αB-crystallin/HSPB2 is critical for hyperactive mTOR-induced cardiomyopathy.

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.

Synergy of hypoxia relief and heat shock protein inhibition for phototherapy enhancement.

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 MnO2 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.

CXL146, a Novel 4H-Chromene Derivative, Targets GRP78 to Selectively Eliminate Multidrug-Resistant Cancer Cells.

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 4H-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 1α, 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.

Thiostrepton Reactivates Latent HIV-1 through the p-TEFb and NF-κB Pathways Mediated by Heat Shock Response.

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.

Morin inhibits Listeria monocytogenes virulence in vivo and in vitro by targeting listeriolysin O and inflammation.

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.

Aberrant MUC1 accumulation in salivary glands of Sjögren's syndrome patients is reversed by TUDCA in vitro.

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-1ß, 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.

Inhibition of SIRT1/2 upregulates HSPA5 acetylation and induces pro-survival autophagy via ATF4-DDIT4-mTORC1 axis in human lung cancer cells.

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.

In Vitro and In Vivo Renoprotective Effects of Telbivudine in Chronic Hepatitis B Patients Receiving Nucleotide Analogue.

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 m2 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).

Involvement of MicroRNA-133a in the Protective Effect of Hydrogen Sulfide against Ischemia/Reperfusion-Induced Endoplasmic Reticulum Stress and Cardiomyocyte Apoptosis.

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.

Sunscreen-induced expression and identification of photosensitive marker proteins in human keratinocytes under UV radiation.

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.

Heat Shock Proteins in Glioblastoma Biology: Where Do We Stand?

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.

Modulation of Heat Shock Response Proteins by ASS234, Targeted for Neurodegenerative Diseases Therapy.

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.

Gene expression profile changes induced by acute toxicity of [C16 mim]Cl in loach (Paramisgurnus dabryanus).

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 ([C16 mim]Cl), a representative IL. Two hundred and fifty-nine differentially expressed candidate genes, whose expression was altered by >2.0-fold by the [C16 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 [C16 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 [C16 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 [C16 mim]Cl-induced toxicity, and that these six candidate genes can be promising biomarkers for detecting [C16 mim]Cl-induced toxicity. Therefore, this study demonstrates the use of a powerful assay to identify genes potentially involved in [C16 mim]Cl toxicity, and it provides a foundation for the further study of related genes and the molecular mechanism of [C16 mim]Cl toxicity. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 404-416, 2017.

MAPK pathway regulated the cardiomyocyte apoptosis in mice with post-infarction heart failure.

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).

[Improving the expression of TP53INP1 in A549 cells enhances the sensitivity of cells to arsenic treatment].

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_2O_3( 5- 40 µmol/L). In addiation, the IC50 of As_2O_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.

Endoplasmic reticulum stress increases brain MAPK signaling, inflammation and renin-angiotensin system activity and sympathetic nerve activity in heart failure.

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-1ß, 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.

Paeoniflorin, a novel heat-shock protein inducing compound, and human myometrial contractility in vitro.

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.

Chalcones from Angelica keiskei: Evaluation of Their Heat Shock Protein Inducing Activities.

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.

Hypertension-mediated enhancement of JNK activation in association with endoplasmic reticulum stress in rat model hippocampus with cerebral ischemia-reperfusion.

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.

AKT inhibition mitigates GRP78 (glucose-regulated protein) expression and contribution to chemoresistance in endometrial cancers.

Overexpression of the unfolded protein response master regulator GRP78 is associated with poor prognosis and therapeutic resistance in numerous human cancers, yet its role in endometrial cancers (EC) is undefined. To better understand the contribution of GRP78 to EC, we examined its expression levels in EC patient samples and EC cell lines. We demonstrate that GRP78 overexpression occurs more frequently in EC tissues compared with that found in normal endometrium, and that GRP78 expression occurs in most EC cell lines examined. Functional analysis demonstrated that GRP78 is inducible by cisplatin in EC cells, and siRNA knockdown of GRP78 augments chemotherapy-mediated cell death. Examination of AKT and GRP78 expression demonstrated that inhibition of AKT activity by MK2206 blocks GRP78 expression in EC cells. SiRNA studies also revealed that knockdown of GRP78 reduces but does not abrogate AKT activity, demonstrating that GRP78 is required for optimal AKT activity. In the presence of MK2206, siRNA knockdown of GRP78 does not augment AKT mediated survival in response to cisplatin treatment, suggesting that GRP78's antiapoptosis functions are part of the AKT survival pathway. Targeted therapies that reduce GRP78 expression or activity in cancers may serve to increase the effectiveness of current therapies for EC patients.