Heat shock protein 90 (Hsp90) inhibitor STA-9090 (Ganetespib) ameliorates inflammation in a mouse model of atopic dermatitis.

Molecular chaperones belonging to the heat shock protein 90 (Hsp90) family are implicated in inflammatory processes and described as potential novel therapeutic targets in autoimmune/inflammatory skin diseases. While the pathological role of circulating Hsp90 has been recently proposed in patients with atopic dermatitis (AD), a chronic inflammatory skin disease characterized by intense itching and recurrent skin lesions, studies aimed at investigating the role of Hsp90 as a potential target of AD therapy have not yet been conducted. Here, the effects of the Hsp90 blocker STA-9090 (Ganetespib) applied systemically or topically were determined in an experimental mouse model of dinitrochlorobenzene (DNCB)-induced AD. Intraperitoneal administration of STA-9090 ameliorated clinical disease severity, histological epidermal thickness, and dermal leukocyte infiltration in AD mice which was associated with reducing the scratching behavior in DNCB-treated animals. Additionally, topically applied STA-9090 led to lowered disease activity in AD mice, reduced serum levels of IgE, and up-regulated filaggrin expression in lesional skin samples. Our observations suggest that Hsp90 may be a promising therapeutic target in atopic dermatitis and potentially other inflammatory or autoimmune dermatoses.

HSP90s are required for hypocotyl elongation during skotomorphogenesis and thermomorphogenesis via the COP1-ELF3-PIF4 pathway in Arabidopsis.

Light and temperature are two key environmental signals that share several molecular components that, in turn, regulate plant growth. Darkness and high ambient temperatures promote skoto- and thermomorphogenesis, including stem elongation. Heat shock proteins 90 (HSP90s) facilitate the adaptation of organisms to various adverse environmental stimuli. Here, we showed that HSP90s are required for hypocotyl elongation during both skoto- and thermomorphogenesis. When HSP90s activities are impaired by the knockdown of HSP90s expression or the application of HSP90 inhibitors, the expression levels and protein abundance of PHYTOCHROME-INTERACTING FACTOR 4 (PIF4) markedly decreased. EARLY FLOWERING 3 (ELF3) deficiency was resistant to the inhibition of HSP90s activities. Furthermore, HSP90s interacted with and destabilized ELF3. In the CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1) mutant, the changes in endogenous PIF4 and ELF3 protein levels caused by the inhibition of HSP90s activities were abolished. HSP90s enhanced the interaction between COP1 and ELF3, reduced ELF3 functional effects on PIF4 and modulated hypocotyl elongation during skoto- and thermomorphogenesis. Our results indicated that HSP90s participate in light and temperature signalling via the COP1-ELF3-PIF4 module to regulate hypocotyl growth in changing environments.

Advances in the role of heat shock protein 90 in prostate cancer.

Prostate cancer is one of the most common tumours in adult men and heat shock proteins play an important biological function in prostate cancer as molecular chaperones involved in the pathogenesis, diagnosis, treatment and prognosis of a wide range of tumours. Among them, increased expression of HSP90, a member of the heat shock protein family, is associated with resistance to prostate cancer denervation and can promote tumour resistance, invasion and bone metastasis, thus making prostate cancer more difficult to treat. Therefore, targeting HSP90 in prostate cancer could be a promising strategy for oncology treatment. This paper reviews the structure and function of HSP90, HSP90-mediated denudation resistance in prostate cancer and HSP90-targeted antitumor therapy, with the aim of providing a new theoretical basis for prostate cancer treatment options in the clinical setting.

Proteomic analysis of lysine acetylation reveals that metabolic enzymes and heat shock proteins may be potential targets for DSS-induced mice colitis.

BACKGROUND: Research on acetylation modification and its modification sites will be of great significance for revealing the mechanism of disease and developing new targeted medicines. In this study, we aim to construct a complete atlas of acetylome in the DSS-induced ulcerative colitis mice model (UC model) METHODS: A high-resolution mass spectrometry-based quantitative approach was employed to identify lysine-acetylated proteins and acetylation sites. Bioinformatics analysis and in vitro experiments verified anti-inflammatory effects of HSP90B1-K142ac. RESULTS: 2597 acetylation events and 1914 sites were quantified, highlighting 140 acetylation site changes in the colitis colon tissue. 91 acetylation sites in 75 proteins were up-regulated, and 49 acetylation sites in 39 proteins were down-regulated in the UC models. The differentially acetylated proteins mainly consisted of non-histone proteins located in the cytoplasm and mitochondria. KEGG and protein-protein interaction networks analysis showed that the differentially acetylated proteins were enriched in the TCA cycle, fatty acid metabolism, and protein processing in the endoplasmic reticulum. 68% of the differentially metabolized enzymes have a down-regulated trend in acetylation levels. The acetylation level of lysine 142 in HSP90B1 was found to be obvious in the UC colon, and point mutation of HSP90B1-K142ac would result in the decreasing secretion of TNF-α and IL-2 in LPS-stimulated cultured cells. CONCLUSION: Our work built a complete atlas of acetylome and revealed the potential role of metabolic enzymes and heat shock proteins in DSS-induced colitis.

Identification and Characterization of HSP90 Gene Family Reveals Involvement of HSP90, GRP94 and Not TRAP1 in Heat Stress Response in Chlamys farreri.

Heat shock proteins 90 (HSP90s) are a class of ubiquitous, highly conserved, and multi-functional molecular chaperones present in all living organisms. They assist protein folding processes to form functional proteins. In the present study, three HSP90 genes, CfHSP90, CfGRP94 and CfTRAP1, were successfully identified in the genome of Chlamys farreri. The length of CfHSP90, CfGRP94 and CfTRAP1 were 7211 bp, 26,457 bp, and 28,699 bp, each containing an open reading frame (ORF) of 2181 bp, 2397 bp, and 2181 bp, and encoding proteins of 726, 798, and 726 amino acids, respectively. A transcriptomic database demonstrated that CfHSP90 and CfGRP94 were the primary functional executors with high expression during larval development and in adult tissues, while CfTRAP1 expression was low. Furthermore, all of the three CfHSP90s showed higher expression in gonads and ganglia as compared with other tissues, which indicated their probable involvement in gametogenesis and nerve signal transmission in C. farreri. In addition, under heat stress, the expressions of CfHSP90 and CfGRP94 were significantly up-regulated in the mantle, gill, and blood, but not in the heart. Nevertheless, the expression of CfTRAP1 did not change significantly in the four tested tissues. Taken together, in coping with heat stress, CfHSP90 and CfGRP94 could help correct protein folding or salvage damaged proteins for cell homeostasis in C. farreri. Collectively, a comprehensive analysis of CfHSP90s in C. farreri was conducted. The study indicates the functional diversity of CfHSP90s in growth, development, and environmental response, and our findings may have implications for the subsequent in-depth exploration of HSP90s in invertebrates.

HSP90 chaperones regulate stomatal differentiation under normal and heat stress conditions.

Stomatal development is tightly connected with the overall plant growth, while changes in environmental conditions, like elevated temperature, affect negatively stomatal formation. Stomatal ontogenesis follows a well-defined series of cell developmental transitions in the cotyledon and leaf epidermis that finally lead to the production of mature stomata. YODA signaling cascade regulates stomatal development mainly through the phosphorylation and inactivation of SPEECHLESS (SPCH) transcription factor, while HSP90 chaperones have a central role in the regulation of YODA cascade. Here, we report that acute heat stress affects negatively stomatal differentiation, leads to high phosphorylation levels of MPK3 and MPK6, and alters the expression of SPCH and MUTE transcription factors. Genetic depletion of HSP90 leads to decreased stomatal differentiation rates. Thus, HSP90 chaperones safeguard the completion of distinct stomatal differentiation steps depending on these two transcription factors under normal and heat stress conditions.

Heat shock transcription factor 1 attenuates TNFα-induced cardiomyocyte death through suppression of NFκB pathway.

Heat shock transcription factor 1 (HSF1), which has been identified as an endogenous cardioprotective factor, possesses potent anti-inflammatory effects. However, the underlying mechanisms have not been fully understood yet. In this study, we investigated the effects of HSF1-regulated RelA, a subunit of NFκB on cardiomyocyte death. Cultured cardiomyocytes were transfected with HSF1 plasmid before the treatment of TNFα. Cell death ratio was determined by cell staining. Additionally, the expression of RelA in the cytoplasm and cytonucleus as well as its subcellular location was detected, and the expression of heat shock proteins (HSP70 and HSP90) in the cardiomyocytes was also examined. Not only did TNFα remarkably enhanced cardiac cell death, but also elevated the expressions of intracellular RelA and elicited its translocation. Overexpression of HSF1 effectively attenuated cell death induced by TNFα. Although HSF1 didn't significantly inhibit the intracellular activation of RelA induced by TNFα at an early stage, HSF1 decreased the levels of RelA and the translocation of RelA in the cytoplasm and cell nucleus at late stage. Besides, the expression of HSP70 and HSP90 was significantly increased when HSF1 was overexpressed. These results suggested that HSF1 attenuated cardiomyocyte death via inhibiting activation of RelA as well as preventing its translocation from the cytoplasm to the cytonucleus, which was partially associated with HSP70 and HSP90 up-regulated by HSF1 overexpression.

Expression changes of HSP90α in cardiac muscles in rats with severs hemorrhagic shock by the treatment of different resuscitating fluid / 重庆医学

Objective To explore the expression changes of HSP90αin cardiac muscles and survival rates in rats by using the different fluids to resuscitate the severs hemorrhagic shocked rats ,and provide reference for the clinical treatment of hemorrhagic shock with different resuscitation fluids .Methods Uncontrolled hemorrhagic shock rats model was established ,using lactic acid salinger liquid ,poly peptide injection gelatin ,hypertonic sodium chloride dextran for fluid resuscitation respectively ,and then checked the HSP90αexpression changes and survival rates in rats .Results the expressions of HSP90αin myocardial tissue and the mortality in rats were different after using different resuscitation fluids in severe hemorrhagic shock rats ,difference was statistically significant(P<0 .05) .Conclusion the expression of HSP90α in cardiac muscles of rats could be induced by severe hemorrhagic shock ,the HSP90αexpressed differently and regularly after using different resuscitating fluids ,it implied that the HSP90α played an important role in the hemorrhagic rats cardiac as a regulating fator .

Roles of heat shock protein 90 in the blockage of H2S against cardiomyocyte injuries induced by chemical hypoxia / 中国病理生理杂志

AIM: To explore the roles of heat shock protein 90 (HSP90) in the blockage of hydrogen sulfide (H2S) against chemical hypoxia-mimetic agent (cobalt chloride, CoCl_2)-induced oxidative stress injuries in H9c2 cardiac cell. METHODS: H9c2 cells were treated with CoCl_2 to set up the chemical hypoxia-induced the model of cardiomyocyte injury. Sodium hydrosulfide (NaHS, a H2S donor) was added into medium for 30 min before CoCl_2 treatment. ATP content was detected by high performance liquid chromatogram (HPLC). Mitochondrial membrane potential (MMP) was measured by rhodamine123 (Rh123) staining and photofluorography. The activity of superoxide dismutase (SOD) was observed using a SOD kit. The expression of heme oxygenase-1 (HO-1) was evaluated by Western blotting. RESULTS: CoCl_2 at concentration of 600 μmol/L significantly reduced SOD activity, ATP level and MMP, and enhanced the expression of HO-1 in H9c2 cells. Pretreatment with 400 μmol/L NaHS dramatically inhibited the cytotoxicity induced by CoCl_2, increased SOD activity, ATP level and MMP, decreased HO-1 expression. 17-allylamino-17 demethoxygeldanamycine(17AAG), an inhibitor of HSP90, obviously blocked the inhibitory effect of H2S on the CoCl_2-induced cytotoxicity, reduced the levels of ATP and MMP, increased HO-1 expression. However, no significantly influence on SOD activity was observed. CONCLUSION: HSP90 may mediate the cardioprotection of H2S via inhibiting the oxidative stress induced by chemical hypoxia.

Effects of macrophage migration inhibitory factor on glucocorticoid release and glucocorticoid receptor in rats / 中华麻醉学杂志

Objective To investigate the effects of macmphage migration inhibitory factor (MIF) on glucocorticoid (GC) re]ease and glucocorticoid recer (GR) in mts.Methods Test Ⅰ Thirty-two male SD rats weighing 250-300 g were randomly divided into 4 groups(n=8 each):control group(C),low dose recombinant MIF (rMIF) group (rMIF-L),middle dose rMIF group (rMIF-M) and high dose rMIF group (rMIF-H).The animals received l ml normal saline via the right femoral vein in group C.The animals received rMIF50.100 and 200 ng in l ml normal saline though right femoral vein in group rMIF-L,rMIF-M or rMIF-H respectively.Blood samples were taken from left femoral artery immediately before inection(T0,baseline),and at 5 min,3 h,6 h.12 h and 24 h after injection of rMIF(T1-5) for determination of serum concentration of corticosterone.Test Ⅱ Primary cultured neonate rat(2-3 days)myocardial ceils were randomly divided into 3 groups(n=24 each):group C,group rMIF-L and group rMIF-M.The ceUs in group C,rMIF-L and rMIF-M wefe incubated with DMEM.rMIF 50 ng+DMEM and rMIF 100 ng+DMEM for 3 h respectively.The expression of GR and HsPg0 wag determined by Western blot.ResuBs Test Ⅰ The serum concentration of corticosterone was signifieemily higher in the other 3 groups than in group C at T1-5(P<0.05).The sertlm concentration of corticostemne was significantly increased at T1-5 in group rMIF-L,rMIF-M and rMIF-H compared with the baseline values(P<0.05).Test Ⅱ HSP90 expresion was significantly lower in the other two groups than in group C(P<0.05).Them was rio signifieanf difference in HSP90 expression between group rMIF-L and group rMIF-M(P>0.05).There was no significant difference in GR expression among the 3 groups ( P > 0.05). Conclusion MIF druing sepsis can weaken GR function through down-regulating HSP9O expression, resulting in CC resistance.

The killing effect of cytotoxic T lymhpocytes on esophageal adenocarcinoma cells mediated by gp96-peptide complexes / 北京大学学报(医学版)

Objective: To study the immunotherapeutic effect on the esopgageal adenocarcinoma mediated by gp96-peptide complexes isolated from the same kind of tumor. Methods: gp96- peptide complexes were purified from nude mice tumors burdened by subcutaneous injection of human esophageal adenocarcinoma cell line SEG-1 . gp96-peptide complexes were carried by the dendritic cells(DC) induced from human peripheral blood mononuclear cells to prepare gp96-DC vaccine. The proliferation of lymphocytes was tested with trypan-blue stain. The quantity of interferon-?(IFN-?) released from cytotoxic T lymphocytes (CTL) was detected with ELISA method. The killing effect of CTL on target cell SEG-1 was measured with MTT. Results: We obtained 120 ?g gp96 from 55 g tumor tissue. DC, gp96, and gp96-DC all could elicit the proliferation of lymphocytes and make them becoming into CTL which released IFN-? and showed different degrees of killing effect on target cell SEG-1. gp96-DC has the strongest eliciting effect among them. At the ratio of E(effect) to T(target) as 40∶1,the killing rate was 68%.No significant difference between the effects of CTL induced by DC alone and of lymphocytes without specific antigen on SEG-1 and K562 cells. Conclusion: The gp96-peptide complexes from tumors can improve the effect of eliciting lymphocyte proliferation of DC and make the lymphocyte becoming into CTL more effectively.These CTLs show prominent killing effect on the target tumor cells.

Tissue-specific expression of an hsc90 gene and nuclear translocation of the HSC90-related protein during amphibian embryogenesis.

Expression and distribution of a constitutive member of the 90 kDa heat-shock protein family, named HSC90, was investigated during amphibian embryonic development. By Northern blot analysis, two hsp90 transcripts (2.5 and 3 kb) which displayed differing developmental regulation were detected during embryogenesis. Expression of the larger transcript (3 kb), which encodes an HSC90-related protein, decreased until the gastrula stage. However, zygotic transcription for this hsc90 gene was found to start from the neurula stage, and the corresponding zygotic hsc90 transcript was specifically located by whole mount in situ hybridization in the anterior neural tube of a late neurula embryo. Later, in a tailbud embryo, hsc90 transcripts were detected in the cephalic region, neural tube, eye vesicles, branchial and mandibular arches and somites. Distribution of the HSC90-related protein was also analysed by immunohistochemistry throughout embryogenesis. As expected, the protein was strongly expressed in the cytoplasm, mainly in the periplasmic area of embryonic tissue cells. Interestingly, HSC90 was also transiently detected in the nuclear area, with this nuclear transfer depending on the chromatin condensation state, up to the blastula stage. During the process of gastrulation, nuclear translocation of HSC90 was also observed at the level of the blastopore dorsal lip, exclusively in cells undergoing invagination.

Establishment of HSP90 overexpressing cell line and effects of HSP90 overexpressing on cell stress responses / 中国病理生理杂志

AIM: To establish a HSP90 highly expressing cell line and study the effect of high level of HSP90 on cell stress response. METHODS: The recombined plasimid pSmycHSP, which contains the full length DNA coding for human HSP90?, was introduced into mouse fibroblast cell line NIH-3T3 by electroporation after being subcloned, purified and identified by limited enzyme digestion. Screened by G418, the positive clones were selected and identified by immunofluorescence, immunocytochemistry and Western-blotting. NIH-3T3 cells transfected with empty plasmid served as control, hyperthermia(44 ℃, 20 min, 40 min)was used to simulate oxidative stress. The activity of lactate dehydrogenase (LDH) in supernatant and damage of DNA were detected by automatic biochemistry analyzer and flow cytometer separately to analyze the effect of high-level HSP90 on cell membrane and DNA injuries under stress condition. RESULTS: The rising level of HSP90 was shown by immunofluorescence, immunocytochemistry and Western-blotting in HSP90 overexpressing cell line. There was no difference in the leakage of LDH between HSP90 overexpressing cell line and control, but the damage of DNA was more severe at 44 ℃for 20 min in HSP90 overexpressing cell line than control. Compared with control, the above indices were relieved at 44 ℃ for 40 min in HSP90 overexpressing cell line. CONCLUSION: The NIH-3T3 derived cell line, which stably expressed high level of HSP90, was established. The effect of high-level HSP90 on cells is complex at different intensity of stress, and the protection may be shown at more severe stress circumstance.

Role of heat shock protein 90 and tubulin in oxidative stress preconditioning / 中国病理生理杂志

AIM: To find the role of heat shock protein 90(HSP90) and tubulin in oxidative stress preconditioning in HepG2 cells.METHODS: The different doses of H2O2 were used to induce cell injury in HepG2 cells.MTT assay,Western blotting and confocal laser microscopy were also used.RESULTS: MTT colorimetry showed that preconditioning(50 mmo1/L H2O2) provided a temporary resistance against subsequent oxidative stress(500 mmol/L H2O2).Western blotting demonstrated that preconditioning increased the levels of HSP90 and tubulin in HepG2 cells,and lessen the declining of HSP90 and tubulin after stress.Tubulin and HSP90's colocalizations in cells with different doses of H2O2 were also observed under laser scanning confocal microscope.CONCLUSION: Tubulin might play important role in oxidative stress preconditioning in HepG2 cells by combining with HSP90.