Background:Glioblastoma is one of the most common primary intracranial tumors of the central nervous system in adults.Although chemotherapy is an important component of glioblastoma treatment,its effectiveness remains...Background:Glioblastoma is one of the most common primary intracranial tumors of the central nervous system in adults.Although chemotherapy is an important component of glioblastoma treatment,its effectiveness remains unsatisfactory.Due to multiple immunosuppressive mechanisms,glioblastoma immunotherapy has not been effective in treating many patients as a result of the clinical breakthroughs in the field.Therefore,the development of cancer immunotherapy relies on the understanding of how tumors interact with the immune system and the analysis of their molecular determinants.This study identified the key interactions between immune cells in the glioma microenvironment using RNA microarrays and single-cell sequencing.Methods:First,we screened differentially expressed genes in tumor and control samples from GSE29796 and GSE50161 datasets using GEO2R.All differentially expressed genes were used to perform enrichment analysis and construct protein-protein interaction topological analysis to analyze the interaction between proteins.Using single-cell RNA sequencing data from the GSE162631 database,we identified immune cell types within the glioblastoma microenvironment,and validated the hub gene expression in these cells.In addition,based on the GEPIA and TIMER databases,hub genes were investigated and compared with immune infiltration to determine differential expression.Finally,CellChat was used to visualize the gene expression distribution and cell-to-cell communication analysis of the proteins between different types of cells.Results:We found that monocytes/macrophages may communicate with each other in the tumor microenvironment through MIF-(CD74+CXCR4)and MIF-(CD74+CD44).In addition,our study indicated that celastrol has the ability to inhibit inflammatory factors expression by MIF/CD74 signaling pathway in U87 cells.Conclusion:This study improved the effectiveness of cancer immunotherapy strategies and developed new ideas for immunotherapy that can be applied to glioblastoma.展开更多
Stroke is the second leading cause of death worldwide,and oxidative stress plays a crucial role.Celastrol exhibits strong antioxidant properties in several diseases;however,whether it can affect oxidation in cerebral ...Stroke is the second leading cause of death worldwide,and oxidative stress plays a crucial role.Celastrol exhibits strong antioxidant properties in several diseases;however,whether it can affect oxidation in cerebral ischemic-reperfusion injury(CIRI)remains unclear.This study aimed to determine whether celastrol could reduce oxidative damage during CIRI and to elucidate the underlying mechanisms.Here,we found that celastrol attenuated oxidative injury in CIRI by upregulating nuclear factor E2-related factor 2(Nrf2).Using alkynyl-tagged celastrol and liquid chromatography-tandem mass spectrometry,we showed that celastrol directly bound to neuronally expressed developmentally downregulated 4(Nedd4)and then released Nrf2 from Nedd4 in astrocytes.Nedd4 promoted the degradation of Nrf2 through K48-linked ubiquitination and thus contributed to astrocytic reactive oxygen species production in CIRI,which was significantly blocked by celastrol.Furthermore,by inhibiting oxidative stress and astrocyte activation,celastrol effectively rescued neurons from axon damage and apoptosis.Our study uncovered Nedd4 as a direct target of celastrol,and that celastrol exerts an antioxidative effect on astrocytes by inhibiting the interaction between Nedd4 and Nrf2 and reducing Nrf2 degradation in CIRI.展开更多
Hepatocellular carcinoma(HCC)is one of most common and deadliest malignancies.Celastrol(Cel),a natural product derived from the Tripterygium wilfordii plant,has been extensively researched for its potential effectiven...Hepatocellular carcinoma(HCC)is one of most common and deadliest malignancies.Celastrol(Cel),a natural product derived from the Tripterygium wilfordii plant,has been extensively researched for its potential effectiveness in fighting cancer.However,its clinical application has been hindered by the unclear mechanism of action.Here,we used chemical proteomics to identify the direct targets of Cel and enhanced its targetability and antitumor capacity by developing a Cel-based liposomes in HCC.We demonstrated that Cel selectively targets the voltage-dependent anion channel 2(VDAC2).Cel directly binds to the cysteine residues of VDAC2,and induces cytochrome C release via dysregulating VDAC2-mediated mitochondrial permeability transition pore(mPTP)function.We further found that Cel induces ROS-mediated ferroptosis and apoptosis in HCC cells.Moreover,coencapsulation of Cel into alkyl glucoside-modified liposomes(AGCL)improved its antitumor efficacy and minimized its side effects.AGCL has been shown to effectively suppress the proliferation of tumor cells.In a xenograft nude mice experiment,AGCL significantly inhibited tumor growth and promoted apoptosis.Our findings reveal that Cel directly targets VDAC2 to induce mitochondria-dependent cell death,while the Cel liposomes enhance its targetability and reduces side effects.Overall,Cel shows promise as a therapeutic agent for HCC.展开更多
Background: Sepsis involves life-threatening organ dysfunction and is caused by a dysregulated host response to infection. No specific therapies against sepsis have been reported. Celastrol(Cel) is a natural anti-infl...Background: Sepsis involves life-threatening organ dysfunction and is caused by a dysregulated host response to infection. No specific therapies against sepsis have been reported. Celastrol(Cel) is a natural anti-inflammatory compound that shows potential against systemic inflammatory diseases. This study aimed to investigate the pharmacological activity and molecular mechanism of Cel in models of endotoxemia and sepsis.Methods: We evaluated the anti-inflammatory efficacy of Cel against endotoxemia and sepsis in mice and macrophage cultures treated with lipopolysaccharide(LPS). We screened for potential protein targets of Cel using activity-based protein profiling(ABPP). Potential targets were validated using biophysical methods such as cellular thermal shift assays(CETSA) and surface plasmon resonance(SPR). Residues involved in Cel binding to target proteins were identified through point mutagenesis, and the functional effects of such binding were explored through gene knockdown.Results: Cel protected mice from lethal endotoxemia and improved their survival with sepsis, and it significantly decreased the levels of pro-inflammatory cytokines in mice and macrophages treated with LPS(P <0.05). Cel bound to Cys424 of pyruvate kinase M2(PKM2), inhibiting the enzyme and thereby suppressing aerobic glycolysis(Warburg effect). Cel also bound to Cys106 in high mobility group box 1(HMGB1) protein, reducing the secretion of inflammatory cytokine interleukin(IL)-1β. Cel bound to the Cys residues in lactate dehydrogenase A(LDHA).Conclusions: Cel inhibits inflammation and the Warburg effect in sepsis via targeting PKM2 and HMGB1 protein.展开更多
Background:Diabetic nephropathy(DN)is a serious complication of diabetes with rising prevalence worldwide.We aimed to explore the anti-DN mechanisms of the compound celastrol derived from the medicinal plant Tripteryg...Background:Diabetic nephropathy(DN)is a serious complication of diabetes with rising prevalence worldwide.We aimed to explore the anti-DN mechanisms of the compound celastrol derived from the medicinal plant Tripterygium wilfordii.Methods:Celastrol-related targets were obtained from Herbal Ingredients’Targets(HIT)and GeneCards databases.DN-related targets were retrieved from GeneCards,DisGeNET,and Therapeutic Targets Database(TTD).A Protein-protein interaction(PPI)network was established using the Search Tool for the Retrieval of Interacting Genes(STRING)database.Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analyses were performed using ClusterProfiler.The cytoHubba plugin was used to select the top 10 hub targets.Molecular docking was performed employing PyMOL and AutoDock software.Cell counting kit-8(CCK-8)and flow cytometry assays were used to detect the viability and apoptosis of NRK-52E cells,respectively.The mRNA expression levels of mitogen-activated protein kinase 3(MAPK3),tumor necrosis factor(TNF),and AKT serine/threonine kinase 1(AKT1)in NRK-52E cells were assessed using quantitative real-time polymerase chain reaction(qRT-PCR).Results:We obtained sixty-six overlapping targets of celastrol and DN.GO and KEGG analyses demonstrated that the core targets of celastrol and DN were mainly involved in the inflammatory and immune response,oxidative stress,advanced glycation end products(AGEs)and their receptors(RAGEs)(AGE-RAGE),TNF,interleukin 17(IL-17),and MAPK signaling pathways.Finally,based on the good binding activity with celastrol,MAPK3,TNF,and AKT1 were identified as the foremost targets of celastrol.We observed that celastrol enhanced the viability of high glucose(HG)-treated NRK-52E cells and inhibited apoptosis in the in vitro assays.Moreover,celastrol decreased the mRNA expression levels of MAPK3,TNF,and AKT1 in high glucose(HG)-treated NRK-52E cells.Conclusion:Celastrol may treat DN by targeting APK3,TNF,and AKT1 and regulating inflammatory responses and oxidative stress through the AGE-RAGE,TNF,IL-17,and MAPK signaling pathways.展开更多
AIM: To investigate the mechanism of celastrol in inhibiting lens epithelial cells(LECs) fibrosis, which is the pathological basis of cataract.METHODS: Human LEC line SRA01/04 was treated with celastrol and transformi...AIM: To investigate the mechanism of celastrol in inhibiting lens epithelial cells(LECs) fibrosis, which is the pathological basis of cataract.METHODS: Human LEC line SRA01/04 was treated with celastrol and transforming growth factor-β2(TGF-β2). Wound-healing assay, proliferation assay, flow cytometry, real-time polymerase chain reaction(PCR), Western blot and immunocytochemical staining were used to detect the pathological changes of celastrol on LECs. Then, we cultured Sprague-Dawley rat lens in medium as a semi-in vivo model to find the function of celastrol further.RESULTS: We found that celastrol inhibited the migration of LECs, as well as proliferation(P<0.05). In addition, it induced the G2/M phase arrest by cell cyclerelated proteins(P<0.01). Moreover, celastrol inhibited epithelial-mesenchymal transition(EMT) by the blockade of TGF-β/Smad and Jagged/Notch signaling pathways.CONCLUSION: Our study demonstrates that celastrol could inhibit TGF-β2-induced lens fibrosis and raises the possibility that celastrol could be a potential novel drug in prevention and treatment of fibrotic cataract.展开更多
Objective Pituitary adrenocorticotropic hormone(ACTH)-secreting adenoma is a relatively intractable endocrine adenoma that can cause a range of severe metabolic disorders and pathological changes involving multiple sy...Objective Pituitary adrenocorticotropic hormone(ACTH)-secreting adenoma is a relatively intractable endocrine adenoma that can cause a range of severe metabolic disorders and pathological changes involving multiple systems.Previous studies have shown that celastrol has antitumor effects on a variety of tumor cells via the AKT/mTOR signaling.However,whether celastrol has pronounced antitumor effects on pituitary ACTH-secreting adenoma is unclear.This study aimed to identify a new effective therapeutic drug for pituitary ACTH-secreting adenoma.Methods Mouse pituitary ACTH-secreting adenoma cells(AtT20 cells)were used as an experimental model in vitro and to establish a xenograft tumor model in mice.Cells and animals were administered doses of celastrol at various levels.The effects of celastrol on cell viability,migration,apoptosis and autophagy were then examined.Finally,the potential involvement of AKT/mTOR signaling in celastrol’s mechanism was assessed.Results Celastrol inhibited the proliferation and migration of pituitary adenoma cells in a time-and concentration-dependent manner.It blocked AtT20 cells in the G0/G1 phase,and induced apoptosis and autophagy by downregulating the AKT/mTOR signaling pathway.Similar results were obtained in mice.Conclusion Celastrol exerts potent antitumor effects on ACTH-secreting adenoma by downregulating the AKT/mTOR signaling in vitro and in vivo.展开更多
AIM:To evaluate celastrol's effect on choroidal neovascularization(CNV).METHODS:In this study,neovascular formation in vitro(tube formation and aortic ring culture)and in vivo(laser induced neovascular in mice)was...AIM:To evaluate celastrol's effect on choroidal neovascularization(CNV).METHODS:In this study,neovascular formation in vitro(tube formation and aortic ring culture)and in vivo(laser induced neovascular in mice)was treated with celastrol to evaluate this natural compound's impact on CNV.Western blot was applied to explore the possible mechanism for it.For in vitro assay,triplicate for each group was repeated at least three times.For in vivo assay,each group contains 5 mice.RESULTS:Celastrol supressed tube formation and aortic ring sprout neovascularization.In vitro assay exhibited that celastrol inhibiting vascular endothelial growth factor(VEGF)-induced proliferation and migration of human umbilical vein endothelial cells and human choroidal endothelial cells,and by blocking VEGF signaling.Furthermore,intraperitoneal administration of celastrol significantly reduced the area of laser-induced CNV in an in vivo mouse model.By day 14,the area of CNV had decreased by 49.15%and 80.26%in the 0.1 mg/kg celastrol-treated group(n=5)and in the 0.5 mg/kg celastrol treated group(n=5),respectively,compared to the vehicle-treated group(n=5).CONCLUSION:Celastrol inhibits CNV by inhibiting VEGF-induced proliferation and migration of vascular endothelial cells,indicating that celastrol is a potent,natural therapeutic compound for the prevention of CNV.展开更多
Objective:Neutrophils play an important role in the pathogenesis of rheumatoid arthritis (RA).In this study,we used the adjuvant-induced arthritis murine model to evaluate the efficacy of celastrol on neutrophil-media...Objective:Neutrophils play an important role in the pathogenesis of rheumatoid arthritis (RA).In this study,we used the adjuvant-induced arthritis murine model to evaluate the efficacy of celastrol on neutrophil-mediated inflammation in RA.Methods:Freund's complete adjuvant-induced arthritis was used as the murine model of RA.Celastrol was intraperitoneally administrated daily after onset of the disease.The joint diameter and inflammatory score were evaluated daily during the treatment period.Myeloperoxidase (MPO) and neutrophil elastase (NE) activities were evaluated by immunohistochemical analyses.Quantitative PCR and enzyme-linked immunoabsorbent assay were used to quantify the expression of cytokines.The expression of apoptosis-related proteins Bcl-2,Bax and caspase-3 were evaluated by western blot.Results:Celastrol suppressed inflammation in joints of arthritic mice and diminished the expression of MPO and NE in the joint tissue.Celastrol significantly inhibited the expression of TNFα and IL-6 induced by LPS in neutrophils in a dose-dependent manner in vitro.Moreover,celastrol induced apoptosis of LPS-stimulated neutrophils by increasing the expression of Bax and cleaved caspase-3 while decreasing Bcl-2 expression.Conclusion:Our findings show that celastrol significantly alleviates murine arthritis by modulating the inflammatory activities of neutrophils.These results indicate that celastrol could serve as an alternative or adjunct modality for the treatment of RA.展开更多
The purpose Celastrol, the main active compound of the Celastrus genus plants, belonging to Celastraceae, has recently marked antitumour potency on solid tumours of various derivations, Methods: We demonstrate here th...The purpose Celastrol, the main active compound of the Celastrus genus plants, belonging to Celastraceae, has recently marked antitumour potency on solid tumours of various derivations, Methods: We demonstrate here that Celastrol also present powerful antileukaemic potency through both growth arrest and apoptosis induction in K562 cells, which was accompanied by typical apoptotic morphological and sharp decreased expression of phosphorylation level of Caspase family members and Akt signaling pathway related proteins were determined by western blot before and after celastrol treatment, and further the effect of AKT signaling pathway on celastrol-induced-apoptosis was analyzed. However, in vitro treatment with Celastrol resulted in significantly reduced expression of phophorylation of Akt, Survivin and Bcl-2 significantly in K562 cells. Results: 25 nmol/L WORT (PI3K-Akt inhibitor) can significantly augmented cell apoptosis induced by Celastrol in K562 cells in dose-dependent manner, Moreover, most Caspase3,8,6 were activated in K562 cells during Celastrol treatment, 50 μmol/Lz-VAD-fmk (Caspase inhibitor) can to enhance the apoptosis induced by Celastrol. Discussion: These results suggest that the fact that Akt signaling pathway might act as new targets of Celastrol, correlates well with the sensitivity to Celastrol, as well as the rate of apoptosis induced by Celastrol, Mechanisms that regulate Akt signaling pathway may be provide novel opportunities for drug development.展开更多
Tripterygium wilfordii has been renowned mostly because of the anticancer effects of its root extracts,which is partly ascribed to the presence of celastrol,a pentacyclic triterpenoid,as one of the main active compone...Tripterygium wilfordii has been renowned mostly because of the anticancer effects of its root extracts,which is partly ascribed to the presence of celastrol,a pentacyclic triterpenoid,as one of the main active components.Celastrol also has recently been reported as an effective prodrug in the treatment of obesity.Despite the promising activities,the pathway leading to celastrol biosynthesis,especially cytochrome P450(CYP)enzyme(s)that occur in its downstream steps,are largely unknown.This study conducted a comparative analysis of the T.wilfordii transcriptome derived from its root and leaf tissues.Differential gene expression analysis identified a number of root-specific CYP genes.Further phylogenetic analysis suggested specific family members of CYPs that may participate in the late steps during celastrol biosynthesis.Root-specific transcription factors(TFs)that may play regulatory roles in celastrol biosynthesis were also discussed.This genetic resource will aid in isolating the celastrol biosynthetic genes as well as engineering the celastrol biosynthesis pathway.展开更多
[Objectives]This study was conducted to investigate the extraction technology of celastrol from Celastrus orbiculatus Thunb.[Methods]Solvent ultrasonic extraction was selected,and with the content of celastrol as the ...[Objectives]This study was conducted to investigate the extraction technology of celastrol from Celastrus orbiculatus Thunb.[Methods]Solvent ultrasonic extraction was selected,and with the content of celastrol as the evaluation index,the effects of different solvents,extraction time,temperatures and material-to-liquid ratios on the extraction rate of celastrol were investigated by single factor and orthogonal experiments.[Results]The optimal extraction conditions were as follows:a solvent ratio of petroleum ether to ethyl acetate at 1∶1,a ratio of solvent to material at 10∶1(v/w),extraction time of 30 min,and an extraction temperature at 30℃.[Conclusions]This method has high extraction rate,and is simple and feasible.展开更多
The endothelial-to-mesenchymal transition(End MT) in endothelial cells contributes to the development of cardiac fibrosis,ultimately leading to cardiac remodeling.In this study,the effects and molecular mechanisms of ...The endothelial-to-mesenchymal transition(End MT) in endothelial cells contributes to the development of cardiac fibrosis,ultimately leading to cardiac remodeling.In this study,the effects and molecular mechanisms of celastrol(CEL) on transforming growth factor-β1(TGF-β1)-induced End MT in human umbilical vein endothelial(HUVEC-12) cells were investigated.The presented data demonstrated that CEL significantly blocked the morphology change of HUVEC-12 cells induced by TGF-β1 without cell cytotoxicity.In accordance with these findings,CEL blocked TGF-β1-induced EndM T as evidenced by the inhibition of the mesenchymal markers,including collagen Ⅰ,Ⅲ,α-SMA,fibronectin m RNA expression,and the increase in the m RNA expression of endothelial cell marker CD31.These changes were also confirmed by double immunofluorescence staining of CD31 and vimentin.The in vitro scratch assay showed that CEL inhibited the migration capacity of the transitioned endothelial cells induced by TGF-β1.Further experiments showed that the beneficial effect of CEL on blocking the End MT in HUVEC-12 cells was associated with the suppression of the TGF-β1/Smads signalling pathway,which was also confirmed by the inhibition of its downstream transcription factor snail1,twist1,twist2,ZEB1 and ZEB2.These results indicate that CEL blocks TGF-β1-induced End MT through TGF-β1/Smads signalling pathway and suggest that it may be a feasible therapy for cardiac fibrosis diseases.展开更多
Excessive oxidative stress impairs cartilage matrix metabolism balance,significantly contributing to osteoarthritis(OA)development.Celastrol(CSL),a drug derived from Tripterygium wilfordii,has recognized applications ...Excessive oxidative stress impairs cartilage matrix metabolism balance,significantly contributing to osteoarthritis(OA)development.Celastrol(CSL),a drug derived from Tripterygium wilfordii,has recognized applications in the treatment of cancer and immune system disorders,yet its antioxidative stress mechanisms in OA remain underexplored.This study aimed to substantiate CSL’s chondroprotective effects and unravel its underlying mechanisms.We investigated CSL’s impact on chondrocytes under both normal and inflammatory conditions.In vitro,CSL mitigated interleukin(IL)-1β-induced activation of proteinases and promoted cartilage extracellular matrix(ECM)synthesis.In vivo,intra-articular injection of CSL ameliorated cartilage degeneration and mitigated subchondral bone lesions in OA mice.Mechanistically,it was found that inhibiting nuclear factor erythroid 2-related factor 2(NRF2)abrogated CSL-mediated antioxidative functions and exacerbated the progression of OA.This study is the first to elucidate the role of CSL in the treatment of OA through the activation of NRF2,offering a novel therapeutic avenue for arthritis therapy.展开更多
Objective To investigate the mechanistic basis for the anti-proliferation and anti-invasion effect of tumor necrosis factor-related apoptosis-induced ligand(TRAIL)and celastrol combination treatment(TCCT)in glioblasto...Objective To investigate the mechanistic basis for the anti-proliferation and anti-invasion effect of tumor necrosis factor-related apoptosis-induced ligand(TRAIL)and celastrol combination treatment(TCCT)in glioblastoma cells.Methods Cell counting kit-8 was used to detect the effects of different concentrations of celastrol(0-16µmol/L)and TRAIL(0-500 ng/mL)on the cell viability of glioblastoma cells.U87 cells were randomly divided into 4 groups,namely control,TRAIL(TRAIL 100 ng/mL),Cel(celastrol 0.5µmol/L)and TCCT(TRAIL 100 ng/mL+celastrol 0.5µmol/L).Cell proliferation,migration,and invasion were detected by colony formation,wound healing,and Transwell assays,respectively.Quantitative reverse transcription polymerase chain reaction and Western blotting were performed to assess the levels of epithelial-mesenchymal transition(EMT)markers(zona occludens,N-cadherin,vimentin,zinc finger E-box-binding homeobox,Slug,and β-catenin).Wnt pathway was activated by lithium chloride(LiCl,20 mol/L)and the mechanism for action of TCCT was explored.Results Celastrol and TRAIL synergistically inhibited the proliferation,migration,invasion,and EMT of U87 cells(P<0.01).TCCT up-regulated the expression of GSK-3β and down-regulated the expression of β-catenin and its associated proteins(P<0.05 or P<0.01),including c-Myc,Cyclin-D1,and matrix metalloproteinase(MMP)-2.In addition,LiCl,an activator of the Wnt signaling pathway,restored the inhibitory effects of TCCT on the expression of β-catenin and its downstream genes,as well as the migration and invasion of glioblastoma cells(P<0.05 or P<0.01).Conclusions Celastrol and TRAIL can synergistically suppress glioblastoma cell migration,invasion,and EMT,potentially through inhibition of Wnt/β-catenin pathway.This underlies a novel mechanism of action for TCCT as an effective therapy for glioblastoma.展开更多
Ferroptosis is a form of regulated cell death, characterized by excessive membrane lipid peroxidation in an iron-and ROS-dependent manner. Celastrol, a natural bioactive triterpenoid extracted from Tripterygium wilfor...Ferroptosis is a form of regulated cell death, characterized by excessive membrane lipid peroxidation in an iron-and ROS-dependent manner. Celastrol, a natural bioactive triterpenoid extracted from Tripterygium wilfordii, shows effective anti-fibrotic and anti-inflammatory activities in multiple hepatic diseases. However, the exact molecular mechanisms of action and the direct protein targets of celastrol in the treatment of liver fibrosis remain largely elusive. Here, we discover that celastrol exerts anti-fibrotic effects via promoting the production of reactive oxygen species(ROS) and inducing ferroptosis in activated hepatic stellate cells(HSCs). By using activity-based protein profiling(ABPP) in combination with bio-orthogonal click chemistry reaction and cellular thermal shift assay(CETSA), we show that celastrol directly binds to peroxiredoxins(PRDXs), including PRDX1, PRDX2, PRDX4 and PRDX6,through the active cysteine sites, and inhibits their anti-oxidant activities. Celastrol also targets to heme oxygenase 1(HO-1) and upregulates its expression in activated-HSCs. Knockdown of PRDX1, PRDX2,PRDX4, PRDX6 or HO-1 in HSCs, to varying extent, elevated cellular ROS levels and induced ferroptosis. Taken together, our findings reveal the direct protein targets and molecular mechanisms via which celastrol ameliorates hepatic fibrosis, thus supporting the further development of celastrol as a promising therapeutic agent for liver fibrosis.展开更多
基金supported by the National Natural Science Foundation of China(No.82204663)the Natural Science Foundation of Shandong Province(No.ZR2022QH058).
文摘Background:Glioblastoma is one of the most common primary intracranial tumors of the central nervous system in adults.Although chemotherapy is an important component of glioblastoma treatment,its effectiveness remains unsatisfactory.Due to multiple immunosuppressive mechanisms,glioblastoma immunotherapy has not been effective in treating many patients as a result of the clinical breakthroughs in the field.Therefore,the development of cancer immunotherapy relies on the understanding of how tumors interact with the immune system and the analysis of their molecular determinants.This study identified the key interactions between immune cells in the glioma microenvironment using RNA microarrays and single-cell sequencing.Methods:First,we screened differentially expressed genes in tumor and control samples from GSE29796 and GSE50161 datasets using GEO2R.All differentially expressed genes were used to perform enrichment analysis and construct protein-protein interaction topological analysis to analyze the interaction between proteins.Using single-cell RNA sequencing data from the GSE162631 database,we identified immune cell types within the glioblastoma microenvironment,and validated the hub gene expression in these cells.In addition,based on the GEPIA and TIMER databases,hub genes were investigated and compared with immune infiltration to determine differential expression.Finally,CellChat was used to visualize the gene expression distribution and cell-to-cell communication analysis of the proteins between different types of cells.Results:We found that monocytes/macrophages may communicate with each other in the tumor microenvironment through MIF-(CD74+CXCR4)and MIF-(CD74+CD44).In addition,our study indicated that celastrol has the ability to inhibit inflammatory factors expression by MIF/CD74 signaling pathway in U87 cells.Conclusion:This study improved the effectiveness of cancer immunotherapy strategies and developed new ideas for immunotherapy that can be applied to glioblastoma.
基金the National Natural Science Foundation of China(Grant No.:81973305)the Science and Technology Planning Project of Guangzhou,China(Grant No.:201904010487)+1 种基金the Natural Science Foundation of Guangdong Province,China(Grant No.:2021A1515010897)the Discipline Construction Fund of Central People’s Hospital of Zhanjiang(Grant Nos.:2020A01 and 2020A02).
文摘Stroke is the second leading cause of death worldwide,and oxidative stress plays a crucial role.Celastrol exhibits strong antioxidant properties in several diseases;however,whether it can affect oxidation in cerebral ischemic-reperfusion injury(CIRI)remains unclear.This study aimed to determine whether celastrol could reduce oxidative damage during CIRI and to elucidate the underlying mechanisms.Here,we found that celastrol attenuated oxidative injury in CIRI by upregulating nuclear factor E2-related factor 2(Nrf2).Using alkynyl-tagged celastrol and liquid chromatography-tandem mass spectrometry,we showed that celastrol directly bound to neuronally expressed developmentally downregulated 4(Nedd4)and then released Nrf2 from Nedd4 in astrocytes.Nedd4 promoted the degradation of Nrf2 through K48-linked ubiquitination and thus contributed to astrocytic reactive oxygen species production in CIRI,which was significantly blocked by celastrol.Furthermore,by inhibiting oxidative stress and astrocyte activation,celastrol effectively rescued neurons from axon damage and apoptosis.Our study uncovered Nedd4 as a direct target of celastrol,and that celastrol exerts an antioxidative effect on astrocytes by inhibiting the interaction between Nedd4 and Nrf2 and reducing Nrf2 degradation in CIRI.
基金support from the National Natural Science Foundation of China(Grants No.82304827,82074098,81841001)the Fundamental Research Funds for the Central public welfare research institutes(ZZ13-ZD-07),the National Key Research and Development Programof China(2020YFA0908000,2022YFC2303600)+7 种基金the Innovation Team and Talents Cultivation Program of National Administration of Traditional Chinese Medicine(No:ZYYCXTD-C-202002)The Shenzhen Medical Research Fund of Shenzhen Medical Academy of Research and Translation(B2302051)the Fundamental Research Funds for the Central Public Welfare Research Institutes(Grants No.ZZ13-YQ-108)the Shenzhen Science and Technology Innovation Commission(Grants No.JCYJ20210324115800001)the Science and Technology Foundation of Shenzhen(Shenzhen Clinical Medical Research Center for Geriatric Diseases),the Distinguished Expert Project of Sichuan Province Tianfu Scholar(CW202002)Supported by Shenzhen Governmental Sustainable Development Fund(KCXFZ20201221173612034)Supported by Shenzhen key Laboratory of Kidney Diseases(ZDSYS201504301616234)Supported by Shenzhen Fund for Guangdong Provincial High-level Clinical Key Specialties(NO.SZGSP001).
文摘Hepatocellular carcinoma(HCC)is one of most common and deadliest malignancies.Celastrol(Cel),a natural product derived from the Tripterygium wilfordii plant,has been extensively researched for its potential effectiveness in fighting cancer.However,its clinical application has been hindered by the unclear mechanism of action.Here,we used chemical proteomics to identify the direct targets of Cel and enhanced its targetability and antitumor capacity by developing a Cel-based liposomes in HCC.We demonstrated that Cel selectively targets the voltage-dependent anion channel 2(VDAC2).Cel directly binds to the cysteine residues of VDAC2,and induces cytochrome C release via dysregulating VDAC2-mediated mitochondrial permeability transition pore(mPTP)function.We further found that Cel induces ROS-mediated ferroptosis and apoptosis in HCC cells.Moreover,coencapsulation of Cel into alkyl glucoside-modified liposomes(AGCL)improved its antitumor efficacy and minimized its side effects.AGCL has been shown to effectively suppress the proliferation of tumor cells.In a xenograft nude mice experiment,AGCL significantly inhibited tumor growth and promoted apoptosis.Our findings reveal that Cel directly targets VDAC2 to induce mitochondria-dependent cell death,while the Cel liposomes enhance its targetability and reduces side effects.Overall,Cel shows promise as a therapeutic agent for HCC.
基金suppor ted by the National Key Research and Development Program of China(2020YFA0908000)the Innovation Team and Talents Cultivation Program of the National Administration of Traditional Chinese Medicine(ZYYCXTD-C-202002)+1 种基金the National Natural Science Foundation of China(82074098,81841001)the Fundamental Research Funds for the Central Public Welfare Research Institutes(ZXKT18003)。
文摘Background: Sepsis involves life-threatening organ dysfunction and is caused by a dysregulated host response to infection. No specific therapies against sepsis have been reported. Celastrol(Cel) is a natural anti-inflammatory compound that shows potential against systemic inflammatory diseases. This study aimed to investigate the pharmacological activity and molecular mechanism of Cel in models of endotoxemia and sepsis.Methods: We evaluated the anti-inflammatory efficacy of Cel against endotoxemia and sepsis in mice and macrophage cultures treated with lipopolysaccharide(LPS). We screened for potential protein targets of Cel using activity-based protein profiling(ABPP). Potential targets were validated using biophysical methods such as cellular thermal shift assays(CETSA) and surface plasmon resonance(SPR). Residues involved in Cel binding to target proteins were identified through point mutagenesis, and the functional effects of such binding were explored through gene knockdown.Results: Cel protected mice from lethal endotoxemia and improved their survival with sepsis, and it significantly decreased the levels of pro-inflammatory cytokines in mice and macrophages treated with LPS(P <0.05). Cel bound to Cys424 of pyruvate kinase M2(PKM2), inhibiting the enzyme and thereby suppressing aerobic glycolysis(Warburg effect). Cel also bound to Cys106 in high mobility group box 1(HMGB1) protein, reducing the secretion of inflammatory cytokine interleukin(IL)-1β. Cel bound to the Cys residues in lactate dehydrogenase A(LDHA).Conclusions: Cel inhibits inflammation and the Warburg effect in sepsis via targeting PKM2 and HMGB1 protein.
基金supported by the Zhejiang Province Chinese Medicine Modernization Program Grant[Number 2020ZX001]the“Pioneer”and“Leading Goose”R&D Program of Zhejiang Grant[Number 2023C03075].
文摘Background:Diabetic nephropathy(DN)is a serious complication of diabetes with rising prevalence worldwide.We aimed to explore the anti-DN mechanisms of the compound celastrol derived from the medicinal plant Tripterygium wilfordii.Methods:Celastrol-related targets were obtained from Herbal Ingredients’Targets(HIT)and GeneCards databases.DN-related targets were retrieved from GeneCards,DisGeNET,and Therapeutic Targets Database(TTD).A Protein-protein interaction(PPI)network was established using the Search Tool for the Retrieval of Interacting Genes(STRING)database.Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analyses were performed using ClusterProfiler.The cytoHubba plugin was used to select the top 10 hub targets.Molecular docking was performed employing PyMOL and AutoDock software.Cell counting kit-8(CCK-8)and flow cytometry assays were used to detect the viability and apoptosis of NRK-52E cells,respectively.The mRNA expression levels of mitogen-activated protein kinase 3(MAPK3),tumor necrosis factor(TNF),and AKT serine/threonine kinase 1(AKT1)in NRK-52E cells were assessed using quantitative real-time polymerase chain reaction(qRT-PCR).Results:We obtained sixty-six overlapping targets of celastrol and DN.GO and KEGG analyses demonstrated that the core targets of celastrol and DN were mainly involved in the inflammatory and immune response,oxidative stress,advanced glycation end products(AGEs)and their receptors(RAGEs)(AGE-RAGE),TNF,interleukin 17(IL-17),and MAPK signaling pathways.Finally,based on the good binding activity with celastrol,MAPK3,TNF,and AKT1 were identified as the foremost targets of celastrol.We observed that celastrol enhanced the viability of high glucose(HG)-treated NRK-52E cells and inhibited apoptosis in the in vitro assays.Moreover,celastrol decreased the mRNA expression levels of MAPK3,TNF,and AKT1 in high glucose(HG)-treated NRK-52E cells.Conclusion:Celastrol may treat DN by targeting APK3,TNF,and AKT1 and regulating inflammatory responses and oxidative stress through the AGE-RAGE,TNF,IL-17,and MAPK signaling pathways.
基金Supported by National Natural Science Foundation of China (No.81300749)Guangdong Province Natural Science Foundation (No.2018A030313628)+1 种基金973 program (No.2015CB964600)the State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University
文摘AIM: To investigate the mechanism of celastrol in inhibiting lens epithelial cells(LECs) fibrosis, which is the pathological basis of cataract.METHODS: Human LEC line SRA01/04 was treated with celastrol and transforming growth factor-β2(TGF-β2). Wound-healing assay, proliferation assay, flow cytometry, real-time polymerase chain reaction(PCR), Western blot and immunocytochemical staining were used to detect the pathological changes of celastrol on LECs. Then, we cultured Sprague-Dawley rat lens in medium as a semi-in vivo model to find the function of celastrol further.RESULTS: We found that celastrol inhibited the migration of LECs, as well as proliferation(P<0.05). In addition, it induced the G2/M phase arrest by cell cyclerelated proteins(P<0.01). Moreover, celastrol inhibited epithelial-mesenchymal transition(EMT) by the blockade of TGF-β/Smad and Jagged/Notch signaling pathways.CONCLUSION: Our study demonstrates that celastrol could inhibit TGF-β2-induced lens fibrosis and raises the possibility that celastrol could be a potential novel drug in prevention and treatment of fibrotic cataract.
基金This work was supported by the National Natural Science Youth Foundation of China(No.81602204).
文摘Objective Pituitary adrenocorticotropic hormone(ACTH)-secreting adenoma is a relatively intractable endocrine adenoma that can cause a range of severe metabolic disorders and pathological changes involving multiple systems.Previous studies have shown that celastrol has antitumor effects on a variety of tumor cells via the AKT/mTOR signaling.However,whether celastrol has pronounced antitumor effects on pituitary ACTH-secreting adenoma is unclear.This study aimed to identify a new effective therapeutic drug for pituitary ACTH-secreting adenoma.Methods Mouse pituitary ACTH-secreting adenoma cells(AtT20 cells)were used as an experimental model in vitro and to establish a xenograft tumor model in mice.Cells and animals were administered doses of celastrol at various levels.The effects of celastrol on cell viability,migration,apoptosis and autophagy were then examined.Finally,the potential involvement of AKT/mTOR signaling in celastrol’s mechanism was assessed.Results Celastrol inhibited the proliferation and migration of pituitary adenoma cells in a time-and concentration-dependent manner.It blocked AtT20 cells in the G0/G1 phase,and induced apoptosis and autophagy by downregulating the AKT/mTOR signaling pathway.Similar results were obtained in mice.Conclusion Celastrol exerts potent antitumor effects on ACTH-secreting adenoma by downregulating the AKT/mTOR signaling in vitro and in vivo.
基金Supported by National Natural Science Foundation of China(No.81570826)。
文摘AIM:To evaluate celastrol's effect on choroidal neovascularization(CNV).METHODS:In this study,neovascular formation in vitro(tube formation and aortic ring culture)and in vivo(laser induced neovascular in mice)was treated with celastrol to evaluate this natural compound's impact on CNV.Western blot was applied to explore the possible mechanism for it.For in vitro assay,triplicate for each group was repeated at least three times.For in vivo assay,each group contains 5 mice.RESULTS:Celastrol supressed tube formation and aortic ring sprout neovascularization.In vitro assay exhibited that celastrol inhibiting vascular endothelial growth factor(VEGF)-induced proliferation and migration of human umbilical vein endothelial cells and human choroidal endothelial cells,and by blocking VEGF signaling.Furthermore,intraperitoneal administration of celastrol significantly reduced the area of laser-induced CNV in an in vivo mouse model.By day 14,the area of CNV had decreased by 49.15%and 80.26%in the 0.1 mg/kg celastrol-treated group(n=5)and in the 0.5 mg/kg celastrol treated group(n=5),respectively,compared to the vehicle-treated group(n=5).CONCLUSION:Celastrol inhibits CNV by inhibiting VEGF-induced proliferation and migration of vascular endothelial cells,indicating that celastrol is a potent,natural therapeutic compound for the prevention of CNV.
基金the National Natural Science Foundation of China(Grant number 81430099)International S&T Cooperation Program of China(Grant number 2014DFA32950)Beijing Municipal Commission of Education(Grant number 521/0101312).
文摘Objective:Neutrophils play an important role in the pathogenesis of rheumatoid arthritis (RA).In this study,we used the adjuvant-induced arthritis murine model to evaluate the efficacy of celastrol on neutrophil-mediated inflammation in RA.Methods:Freund's complete adjuvant-induced arthritis was used as the murine model of RA.Celastrol was intraperitoneally administrated daily after onset of the disease.The joint diameter and inflammatory score were evaluated daily during the treatment period.Myeloperoxidase (MPO) and neutrophil elastase (NE) activities were evaluated by immunohistochemical analyses.Quantitative PCR and enzyme-linked immunoabsorbent assay were used to quantify the expression of cytokines.The expression of apoptosis-related proteins Bcl-2,Bax and caspase-3 were evaluated by western blot.Results:Celastrol suppressed inflammation in joints of arthritic mice and diminished the expression of MPO and NE in the joint tissue.Celastrol significantly inhibited the expression of TNFα and IL-6 induced by LPS in neutrophils in a dose-dependent manner in vitro.Moreover,celastrol induced apoptosis of LPS-stimulated neutrophils by increasing the expression of Bax and cleaved caspase-3 while decreasing Bcl-2 expression.Conclusion:Our findings show that celastrol significantly alleviates murine arthritis by modulating the inflammatory activities of neutrophils.These results indicate that celastrol could serve as an alternative or adjunct modality for the treatment of RA.
文摘The purpose Celastrol, the main active compound of the Celastrus genus plants, belonging to Celastraceae, has recently marked antitumour potency on solid tumours of various derivations, Methods: We demonstrate here that Celastrol also present powerful antileukaemic potency through both growth arrest and apoptosis induction in K562 cells, which was accompanied by typical apoptotic morphological and sharp decreased expression of phosphorylation level of Caspase family members and Akt signaling pathway related proteins were determined by western blot before and after celastrol treatment, and further the effect of AKT signaling pathway on celastrol-induced-apoptosis was analyzed. However, in vitro treatment with Celastrol resulted in significantly reduced expression of phophorylation of Akt, Survivin and Bcl-2 significantly in K562 cells. Results: 25 nmol/L WORT (PI3K-Akt inhibitor) can significantly augmented cell apoptosis induced by Celastrol in K562 cells in dose-dependent manner, Moreover, most Caspase3,8,6 were activated in K562 cells during Celastrol treatment, 50 μmol/Lz-VAD-fmk (Caspase inhibitor) can to enhance the apoptosis induced by Celastrol. Discussion: These results suggest that the fact that Akt signaling pathway might act as new targets of Celastrol, correlates well with the sensitivity to Celastrol, as well as the rate of apoptosis induced by Celastrol, Mechanisms that regulate Akt signaling pathway may be provide novel opportunities for drug development.
基金This work was supported in part by a grant from the National Key R&D Program of China(SQ2018YFC170017)a grant from the National Natural Science Foundation of China(31670300).
文摘Tripterygium wilfordii has been renowned mostly because of the anticancer effects of its root extracts,which is partly ascribed to the presence of celastrol,a pentacyclic triterpenoid,as one of the main active components.Celastrol also has recently been reported as an effective prodrug in the treatment of obesity.Despite the promising activities,the pathway leading to celastrol biosynthesis,especially cytochrome P450(CYP)enzyme(s)that occur in its downstream steps,are largely unknown.This study conducted a comparative analysis of the T.wilfordii transcriptome derived from its root and leaf tissues.Differential gene expression analysis identified a number of root-specific CYP genes.Further phylogenetic analysis suggested specific family members of CYPs that may participate in the late steps during celastrol biosynthesis.Root-specific transcription factors(TFs)that may play regulatory roles in celastrol biosynthesis were also discussed.This genetic resource will aid in isolating the celastrol biosynthetic genes as well as engineering the celastrol biosynthesis pathway.
文摘[Objectives]This study was conducted to investigate the extraction technology of celastrol from Celastrus orbiculatus Thunb.[Methods]Solvent ultrasonic extraction was selected,and with the content of celastrol as the evaluation index,the effects of different solvents,extraction time,temperatures and material-to-liquid ratios on the extraction rate of celastrol were investigated by single factor and orthogonal experiments.[Results]The optimal extraction conditions were as follows:a solvent ratio of petroleum ether to ethyl acetate at 1∶1,a ratio of solvent to material at 10∶1(v/w),extraction time of 30 min,and an extraction temperature at 30℃.[Conclusions]This method has high extraction rate,and is simple and feasible.
文摘The endothelial-to-mesenchymal transition(End MT) in endothelial cells contributes to the development of cardiac fibrosis,ultimately leading to cardiac remodeling.In this study,the effects and molecular mechanisms of celastrol(CEL) on transforming growth factor-β1(TGF-β1)-induced End MT in human umbilical vein endothelial(HUVEC-12) cells were investigated.The presented data demonstrated that CEL significantly blocked the morphology change of HUVEC-12 cells induced by TGF-β1 without cell cytotoxicity.In accordance with these findings,CEL blocked TGF-β1-induced EndM T as evidenced by the inhibition of the mesenchymal markers,including collagen Ⅰ,Ⅲ,α-SMA,fibronectin m RNA expression,and the increase in the m RNA expression of endothelial cell marker CD31.These changes were also confirmed by double immunofluorescence staining of CD31 and vimentin.The in vitro scratch assay showed that CEL inhibited the migration capacity of the transitioned endothelial cells induced by TGF-β1.Further experiments showed that the beneficial effect of CEL on blocking the End MT in HUVEC-12 cells was associated with the suppression of the TGF-β1/Smads signalling pathway,which was also confirmed by the inhibition of its downstream transcription factor snail1,twist1,twist2,ZEB1 and ZEB2.These results indicate that CEL blocks TGF-β1-induced End MT through TGF-β1/Smads signalling pathway and suggest that it may be a feasible therapy for cardiac fibrosis diseases.
基金supported by the Key Research and Development Plan of Jiangsu Province(Social Development Project,No.BE2021679)the National Natural Science Foundation Project(No.82171762).
文摘Excessive oxidative stress impairs cartilage matrix metabolism balance,significantly contributing to osteoarthritis(OA)development.Celastrol(CSL),a drug derived from Tripterygium wilfordii,has recognized applications in the treatment of cancer and immune system disorders,yet its antioxidative stress mechanisms in OA remain underexplored.This study aimed to substantiate CSL’s chondroprotective effects and unravel its underlying mechanisms.We investigated CSL’s impact on chondrocytes under both normal and inflammatory conditions.In vitro,CSL mitigated interleukin(IL)-1β-induced activation of proteinases and promoted cartilage extracellular matrix(ECM)synthesis.In vivo,intra-articular injection of CSL ameliorated cartilage degeneration and mitigated subchondral bone lesions in OA mice.Mechanistically,it was found that inhibiting nuclear factor erythroid 2-related factor 2(NRF2)abrogated CSL-mediated antioxidative functions and exacerbated the progression of OA.This study is the first to elucidate the role of CSL in the treatment of OA through the activation of NRF2,offering a novel therapeutic avenue for arthritis therapy.
基金Supported by Scientific and Technological Research Programme of Chongqing Municipal Education Commission,China(No.KJ130320)。
文摘Objective To investigate the mechanistic basis for the anti-proliferation and anti-invasion effect of tumor necrosis factor-related apoptosis-induced ligand(TRAIL)and celastrol combination treatment(TCCT)in glioblastoma cells.Methods Cell counting kit-8 was used to detect the effects of different concentrations of celastrol(0-16µmol/L)and TRAIL(0-500 ng/mL)on the cell viability of glioblastoma cells.U87 cells were randomly divided into 4 groups,namely control,TRAIL(TRAIL 100 ng/mL),Cel(celastrol 0.5µmol/L)and TCCT(TRAIL 100 ng/mL+celastrol 0.5µmol/L).Cell proliferation,migration,and invasion were detected by colony formation,wound healing,and Transwell assays,respectively.Quantitative reverse transcription polymerase chain reaction and Western blotting were performed to assess the levels of epithelial-mesenchymal transition(EMT)markers(zona occludens,N-cadherin,vimentin,zinc finger E-box-binding homeobox,Slug,and β-catenin).Wnt pathway was activated by lithium chloride(LiCl,20 mol/L)and the mechanism for action of TCCT was explored.Results Celastrol and TRAIL synergistically inhibited the proliferation,migration,invasion,and EMT of U87 cells(P<0.01).TCCT up-regulated the expression of GSK-3β and down-regulated the expression of β-catenin and its associated proteins(P<0.05 or P<0.01),including c-Myc,Cyclin-D1,and matrix metalloproteinase(MMP)-2.In addition,LiCl,an activator of the Wnt signaling pathway,restored the inhibitory effects of TCCT on the expression of β-catenin and its downstream genes,as well as the migration and invasion of glioblastoma cells(P<0.05 or P<0.01).Conclusions Celastrol and TRAIL can synergistically suppress glioblastoma cell migration,invasion,and EMT,potentially through inhibition of Wnt/β-catenin pathway.This underlies a novel mechanism of action for TCCT as an effective therapy for glioblastoma.
基金supported by the National Key Research and Development Program of China (2020YFA0908000)the Innovation Team and Talents Cultivation Program of National Administration of Traditional Chinese Medicine (ZYYCXTD-C-202002,China)+1 种基金the National Natural Science Foundation of China(81903588,81803456,82074098 and 81841001,China)the Fundamental Research Funds for the Central Public Welfare Research Institutes (ZXKT18003 and ZZ15-YQ-063,China)。
文摘Ferroptosis is a form of regulated cell death, characterized by excessive membrane lipid peroxidation in an iron-and ROS-dependent manner. Celastrol, a natural bioactive triterpenoid extracted from Tripterygium wilfordii, shows effective anti-fibrotic and anti-inflammatory activities in multiple hepatic diseases. However, the exact molecular mechanisms of action and the direct protein targets of celastrol in the treatment of liver fibrosis remain largely elusive. Here, we discover that celastrol exerts anti-fibrotic effects via promoting the production of reactive oxygen species(ROS) and inducing ferroptosis in activated hepatic stellate cells(HSCs). By using activity-based protein profiling(ABPP) in combination with bio-orthogonal click chemistry reaction and cellular thermal shift assay(CETSA), we show that celastrol directly binds to peroxiredoxins(PRDXs), including PRDX1, PRDX2, PRDX4 and PRDX6,through the active cysteine sites, and inhibits their anti-oxidant activities. Celastrol also targets to heme oxygenase 1(HO-1) and upregulates its expression in activated-HSCs. Knockdown of PRDX1, PRDX2,PRDX4, PRDX6 or HO-1 in HSCs, to varying extent, elevated cellular ROS levels and induced ferroptosis. Taken together, our findings reveal the direct protein targets and molecular mechanisms via which celastrol ameliorates hepatic fibrosis, thus supporting the further development of celastrol as a promising therapeutic agent for liver fibrosis.
