OBJECTIVE To investigate the effects of T-006(tetramethylpyrazine derivative)in promotingα-Synuclein(α-Syn)degradation and evaluated the neuroprotective effects in cellular and animalα-Syn model of Parkinson diseas...OBJECTIVE To investigate the effects of T-006(tetramethylpyrazine derivative)in promotingα-Synuclein(α-Syn)degradation and evaluated the neuroprotective effects in cellular and animalα-Syn model of Parkinson disease(PD).METHODS The inducible PC12 cells overexpressingα-syn and the homozygous transgenic(Tg)mice expressing A53T humanα-syn were used to evaluate the neuroprotective effects of T-006.For cellular study,MTT,Western blotting,proteasomal activity assay and qRT-PCR were applied to analyze the pharmacological effects and underlying mecha⁃nisms.The gene knock-down and overexpression approaches were used to dissect the molecular signaling pathways.For animal study,ten-month-old homozygousα-Syn Tg mice were treated with T-006(3 mg·kg-1)daily by gavage for four weeks.The Western blotting,immunohistochemistry and behavioral tests were applied to determine the neuropatho⁃logical changes.RESULTS T-006 promoted the degradation of WT and mutantα-Syn in PC12α-Syn inducible cells via an ubiquitin-proteasome system(UPS)dependent and autophagy-lysosome pathway independent manner.The mecha⁃nism of action involved the upregulation of 20S proteasome subunit LMP7 expression,which leads to activation of the chymotrypsin-like proteasomal activity for protein degradation.Mechanistically,we demonstrated that T-006 activated PKA/Akt/mTOR pathway upstream for LMP 7 up-regulation and UPS activation.Finally,we illustrated that T-006 promoted both Triton-soluble and-insoluble forms ofα-syn and protected againstα-Syn-induced neurotoxicity in A53Tα-Syn Tg mice.CONCLUSION T-006 is a potent UPS activator which promotes the degradation of pathogenic proteinα-Syn in cellular and animal PD models.Our study thus high-lights the therapeutic potential of small molecular UPS activator like T-006 in the treatment of PD and related conditions.展开更多
To compare liver proteolysis and proteasome activation in steatotic liver grafts conserved in University of Wisconsin(UW)and Institut Georges Lopez-1(IGL-1)solutions.METHODSFatty liver grafts from male obese Zück...To compare liver proteolysis and proteasome activation in steatotic liver grafts conserved in University of Wisconsin(UW)and Institut Georges Lopez-1(IGL-1)solutions.METHODSFatty liver grafts from male obese Zücker rats were conserved in UW and IGL-1 solutions for 24 h at 4°Cand subjected to“ex vivo”normo-thermic perfusion(2 h;37°C).Liver proteolysis in tissue specimens and perfusate was measured by reverse-phase high performance liquid chromatography.Total free amino acid release was correlated with the activation of the ubiquitin proteasome system(UPS:measured as chymotryptic-like activity and 20S and 19S proteasome),the prevention of liver injury(transaminases),mitochondrial injury(confocal microscopy)and inflammation markers(TNF 1 alpha,high mobility group box-1(HGMB-1)and PPAR gamma),and liver apoptosis(TUNEL assay,cytochrome c and caspase 3).RESULTSProfiles of free AA(alanine,proline,leucine,isoleucine,methionine,lysine,ornithine,and threonine,among others)were similar for tissue and reperfusion effluent.In all cases,the IGL-1 solution showed a significantly higher prevention of proteolysis than UW(P<0.05)after cold ischemia reperfusion.Livers conserved in IGL-1 presented more effective prevention of ATP-breakdown and more inhibition of UPS activity(measured as chymotryptic-like activity).In addition,the prevention of liver proteolysis and UPS activation correlated with the prevention of liver injury(AST/ALT)and mitochondrial damage(revealed by confocal microscopy findings)as well as with the prevention of inflammatory markers(TNF1alpha and HMGB)after reperfusion.In addition,the liver grafts preserved in IGL-1 showed a significant decrease in liver apoptosis,as shown by TUNEL assay and the reduction of cytochrome c,caspase 3 and P62 levels.CONCLUSIONOur comparison of these two preservation solutions suggests that IGL-1 helps to prevent ATP breakdown more effectively than UW and subsequently achieves a higher UPS inhibition and reduced liver proteolysis.展开更多
Cancer is a major global health issue.Effective therapeutic strategies can prolong patients’survival and reduce the costs of treatment.Drug repurposing,which identifies new therapeutic uses for approved drugs,is a pr...Cancer is a major global health issue.Effective therapeutic strategies can prolong patients’survival and reduce the costs of treatment.Drug repurposing,which identifies new therapeutic uses for approved drugs,is a promising approach with the advantages of reducing research costs,shortening development time,and increasing efficiency and safety.Disulfiram(DSF),a Food and Drug Administration(FDA)-approved drug used to treat chronic alcoholism,has a great potential as an anticancer drug by targeting diverse human malignancies.Several studies show the antitumor effects of DSF,particularly the combination of DSF and copper(DSF/Cu),on a wide range of cancers such as glioblastoma(GBM),breast cancer,liver cancer,pancreatic cancer,and melanoma.In this review,we summarize the antitumor mechanisms of DSF/Cu,including induction of intracellular reactive oxygen species(ROS)and various cell death signaling pathways,and inhibition of proteasome activity,as well as inhibition of nuclear factor-kappa B(NF-κB)signaling.Furthermore,we highlight the ability of DSF/Cu to target cancer stem cells(CSCs),which provides a new approach to prevent tumor recurrence and metastasis.Strikingly,DSF/Cu inhibits several molecular targets associated with drug resistance,and therefore it is becoming a novel option to increase the sensitivity of chemo-resistant and radio-resistant patients.Studies of DSF/Cu may shed light on its improved application to clinical tumor treatment.展开更多
基金Science and Technology Development Fund(FDCT)of Macao SAR(069/2015/A2134/2014/A3+4 种基金062-2017-AIR)Research Committee,University of Macao(MYRG2015-00182-ICMS-QRCMMYRG2015-00214-ICMSQRCMMYRG139(Y1-L4)-ICMS12-LMYand MYRG2016-00129-ICMS-QRCM)
文摘OBJECTIVE To investigate the effects of T-006(tetramethylpyrazine derivative)in promotingα-Synuclein(α-Syn)degradation and evaluated the neuroprotective effects in cellular and animalα-Syn model of Parkinson disease(PD).METHODS The inducible PC12 cells overexpressingα-syn and the homozygous transgenic(Tg)mice expressing A53T humanα-syn were used to evaluate the neuroprotective effects of T-006.For cellular study,MTT,Western blotting,proteasomal activity assay and qRT-PCR were applied to analyze the pharmacological effects and underlying mecha⁃nisms.The gene knock-down and overexpression approaches were used to dissect the molecular signaling pathways.For animal study,ten-month-old homozygousα-Syn Tg mice were treated with T-006(3 mg·kg-1)daily by gavage for four weeks.The Western blotting,immunohistochemistry and behavioral tests were applied to determine the neuropatho⁃logical changes.RESULTS T-006 promoted the degradation of WT and mutantα-Syn in PC12α-Syn inducible cells via an ubiquitin-proteasome system(UPS)dependent and autophagy-lysosome pathway independent manner.The mecha⁃nism of action involved the upregulation of 20S proteasome subunit LMP7 expression,which leads to activation of the chymotrypsin-like proteasomal activity for protein degradation.Mechanistically,we demonstrated that T-006 activated PKA/Akt/mTOR pathway upstream for LMP 7 up-regulation and UPS activation.Finally,we illustrated that T-006 promoted both Triton-soluble and-insoluble forms ofα-syn and protected againstα-Syn-induced neurotoxicity in A53Tα-Syn Tg mice.CONCLUSION T-006 is a potent UPS activator which promotes the degradation of pathogenic proteinα-Syn in cellular and animal PD models.Our study thus high-lights the therapeutic potential of small molecular UPS activator like T-006 in the treatment of PD and related conditions.
基金Supported by Instituto de Salud Carlos III(ISCIII)through the FIS project PI12/0056,co-funded by FEDER from Regional Development European Funds(European Union)
文摘To compare liver proteolysis and proteasome activation in steatotic liver grafts conserved in University of Wisconsin(UW)and Institut Georges Lopez-1(IGL-1)solutions.METHODSFatty liver grafts from male obese Zücker rats were conserved in UW and IGL-1 solutions for 24 h at 4°Cand subjected to“ex vivo”normo-thermic perfusion(2 h;37°C).Liver proteolysis in tissue specimens and perfusate was measured by reverse-phase high performance liquid chromatography.Total free amino acid release was correlated with the activation of the ubiquitin proteasome system(UPS:measured as chymotryptic-like activity and 20S and 19S proteasome),the prevention of liver injury(transaminases),mitochondrial injury(confocal microscopy)and inflammation markers(TNF 1 alpha,high mobility group box-1(HGMB-1)and PPAR gamma),and liver apoptosis(TUNEL assay,cytochrome c and caspase 3).RESULTSProfiles of free AA(alanine,proline,leucine,isoleucine,methionine,lysine,ornithine,and threonine,among others)were similar for tissue and reperfusion effluent.In all cases,the IGL-1 solution showed a significantly higher prevention of proteolysis than UW(P<0.05)after cold ischemia reperfusion.Livers conserved in IGL-1 presented more effective prevention of ATP-breakdown and more inhibition of UPS activity(measured as chymotryptic-like activity).In addition,the prevention of liver proteolysis and UPS activation correlated with the prevention of liver injury(AST/ALT)and mitochondrial damage(revealed by confocal microscopy findings)as well as with the prevention of inflammatory markers(TNF1alpha and HMGB)after reperfusion.In addition,the liver grafts preserved in IGL-1 showed a significant decrease in liver apoptosis,as shown by TUNEL assay and the reduction of cytochrome c,caspase 3 and P62 levels.CONCLUSIONOur comparison of these two preservation solutions suggests that IGL-1 helps to prevent ATP breakdown more effectively than UW and subsequently achieves a higher UPS inhibition and reduced liver proteolysis.
基金supported by grants from the Undergraduate Research and Innovation Project of University of South China(Nos.X202110555528,S202210555245,and X202210555136)
文摘Cancer is a major global health issue.Effective therapeutic strategies can prolong patients’survival and reduce the costs of treatment.Drug repurposing,which identifies new therapeutic uses for approved drugs,is a promising approach with the advantages of reducing research costs,shortening development time,and increasing efficiency and safety.Disulfiram(DSF),a Food and Drug Administration(FDA)-approved drug used to treat chronic alcoholism,has a great potential as an anticancer drug by targeting diverse human malignancies.Several studies show the antitumor effects of DSF,particularly the combination of DSF and copper(DSF/Cu),on a wide range of cancers such as glioblastoma(GBM),breast cancer,liver cancer,pancreatic cancer,and melanoma.In this review,we summarize the antitumor mechanisms of DSF/Cu,including induction of intracellular reactive oxygen species(ROS)and various cell death signaling pathways,and inhibition of proteasome activity,as well as inhibition of nuclear factor-kappa B(NF-κB)signaling.Furthermore,we highlight the ability of DSF/Cu to target cancer stem cells(CSCs),which provides a new approach to prevent tumor recurrence and metastasis.Strikingly,DSF/Cu inhibits several molecular targets associated with drug resistance,and therefore it is becoming a novel option to increase the sensitivity of chemo-resistant and radio-resistant patients.Studies of DSF/Cu may shed light on its improved application to clinical tumor treatment.
