Effect Of Heat Shock Protein Research Articles

Heat shock protein 22 alleviates doxorubicin-induced kidney injury by suppressing oxidative stress and apoptosis

This study investigated the effects of heat shock protein 22 (HSP22) against doxorubicin (DOX)-induced kidney injury. Mice were randomly assigned to four groups: CON, ad-HSP22, DOX, and ad-HSP22 + DOX. Adeno-associated virus carrying the HSP22 gene (ad-HSP22) was administered via tail vein injection for four weeks, followed by intraperitoneal simulation with DOX (20 mg/kg) for another five days. Upon euthanasia, ELISA, histological staining (H&E, IHC, DHE, and TUNEL), and western blot analyses were employed to assess relevant markers. Serum biomarkers of kidney injury, SCr, and BUN, were upregulated after DOX administration but normalized with HSP22 overexpression. Pathological changes induced by DOX were also reversed by HSP22 overexpression in H&E, IHC, DHE, and TUNEL stains. DOX-induced upregulation of NOX-2 and NOX-4 and downregulation of SOD-1 and SOD-2 were reversed by HSP22 overexpression. Similarly, DOX-induced increases in Bax and decrease in Bcl-2 were attenuated by HSP22 overexpression. The study further demonstrated that the Nrf2/HO-1 signaling pathway was activated by HSP22 overexpression. In vitro experiments corroborated the findings from in vivo experiments. In conclusion, HSP22 alleviates DOX-induced kidney injury by suppressing oxidative stress and apoptosis, primarily through the activation of the Nrf2/HO-1 signaling pathway. These results suggest HSP22 as a potential therapeutic target for DOX-induced kidney injury.

PSLBI-15 Effects of heat shock protein 70 on cell viability and protective capacity of serum against histone cytotoxicity

Abstract Bovine respiratory disease (BRD) is the most significant health problem associated with newly received cattle in the beef industry. Serum concentrations of extracellular histones increase in a variety of diseases and histones are known to be cytotoxic to mammalian cells. Work in our lab indicated that failure to protect against cytotoxic effects of extracellular histones may be a component of severe cases of respiratory disease in cattle. The mechanisms or proteins that neutralize cytotoxic effects of extracellular histones are not fully elucidated, but C-reactive protein and complement activity have been associated with a reduced cytotoxic effect of extracellular histones. Work in the literature suggests that heat shock protein 70 (HSP70) concentrations in the serum is increased in disease and may directly activate complement. Unpublished observations in our lab indicated that HSP70 potentially interacts with histones to exacerbate cytotoxicity leading to our hypothesis that increased concentrations of HSP70 will decrease the protective capacity against histone cytotoxicity. Therefore, the objectives of the current study were to evaluate serum concentrations of HSP70 in weaned bulls entering the feedlot and the effects of increasing HSP70 concentrations on histone cytotoxicity in vitro. Weaned seedstock bulls (n = 130) composed of various breeds were fed using the GrowSafe System at the Tucumcari Bull Test for 122 d. At receiving, blood samples were obtained via coccygeal venipuncture, centrifuged, and serum immediately frozen on dry ice. Serum was analyzed for complement activity (CH50 assay) and protective capacity against extracellular histones using our novel histone toxicity assay. The ratio value of histone toxicity to CH50 was used to separate a subset of bulls as protective (n = 10) and non-protective (n = 10). Concentrations of HSP70 were quantified utilizing a commercially available ELISA kit. Data were analyzed using the GLM procedure of SAS. Serum concentrations of HSP70 were similar (P > 0.10) for protective and non-protective bulls. To test the responses to increased concentrations of HSP70 to mimic disease, protective and non-protective serum was tested against histones alone and in combination with 0.5 or 40 ng/mL recombinant bovine HSP70. Complement activity from protective and non-protective bulls was not influenced (P > 0.10) in the presence and absence of recombinant bovine HSP70. Exogenous HSP70 at 0.5 or 40 ng/mL did not influence protective capacity in non-protective animals (P = 0.95). However, HSP70 at 0.5 ng/mL did not (P = 0.18) reduce protective capacity and reduced protective capacity (P < 0.01) at 40 ng/mL in serum from bulls that were assigned to the protective group. Results indicate that weaned bulls entering the feedlot display varied capacity to protect against histone toxicity and increased concentrations of HSP70 can reduce the protective capacity indicating that circulating HSP70 may play a role in bovine respiratory disease pathogenesis.

Effects of cold stress on protein metabolism of certain walnut cultivars

To investigate the effects of heat shock proteins (HSPs) on walnut (Juglans regia L.) plants under low-temperature stress, first of all, low-temperature tolerances of ten walnut cultivars (Chandler, Fernor, Franquette, Pedro, Bilecik, Kaman-I, Kaman-II, Kaman-III, Sebin, and Yalova I) were determined. One-year-old shoot samples were taken from the plants in two different periods, cold-acclimated (CA) and non-acclimated (NA), and were exposed to +5°C, -5°C, -15°C and -25°C for 12 h. Cold injury was determined by ion leakage analysis in the thawed bark tissues. According to the results of this analysis, two cultivars were determined as cold-sensitive (Chandler) and cold-tolerant (Fernor) in terms of low-temperature tolerance. To examine the effects of HSPs on cold tolerance in walnut plants, the protein profiles, the amount of total protein, and the HSPs of these two cultivars were determined. As a result of the immunoblot analysis, it was determined that 44 kDa HSP23 and 59 kDa HSP60 are responsible for low-temperature tolerance in walnut plants.

Molecular mechanism of anticancer effect of heat shock protein 90 inhibitor BIIB021 in human bladder cancer cell line

Bladder cancer is a type of urologic malignancy that exhibits significant morbidity, mortality, and treatment costs. Inhibition of heat shock protein 90 (HSP90) activity has been a promising pharmacological strategy for blocking of bladder cancer pathogenesis. BIIB021 is a next-generation HSP90 inhibitor which interrupts ATP hydrolysis process of HSP90 and inhibits the stabilization and correct folding of client proteins. In current study, we aimed to investigate the molecular mechanism of the anticancer activity of BIIB021 in human bladder cancer T24 cells. Our results revealed that nanomolar concentration of BIIB021 decreased viability of T24 cell. BIIB021 downregulated HSP90 expression in T24 cells and inhibited the refolding activity of luciferase in the presence of T24 cell lysate. PCR array data indicated a significant alteration in transcript levels of cancer-related genes involved in metastases, apoptotic cell death, cell cycle, cellular senescence, DNA damage and repair mechanisms, epithelial-to-mesenchymal transition, hypoxia, telomeres and telomerase, and cancer metabolism pathways in T24 cells. All findings hypothesize that BIIB021 could exhibit as effective HSP90 inhibitor in the future for treatment of bladder cancer patients.

Therapeutic potential of earthworm-derived proteins: Targeting NEDD4 for cardiovascular disease intervention

Targeting neuronal precursor cell-expressed developmentally down-regulated 4 (NEDD4) offers a promising strategy for cardiovascular therapies. NEDD4, a ubiquitin ligase enzyme, is crucial in protein degradation and cellular signaling. Earthworms (Lumbricus genus) are noteworthy for their rich biochemical composition and pharmacological properties. This study investigated the interactions between Lumbricus-derived proteins and NEDD4 to identify potential cardiovascular therapeutic candidates. Using advanced computational techniques, including 3D structure modeling, protein-protein docking simulations, 100 ns molecular dynamics (MD) simulations, and Molecular mechanics/Poisson–Boltzmann surface area (MM/PBSA) calculations, we assessed the binding affinities and functional impacts of these proteins on NEDD4 activity. The findings indicated that Lumbricus-derived proteins such as Lumbrokinase, heat shock protein, and elongation factor 1 (EF-1)-alpha showed activities similar to standard antagonists in modulating NEDD4. These results aligned with previous studies showing the inhibitory effects of heat shock protein and EF-1 on NEDD4 ubiquitination and ligase activity. Additionally, MM/PBSA calculations revealed favorable binding free energies for these compounds, underscoring their therapeutic potential in cardiovascular diseases. In conclusion, this study highlighted the potential of Lumbricus-derived compounds in cardiovascular disease therapy via the NEDD4 pathway, warranting further biochemical and preclinical validation and exploring broader therapeutic applications.

Inhibition of Hsp110-STAT3 interaction in endothelial cells alleviates vascular remodeling in hypoxic pulmonary arterial Hypertension model

BackgroundPulmonary arterial hypertension (PAH) is a progressive and devastating disease characterized by pulmonary vascular remodeling which is associated with the malignant phenotypes of pulmonary vascular cells. Recently, the effects of heat shock protein 110 (Hsp110) in human arterial smooth muscle cells were reported. However, the underlying roles and mechanisms of Hsp110 in human pulmonary arterial endothelial cells (HPAECs) that was disordered firstly at the early stage of PAH remain unknown.MethodsIn this research, the expression of Hsp110 in PAH human patients and rat models was investigated, and the Hsp110 localization was determined both in vivo and in vitro. The roles and mechanism of elevated Hsp110 in excessive cell proliferation and migration of HPAECs were assessed respectively exposed to hypoxia. Small molecule inhibitors targeting Hsp110-STAT3 interaction were screened via fluorescence polarization, anti-aggregation and western blot assays. Moreover, the effects of compound 6 on HPAECs abnormal phenotypes in vitro and pulmonary vascular remodeling of hypoxia-indued PAH rats in vivo by interrupting Hsp110-STAT3 interaction were evaluated.ResultsOur studies demonstrated that Hsp110 expression was increased in the serum of patients with PAH, as well as in the lungs and pulmonary arteries of PAH rats, when compared to their respective healthy subjects. Moreover, Hsp110 levels were significantly elevated in HPAECs under hypoxia and mediated its aberrant phenotypes. Furthermore, boosted Hsp110-STAT3 interaction resulted in abnormal proliferation and migration via elevating p-STAT3 and c-Myc in HPAECs. Notably, we successfully identified compound 6 as potent Hsp110-STAT3 interaction inhibitor, which effectively inhibited HPAECs proliferation and migration, and significantly ameliorated right heart hypertrophy and vascular remodeling of rats with PAH.ConclusionsOur studies suggest that elevated Hsp110 plays a vital role in HPAECs and inhibition of the Hsp110-STAT3 interaction is a novel strategy for improving vascular remodeling. In addition, compound 6 could serve as a promising lead compound for developing first-in-class drugs against PAH.

Hsp90 is a potential risk factor for ovarian cancer prognosis: an evidence of a Chinese clinical center

BackgroundThe potential treatment effects of heat shock protein 90 (Hsp90) inhibitors in ovarian cancer (OC) are controversial. This research aims to investigate the relationship between the level of Hsp90 in peripheral blood and the prognosis of OC patients, as well as the clinicopathological indicators.Materials and methodsWe retrospectively collected the clinicopathological indicators of OC patients who were admitted to the Department of Obstetrics and Gynecology of the First Affiliated Hospital of Anhui Medical University from 2017 to 2022. Hsp90 level in patient blood was detected by enzyme-linked immunosorbent assay, and the correlation between Hsp90 level and OC prognosis was systematically investigated. Kaplan–Meier method was used to draw the survival curve, and the average survival time and survival rate were calculated. The log-rank test and Cox model were used for univariate survival analysis, and the Cox proportional hazards model was applied for multivariate survival analysis. Based on the TCGA dataset of OC obtained by cBioPortal, Pearson’s correlation coefficients between Hsp90 level values and other mRNA expression values were calculated to further conduct bioinformatics analysis. GSEA and GSVA analysis were also conducted for gene functional enrichment. The expression of Hsp90 in OC tissues were evaluated and compared by Immunohistochemical staining.ResultsAccording to the established screening criteria, 106 patients were selected. The enzyme-linked immunosorbent assay results showed that 50.94% OC patients with abnormal Hsp90 level. According to the outcome of Kaplan–Meier curves, the results revealed that the abnormal level of Hsp90 was suggested to poor prognosis (P = 0.001) of OC patients. Furthermore, the result of multivariate Cox proportional hazards regression model analysis also predicted that abnormal Hsp90 level (HR = 2.838, 95%CI = 1.139–7.069, P = 0.025) was linked to poor prognosis, which could be an independent prognostic factor for the prognosis of OC patients. Moreover, top 100 genes screened by Pearson’s value associated with Hsp90, indicating that Hsp90 participated in the regulation of ATF5 target genes, PRAGC1A target genes and BANP target genes and also enriched in the metabolic processes of cell response to DNA damage stimulus, response to heat and protein folding.ConclusionHsp90 level is positively associated with OC mortality and is a potential prognostic indicator of OC.

Heat shock protein 70 reflected the state of inhabited fish response to water quality within lake ecosystem

Heat shock proteins are natural biomarker that plays a vital role in genetic variability and adaptive survival in organisms under environmental stress. We assessed the effect of the stressor using Heat shock protein 70 in Hemibarbus labeo and Lepomis macrochirus at three geological points of Juam and Yedang lake. The study focused on the stress-relieving effect of heat shock protein 70 expressed in fish associated with physicochemical factors in the lake ecosystem. We recorded significant upregulation (p < 0.05) of the heat shock protein 70 gene in L. macrochirus in Juam (0.99-fold) and Yedang (1.44-fold). The upregulation in H. labeo indicated 0.83-fold in Juam and 1.21-fold in Yedang. The water quality index determined using physicochemical factors was low in the middle regions of both lakes. However, the maximum gene expression distribution was found in Yedang. The two-way clustering analysis revealed ammonia, total phosphorus, and suspended solids with maximum stress threshold. Dissolved oxygen, pH, nitrate, and organic carbon were the stress factors with minimal concentrations induced expression. The key observations are (i) the vital influence of molecular markers regulated tolerance at initial stress levels, (ii) the synergistic effect of the stressors and Heat shock protein 70 induction impacted adaptive strategy in fishes to regain tolerance (iii) fish population (L. macrochirus) with elevated response exhibited maximum distribution, correlating with the practical scenario. Heat shock protein 70 expressions through metabolic compensation strategy thus influenced the size of inhabited fish population retarding biodiversity of the lake ecosystem.

Effects of heat shock protein 90 on complement activation in myocardial ischemia/reperfusion injury after pioglitazone preconditioning.

Heat shock protein 90 (HSP90) appears to have a pivotal function in ischemic preconditioning. Pioglitazone preconditioning (PioC) attenuates myocardial ischemia/reperfusion (I/R) injuries. The current study aims to investigate the role of HSP90, complement C3 and C5a, and nuclear factor kappa-B (NF-κB) in PioC-induced cardioprotection. A total of 80 rats were randomly categorized into 4 groups, as follows: sham, I/R, PioC, and PioC+HSP90 inhibitor geldanamycin (PioC+GA). The sham group rats had a thoracotomy, in which the ligature was passed by the heart with no ligation (150 min). The other 3 groups were exposed to ischemia (30 min) followed by reperfusion (2 h). In the PioC group, pioglitazone (3 mg/kg) was administered intravenously 24 h before ischemia. In the PioC+GA group, after being pretreated with pioglitazone, GA was administered (intraperitoneally, 1 mg/kg) 30 min before ischemia. Myocardial infarct sizes (ISs), apoptosis rates, creatine kinase-MB (CK-MB), lactate dehydrogenase (LDH), and cardiac troponin I (cTnI) serum levels were determined. The HSP90, C3, NF-κB, C5a, B-cell lymphoma-2 (Bcl-2), and Bax expression levels, as well as interleukin (IL)-1β, IL-6, intercellular cell adhesion molecule-1 (ICAM-1), and tumor necrosis factor alpha (TNF-α) mRNA levels were measured. The myocardial ISs, serum CK-MB, cTnI and LDH levels, apoptosis rates, IL-1β, IL-6, TNF-α, ICAM-1 release, as well as Bax, C5a, C3, and NF-κB protein expression were considerably lower in the PioC group than in the I/R group (p < 0.05). The Bcl-2 and HSP90 expression was higher in the PioC group than in the I/R group (p < 0.05). Geldanamycin inhibited the effects of PioC. These data strongly demonstrate that the PioC-induced is dependent upon HSP90 activity. The HSP90 is indispensable for PioC-mediated cardioprotection. The HSP90 attenuates I/R-induced ISs, apoptosis of cardiomyocytes and myocardial inflammation through C3, C5a and NF-κB activation inhibition.

Effects of heat shock protein 90 on complement activation of myocardial ischemia/reperfusion injury in pioglitazone preconditioning

Effects of heat shock protein 90 on complement activation of myocardial ischemia/reperfusion injury in pioglitazone preconditioning

Effects of heat shock protein 90 on complement activation of myocardial ischemia/reperfusion injury in pioglitazone preconditioning

Effects of heat shock protein 90 on complement activation of myocardial ischemia/reperfusion injury in pioglitazone preconditioning

Effects of heat shock protein 90 on complement activation of myocardial ischemia/reperfusion injury in pioglitazone preconditioning

Effects of heat shock protein 90 on complement activation of myocardial ischemia/reperfusion injury in pioglitazone preconditioning

Effects of heat shock protein 90 on complement activation of myocardial ischemia/reperfusion injury in pioglitazone preconditioning

Effects of heat shock protein 90 on complement activation of myocardial ischemia/reperfusion injury in pioglitazone preconditioning

Effects of heat shock protein 90 on complement activation in myocardial ischemia/reperfusion injury after pioglitazone preconditioning

Effects of heat shock protein 90 on complement activation in myocardial ischemia/reperfusion injury after pioglitazone preconditioning

Unfolded protein response in endothelial injury

ABSTRACT Endothelial barrier dysfunction is associated with sepsis and lung injury, both direct and indirect. We discuss the involvement of unfolded protein response in the protective effects of heat shock protein 90 inhibitors and growth hormone releasing hormone antagonists in the vascular barrier, to reveal new possibilities in acute respiratory distress syndrome treatment.

Study on the role of heat shock protein 90 (HSP90) gene in chicken preadipocytes proliferation and differentiation

In this study, we examined the effects of Heat Shock Protein 90 (HSP90) on adipocyte proliferation and differentiation in chickens. To achieve this, we constructed RNA interference (RNAi) vectors to target HSP90 and transfected the vectors into primary adipocytes. After transfection, oil red O staining was performed to determine the status of triglyceride accumulation in the cells, whereas the CCK-8 cell kit and 5-Ethynyl-2'-Deoxyuridine (EdU) assays were used to determine cell proliferation. Thereafter, the mRNA and protein expression levels of PPARγ, FAS, SREBP-1c, and HSP90 were determined, and the results showed that after the interference of HSP90, the mRNA and protein expression levels of HSP90 in the chicken adipocytes decreased significantly compared to the control and blank groups (p < 0.05). The decreased mRNA and protein expression of PPARγ, FAS, and SREBP-1c was related to adipocyte differentiation (p < 0.05). However, HSP90 interference had no effect on adipocyte proliferation (p > 0.05). Taken together, the results of this study showed that HSP90 influenced the expression of PPARγ and adipose-differentiation-related genes, thereby regulating triglyceride accumulation and adipocyte differentiation in chickens.

Climate resilience in small ruminant and immune system: An old alliance in the new sustainability context

Physiological and immunological responses of small ruminants are affected by heat stress in the attempt to recover animals’ thermal and cellular homeostasis. This review will report scientific investigations related to the complex mechanisms triggered by small ruminants to cope with heat stress that causes perturbation at different level and organ, including endocrinal and cellular response with a bidirectional crosstalk. In particular, during heat stress exposition the role of hormones (prolactin, triiodothyronine, thyroxine, and cortisol), the immune cell reactivity, the initiation of the inflammatory response in terms of cytokines profile, the heat shock cellular response, and oxidative stress will be discussed. Some critical issues for the evaluation of biomarkers of stress resiliency and thermo-tolerance at endocrinal level, immune cell response and heat shock cellular response will be examined. A scientific debate on the effect of heat shock proteins and their use as biomarker of thermo-tolerance for future breeding programs in a context of climate resilient ruminant production will be proposed. Moreover, based on the latest advanced comprehension of the mechanisms involved in heat stress cellular regulation, the circulating miRNAs will be introduced as promising biomarkers for their potential crosstalking with the other inflammatory biomarkers.

Heat Shock Protein 70 Inhibition Effect on Inflammatory Receptors via Nuclear Factor Kappa B on Melanoma Cell Lines, A-375 and C8161

The aim of this study was evaluation of effect of heat shock protein 70 on toll-like receptor 4 and toll-like receptor 2 in melanoma cell line, A-375. The melanoma cell lines were stimulated in time course with heat shock protein 70 protein and incubation was performed. After 48 h, the cells were collected and total messenger ribonucleic acid was extracted and the expression of toll-like receptor 2 and toll-like receptor 4 was determined by quantitative reverse transcription polymerase chain reaction. The results showed that the appearance of toll-like receptor 2, toll-like receptor 4 in melanoma cell line, A-375 decreased in exposure to heat shock protein 70. The results showed the expression of nuclear factor kappa B in melanoma cell line. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide evaluation results of melanoma cell death were compared at different times and statistical analysis showed a significant difference (p<0.001). The role of heat shock protein 70 has been identified as a tumor suppressor in melanoma cancer. Heat shock protein 70 is a new treatment for melanoma in the near future. Heat shock protein 70 can be considered as a potential target in gene therapy for patients with melanoma cancer and the advantages of gene therapy by heat shock protein 70 in this type of cancer over chemotherapy can reduce cytotoxic effects and its low invasiveness.

Effects of heat shock protein 70 knockdown on the tolerance of the brine shrimp Artemia franciscana to aquaculture-related stressors: Implications for aquatic animal health and production

Temperature, pH, salinity and ammonia stress adversely affect the performance and well-being of aquatic organisms, especially those used in aquaculture, a major food production sector. Management strategy to mitigate stress caused by abiotic factors in farmed aquatic organisms is scarce but is highly warranted considering the negative impact of these factors on the welfare and economic production of the farmed animals. The induction of heat shock proteins, mainly 70 kDa Hsp (Hsp70), in farmed fish and shellfish, including the live feed brine shrimp Artemia franciscana, by exposure to a non-lethal heat shock (NLHS) of 37 °C for 30 min followed by 6 h recovery period caused a significant improvement in the tolerance of the animals towards subsequent challenge with an abiotic stress. Although the protective effect of NLHS was associated with enhanced Hsp70 accumulation in these species, the induction of other members of Hsp and their collective involvement in the stress tolerance could not be ignored. This triggered an argument as to whether only Hsp70 or its combination with other Hsps was responsible for inducing stress tolerance phenotype in these species. In this study, we employed RNAi technology in the biological model organism Artemia to gain insight into the functional role of Hsp70 in stress tolerance. The results showed that upon exposure to a NLHS, the wild-type Artemia nauplii capable of producing Hsp70 exhibited significantly higher survival than the unexposed control in all the abiotic stress tolerance tests performed in this study. However, the Hsp70-deficient mutant nauplii did not exhibit increased stress-tolerant phenotypes as the survival percentage of the nauplii decreased by 29, 19, and 15%, respectively, upon challenge with a median lethal concentration of salinity, pH, and ammonia. These results suggest that Hsp70 was involved in inducing tolerance against stress caused by the abiotic factors, and hence can be a potential target for developing strategies to protect economically important aquaculture species from environmental disturbances.

Hsp70 Promotes SUMO of HIF-1α and Promotes Lung Cancer Invasion and Metastasis

Objective This study aims to investigate the effect of heat shock protein-70 (Hsp70) on epithelial-mesenchymal transition (EMT) of lung cancer cells under heat stimulation and to explore its possible molecular mechanism. Methods qRT-PCR and immunohistochemistry assay were used to detect the expression of Hsp70 in lung cancer tissues and adjacent tissues. EdU assay was used to detect the cell activity. The effect of Hsp70 on the migration and invasion of A549 and NCI-H446 cells was detected by the wound-healing assay and Transwell assay. A tumor transplantation animal model was established to detect the effect of overexpression of Hsp70 on proliferation and metastasis of lung cancer cells. Western blot assay was used to detect the effect of thermal stimulation and overexpression of Hsp70 on SUMO modification of HIF-1α. Results The wound-healing rate of A549 and NCI-H446 cells under Hsp70 stimulation was significantly higher than blank control group. At the same time, the number of cells passing through the membrane increased significantly. Hypodermic tumor transplantation in nude mice proved that knockout Hsp70 can inhibit proliferation and metastasis of lung cancer cells. Thermal stimulation upregulated the expression of Hsp70 and promoted SUMO modification of HIF-1α, ultimately promoting the proliferation and metastasis of lung cancer. Inhibition of Hsp70 reverses the effect of thermal stimulation on lung cancer by reducing the SUMO modification of HIF-1α. Conclusion Thermal stimulation can promote EMT in A549 and NCI-H446 cells and promote cell migration and invasion in vitro and in vivo by upregulation of Hsp70. This process is associated with the promotion of SUMO modification of HIF-1α.