is a complex, multifactorial disease affecting a number of cell types and intracellular pathways. Cells and pathways affected by ALS pathology and which may be targeted by a heat shock protein-based therapy are also discussed in this review. For example, protein aggregation is a characteristic pathological feature of neurodegenerative diseases including ALS. Enhanced heat shock protein expression not only affects protein aggregation directly, but can also lead to more effective clearance of protein aggregates via the unfolded protein response, the proteasome-ubiquitin system or by autophagy. However, compounds such as Arimoclomol have effects beyond targeting protein mis-handling and can also affect additional pathological mechanisms such as oxidative stress. Therefore, by targeting multiple pathological mechanisms, compounds such as Arimoclomol may be particularly effective in the development of a disease-modifying therapy for ALS and other neurodegenerative disorders., (© 2013.)