Heat shock pretreatment improves mesenchymal stem cell viability by heat shock proteins and autophagy to prevent cisplatin-induced granulosa cell apoptosis

Abstract Background Bone marrow mesenchymal stem cells (BMSCs) can partially repair chemotherapy-induced ovarian damage. However, low survival rate after transplantation hampers the therapeutic efficiency of BMSCs. Heat shock pretreatment (HSP) effectively improves the cell survival. This study attempted to investigate the mechanisms of HSP on BMSCs survival and the effects of heat shock-pretreated BMSCs (HS-MSCs) on cisplatin-induced granulosa cell (GC) apoptosis. Methods BMSCs were isolated, cultured, and identified. After receiving HSP for different duration times in a 42 °C water bath, the apoptotic rates of BMSCs were detected by Annexin V-FITC/PI to determine the optimal condition of HSP. Cisplatin was added to the medium of HS-MSCs to simulate chemotherapy environment. The proliferative curve, apoptotic rate, and viability of HS-MSCs were determined by CCK-8, Annexin V-FITC/PI, and Hoechst33342/PI respectively to explore the alteration of biological characteristics. The levels of heat shock protein 70 and 90 (HSP70 and HSP90) and the expressions of autophagy-related markers (Beclin1 and LC3B) were detected by Western blot. In addition, the autophagosomes were observed by transmission electronic microscopy to discuss the possible mechanisms. The GCs were isolated, cultured, and identified. The HS-MSCs were co-cultured with GCs before and after the addition of cisplatin. Then, the apoptotic rate and viability of GCs were detected to investigate the therapeutic and preventive effects of HS-MSCs on GC apoptosis. Results After receiving HSP at 42 °C for 1 h, BMSCs represented the lowest apoptotic rate. After the addition of cisplatin, the apoptotic rate of HS-MSCs (11.94% ± 0.63%) was lower than that of BMSCs (14.30% ± 0.80%) and the percentage of HS-MSCs expressing bright blue/dull red fluorescence was lower than that of BMSCs. The expression of HSP70 and HSP90 increased, while the number of autophagosomes, the expression of Beclin1, and the LC3BII/LC3BI ratio decreased in HS-MSCs. The apoptotic rates of GCs co-cultured with HS-MSCs before and after the addition of cisplatin were 39.88% ± 1.65% and 36.72% ± 0.96%, both lower than those of cisplatin-induced GCs (53.81% ± 1.89%). Conclusion HSP can alleviate the apoptosis and improve the survival of BMSCs under chemotherapy environment. The mechanism may be associated with the elevated expression of HSP70 and HSP90 and the attenuation of autophagy. Moreover, HS-MSCs have both therapeutic and preventive effects on cisplatin-induced GC apoptosis.

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Distinct Gene Expression Patterns of Two Heat Shock Protein 70 Members During Development, Diapause, and Temperature Stress in the Freshwater Crustacean Daphnia magna

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.692517 ◽

2021 ◽

Vol 9 ◽

Author(s):

Luxi Chen ◽

Rocío Gómez ◽

Linda C. Weiss

Keyword(s):

Heat Shock ◽

Family Member ◽

Daphnia Magna ◽

Nuclear Translocation ◽

Protein Localization ◽

Expression Patterns ◽

Family Members ◽

Hsp70 Family ◽

Before And After ◽

Shock Proteins

Dormancy is a lifecycle delay that allows organisms to escape suboptimal environmental conditions. As a genetically programmed type of dormancy, diapause is usually accompanied by metabolic depression and enhanced tolerance toward adverse environmental factors. However, the drivers and regulators that steer an organism’s development into a state of suspended animation to survive environmental stress have not been fully uncovered. Heat shock proteins 70 (HSP70s), which are often produced in response to various types of stress, have been suggested to play a role in diapause. Considering the diversity of the Hsp70 family, different family members may have different functions during diapause. In the present study, we demonstrate the expression of two hsp70 genes (A and B together with protein localization of B) throughout continuous and diapause interrupted development of Daphnia magna. Before and after diapause, the expression of Dmhsp70-A is low. Only shortly before diapause and during diapause, Dmhsp70-A is significantly upregulated and may therefore be involved in diapause preparation and maintenance. In contrast, Dmhsp70-B is expressed only in developing embryos but not in diapausing embryos. During continuous development, the protein of this Hsp70 family member is localized in the cytosol. When we expose both embryo types to heat stress, expression of both hsp70 genes increases only in developing embryos, and the protein of family member B is translocated to the nucleus. In this stress formation, this protein provides effective protection of nucleoplasmic DNA. As we also see this localization in diapausing embryos, it seems that Daphnia embryo types share a common subcellular strategy when facing dormancy or heat shock, i.e., they protect their DNA by HSP70B nuclear translocation. Our study underlines the distinctive roles that different Hsp70 family members play throughout continuous and diapause interrupted development.

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Inhibition of human head and neck squamous cell cancer growth by modulation of heat shock proteins and induction of the mitochondrial apoptotic pathway with withaferin A

Journal of Clinical Oncology ◽

10.1200/jco.2009.27.15_suppl.e17003 ◽

2009 ◽

Vol 27 (15_suppl) ◽

pp. e17003-e17003

Author(s):

M. S. Cohen ◽

B. N. Timmermann ◽

G. O'Donnell ◽

A. K. Samadi

Keyword(s):

Flow Cytometry ◽

Membrane Potential ◽

Heat Shock ◽

Heat Shock Proteins ◽

Cell Viability ◽

Mitochondrial Membrane ◽

Annexin V ◽

Withaferin A ◽

Shock Proteins ◽

Mts Assay

e17003 Background: Epithelial cancers, particularly lung cancer and head and neck squamous cell carcinoma (HNSCC), continue to pose formidable challenges in clinical practice. Novel chemotherapeutic agents have been developed, but even those have limited long-term benefits in the treatment of these tumors, illustrating the need to continue to improve systemic therapy for affected patients. The objective of the present study was to investigate the effect of withaferin A (WA), a plant-derived small molecule, on cancer cell growth, heat shock protein expression and induction of apoptosis in human HNSCCs. Methods: MDA1986 and JMAR HNSCC cells were used in all experiments. The effect of WA on cell viability was determined by MTS assay. Apoptosis and mitochondrial membrane potential changes were assessed by annexin V/propidium iodide and JC-1 staining respectively using standard flow cytometry methods. Effect of WA on modulation of heat shock proteins was determined by Western blot analysis. Results: Withaferin A reduces cell viability in both MDA1986 and JMAR cells with IC50 levels of 265 ± 5 nM by MTS assay, which is 5 fold higher than its reported activity in breast cancer cells. WA completely down-regulates HSP90beta, GRP94, and TRAP-1 expression at 250 nM concentration in HNSCC cells at 24 hours treatment. In addition, WA markedly increased HSP70 levels and mildly increased HSP27 levels in a dose-dependent manner at 24 hours treatment. Flow cytometry with Annexin V/PI staining shows that 5 μM WA treatment for 24 hours induced apoptosis in 63% of MDA1986 and 60% of JMAR cells. WA at 5 μM also reduced mitochondrial membrane potential by JC-1 staining with flow cytometry to less than 10% of controls in both JMAR and MDA1986 cells at 24 hours treatment. Conclusions: Withaferin A is a potent novel inhibitor of HSP90 in human HNSCCs. In addition to HSP modulation, its anticancer mechanistic effects involve apoptotic cell death through the mitochondrial pathway. This molecule shows promise for further in vivo studies to establish preclinical proof of concept as a novel anticancer therapy in this disease. No significant financial relationships to disclose.

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Heat Shock Proteins in Germinating Pollen of Nicotiana tabacum Before and After Heat Shock

Sexual Reproduction in Higher Plants ◽

10.1007/978-3-642-73271-3_44 ◽

1988 ◽

pp. 277-282 ◽

Cited By ~ 3

Author(s):

M. M. A. Van Herpen ◽

W. H. Reijnen ◽

J. A. M. Schrauwen ◽

P. F. M. De Groot ◽

G. J. Wullems

Keyword(s):

Nicotiana Tabacum ◽

Heat Shock ◽

Heat Shock Proteins ◽

Before And After ◽

Shock Proteins ◽

Germinating Pollen

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Carbenoxolone Could Deteriorate Streptozotocin-induced Diabetes through Induction of Heat Shock Protein 70 and IFN-γ in C57BL/6 Mice

Iranian Journal of Allergy Asthma and Immunology ◽

10.18502/ijaai.v17i6.621 ◽

2019 ◽

Author(s):

Mehdi Rasouli ◽

Yaser Jafari-khataylou ◽

Javad Ashrafi-Helan

Keyword(s):

Heat Shock ◽

Interleukin 10 ◽

Positive Control ◽

Diabetic Mice ◽

Before And After ◽

Disease Induction ◽

Streptozotocin Induced Diabetes ◽

Ifn Γ ◽

The Difference ◽

Shock Proteins

Type 1 diabetes (T1D), a spontaneous autoimmune disease, is associated with destruction of insulin-producing β-cells in the pancreas. Since some heat shock proteins (HSP), such as HSP70 exert a protective effect in both tissues and cells, the present study was conducted to elucidate the effects of carbenoxolone (CBX) as an HSP70 inducer on T1D. The disease was induced in male C57BL/6 mice using streptozotocin (STZ) and subjects were allocated to therapeutic 1 and therapeutic 2 groups, as well as negative and positive control groups. The treated mice (therapeutic 1 and therapeutic 2 groups) received 50 mg/kg CBX intraperitoneally every 24 hours, in the therapeutic 1 group the drug was injected before and after disease induction whereas in the therapeutic 2 group the drug was injected only after disease induction. Serum fasting blood sugar (FBS) level, cytokines production (Interferon-gamma (IFN-γ), Interleukin 10 (IL-10), and IL-17), serum HSP70 level and CD4+CD25+Foxp3+ regulatory T cell (Treg) frequency measurements were outperformed 14 days after the last STZ injection. Our results showed that in the treated groups, serum HSP70, IFN-γ, and IL-17 levels were increased in contrast to the untreated groups. The IL-10 level was markedly decreased in comparison to untreated diabetic mice (p<0.05). Moreover, it was found that the frequency of Tregs in treated mice was lower in comparison to the untreated mice but the difference was not significant (p>0.05). Our results confirm that CBX might through HSP70 induction, followed by increasing IFN-γ level leads to suppression of IL-10 production in diabetic mice resulted in toxic effects on pancreatic islet beta cells and deteriorating of disease.

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Apoptotic Effect of Homobrassinin and Thiazino[6,5-b]indol is Associated with Downregulation of Heat Shock Proteins in Human Ovarian Adenocarcinoma Cells

Acta Chimica Slovenica ◽

10.17344/acsi.2020.6281 ◽

2021 ◽

Vol 68 (1) ◽

pp. 151-158

Author(s):

Zuzana Solárová ◽

Martin Kello ◽

Peter Solár

Keyword(s):

Heat Shock ◽

Heat Shock Proteins ◽

Flow Cytometric Analysis ◽

Antimicrobial Properties ◽

Annexin V ◽

Phytopathogenic Fungi ◽

Cytochrome C Release ◽

Ovarian Adenocarcinoma ◽

Apoptotic Effect ◽

Shock Proteins

Phytoalexins are substances with antimicrobial properties produced by plants after being attacked by microorganisms, especially phytopathogenic fungi and viruses. They are also currently being studied for their antitumor effect. We aimed to study the apoptosis-stimulating effect of homobrassinin and thiazino[6,5-b]indol in human ovarian adenocarcinoma A2780 and A2780cis cells via flow cytometric analysis of annexin V/PI, caspase 3 and 9 activity, cytochrome C release, and smac-diablo accumulation. Using the western blot technique, we also monitored the effect of both indoles on the response of heat shock proteins in these cells. Thiazino[6,5-b]indol showed more pronounced sensitizing and/or pro-apoptotic effect compared to homobrassinin accompanied by increased smac-diablo accumulation at earlier time intervals and pronounced externalization of phosphatidylserine at 72 h in A2780cis compared to A2780 cells. The apoptosis stimulating effect of thiazino[6,5-b]indol in A2780cis cells was associated with significant irreversible downregulation of HSP70 and HSP90 and partly with a decrease of HSP40. On the other hand, cisplatin-induced the apoptosis of sensitive A2780 cells with reversible downregulation of HSP40 and HSP57. In conclusion, the effect of thiazino[6,5-b]indol on resistant A2780cis cells could have a great utility in both the potential prevention and the treatment of other cisplatin-resistant tumor cells.

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Action of inhibitors Heat shock proteins 90 and 27 on dexamethasone-induced apoptosis of tumor cells

Bulletin of Siberian Medicine ◽

10.20538/1682-0363-2010-3-68-71 ◽

2010 ◽

Vol 9 (3) ◽

pp. 68-71 ◽

Cited By ~ 1

Author(s):

Ye. V. Kaigorodova ◽

N. V. Ryazantseva ◽

V. V. Novitsky ◽

A. N. Maroshkina ◽

M. V. Belkina ◽

...

Keyword(s):

Cell Death ◽

Heat Shock ◽

Programmed Cell Death ◽

Heat Shock Protein ◽

Tumor Cells ◽

Propidium Iodide ◽

Fluorescent Microscopy ◽

Annexin V ◽

Induced Apoptosis ◽

Shock Proteins

Programmed cell death of tumor cells of line Jurkat in conditions of cultivation with various concentration of dexamethasone, selective inhibitors of Hsp90 (Heat shock protein — Hsp) (17-AAG) and Hsp27 (KRIBB3) was investigated. An estimation of realisation apoptosis spent by method of fluorescent microscopy with use annexin V-FITC and propidium iodide. Inhibition of Hsp90 and Hsp27 leads to activation of tumor cells Jurkat apoptotic program and strengthening of dexamethasone-induced apoptosis. Hsp27 and Hsр90 play an antiapoptotic role in tumor cells of line Jurkat.

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