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Background: The aim of this study was to determine how Stathmin-1 and Heat Shock Protein 27 (HSP27) can be used as adjunctive biomarkers to differentiate high-grade dysplasia from benign/reactive lesions in cervical tissues. In addition, we aimed to see if any of these markers can differentiate endometrial from endocervical adenocarcinomas., Methods: Fifty cases including benign cervical tissue, low-grade squamous intraepithelial lesion (LSIL), high-grade squamous intraepithelial lesion (HSIL), adenocarcinoma in situ of the endocervix, invasive endocervical adenocarcinoma, and endometrial adenocarcinoma were selected. Stathmin-1 and HSP27 immunohistochemistry (IHC) were performed for each case and the results were compared to the previously available p16 IHC stains., Results: p16 stained positively in 100% of HSIL, endocervical adenocarcinoma in situ, and invasive endocervical cases. Stathmin-1 stained positively in 43% of HSIL and 90% of endocervical adenocarcinoma in situ and all invasive endocervical cases. Stathmin-1 and p16 were negative in all benign cervical samples. Stathmin-1, HSP27, and p16 stained 100% of LSIL cases. HSP27 stained indiscriminately, including 100% of benign cervical tissue. 87% of the endometrial adenocarcinomas stained positively for p16, Stathmin-1, and HSP27., Conclusion: p16 remains superior to both Stathmin-1 and HSP27 in differentiating dysplasia from benign, reactive changes of the cervix., (© 2021. The Author(s).)

Vaccinations are widely credited with reducing death rates from COVID-19, but the underlying host-viral mechanisms/interactions for morbidity and mortality of SARS-CoV-2 infection remain poorly understood. Acute respiratory distress syndrome (ARDS) describes the severe lung injury, which is pathologically associated with alveolar damage, inflammation, non-cardiogenic edema, and hyaline membrane formation. Because proteostatic pathways play central roles in cellular protection, immune modulation, protein degradation, and tissue repair, we examined the pathological features for the unfolded protein response (UPR) using the surrogate biomarker glucose-regulated protein 78 (GRP78) and co-receptor for SARS-CoV-2. At autopsy, immunostaining of COVID-19 lungs showed highly elevated expression of GRP78 in both pneumocytes and macrophages compared with that of non-COVID control lungs. GRP78 expression was detected in both SARS-CoV-2-infected and un-infected pneumocytes as determined by multiplexed immunostaining for nucleocapsid protein. In macrophages, immunohistochemical staining for GRP78 from deceased COVID-19 patients was increased but overlapped with GRP78 expression taken from surgical resections of non-COVID-19 controls. In contrast, the robust in situ GRP78 immunostaining of pneumocytes from COVID-19 autopsies exhibited no overlap and was independent of age, race/ethnicity, and gender compared with that from non-COVID-19 controls. Our findings bring new insights for stress-response pathways involving the proteostatic network implicated for host resilience and suggest that targeting of GRP78 expression with existing therapeutics might afford an alternative therapeutic strategy to modulate host-viral interactions during SARS-CoV-2 infections., (© 2021. Cell Stress Society International.)

Introduction: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), mainly transmitted by droplets and close contact, has caused a pandemic worldwide as of March 2020. According to the current case reports and cohort studies, the symptoms of pregnant women infected with SARS-CoV-2 were similar to normal adults and may cause a series of adverse consequences of pregnancy (placental abruption, fetal distress, epilepsy during pregnancy, etc.). However, whether SARS-CoV-2 can be transmitted to the fetus through the placental barrier is still a focus of debate., Methods: In this study, in order to find out whether SARS-CoV-2 can infect fetus through the placental barrier, we performed qualitative detection of virus structural protein (spike protein and nucleoprotein) and targeted receptor protein Angiotensin Converting Enzyme 2 (ACE2), Basigin (CD147) and molecular chaperone GRP78 expression on the placental tissue of seven pregnant women diagnosed with COVID-19 through immunohistochemistry. Amniotic fluid, neonatal throat, anal swab and breastmilk samples were collected immediately in the operating room or delivery room for verification after delivery, which were all tested for SARS-CoV-2 by reverse transcriptionpolymerase chain reaction (RT-PCR)., Results/discussion: The result showed that CD147 was expressed on the basal side of the chorionic trophoblast cell membrane and ACE2 was expressed on the maternal side, while GRP78 was strongly expressed in the cell membrane and cytoplasm. The RT-PCR results of Amniotic fluid, neonatal throat, anal swab and breastmilk samples were all negative. On the basis of these findings, we speculated that it may be due to the placental barrier between mother and baby, for example, villous matrix and interstitial blood vessels have low expression of virus-related receptors (ACE2, CD147, GRP78), the probability of vertical transmission of SARS-CoV-2 through the placenta is low., (Copyright © 2021. Published by Elsevier Ltd.)

Atherosclerosis is a multifactorial chronic inflammatory disease of the arterial wall, and an important pathological basis of coronary heart disease. Endothelial cells, vascular smooth muscle cells, and macrophages play important roles in the development of atherosclerosis. Of note, apoptosis and autophagy, two types of programmed cell death, influence the development and progression of atherosclerosis via the modulation of such cells. The small heat shock protein Hsp27 is a multifunctional protein induced by various stress factors and has a protective effect on cells. A large number of studies have demonstrated that Hsp27 plays an important role in regulating apoptosis. Recently, some studies have suggested that Hsp27 also participates in the autophagic process. Moreover, Hsp27 is closely related to the occurrence and development of atherosclerosis. Here, we summarize the molecular mechanisms of apoptosis and autophagy and discuss their effects on endothelial cells, vascular smooth muscle cells, and macrophages in the context of atherosclerotic procession. We further explore the involvement of Hsp27 in apoptosis, autophagy, and atherosclerosis. We speculate that Hsp27 may exert its anti-atherosclerotic role via the regulation of apoptosis and autophagy; this may provide the basis for the development of new approaches for the prevention and treatment of atherosclerosis., (Copyright © 2020. Published by Elsevier Ltd.)

Vaspin is a novel adipokine mainly expressed in visceral adipose tissue and closely related to obesity and insulin-resistance. Currently, data about its ovarian expression are limited to animal models and its role in human reproduction is largely unexplored. Our study's aims were then to characterise vaspin expression in the human ovary and to study in vitro its effects on granulosa cells physiology. Secondly, we assessed vaspin and its receptor GRP78 variations in granulosa cells and follicular fluid of a cohort of 112 infertile women undergoing an in vitro fertilisation procedure and allocated to three groups, each including normal-weight and obese subjects: 34 PCOS patients, 33 women with isolated polycystic ovary morphology (ECHO group) and 45 controls. Vaspin and GRP78 expression in the ovary was assessed by immunohistochemistry, RT-qPCR and Western blot. Granulosa cells and follicular fluid were analysed by RT-qPCR and ELISA, respectively. In vitro, granulosa cells metabolism was studied after stimulation with recombinant human vaspin, with and without a siRNA directed against GRP78. Vaspin was highly expressed in the human ovary and concentration-dependently enhanced granulosa cells steroidogenesis, proliferation and viability through GRP78 (P < 0.0001). Vaspin levels in both granulosa cells and follicular fluid were significantly higher in obese women (P < 0.0001) and in the normal-weight ECHO group (P < 0.001), which also had the highest expression rates of GRP78 (P < 0.05). Although further investigation is needed, vaspin appears as a novel modulator of human granulosa cells physiology and possibly plays a role in PCOS pathogenesis, notably protecting from insulin-resistance induced complications.

Background/aim: We aimed to investigate the synergistic effects of apigenin and curcumin on the cross-talk between apoptosis and autophagic cell death, as well as on paraptosis in HeLa cells., Materials and Methods: Cell viability was measured using the MTT assay. Synergistic effects were measured using the Bliss independence model. qRT-PCR was used to study the expression of genes related to apoptosis, autophagic cell death, and cross-talk. GRP78/BiP immunostaining was used to identify endoplasmic reticulum (ER) stress., Results: Treatment with a combination of apigenin and curcumin increased the expression levels of genes related to cell death in HeLa cells 1.29- to 27.6-fold. The combination of curcumin and apigenin showed a synergistic anti-tumor effect via cross-talk between processes leading to apoptosis and autophagic cell death, as well as ER stress-associated paraptosis. GRP78 expression was down-regulated, and massive cytoplasmic vacuolization was observed in HeLa cells., Conclusion: The combination of curcumin and apigenin is an effective potential therapeutic for cervical cancers., (Copyright © 2021 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Our cluster analysis of the Cancer Genome Atlas for co-expression of HSP27 and CRYAB in breast cancer patients identified three patient groups based on their expression level combination (high HSP27 + low CRYAB; low HSP27 + high CRYAB; similar HSP27 + CRYAB). Our analyses also suggest that there is a statistically significant inverse relationship between HSP27 and CRYAB and known clinicopathological markers in breast cancer. Screening an unbiased 248 breast cancer patient tissue microarray (TMA) for the protein expression of HSP27 and phosphorylated HSP27 (HSP27-82pS) with CRYAB also identified three patient groups based on HSP27 and CRYAB expression levels. TMA24 also had recorded clinical-pathological parameters, such as ER and PR receptor status, patient survival, and TP53 mutation status. High HSP27 protein levels were significant with ER and PR expression. HSP27-82pS associated with the best patient survival (Log Rank test). High CRYAB expression in combination with wild-type TP53 was significant for patient survival, but a different patient outcome was observed when mutant TP53 was combined with high CRYAB expression. Our data suggest that HSP27 and CRYAB have different epichaperome influences in breast cancer, but more importantly evidence the value of a cluster analysis that considers their coexpression. Our approach can deliver convergence for archival datasets as well as those from recent treatment and patient cohorts and can align HSP27 and CRYAB expression to important clinical-pathological features of breast cancer., (© 2022. The Author(s).)

Background: Heat-shock proteins (HSP) is a key chaperone protein which maintains intracellular proteostasis and is expressed on the surface of solid and hematological malignancies. Several studies have reported paradoxical evidence of the association between HSP expression and prognosis of oral cancer. To address the discrepancy, we carried out the meta-analysis to assess the role of HSP such as: HSP70, HSP90, HSP27, HSP60, and HSP105 in susceptibility, progression, and prognosis of oral cancer., Materials and Methods: We retrieved the PubMed, Embase, Web of science, China National Knowledge Infrastructure (CNKI), and Wanfang databases to acquire the eligible studies which were associated with HSP70, HSP90, HSP27, HSP60, and HSP105 protein expression and oral cancer. We applied hazard ratio (HR) and its 95% confidence interval (95% CI) to assess the value of HSP protein expression in overall survival of oral cancer; odds ratio (OR) and its 95% CI were used to evaluate the association of risk and clinical features of oral cancer. Funnel plot, Begg test, and Egger line regression test were utilized to observe publication bias among studies. All statistical analysis was performed with Stata 14.0 software (Stata Corporation, College Station, TX)., Results: A total of 26 studies were included in the present meta-analysis. On based of the results, HSP70 and HSP27 had no significant association with progression of oral cancer. However, the pooled HR and 95% CI revealed a significant well effects of HSP70 and HSP27 expression on survival of oral cancer. Moreover, the susceptibility of oral cancer was significantly associated with HSP70 and HSP60 overexpression., Conclusion: HSP70 and HSP27 protein overexpression might be valuable biomarkers for the prognosis of oral cancer. And HSP70 and HSP60 might have potential predictive effects on the risk of oral cancer., Competing Interests: The authors have no conflicts of interest to disclose., (Copyright © 2021 the Author(s). Published by Wolters Kluwer Health, Inc.)

The Egyptian cotton leaf worm, Spodoptera littoralis (Boisd.), is a major agricultural lepidopterous pest causing extensive damage in a variety of crops including vegetable, cotton, fodder, and fiber crops. Heat shock protein (HSP) family members play important roles in protecting insects against environmental stressors. In this study, we characterized three putative heat shock proteins (SpliHsp70, SpliHsp90, and SpliHSF) from S. littoralis and analyzed their expression levels in response to heat, cold, ultraviolet irradiation, Bacillus thuringiensis, and Spodoptera littoralis nucleopolyhedrovirus treatments. Significant upregulation of SpliHsp70 was observed in female pupae, while the highest expression levels of SpliHsp90 and SpliHSF were found in female adults. Heat shock triggered increases in SpliHsp levels compared to cold treatment. SpliHsp90 exhibited the highest expression levels during the first 30 min of UV treatment. Both bacterial and viral pathogenic agents effected the regulation of Hsps in S. littoralis. These findings suggest that SpliHsp genes might play significant roles in the response to biotic and abiotic stress, as well as in the regulation of developmental stages.

Background/aims: The imbalance of T helper 17 (Th17) and regulatory T (Treg) cells exists in the occurrence and development of various diseases. Endoplasmic reticulum stress (ERS) is an important self-protective cellular response to harmful stimuli, such as uremic environment. The objective of this study was to investigate the Th17/Treg cell balance and ERS in a uremic environment and analyze the relationship between them., Methods: (1) The rat spleen lymphocytes were extracted and treated with thapsigargin (inducer of ERS) and sodium citrate. The proportion of Th17 and Treg cells were then detected. (2) The uremic serum-cultured lymphocytes were used and divided into three groups: non-uremic serum group, uremic serum group, and uremic serum + sodium citrate group. Afterward, the proportion of Th17/Treg cells and the expression of ERS-related proteins (GRP78 and CHOP) were detected., Results: Thapsigargin had no significant effect on the proportion of Th17 cells within a limited concentration range, but it could reduce the proportion of Treg cells, sodium citrate had a negative influence on the deviation of Th17/Treg cells treated with thapsigargin. Uremic serum treatment reduced the proportion of Treg cells, resulting in an increase of the Th17/Treg ratio. However, sodium citrate had no influence on the deviation of Th17/Treg cells treated by uremic serum. Sodium citrate reduced the elevation of ERS-related proteins induced by uremic serum., Conclusions: Uremic serum can lead to the imbalance of Th17/Treg cells as well as ERS, suggesting that ERS is one of the mechanisms of the imbalance of Th17/Treg cells induced by uremic serum. Sodium citrate can inhibit ERS induced by uremic serum., (Copyright © 2020 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Endoplasmic reticulum stress (ER stress) just like a double-edged sword depending on different conditions in the development of multiple hepatic diseases. But the molecular mechanisms of functional conversion during ER stress have not been fully elucidated. In this study, we aim to illustrate the role of PPARα and the subtle mechanism in the functional conversion of ER stress. Tunicamycin (TM) and thapsigargin (TG), as ER stress inducers, were used to induce ER stress in AML12 cells. During the ER stress, qRT-PCR and immunoblotting was used to measure the expression levels of GRP78 and CHOP which show a gradually increasing trend, while PPARα and autophagy was significantly activated in the early stage but was inhibited in the late stage. Moreover, PPARα inhibition by siRNA promoted cell injury in the mild-ER stress and PPARα activation by WY-14643 reduced cell apoptosis in the serious ER stress. In the mild-ER stress with PPARα knocked down, activation of autophagy by rapamycin significantly improved cell survival, in the serious ER stress with PPARα activation, inhibition of autophagy by 3-MA aggravate cell injury. In addition, in the mild-ER stress with PPARα knocked down, CHOP knocked down by siRNA reduced cell apoptosis, in the serious ER stress activated PPARα, CHOP over-expression mediated by lentiviral vector contributed to serious cell injury. Furthermore, C57BL/6 mice was used to induce ER stress with TM intraperitoneal injection, PPARα and autophagy was upregulated in the mild-ER stress while downregulated in the serious ER stress, measured by qRT-PCR and immunoblotting, further confirmed the finding in vitro. Our results firstly demonstrated that PPARα is a key molecule in the functional conversion of ER stress: protective effects in the mild ER stress was mediated by PPARα-autophagy pathway and destructive effects in the serious ER stress was mediated by PPARα-CHOP pathway.

In aquaculture, fish species may experience stressful episodes caused by poor farming conditions. The exponential increase of global aquaculture has raised the number of research studies aimed at demonstrating the sensitivity of aquatic animals in confined environments. The development of a real-time PCR and immunohistochemistry methods were investigated to evaluate the presence, localization, and quantity of biomarkers of oxidative stress in European sea bass (Dicentrarchus labrax). In particular, stress tests such as manipulation and temperature changes were conducted through molecular methods to identify the expression level of heat shock protein 70 (HSP70) in stressed animals compared with a control group. The immunohistochemical technique was also applied to locate and study the trends-levels of nitrotyrosine (NT), heat shock protein 70 (HSP70), malondialdehyde (MDA), and 4-hydroxy-2-nonenal (HNE) in different tissues from stressed animals and control group. The presence of the rodlet cell (RCs) was evaluated by histology in both a control and stressed group. Our results show that the real-time PCR method developed is specific for the evaluated target gene and that manipulation and temperature increase are strong stressors for animals. Relative quantification data revealed a gene expression increase of HSP70 in the stressed group of animals compared to the control group. The antibodies used for the immunohistochemical staining were efficient, and it was possible to appreciate the increase of immunoprecipitates in European sea bass either manipulated or stressed by temperature increase. The present study can be a starting point to allow the quantification of HSP70 and the identification of other stress biomarkers in D. labrax.

To optimize treatment decisions in advanced bladder cancer (BC), we aimed to assess the therapy predictive value of STIP1 with regard to cisplatin therapy. Cisplatin-based chemotherapy represents the standard first-line systemic treatment of advanced bladder cancer. Since novel immunooncologic agents are already available for cisplatin-resistant or ineligible patients, biological markers are needed for the prediction of cisplatin resistance. STIP1 expression was analyzed in paraffin-embedded bladder cancer tissue samples of 98 patients who underwent adjuvant or salvage cisplatin-based chemotherapy by using immunohistochemistry. Furthermore, pre-chemotherapy serum STIP1 concentrations were determined in 48 BC patients by ELISA. Results were correlated with the clinicopathological and follow-up data. Stronger STIP1 nuclear staining was associated with worse OS in both the whole patient group (p = 0.034) and the subgroup of patients who received at least 2 cycles of chemotherapy (p = 0.043). These correlations remained significant also in the multivariable analyses (p = 0.035 and p = 0.040). Stronger STIP1 cytoplasmatic immunostaining correlated with shorter PFS both in the whole cohort (p = 0.045) and in the subgroup of patients who received at least 2 cycles of chemotherapy (p = 0.026). Elevated STIP1 serum levels were associated with older patient's age, but we found no correlation between STIP1 serum levels and patients' outcome. Our results suggest that tissue STIP1 analysis might be used for the prediction of cisplatin-resistance in BC. In contrast, pretreatment STIP1 serum levels showed no predictive value for chemotherapy response and survival.

The objective of this study was to determine the frequency of expression of GRP78 and BIK/NBK proteins, as well as to evaluate their correlation with prognostic factors for clinical use in patients with locally advanced breast cancer (LABC) or preoperative chemotherapy (PC). An observational, analytical and retrospective study that evaluated the expression of BIK/NBK and GRP78 by means of immunohistochemistry in paraffin-embedded tumour tissue samples obtained before the start of PC was executed. GRP78 was positive in 93.3% and BIK/NBK was positive in 40% of the samples analysed. Correlation of GRP78 expression with Ki67 expression (Rho = 0.732, p  = .039), and of BIK/NBK expression with Ki67 expression (Rho = 0.777, p  = .023) and number of residual lymph nodes (Rho = 0.619, p  = .014) was observed. Given the correlation observed, association studies of the expression of GRP78 and BIK/NBK with clinical and pathological response, recurrence and survival should be carried out in a greater number of patients with LABC or PC.Impact Statement What is already known on this subject? There are reports of the overexpression of GRP78 in various types of cancer cell lines and tumour biopsies, including breast cancer, with participation in resistance to adjuvant chemotherapeutic agents (topoisomerase inhibitors including Adriamycin). On the other hand, BIK/NBK has been observed expressed associated with poor outcomes in breast cancer. There are no studies of the expression of both biomarkers in patients with preoperative chemotherapy, including locally advanced breast cancer (LABC) or their correlation with prognosis markers of clinical use. What the results of this study add? We observed a correlation of higher expression percentage of GRP78 and BIK/NBK with higher expression of Ki67, a marker of poor prognosis in breast cancer, in addition to a significant correlation of higher percentage of BIK/NBK expression with a greater number of residual nodes in patients with LABC or PC. What the implications are of these findings for clinical practice and/or further research? There is a potential prognostic value of GRP78 and BIK/NBK in the pathological response that merits further study addressing their interaction with other proteins, their cellular location in relation to the clinical stages of breast cancer, and their association with prognosis (clinical and pathological response, recurrence, disease free period and survival) in a greater number of patients with LABC or PC.

Invertebrates are becoming more popular models for research on the immune system. The innate immunity possessed by insects shows both structural and functional similarity to the resistance displayed by mammals, and many processes occurring in insect hemocytes are similar to those that occur in mammals. The humoral immune response in insects acts by melanization, clotting and the production of reactive oxygen species and antimicrobial peptides, while the cellular immunity system is based on nodulation, encapsulation and phagocytosis. An increasingly popular insect model in biological research is Galleria mellonella, whose larvae are sensitive to infection by the entomopathogenic fungus Conidiobolus coronatus, which can also be dangerous to humans. One group of factors that modulate the response of the immune system during infection in mammals are heat shock proteins (HSPs). The aim of this study was to investigate whether infection by C. coronatus in G. mellonella hemolymph is accompanied by an increase of HSP90, HSP70, HSP60 and HSP27. Larvae (five-day-old last instar) were exposed for 24 hours to fully-grown and sporulating fungus. Hemolymph was collected either immediately after termination of exposure (F24) or 24 hours later (F48). The concentration of the HSPs in hemolymph was determined using ELISA. Immunolocalization in hemocytes was performed using fluorescence microscopy and flow cytometry. HSP90, HSP70, HSP60 and HSP27 were found to be present in the G. mellonella hemocytes. HSP60 and HSP90 predominated in healthy insects, with HSP70 and HSP27 being found in trace amounts; HSP60 and HSP27 were elevated in F24 and F48, and HSP90 was elevated in F48. The fungal infection had no effect on HSP70 levels. These findings shed light on the mechanisms underlying the interaction between the innate insect immune response and entomopathogen infection. The results of this innovative study may have a considerable impact on research concerning innate immunology and insect physiology., Competing Interests: The authors have read the journal’s policy and have the following conflicts: MIB is the President of Biombio. This does not alter our adherence to all the PLOS ONE policies on sharing data and materials. There are no patents, products in development or market products to declare.

Introduction: Parallel with the progression of Chronic Lymphocytic Leukemia (CLL), the levels of 78KDa Glucose-Regulated Protein (GRP78) and Hypoxia-Inducible Factor 1 alpha (HIF-1α) are increased as they may activate the induction of anti-apoptotic proteins such as BCL2 Associated Athanogene 3 (BAG3). Previous studies have indicated that there is a positive correlation among GRP78, HIF-1α and BAG3., Objective: This study aimed to evaluate the effect of metabolic factors involved in invasive CLL on apoptotic factors., Methods: A case-control study was conducted on 77 patients diagnosed with CLL along with 100 healthy individuals. Cell blood count was performed for all participants. According to Binet's classification, CLL patients were divided into different groups. B cells were isolated from the peripheral blood of CLL patients by binding to anti-CD19 beads. The expression of BAG3, GRP78 and HIF-1α genes was analyzed using the RT-PCR method. To confirm the results of RT-PCR, western blot analysis was carried out., Results: The results showed that there was a strong association among the expression of BAG3, GRP78 and HIF-1α. The stage of CLL in patients was highly correlated with the expression rate of each gene (p<0.001). Accordingly, the western blot analysis indicated that the concentrations of GRP78 and HIF-1α were significantly higher than the expression of BAG3, considering the stage of CLL., Conclusion: It was shown that increased expression of GRP78 and HIF-1α could result in the elevation of BAG3, as well as the disease progression. Therefore, the role of these metabolic factors might be more pronounced compared with the anti-apoptotic agents to monitor disease progression in CLL patients., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Aim: To evaluate the over expression of Glucose-regulated protein 78 kda (GRP78) in recurrent glioblastoma multiforme (GBM) and explore its chemoresatance., Material and Methods: Western blot analysis and real-time PCR were done to detect the expression of GRP78 in GBM tissues., Results: It was detected that GRP78 is up-expressed in GBM, especially in recurrent GBM. Radiotherapy (RT) plus temozolomide chemotherapy after primary tumor removal in relapsed patients induces C/EBP homologous protein (CHOP)., Conclusion: The results mentioned above illustrate an essential function of GRP78 in the pathogenesis of GBM and suggest this receptor as the latest prognostic marker for further GRP78-targeted molecular cancer therapy and potential diagnosis of GMB.

The aim of the present study was to investigate the effect of bortezomib on heat shock protein 27 (HSP27) in multiple myeloma (MM) and provide a potential new target for clinical treatment. Peripheral blood was collected from 50 normal subjects and 50 patients with newly diagnosed MM and the expression of HSP27 was detected by ELISA. The changes of HSP27 after conventional vincristine, doxorubicin and dexamethasone (VAD) chemotherapy, and bortezomib plus VAD were compared. The effect of bortezomib on U266 cell proliferation and apoptosis was detected using a Cell Counting Kit‑8 assay and Annexin V‑FITC/propidium iodide double staining with flow cytometry. The content of HSP27 following bortezomib treatment was determined by ELISA. Western blot analysis and reverse transcription‑quantitative PCR were used to detect the mRNA and protein expression of HSP27, Bax and Bcl‑2. HSP27 expression was increased in patients with MM compared with healthy control subjects, and the expression was increased as the cancer progressed (P<0.05). Compared with the VAD chemotherapy group, the bortezomib plus VAD chemotherapy regimen significantly inhibited the expression of HSP27 (P<0.05), and the content of HSP27 was decreased in patients in which treatment was effective compared to those patients that exhibited disease progression (P<0.05). The efficacy of the treatment regimes was not associated with age or gender. Compared with the control group, bortezomib or OGX‑427 (HSP27 inhibitor) treatment inhibited U266 cell proliferation, promoted U266 cell apoptosis (P<0.05) and significantly decreased HSP27 expression (P<0.05). Furthermore, the expression of HSP27 and Bcl‑2 was significantly decreased, while the expression of Bax was increased by bortezomib and OGX‑427 (P<0.05). There was no significant difference between the bortezomib and OGX‑427 group in the in vitro analysis. HSP27 was positively correlated with Bcl‑2 expression and negatively correlated with Bax expression in U266 cells. In conclusion, bortezomib promotes the apoptosis of MM cells, potentially by downregulating the expression of HSP27, providing a potential novel target for the clinical treatment of multiple myeloma.

The objective of this study was to create various pH/temp decline rates in hot-boned bull beef M. longissimus lumborum (LL) through a combination of electrical stimulation (ES) and pre-rigor holding temperature. The relationship between the pre-rigor interventions, the activities of µ-calpain and small heat shock proteins (sHSP), and the impacts on meat product quality were determined. Paired LL loins from 13 bulls were hot-boned within 40 min of slaughter, immediately ES and subjected to various holding temperatures (5, 15, 25, and 35°C) for 3 hr. The rate of muscle pH decline, sarcomere length, shear force, and proteolysis of muscle proteins were measured. ES-25°C had a longer sarcomere length compared to non-electrical stimulation samples. ES-25°C and ES-35°C samples had lower shear force values, higher µ-calpain activity and higher desmin, troponin-T, and sHSP degradation. The above findings suggest that pH/temp decline rates created in hot-boned muscle impacted muscle protein proteolysis by increasing the activity of proteases and degradation of sHSP., (© 2019 Japanese Society of Animal Science.)

Lead (Pb) is an environmental pollutant and can damage organisms. Selenium (Se) can alleviate Pb poisoning. The present study aimed to investigate the alleviative effect of Se on Pb-induced immune toxicity in chicken hearts. One-hundred-and-eighty Hy-line male chickens were randomly divided into four groups at 7 days of age. The control group was offered a standard commercial diet (SD) and drinking water (DW); the Se group was offered SD supplemented with sodium selenite (SeSD) and DW; the Pb + Se group was offered SeSD and DW supplemented with lead acetate (PbDW); and the Pb group was offered SD and PbDW. Relative mRNA expression of inducible nitric oxide synthase (iNOS), interleukins (IL-2, IL-4, IL-6, IL-12β, IL-17 and IFN-γ), and heat shock proteins (HSP27, HSP40, HSP60, HSP70, and HSP90) were determined by means of quantitative real-time PCR. Relative protein expression of iNOS, HSP60, HSP70, and HSP90 was assessed, as well as nitric oxide (NO) content and iNOS activity in heart tissue. The results indicated a down-regulation of interleukin (IL)-2 and IFN-γ and an up-regulation of NO, iNOS, interleukins (IL-4, IL-6, IL-12β, IL-17), and heat shock proteins (HSP27, HSP40, HSP60, HSP70, and HSP90) in Pb-damaged hearts. Se alleviated all of the above Pb-induced changes. There were time-dependent effects on NO content, iNOS activity, and mRNA levels of iNOS, IL-2, IL-4, IL-6, IL-17, HSP27, HSP40, HSP60, HSP70, and HSP90 after Pb treatment in the chicken hearts. Se alleviated Pb-induced immune toxicity in the chicken hearts.

BACKGROUND Gambogic acid (AG) is believed to be a potent anti-cancer agent. ER (endoplasmic reticulum) stress-induced cell apoptosis was identified as one of the anti-proliferative mechanisms of several anti-cancer agents. In this study, we investigated the involvement of ER stress-induced apoptosis in the anti-proliferative effect of GA on NSCLC (non-small cell lung cancer) cells. MATERIAL AND METHODS GA at 0, 0.5, and 1.0 μmol/l was used to treat A549 cells. We also used the ER stress-specific inhibitor 4-PBA (4-phenylbutyric acid) (1 μmol/l) to co-treat the cells incubated with GA. Cell viability was assessed by MTT (methyl thiazolyl tetrazolium) assay. Cell apoptosis was evaluated by MTT (methyl thiazolyl tetrazolium) assay. Intracellular ROS (reactive oxygen species) production was detected by DCFH-DA (2,7- dichloro-dihydrofluorescein diacetate) florescent staining. Western blotting was used to assess the expression and phosphorylation levels of protein. RESULTS GA treatment significantly reduced cell viabilities of NSCLC cells in a concentration-dependent manner. GA treatment increased intracellular ROS level, expression levels of GRP (glucose-regulated protein) 78, CHOP (C/EBP-homologous protein), ATF (activating transcription factor) 6 and caspase 12, as well as the phosphorylation levels of PERK (protein kinase R-like ER kinase) and IRE (inositol-requiring enzyme) 1alpha. Co-treatment of 4-PBA dramatically impaired the inhibitory effect of GA on cell viability. 4PBA co-treatment also decreased expression levels of GRP78, CHOP, ATF6, and caspase12, as well as the phosphorylation levels of PERK and IRE1alpha, in GA-treated NSCLC cells, without affecting ROS levels. CONCLUSIONS GA inhibited NSCLC cell proliferation by inducing ROS-induced ER stress-medicated apoptosis of NSCLC cells.

Both endoplasmic reticulum (ER) stress and autophagy involve in the pathological process of Parkinson's disease (PD). But the relationship between them is not clear in PD. A 6-OHDA-induced parkinsonian rat is recognized as a standard model for many years, and it can be used in experimental study. The glucose regulated protein 78 (GRP78) is a master regulator of ER stress, and the C/EBP homologous binding protein (CHOP) is an indicator of the UPR signaling. Besides, the Beclin-1 is also well known as a regulator of autophagy, and P62 is a specific marker to monitor autophagy. Therefore, we investigated the expressions of GRP78, CHOP, Beclin-1 and P62 in 6-OHDA-induced parkinsonian rat. Unilateral 6-OHDA injection into medial forebrain bundle was used except sham-operated rats. The rats were randomly divided into 6 groups: a sham-operated group; a model group; a 3-methyladenine (3-MA) group, administered 3-MA---autophagy inhibitor; a rapamycin group, administered rapamycin---autophagy inducer; a 4-phenylbutyric acids (4-PBA) group, administered 4-PBA---ER stress inhibitor; a tunicamycin (TM) group, administered TM---ER stress inducer. The results showed that the expressions of GRP78, CHOP and Beclin-1 increased, P62 decreased in model group; the expressions of GRP78 and CHOP were unchanged in 3-MA group and rapamycin group; but the expression of Beclin-1 decreased and P62 increased in 4-PBA group, while the expression of Beclin-1 increased and P62 decreased in TM group. These data suggest that ER stress and autophagy occurred in 6-OHDA-induced parkinsonian rat, and ER stress might induce autophagy. The result is important for the pathological mechanism of PD., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The periplasm of food-borne enteric pathogens is perhaps the only internal space of living species that can be severely acidified (pH 1-3), which occurs when these pathogens pass through the acidic human stomach. Whereas the periplasmic chaperoning systems are known to deal with such a harsh unfolding stress, other protein quality control mechanisms remain unexplored. Here we report a protease-mediated degradation mechanism that facilitates bacterial acid resistance. The genetic analysis revealed that mutant in degP encoding an HtrA family serine protease rendered Escherichia coli highly acid vulnerable. Combining genetically encoded trifunctional probe with 2D-based comparative proteomics, we identified its substrates. We further demonstrated that DegP directly bound to diverse aggregation-prone periplasmic proteins upon acid stress and these pre-mixed DegP-substrate co-aggregates were subsequently digested by proteolytic-rescued DegP during acid recovery. DegP represents an unprecedented "acid protease" that maintains protein homeostasis in coping with acid-induced protein unfolding stress within E. coli periplasm., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Arsenic and copper, two common environmental contaminants, presented potentially harmful to the immune system. In order to examine whether simultaneous exposure to arsenic and copper has the potential effects on immunity and inflammatory response in chicken thymus, 30 mg/kg arsenic trioxide and/or 300 mg/kg copper sulfate were added to chicken basal diet, respectively. After 12 weeks of administrations, we observed significantly decreased thymus weight and thymus index, inflammatory cell infiltration and hyperemia visible to the unaided eye. Concurrent administration arsenic and copper markedly increased levels of NF-κB, COX-2, iNOS and PTGEs and pro-inflammatory cytokines in concomitant with a tendency of Th1 bias immune response. Additionally, heat shock proteins levels were noticeably elevated in all treated groups. Above mentioned indicators were more pronounced among co-administrated groups. Collectively, arsenic and/or copper exposure engendered immunotoxicity of chicken thymus, resulting in inflammatory responses and imbalance of the immune response. Concurrent exposure of arsenic and copper was likely to potentiate the toxicity of chicken thymus and detailed mechanisms required further exploration. Heat shock proteins may play protective roles in thymus damage, and HSP60 combines with the IL-17/IL-10 ratio may be potential predictive marker of arsenic and/or copper-subchronic toxicity in chicken thymus., (Copyright © 2018. Published by Elsevier B.V.)

Objectives: Wound healing of the oral mucosal epithelium through the application of far infrared radiation emitted by isotropic high-density carbon was investigated in order to clarify the preventive and therapeutic effects of isotropic high-density carbon on oral mucosal injury., Materials and Methods: A carbon massager with an isotropic high-density carbon tip was used. Far infrared radiation was applied to the human buccal mucosal squamous cell carcinoma cell line, HO-1-N-1 using a carbon massager, and cell growth factors and heat shock protein levels were measured using real-time RT-PCR and ELISA. Far infrared radiation was applied to oral mucosal injury in SD rats over time using the carbon massager, and its effects were examined by HE staining and immunostaining. The immunostaining positive rate was measured and analyzed using image analysis software., Results: Far infrared radiation induced stronger mRNA expression and higher HSP27 and HSP70 protein levels on real-time RT-PCR and ELISA than in the control group. The far infrared radiation of oral mucosal injury in rats induced strong positive reactions, and positive rates for Ki67, HSP27, and HSP70 were higher than those in the control group., Conclusions: The treatment of oral mucosal injury with far infrared radiation emitted by isotropic high-density carbon appears to have promoted heat shock protein production and induced regenerative reactions more strongly than in the control group., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Hemorrhage shock could initiate endoplasmic reticulum stress (ERS) and then induce neuronal apoptosis. The aim of this study was to investigate whether sevoflurane postconditioning could attenuate brain injury via suppressing apoptosis induced by ERS. Seventy male rats were randomized into five groups: sham, shock, low concentration (sevo1, 1.2%), middle concentration (sevo2, 2.4%) and high concentration (sevo3, 3.6%) of sevoflurane postconditioning. Hemorrhage shock was induced by removing 40% of the total blood volume during an interval of 30 min. 1 h after the completion of bleeding, the animals were reinfused with shed blood during the ensuing 30 min. The spatial learning and memory ability of rats were measured by Morris water maze (MWM) test three days after the operation. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) positive cells in the hippocampus CA1 region were assessed after the MWM test. The expression of C/EBP-homologousprotein (CHOP) and glucose-regulated protein 78 (GRP78) in the hippocampus were measured at 24 h after reperfusion. We found that sevoflurane postconditioning with the concentrations of 2.4% and 3.6% significantly ameliorated the spatial learning and memory ability, decreased the TUNEL-positive cells, and reduced the GRP78 and CHOP expression compared with the shock group. These results suggested that sevoflurane postconditioning with the concentrations of 2.4% and 3.6% could ameliorate spatial learning and memory deficit after hemorrhage shock and resuscitation injury via suppressing apoptosis induced by ERS., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Stress-induced phosphoprotein 1 (STIP1) is an adaptor protein that bridges HSP70 and HSP90 folding and a secretory protein that regulates malignant tumor progression. The aim of the present study was to demonstrate the clinicopathological significance and prognostic role of STIP1 in colorectal cancer (CRC). We used data from The Cancer Genome Atlas (TCGA) to analyze STIP1 expression in CRC and utilized 8 pairs of fresh-frozen tissue samples to investigate STIP1 expression in CRC tissues and adjacent normal tissues using quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot assays. We also used immunohistochemical staining to detect STIP1 expression in 144 formalin-fixed, paraffin-embedded (FFPE) CRC tissue samples and determine the clinical significance of STIP1 expression in CRC. The results of bioinformatics analysis, qRT-PCR, and Western blot showed that STIP1 expression was higher in CRC tissues than in adjacent normal tissues. High STIP1 expression was significantly correlated with advanced T stage (P = .01), N stage (P = .001), M stage (P < .001), and TNM stage (P < .001). Moreover, Kaplan-Meier analyses indicated that higher STIP1 expression predicted a worse prognosis in patients with CRC, and Cox regression analysis revealed that STIP1 was an independent prognostic factor for overall survival and disease-free survival in patients with CRC. In conclusion, our results suggest that STIP1 acts as an oncogene in CRC and can therefore serve as a biomarker for the prognosis of patients with CRC., (Copyright © 2018. Published by Elsevier Inc.)

Background: Heat stress (HS) has an adverse effect on meat quality; however, the underlying molecular mechanisms altering meat quality due to muscle responses to stress remain unclear. Sixteen castrated male crossbreeds between Landrace × Yorkshire sows and Duroc boars (79.00 ± 1.50 kg body weight) were exposed to either thermal neutral (22 °C, n = 8) or HS (30 °C, n = 8) conditions for 3 weeks. Subsequently, the longissimus dorsi (LD) muscle of all pigs was assayed for meat quality parameters and proteome analysis., Results: HS decreased post mortem (24 h) pH and intramuscular fat, changed ultimate L*, a* and b* values and increased drip loss and shear force. Proteome analysis of the LD was conducted by two-dimensional gel electrophoresis and mass spectrometry. A total of 23 differentially expressed proteins were identified, of which three were verified by western blotting analysis. The identified proteins were involved in six types of biological process: carbohydrate metabolism, myofibrillar and cytoskeleton structure, stress response, antioxidant and detoxification, calcium binding and cellular apoptosis. Interestingly, HS induced higher levels of heat shock protein, antioxidants and calcium binding proteins, which are involved in the mechanisms of defense and homeostasis., Conclusion: The results indicate that HS-induced changes in the expression of myofibrillar proteins, glucose and energy metabolism-related proteins, heat shock protein and antioxidant enzymes might, at least partly, contribute to increase in meat tenderness. These findings will provide the foundation for developing future mitigating solutions and preventative therapies to reduce the detrimental effects of chronic HS on muscle function, metabolism and meat quality. © 2017 Society of Chemical Industry., (© 2017 Society of Chemical Industry.)

The 78-kDa glucose-regulated protein (GRP78), a chaperone protein located in the endoplasmic reticulum (ER), has been reported to have neuroprotective effects in the injured central nervous system. Our aim was to examine the expression profiles and subcellular distributions of GRP78 and its association with the neuroglial reaction in the rat striatum after transient, focal cerebral ischemia. In sham-operated rats, constitutive, specific immunoreactivity for GRP78 was almost exclusively localized to the rough ER of striatal neurons, with none in the resting, ramified microglia or astrocytes. At 1 day post reperfusion, increased expression was observed in ischemia-resistant cholinergic interneurons, when most striatal neurons had lost GRP78 expression (this occurred earlier than the loss of other neuronal markers). By 3 days post reperfusion, GRP78 expression had re-emerged in association with the activation of glial cells in both infarct and peri-infarct areas but showed different patterns in the two regions. Most of the expression induced in the infarct area could be attributed to brain macrophages, while expression in the peri-infarct area predominantly occurred in neurons and reactive astrocytes. A gradual, sustained induction of GRP78 immunoreactivity occurred in reactive astrocytes localized to the astroglial scar, lasting for at least 28 days post reperfusion. Using correlative light- and electron-microscopy, we found conspicuous GRP78 protein localized to abnormally prominent, dilated rough ER in both glial cell types. Thus, our data indicate a link between GRP78 expression and the activated functional status of neuroglial cells, predominantly microglia/macrophages and astrocytes, occurring in response to ischemia-induced ER stress.

Background: Centrosomes contain two centrioles: a pre-existing mature centriole and a newly formed immature centriole. Each centriole is duplicated once within a cell cycle, which is crucial for proper centrosome duplication and cell division., Objective: To describe the centrosome duplication cycle in human epidermis, Bowen's disease (BD), and squamous cell carcinoma (SCC)., Methods: Immunofluorescent staining of centriolar proteins and Ki-67 was used to evaluate cell cycles and the number of centrioles. Centrobin and Outer dense fiber of sperm tails 2 (ODF2) were used as markers for immature and mature centrioles, respectively., Results: Normal human primary epidermal keratinocytes in a monolayered culture have one centrobin + centriole (CTRB 1+ cells) supposed in G0/G1 phases or have two centrobin + centrioles (CTRB 2+ cells) supposed in S-G2 phase. In a three-dimensional culture and in vivo human epidermis, the majority of suprabasal cells were CTRB 2+ cells, in spite of their non-proliferative Ki-67 - nature. The tumor mass of BD and SCC contained CTRB 1+ cells and Ki-67 + proliferating and Ki-67 - non-proliferative CTRB 2+ cells. Clumping cells in BD had increased numbers of centrioles, with an approximate 1:1 to 2:1 ratio of centrobin + to ODF2 + centrioles., Conclusions: The cell cycle arrest of suprabasal cells is distinct from the G0 arrest of monolayered epithelial cells. Tumor mass of BD and SCC contained non-proliferative cells with the characteristics of the suprabasal cells of normal epidermis. A constant ratio of the number of centrobin + to ODF2 + centrioles indicates that multiple centrioles were induced by cell division failure rather than centriole overduplication in clumping cells., (Copyright © 2018. Published by Elsevier B.V.)

The heat shock proteins are endogenous proteins with the ability to act as molecular chaperones. Methods that provide cell protection by way of some damage can positively influence the results of surgery. The present review summarizes current knowledge concerning the cardioprotective role of the heat shock proteins as occurs in heart damage, including relevant information about the stresses that regulate the expression of these proteins and their potential role as biomarkers of heart disease.

In the present study, 155 Nellore cattle were genotyped for the CAPN4751 and UOGCAST polymorphisms and phenotyped for shear force (SF) at 1, 7 and 14days aging. The effects of different genotypic combinations were evaluated on the Longissimus muscle proteomic profile using 2DE and mass spectrometry. A significant association was found between genotypes for UOGCAST and CAPN4751 and meat tenderness. The CC genotype for both markers was favorable for lesser shear force than TT. A total of 40 spots showed significant differential expression profiles (P<0.05), of which eight had a main effect for the CAPN4751 marker, 11 for UOGCAST, two for both markers, and 19 had interactions between markers, including myosin (MYL1, MYL2, MYLPF and MYL6B), actin (ACTA1 and CAPZβ), troponin (TNNT1 and TNNT3) and heat shock proteins (HSPB6, HSPB1 and HSP70-2). The results demonstrated that UOGCAST and CAPN4751 genotypes led to variability on the expression of proteins that are involved in muscle metabolism, and consequently affect meat tenderness., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Objective: To investigate the protective effect of prostaglandin E1 (PGE-1) against brain injury induced by hyperoxia in neonatal rats and observe the changes in the expression of glucose-regulated protein 78 (GRP78) and cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP), and to provide a theoretical basis for the clinical application of PGE-1 in the treatment of neonatal brain injury induced by hyperoxia., Methods: Sixty neonatal Wistar rats were randomly divided into air control group, hyperoxic brain injury model group, and hyperoxic brain injury+PGE-1 group. All rats except those in the air control group were treated to establish a hyperoxic brain injury model. From the first day of modeling, the rats in the hyperoxia brain injury+PGE-1 group were intraperitoneally injected with PGE-1 2 μg/kg daily for 7 consecutive days, while the other two groups were treated with normal saline instead. The water content of brain tissue was measured; the pathological changes of brain tissue were evaluated by hematoxylin-eosin staining; the apoptosis of brain cells was assessed by nuclear staining combined with TUNEL staining; the protein expression of GRP78 and CHOP in brain tissue was measured by Western blot., Results: The water content of brain tissue in the hyperoxic brain injury model group was significantly higher than that in the hyperoxic brain injury+PGE-1 group and air control group (P<0.05); the water content of brain tissue in the hyperoxic brain injury+PGE-1 group was significantly higher than that in the air control group (P<0.05). The pathological section of brain tissue showed inflammatory cell infiltration and mild cerebrovascular edema in the brain parenchyma in the hyperoxic brain injury model group; the periparenchymal inflammation and edema in the hyperoxic brain injury+PGE-1 group were milder than those in the hyperoxic brain injury model group. The apoptosis index of brain tissue in the hyperoxic brain injury model group was significantly higher than that in the hyperoxic brain injury+PGE-1 group and air control group (P<0.05); the apoptosis index of brain tissue in the hyperoxic brain injury+PGE-1 group was significantly higher than that in the air control group (P<0.05). The protein expression of GRP78 and CHOP in brain tissue was significantly higher in the hyperoxic brain injury model group than in the hyperoxic brain injury+PGE-1 group and air control group (P<0.05); the protein expression of GRP78 and CHOP was significantly higher in the hyperoxic brain injury+PGE-1 group than in the air control group (P<0.05)., Conclusions: PGE-1 has a protective effect against hyperoxia-induced brain injury in neonatal rats, which may be related to the inhibition of cell apoptosis by down-regulating the expression of GRP78 and CHOP.

Studies have suggested that the phenomenon of dark cutting (Canada B4) beef may also be related to muscle glycolytic proteins. The objective of this study, therefore, was to analyze longissimus thoracis (LT; n=23), from Canada AA (n=8), atypical (AB4; pH<5.9, n=8) and typical (TB4; pH>5.9, n=7) B4 heifer and steer carcasses, for sarcoplasmic and myofibrillar proteins using 2-D gel electrophoresis and LC-MS/MS mass spectrometry. Results indicated that AB4 LT had intramuscular pH and lactate concentration similar to Canada AA but lower (P<0.05) L* and b*. Moreover, AB4 LT were tougher than Canada AA even at 21days post-mortem, unlike TB4. Canada AB4 LT had reduced (P<0.05) levels of creatine kinase, glyceraldehyde-3-phosphate dehydrogenase, lactate dehydrogenase and glycerol-3-phosphate dehydrogenase [NAD(+)], indicating a compromised glycolytic capacity in AB4. Canada AB4 LT had increased (P<0.05) abundances of phosphatidylethanolamine-binding protein 1 and small heat shock proteins., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Objective: Biomarkers reflecting the etiology and pathophysiology of inner ear diseases are limited. Evaluation of proteins in the perilymph may improve our understanding of inner ear disease. Heat shock proteins (HSP) belong to a superfamily of stress proteins and promote refolding of denatured proteins. The aim of the study was to analyze HSP in human perilymph and to identify possible correlation with audiological and etiologic data., Methods: Sampling of the perilymph was performed during cochlear implantation and vestibular schwannoma removal. Individual proteins were identified by a shot-gun proteomics approach by orbitrap mass spectrometry. Expression of HSP genes was determined in human cochlear tissue that was obtained during transcochlear surgeries., Results: Ten subgroups of HSP were identified in human perilymph samples. Increased levels of HSP were detected in a higher percentage in the perilymph of patients with residual hearing when compared with patients with no residual hearing in cochlear implantation. In patients with complete preservation of residual hearing, HSP 90 is identified in a lower percentage whereas HSP 70 1A/1B and 6 was identified in all the samples. Constitutive expression of HSP family members was verified in normal cochlear tissue., Conclusion: The 10 HSP variants are not identified in all the perilymph samples, but in a higher proportion in patients with residual hearing compared with patients with no residual hearing. In-depth proteome analysis of perilymph samples in correlation to patients' audiogram data shows an increased concentration of HSP in patients with residual hearing. An increase in specific HSP in patients with loss of residual hearing after cochlear implantation was not observed.

Purpose: To investigate the effect of dexmedetomidine (Dex) in a rat ex vivo lung model of ischemia-reperfusion injury., Methods: An IL-2 ex vivo lung perfusion system was used to establish a rat ex vivo lung model of ischemia-reperfusion injury. Drugs were added to the perfusion solution for reperfusion. Lung injury was assessed by histopathological changes, airway pressure (Res), lung compliance (Compl), perfusion flow (Flow), pulmonary venous oxygen partial pressure (PaO2), and lung wet/dry (W/D) weight ratio. The levels of superoxide dismutase (SOD), malondialdehyde (MDA), 78 kDa glucose-regulated protein (GRP78) and CCAAT/enhancer-binding protein homologous protein (CHOP) were measured, respectively., Results: The introduction of Dex attenuated the post-ischemia-reperfusion lung damage and MDA level, improved lung histology, W/D ratio, lung injury scores and SOD activity. Decreased mRNA and protein levels of GRP78 and CHOP compared with the IR group were observed after Dex treatment. The effect of Dex was dosage-dependence and a high dose of Dex (10 nM) was shown to confer the strongest protective effect against lung damage (P<0.05). Yohimbine, an α2 receptor antagonist, significantly reversed the protective effect of Dex in lung tissues (P<0.05)., Conclusion: Dex reduced ischemia-reperfusion injury in rat ex vivo lungs.

Aims: Heat shock proteins (HSPs) are a group of molecules induced by a variety of environmental and pathophysiological stresses, including cancer. HSPs are implicated in the regulation of apoptosis and immunity in neoplasm. Transcription factor heat shock factor 1 (HSF1) acts as the master regulator to control HSP expression, and is therefore involved in tumorigenesis. The purpose of this study was to evaluate the expression and clinicopathological relevance of HSPs and HSF1 in clear cell renal cell carcinoma (ccRCC)., Methods and Results: The expression of HSP27, HSP60, HSP70, HSP90 and HSF1 was assessed in 428 cases of ccRCC using immunohistochemistry. High expression of HSP60 and HSP70 was correlated positively with grade and stage. High expression of HSF1 was correlated positively with stage. Univariate and multivariate analyses demonstrated that 216 patients (52%) with tumour expressing three or four markers in a panel of HSP60, HSP70, HSP90 and HSF1 had a significantly heightened risk for cancer-specific mortality than tumours expressing fewer than three markers (P < 0.0001; concordance index, 0.81)., Conclusions: Immunohistochemical examination of HSPs and HSF1 provides useful prognostic information that may contribute to the design of therapeutic strategies for patients with ccRCC., (© 2017 John Wiley & Sons Ltd.)

The small heat shock protein (sHsp) chaperones are crucial for cell survival and can prevent aggregation of client proteins that partially unfold under destabilizing conditions. Most investigations on the chaperone activity of sHsps are based on a limited set of thermosensitive model substrate client proteins since the endogenous targets are often not known. There is a high diversity among sHsps with a single conserved β-sandwich fold domain defining the family, the α-crystallin domain, whereas the N-terminal and C-terminal regions are highly variable in length and sequence among various sHsps and conserved only within orthologues. The endogenous targets are probably also varying among various sHsps, cellular compartments, cell type and organism. Here we have investigated Hsp21, a non-metazoan sHsp expressed in the chloroplasts in green plants which experience huge environmental fluctuations not least in temperature. We describe how Hsp21 can also interact with the chloroplast thylakoid membranes, both when isolated thylakoid membranes are incubated with Hsp21 protein and when plants are heat-stressed. The amount of Hsp21 associated with the thylakoid membranes was precisely determined by quantitative mass spectrometry after metabolic 15 N-isotope labeling of either recombinantly expressed and purified Hsp21 protein or intact Arabidopsis thaliana plants. We found that Hsp21 is among few proteins that become associated with the thylakoid membranes in heat-stressed plants, and that approximately two thirds of the pool of chloroplast Hsp21 is affected. We conclude that for a complete picture of the role of sHsps in plant stress resistance also their association with the membranes should be considered., (© 2017 The Authors Protein Science published by Wiley Periodicals, Inc. on behalf of The Protein Society.)

Macrophages are the main immune-competent cells that infiltrate in tumors. Tumor-associated macrophages (TAMs), termed M2 macrophages, facilitate tumor progress and promote metastasis. However, M2 macrophages always display an immunosuppressive phenotype, which is not in accordance with the tumor inflammatory microenvironment and inflammation-related metastasis. In this study, we established a macrophage polarization model with human monocytes and found that the conditioned medium from M2 macrophages increased GRP78 expression in tumor cells and facilitated tumor cell migration. Mechanistically, excessive GRP78 formed a protein complex with STAT3 and JAK2 to promote STAT3 phosphorylation. Furthermore, p-STAT3 facilitated the high expression of inflammatory factors IL-1β and TNF-α in tumor cells, which was important in M2 macrophage-induced metastasis. The present data demonstrate that M2 macrophages elevate tumor cell GRP78 expression to trigger an inflammatory response, which further facilitates tumor metastasis. Therefore, our study not only uncovered a new cause of GRP78 overexpression in tumor cell, but also, explained the antinomy of TAMs immunosuppressive properties and inflammation-related tumor metastasis., (Copyright © 2017 Elsevier Inc. All rights reserved.)

It is widely accepted that mature sperm contains RNA. The first hypothesis was that sperm RNAs have no functions of their own but are simply residues of spermatogenesis reflecting the events that occurred during their formation in the testes. More recently new discoveries have essentially expanded these views, showing that sperm mRNAs constitute a population of stable full-length transcripts, many of which are selectively retained during spermatogenesis and delivered to oocytes contributing to early embryo development. It is well known that semen quality can be influenced by occasional physical stress, infection, and variation in temperature and the definition of new markers for evaluation of semen could offer knowledge about the fertility potential of a semen sample. The aim of the present study was to evaluate the presence and the relative quantity of transcripts and protein of heat shock protein 70 (HSP70), 90 (HSP90) and clusterin (CLU) in Percoll-selected spermatozoa collected from seven adult boars of proven fertility routinely employed for artificial insemination. Our results showed the presence of HSP70, HSP90 and CLU transcripts with different level of expression: high for HSPs and low for CLU transcripts. The transcript level of both HSPs are similar among selected spermatozoa derived from high quality sperm with the exception of one boar that showed a reduced content of HSP70 and HSP90 mRNA together with a lower semen quality. At protein level, both HSPs were detected with similar amount among all seven boars whilst no band was evidenced for CLU protein.

We investigated the role of endoplasmic reticulum stress (ERS) in chronic intermittent hypobaric hypoxia (CIHH)-induced cardiac protection. Adult male Sprague-Dawley rats were exposed to CIHH treatment simulating 5000 m altitude for 28 days, 6 hours per day. The heart was isolated and perfused with Langendorff apparatus and subjected to 30-min ischemia followed by 60-min reperfusion. Cardiac function, infarct size, and lactate dehydrogenase (LDH) activity were assessed. Expression of ERS molecular chaperones (GRP78, CHOP and caspase-12) was assayed by western blot analysis. CIHH treatment improved the recovery of left ventricular function and decreased cardiac infarct size and activity of LDH after I/R compared to control rats. Furthermore, CIHH treatment inhibited over-expression of ERS-related factors including GRP78, CHOP and caspase-12. CIHH-induced cardioprotection and inhibition of ERS were eliminated by application of dithiothreitol, an ERS inducer, and chelerythrine, a protein kinase C (PKC) inhibitor. In conclusion CIHH treatment exerts cardiac protection against I/R injury through inhibition of ERS via PKC signaling pathway.

Growth temperature is a critical factor that affects cultivation of Arthrospira platensis which is a type of cyanobacterium widely known as Spirulina that has significant commercial value. To investigate the molecular mechanism underlying the thermotolerance of Spirulina, differential protein expression profiling was carried out using iTRAQ-based quantitative proteomic analysis. This study only analyzed changes in thylakoids. Among the 2085 proteins quantified, 43 differentially expressed proteins were selected based on the fold change cutoff scores of ≥2 or ≤0.5 for up-regulation or down-regulation, respectively. An analysis of these 43 proteins found that 23% of them are photosynthetic system proteins which include photosynthetic enzymes and pigment proteins. The dynamic change of these proteins indicates that photosynthetic system functions were profoundly affected under heat stress and the light-dependent reactions were probably the most sensitive to temperature changes. Meanwhile, to cope with the low energy production due to impaired photosynthesis there was a considerable down-shift in protein synthesis which is a very energy demanding process. The impaired photosynthesis led to low energy generation that was compensated by a down-shift in translation (the most energy-demanding process) and an up-shift of glycolysis. The reduction of many ribosome proteins may lead to a loss in translation efficiency; therefore, Spirulina may adopted a different mechanism to increase translational elongation under heat stress to compensate for this loss, such as elevate L7/L12 proteins. Changes were also found in the classical heat shock proteins, the ROS scavenging system, DNA-binding proteins, and some membrane proteins. In conclusion, this research demonstrate that heat stress induces profound changes in cellular physiology and shed light on the mechanism of the heat stress response and thermotolerance of Arthrospira platensis., Biological Significance: Arthrospira platensis, widely known as Spirulina, is a type of cyanobacteria which is cultivated at large scale for it great commercial value. It has become a consensus that continually increasing temperature due to global warming is bringing serious threat to agriculture, including the Spirulina cultivation. High temperature not only limits biomass accumulation by Spirulina, but also changes the composition of nutrition. Therefore there is a greater need than ever before to understand how Spirulina tolerates and cope with high temperature. In this study, we for the first time applied the iTRAQ-based quantitative proteomic technology to investigate the thermotolerance of Spirulina. Our results showed that many biological processes were altered by heat stress. Most significantly, we found that heat stress harmed the photosynthesis ability and caused low energy production, and to deal with this situation, energy demanding processes like protein synthesis were down-shifted and the alternative energy metabolism process glycolysis was up-shifted. Our results also show other important proteins, like the classical heat shock proteins and some antioxidant proteins, are also increased. Thus our study sheds light to our understanding of the mechanism underlying the thermotolerance of Spirulina., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Using immune cells of sea urchin Strongylocentrotus droebachiensis in early development as a model, the cellular protective mechanisms against ionic and poly(allylamine)-coated silver nanoparticle (AgNPs; 14 ± 6 nm) treatments at 100 μg L -1 were investigated. Oxidative stress, heat shock protein expression, and pigment production by spherulocytes were determined as well as AgNP translocation pathways and their multiple effects on circulating coelomocytes. Sea urchins showed an increasing resilience to Ag over time because ionic Ag is accumulated in a steady way, although nanoAg levels dropped between 48 h and 96 h. A clotting reaction emerged on tissues injured by dissolved Ag (present as chloro-complexes in seawater) between 12 h and 48 h. Silver contamination and nutritional state influenced the production of reactive oxygen species. After passing through coelomic sinuses and gut, AgNPs were found in coelomocytes. Inside blood vessels, apoptosis-like processes appeared in coelomocytes highly contaminated by poly(allylamine)-coated AgNPs. Increasing levels of Ag accumulated by urchins once exposed to AgNPs pointed to a Trojan-horse mechanism operating over 12-d exposure. However, under short-term treatments, physical interactions of poly(allylamine)-coated AgNPs with cell structures might be, at some point, predominant and responsible for the highest levels of stress-related proteins detected. The present study is the first report detailing nano-translocation in a marine organism and multiple mechanisms by which sea urchin cells can deal with toxic AgNPs. Environ Toxicol Chem 2017;36:1872-1886. © 2016 SETAC., (© 2016 SETAC.)

The small heat shock protein HSPB8 and its co-chaperone BAG3 are proposed to regulate cytoskeletal proteostasis in response to mechanical signaling in muscle cells. Here, we show that in dividing cells, the HSPB8-BAG3 complex is instrumental to the accurate disassembly of the actin-based contractile ring during cytokinesis, a process required to allow abscission of daughter cells. Silencing of HSPB8 markedly decreased the mitotic levels of BAG3 in HeLa cells, supporting its crucial role in BAG3 mitotic functions. Cells depleted of HSPB8 were delayed in cytokinesis, remained connected via a disorganized intercellular bridge, and exhibited increased incidence of nuclear abnormalities that result from failed cytokinesis (i.e., bi- and multi-nucleation). Such phenotypes were associated with abnormal accumulation of F-actin at the intercellular bridge of daughter cells at telophase. Remarkably, the actin sequestering drug latrunculin A, like the inhibitor of branched actin polymerization CK666, normalized F-actin during cytokinesis and restored proper cell division in HSPB8-depleted cells, implicating deregulated actin dynamics as a cause of abscission failure. Moreover, this HSPB8-dependent phenotype could be corrected by rapamycin, an autophagy-promoting drug, whereas it was mimicked by drugs impairing lysosomal function. Together, the results further support a role for the HSPB8-BAG3 chaperone complex in quality control of actin-based structure dynamics that are put under high tension, notably during cell cytokinesis. They expand a so-far under-appreciated connection between selective autophagy and cellular morphodynamics that guide cell division.

The present study for the first time evaluates the serodiagnostic efficacy of two recombinant antigens namely, listeriolysin O (rLLO) and phosphatidyl-inositol phospholipase C (rPI-PLC). Indirect ELISA with the above recombinant antigens was used on samples collected from bovines (n=106), goats (n=138) and pigs (n=92) having either a history of abortion, emaciation and/or apparently healthy animals. Isolation of Listeria was attempted from the blood samples using USDA-FSIS method. On screening of test sera by rLLO-based ELISA, antibodies against anti-listeriolysin O (ALLO) were observed in goats (22.46%), bovines (15.10%) and pigs (16.31%). As advocated, after adsorption of positive serum samples with streptolysin O (SLO), the seropositivity for ALLO was marginally reduced (p>0.05) in goats (21.73%) and bovines (10.38%), whereas, in pigs the reduction (5.43%) was significant (p<0.05). On the contrary, rPI-PLC-based ELISA revealed higher non-specific seropositivity for antilisterial antibodies in goats (45.65%), bovines (31.13%) and pigs (8.69%). Further, on comparing the seropositivity with isolation rate, of the 16 animals that were culturally-positive for L. monocytogenes, 15 showed ALLO positivity in unadsorbed as well as SLO-adsorbed sera by rLLO-based ELISA, however, rPI-PLC-based ELISA could detect seropositivity in only 5 animals. Moreover, rPI-PLC-based ELISA also showed seropositivity in those animals (7/30) that were culturally positive for other Listeria spp. In conclusion, rLLO can serve as a better antigen than rPI-PLC in ELISA for the serodiagnosis of listeriosis in animals; however, prior adsorption of test sera with SLO is required to avoid false positive results., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Heat shock proteins (HSPs) are abundant cellular proteins involved with protein homeostasis. They have both constitutive and inducible isoforms, whose expression levels are further increased by stress conditions, such as temperature elevation, reduced oxygen levels, infection, inflammation and exposure to toxic substances. In these situations, HSPs exert a pivotal role in offering protection, preventing cell death and promoting cell recovery. Although the majority of HSPs functions are exerted in the cytoplasm and organelles, several lines of evidence reveal that HSPs are able to induce cell responses in the extracellular milieu. HSPs do not possess secretion signal peptides, and their secretion was subject to widespread skepticism until the demonstration of the role of unconventional secretion forms such as exosomes. Secretion of HSPs may confer immune system modulation and be a cell-to-cell mediated form of increasing stress resistance. Thus, there is a wide potential for secreted HSPs in resistance of cancer therapy and in the development new therapeutic strategies.

Many biochemical events occur inside grains during post-harvest processes. Several methods have been developed to relate the chemical composition of the coffee grain to the beverage quality, including identification of possible molecular markers for flavor characterizing. This study was aimed at evaluating the changes in the proteomic profile of pulped and natural C. arabica grains dried in a yard or dryer at 60°C. It was observed that fruits dried in a dryer at 60°C showed an altered proteomic profile, with a reduction in the most abundant proteins compared to those yard-dried grains. Among the identified proteins, those involved in the metabolism of sugars and stress response were highlighted. Results have shown that post-harvest processes that impact coffee quality are related to changes in protein abundance, indicating that proteomic analysis may be effective in the identification of biochemical changes in coffee grains subjected to different post-harvest processes., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Background: While there is compelling rationale to use heat shock protein 90 (Hsp90) inhibitors for treatment of advanced prostate cancer, agents that target the N-terminal ATP-binding site of Hsp90 have shown little clinical benefit. These N-terminal binding agents induce a heat shock response that activates compensatory heat shock proteins, which is believed to contribute in part to the agents' lack of efficacy. Here, we describe the functional characterization of two novel agents, SM253 and SM258, that bind the N-middle linker region of Hsp90, resulting in reduced client protein activation and preventing C-terminal co-chaperones and client proteins from binding to Hsp90., Methods: Inhibition of Hsp90 activity in prostate cancer cells by SM253 and SM 258 was assessed by pull-down assays. Cell viability, proliferation and apoptosis were assayed in prostate cancer cell lines (LNCaP, 22Rv1, PC-3) cultured with N-terminal Hsp90 inhibitors (AUY922, 17-AAG), SM253 or SM258. Expression of HSR heat shock proteins, Hsp90 client proteins and co-chaperones was assessed by immunoblotting. Efficacy of the SM compounds was evaluated in human primary prostate tumors cultured ex vivo by immunohistochemical detection of Hsp70 and Ki67., Results: SM253 and SM258 exhibit antiproliferative and pro-apoptotic activity in multiple prostate cancer cell lines (LNCaP, 22Rv1, and PC-3) at low micromolar concentrations. Unlike the N-terminal inhibitors AUY922 and 17-AAG, these SM agents do not induce expression of Hsp27, Hsp40, or Hsp70, proteins that are characteristic of the heat shock response, in any of the prostate cell lines analyzed. Notably, SM258 significantly reduced proliferation within 2 days in human primary prostate tumors cultured ex vivo, without the significant induction of Hsp70 that was caused by AUY922 in the tissues., Conclusions: Our findings provide the first evidence of efficacy of this class of C-terminal modulators of Hsp90 in human prostate tumors, and indicate that further evaluation of these promising new agents is warranted. Prostate 76:1546-1559, 2016. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

Cold atmospheric pressure plasma (CAP) has potential to be utilized as an alternative method for sterilization in food industries without thermal damage or toxic residues. In contrast to the bactericidal effects of CAP, information regarding the efficacy of CAP against eukaryotic microorganisms is very limited. Therefore, herein we investigated the effects of CAP on the budding yeast Saccharomyces cerevisiae, with a focus on the cellular response to CAP. The CAP treatment caused oxidative stress responses including the nuclear accumulation of the oxidative stress responsive transcription factor Yap1, mitochondrial fragmentation, and enhanced intracellular oxidation. Yeast cells also induced the expression of heat shock protein (HSP) genes and formation of Hsp104 aggregates when treated with CAP, suggesting that CAP denatures proteins. As phenomena unique to eukaryotic cells, the formation of cytoplasmic mRNP granules such as processing bodies and stress granules and changes in the intracellular localization of Ire1 were caused by the treatment with CAP, indicating that translational repression and endoplasmic reticulum (ER) stress were induced by the CAP treatment. These results suggest that the fungicidal effects of CAP are attributed to the multiple severe stresses.