Rheumatoid arthritis(RA)is a progressive autoimmune disease characterized by bone destruction that is primarily caused by the overactivation of osteoclasts(OCs),which are critical therapeutic targets.Triptolide(TP)has...Rheumatoid arthritis(RA)is a progressive autoimmune disease characterized by bone destruction that is primarily caused by the overactivation of osteoclasts(OCs),which are critical therapeutic targets.Triptolide(TP)has strong anti-RA effects but is limited by its narrow therapeutic window and associated toxicity,necessitating combination therapy to increase its efficacy and reduce side effects.Medicarpin(Med),a flavonoid with anti-inflammatory and anti-bone destruction properties,has shown potential in reducing osteoclastogenesis.However,the mechanisms underlying the synergistic effects of TP and Med on RA treatment remain unclear.We addressed this issue by evaluating the effects of TP,Med,and their combination on a collagen-induced arthritis(CIA)rat model,with a focus on bone erosion as the primary research endpoint.We subsequently performed experimental validation in an in vitro OC dif-ferentiation model to assess the impacts of these treatments on OC formation and function.Based on polymerase chain reaction(PCR)microarray data from RA patients,further investigations focused on N^(6)-methyladenosine(m^(6)A)methylation and its regulatory factors,methyltransferase-like 3(METTL3)and YT521-B homology domain family protein 1(YTHDF1),which have been identified as potential tar-gets of TP and Med.Key findings revealed that the TP and Med combination significantly alleviated bone destruction and inhibited OC differentiation,exerting stronger effects at lower doses than either drug alone.Mechanistically,TP and Med synergistically modulated METTL3 and YTHDF1 to suppress osteo-clastogenesis through distinct m6 A methylation pathways,contributing to the mitigation of RA-associated bone destruction.Overall,our data highlight the potential of the m^(6)A modification as a ther-apeutic mechanism for the combined use of TP and Med for RA treatment,providing a theoretical basis for the clinical application of herbal active ingredient combinations.展开更多
Background Liver ischemia/reperfusion(I/R)injury is usually caused by hepatic inflow occlusion during liver surgery,and is frequently observed during war wounds and trauma.Hepatocyte ferroptosis plays a critical role ...Background Liver ischemia/reperfusion(I/R)injury is usually caused by hepatic inflow occlusion during liver surgery,and is frequently observed during war wounds and trauma.Hepatocyte ferroptosis plays a critical role in liver I/R injury,however,it remains unclear whether this process is controlled or regulated by members of the DEAD/DExH-box helicase(DDX/DHX)family.Methods The expression of DDX/DHX family members during liver I/R injury was screened using transcriptome analysis.Hepatocyte-specific Dhx58 knockout mice were constructed,and a partial liver I/R operation was performed.Single-cell RNA sequencing(scRNA-seq)in the liver post I/R suggested enhanced ferroptosis by Dhx58hep−/−.The mRNAs and proteins associated with DExH-box helicase 58(DHX58)were screened using RNA immunoprecipitation-sequencing(RIP-seq)and IP-mass spectrometry(IP-MS).Results Excessive production of reactive oxygen species(ROS)decreased the expression of the IFN-stimulated gene Dhx58 in hepatocytes and promoted hepatic ferroptosis,while treatment using IFN-αincreased DHX58 expression and prevented ferroptosis during liver I/R injury.Mechanistically,DHX58 with RNA-binding activity constitutively associates with the mRNA of glutathione peroxidase 4(GPX4),a central ferroptosis suppressor,and recruits the m6A reader YT521-B homology domain containing 2(YTHDC2)to promote the translation of Gpx4 mRNA in an m6A-dependent manner,thus enhancing GPX4 protein levels and preventing hepatic ferroptosis.Conclusions This study provides mechanistic evidence that IFN-αstimulates DHX58 to promote the translation of m6A-modified Gpx4 mRNA,suggesting the potential clinical application of IFN-αin the prevention of hepatic ferroptosis during liver I/R injury.展开更多
基金supported by the National Natural Science Foundation of China(U22A20374).
文摘Rheumatoid arthritis(RA)is a progressive autoimmune disease characterized by bone destruction that is primarily caused by the overactivation of osteoclasts(OCs),which are critical therapeutic targets.Triptolide(TP)has strong anti-RA effects but is limited by its narrow therapeutic window and associated toxicity,necessitating combination therapy to increase its efficacy and reduce side effects.Medicarpin(Med),a flavonoid with anti-inflammatory and anti-bone destruction properties,has shown potential in reducing osteoclastogenesis.However,the mechanisms underlying the synergistic effects of TP and Med on RA treatment remain unclear.We addressed this issue by evaluating the effects of TP,Med,and their combination on a collagen-induced arthritis(CIA)rat model,with a focus on bone erosion as the primary research endpoint.We subsequently performed experimental validation in an in vitro OC dif-ferentiation model to assess the impacts of these treatments on OC formation and function.Based on polymerase chain reaction(PCR)microarray data from RA patients,further investigations focused on N^(6)-methyladenosine(m^(6)A)methylation and its regulatory factors,methyltransferase-like 3(METTL3)and YT521-B homology domain family protein 1(YTHDF1),which have been identified as potential tar-gets of TP and Med.Key findings revealed that the TP and Med combination significantly alleviated bone destruction and inhibited OC differentiation,exerting stronger effects at lower doses than either drug alone.Mechanistically,TP and Med synergistically modulated METTL3 and YTHDF1 to suppress osteo-clastogenesis through distinct m6 A methylation pathways,contributing to the mitigation of RA-associated bone destruction.Overall,our data highlight the potential of the m^(6)A modification as a ther-apeutic mechanism for the combined use of TP and Med for RA treatment,providing a theoretical basis for the clinical application of herbal active ingredient combinations.
基金National Key Research and Development Program of China(2023YFC2505900)National Natural Science Foundation of China(92269204,82171755,92369106,82171749,82171811,82073184)+1 种基金Military Outstanding Youth Program(2020QN06119,01-SWK JYCJJ07,23SWAQ53)Program of Leading Talents in Shanghai,and Shanghai Shuguang Program(20SG39)。
文摘Background Liver ischemia/reperfusion(I/R)injury is usually caused by hepatic inflow occlusion during liver surgery,and is frequently observed during war wounds and trauma.Hepatocyte ferroptosis plays a critical role in liver I/R injury,however,it remains unclear whether this process is controlled or regulated by members of the DEAD/DExH-box helicase(DDX/DHX)family.Methods The expression of DDX/DHX family members during liver I/R injury was screened using transcriptome analysis.Hepatocyte-specific Dhx58 knockout mice were constructed,and a partial liver I/R operation was performed.Single-cell RNA sequencing(scRNA-seq)in the liver post I/R suggested enhanced ferroptosis by Dhx58hep−/−.The mRNAs and proteins associated with DExH-box helicase 58(DHX58)were screened using RNA immunoprecipitation-sequencing(RIP-seq)and IP-mass spectrometry(IP-MS).Results Excessive production of reactive oxygen species(ROS)decreased the expression of the IFN-stimulated gene Dhx58 in hepatocytes and promoted hepatic ferroptosis,while treatment using IFN-αincreased DHX58 expression and prevented ferroptosis during liver I/R injury.Mechanistically,DHX58 with RNA-binding activity constitutively associates with the mRNA of glutathione peroxidase 4(GPX4),a central ferroptosis suppressor,and recruits the m6A reader YT521-B homology domain containing 2(YTHDC2)to promote the translation of Gpx4 mRNA in an m6A-dependent manner,thus enhancing GPX4 protein levels and preventing hepatic ferroptosis.Conclusions This study provides mechanistic evidence that IFN-αstimulates DHX58 to promote the translation of m6A-modified Gpx4 mRNA,suggesting the potential clinical application of IFN-αin the prevention of hepatic ferroptosis during liver I/R injury.
