Background Lactate is a classical byproduct of glucose metabolism,and the main lactate production pathway depends on glycolysis.Lactate stabilized HIF1αby inhibiting PHD activity,leading to hypoxic stress response an...Background Lactate is a classical byproduct of glucose metabolism,and the main lactate production pathway depends on glycolysis.Lactate stabilized HIF1αby inhibiting PHD activity,leading to hypoxic stress response and exacerbating glycolysis in multiple tissues.However,the redox induction mechanism of lactate in mammary gland has not been understood yet.Herein,we describe a lactate-responsive HIF1α/circadian control mechanism in oxidative stress in the mammary glands of dairy cows.Results The in vivo study showed that dairy cows with high lactate concentrations are associated with reduced milk yield and more ROS accumulation in mammary gland.Western blot results in MAC-T cells showed positive correlation between lactate concentrations,expression of HIF1αand oxidative stress indicators,but not circadian core components.To test how lactate-mediated HIF1αdysfunction leads to cell protection process,we investigated altered expression of circadian core related genes following HIF1αstabilization.We found that stabilized HIF1αby lactate inhibited stimulated expression of circadian core components due to the similarity of HRE and E-box transcription elements.Furthermore,we found that lactate treatment strengthened the binding of HIF1αwith BMAL1,HMOX1 and FOXO3 in MAC-T cells.Moreover,HIF1αknockdown altered expression of circadian rhythm related genes and reduced oxidative stress state.Conclusion In summary,our study highlights the central role of competitive transcriptional element occupancy in lactate-mediated oxidative stress of mammary gland,which is caused by HIF1αstabilization and circadian rhythm dysfunction.Our findings introduce a novel nutritional strategy with potential applications in dairy farming for optimizing milk production and maintaining mammary gland health.展开更多
Background Hypoxic stimuli induce follicular atresia by regulating granulosa cell(GC)apoptosis.Notably,mature follicles can still develop and ovulate under hypoxic conditions,highlighting the importance of the hypoxic...Background Hypoxic stimuli induce follicular atresia by regulating granulosa cell(GC)apoptosis.Notably,mature follicles can still develop and ovulate under hypoxic conditions,highlighting the importance of the hypoxic adaptation in ovarian follicular selection.To date,the role and mechanism of hypoxia-inducible factor 1 subunit alpha(HIF1A)-mediated hypoxic responses in follicular atresia are unclear.This study aimed to investigate whether and how HIF1A regulates follicular atresia via the modulation of O-linked N-acetylglucosamine(O-GlcNAc)protein modification(O-GlcNAcylation).Results Our findings revealed that HIF1A was highly expressed in pig ovaries.Compared with that in healthy follicles,its expression was significantly downregulated in atretic follicles.Under hypoxic conditions,pharmacological inhibition or siRNA-mediated knockdown of HIF1A increased porcine GC apoptosis.Mechanistically,HIF1A knockdown Suppressed O-GlcNAc transferase degradation,leading to increased global O-GlcNAcylation.Using 4D labelfree quantitative proteomics,we identified 53 O-GlcNAcylated proteins.Importantly,O-GlcNAcylation stabilized vascular endothelial zinc finger 1(VEZF1),and HIF1A knockdown upregulated VEZF1 protein levels by promoting O-GlcNAcylation.The HIF1A-VEZF1 axis modulates forkhead box O1(FOXO1)expression by regulating endothelin-1.As a transcription factor,FOXO1 directly binds to the Bcl-2 associated X(BAX)promoter,activating its transcription and ultimately inducing porcine GC apoptosis and follicular atresia.Conclusion Overall,our study elucidates a novel molecular mechanism by which HIF1A deficiency modulates follicular atresia through O-GlcNAcylation-mediated VEZF1 expression.These results not only clarify the molecular mechanism of ovarian follicular development under hypoxic conditions but also offer potential targets for improving follicular selection efficiency in pig breeding.展开更多
Osteocytes are the main cells in mineralized bone tissue.Elevated osteocyte apoptosis has been observed in lytic bone lesions of patients with multiple myeloma.However,their precise contribution to bone metastasis rem...Osteocytes are the main cells in mineralized bone tissue.Elevated osteocyte apoptosis has been observed in lytic bone lesions of patients with multiple myeloma.However,their precise contribution to bone metastasis remains unclear.Here,we investigated the pathogenic mechanisms driving melanoma-induced osteocyte death.Both in vivo models and in vitro assays were combined with untargeted RNA sequencing approaches to explore the pathways governing melanoma-induced osteocyte death.We could show that ferroptosis is the primary mechanism behind osteocyte death in the context of melanoma bone metastasis.HMOX1 was identified as a crucial regulatory factor in this process,directly involved in inducing ferroptosis and affecting osteocyte viability.We uncover a non-canonical pathway that involves excessive autophagy-mediated ferritin degradation,highlighting the complex relationship between autophagy and ferroptosis in melanoma-induced osteocyte death.In addition,HIF1αpathway was shown as an upstream regulator,providing a potential target for modulating HMOX1 expression and influencing autophagy-dependent ferroptosis.In conclusion,our study provides insight into the pathogenic mechanisms of osteocyte death induced by melanoma bone metastasis,with a specific focus on ferroptosis and its regulation.This would enhance our comprehension of melanoma-induced osteocyte death.展开更多
Cervical cancer is a major malignancy that poses a significant threat to women's health[1].In 2020,an estimated 604,000 new cases and 342,000 deaths were reported globally[2].The most common pathological subtype i...Cervical cancer is a major malignancy that poses a significant threat to women's health[1].In 2020,an estimated 604,000 new cases and 342,000 deaths were reported globally[2].The most common pathological subtype is squamous cell carcinoma[3,4].However,treatment options for advanced cervical squamous cell carcinoma(CSCC)are limited.Surgery is often not feasible at this stage,resulting in poor prognosis[5,6].Therefore,identifying novel molecular markers and elucidating the mechanisms that drive CSCC growth and metastasis are crucial for improving treatment outcomes.展开更多
Objective To investigate the chemical compositions of Maxing Shigan Decoction(麻杏石甘汤,MXSGD)and elucidate its anti-influenza A virus(IAV)mechanism from prediction to validation.Methods Ultra high-performance liquid...Objective To investigate the chemical compositions of Maxing Shigan Decoction(麻杏石甘汤,MXSGD)and elucidate its anti-influenza A virus(IAV)mechanism from prediction to validation.Methods Ultra high-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS)was employed to analyze the chemical compositions of MXSGD.Network pharmacology theories were used to screen and identify shared targets of both the potential targets of active ingredients of MXSGD and IAV.A protein-protein interaction(PPI)network was then constructed,followed by Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analyses.The binding stability between core bioactive compounds and key targets was validated by molecular docking and dynamic simulations.A total of 24 BALB/c mice were infected with IAV to build IAV mouse models.After successful modelling,the mouse models were randomly divided into model,MXSGD high-dose(2.8 g/kg),MXSGD low-dose(1.4 g/kg),and oseltamivir(20.14 mg/kg)groups,with an additional normal mice as control group(n=6 per group).The treatments were administered by gavage daily between 8:00 a.m.and 10:00 a.m.for five consecutive days.Upon completion of the administration,the body weight ratio,lung index,protein content in the bronchoalveolar lavage fluid(BALF),and the levels of inflammatory factors including interleukin(IL)-6 and tumor necrosis factor(TNF)-αin mice were measured to preliminarily analyze the therapeutic efficacy of MXSGD against IAV infection.Furthermore,the expression levels of mechanistic target of rapamycin(mTOR),hypoxia inducible factor(HIF)-1α,and vascular endothelial growth factor(VEGF)proteins in the HIF-1 signaling pathway,which was enriched by network pharmacology,were detected by Western blot.Results A total of 212 chemical components in MXSGD were identified by the UPLC-MS/MS method.These chemical components can be classified into 9 primary categories and 31 secondary categories.After intersecting the chemical component targets with IAV-related targets,a total of 567 potential MXSGD components targeting IAV were identified.The construction of PPI network and the results of both GO and KEGG enrichment analyses revealed that the anti-IAV effects of MXSGD were associated with multiple pathways,including apoptosis,TNF,HIF-1,and IL-17 signaling pathways.The results of molecular docking demonstrated that the binding energies between the core compound 1-methoxyphaseollin and key targets including HIF-1α,mTOR,and VEGF were all lower than–5.0 kcal/mol.Furthermore,molecular dynamics simulations confirmed the structural stability of the resulting complexes.Animal experiments showed that compared with the normal controls,IAV-infected mice showed significantly reduced body weight ratio,markedly increased lung index,protein content in BALF,and the levels of inflammatory factors such as IL-6 and TNF-α(P<0.01),thereby causing damage to the lung tissue;consequently,the expression levels of mTOR,HIF-1α,and VEGF proteins in the lung tissues of these mice were significantly elevated(P<0.01).However,after MXSGD treatment,the mouse models presented a significant increase in body weight ratio,as well as marked decreases in lung index,protein content in BALF,and the levels of inflammatory factors including IL-6 and TNF-α(P<0.01).Furthermore,the therapy alleviated IAV-induced injuries and significantly downregulated the expression levels of mTOR,HIF-1α,and VEGF proteins in lung tissues(P<0.01 or P<0.05).Conclusion MXSGD exerts anti-IAV effects through multi-component,multi-target,and multi-pathway synergism.Among them,1-methoxyphaseollin is identified as a potential key component,which alleviates virus-induced lung injury and inflammatory response via the regulation of HIF-1 signaling pathway,providing experimental evidence for the clinical application of MXSGD.展开更多
基金supported by National Nature Science Foundation of China(32102552 and 32172741).
文摘Background Lactate is a classical byproduct of glucose metabolism,and the main lactate production pathway depends on glycolysis.Lactate stabilized HIF1αby inhibiting PHD activity,leading to hypoxic stress response and exacerbating glycolysis in multiple tissues.However,the redox induction mechanism of lactate in mammary gland has not been understood yet.Herein,we describe a lactate-responsive HIF1α/circadian control mechanism in oxidative stress in the mammary glands of dairy cows.Results The in vivo study showed that dairy cows with high lactate concentrations are associated with reduced milk yield and more ROS accumulation in mammary gland.Western blot results in MAC-T cells showed positive correlation between lactate concentrations,expression of HIF1αand oxidative stress indicators,but not circadian core components.To test how lactate-mediated HIF1αdysfunction leads to cell protection process,we investigated altered expression of circadian core related genes following HIF1αstabilization.We found that stabilized HIF1αby lactate inhibited stimulated expression of circadian core components due to the similarity of HRE and E-box transcription elements.Furthermore,we found that lactate treatment strengthened the binding of HIF1αwith BMAL1,HMOX1 and FOXO3 in MAC-T cells.Moreover,HIF1αknockdown altered expression of circadian rhythm related genes and reduced oxidative stress state.Conclusion In summary,our study highlights the central role of competitive transcriptional element occupancy in lactate-mediated oxidative stress of mammary gland,which is caused by HIF1αstabilization and circadian rhythm dysfunction.Our findings introduce a novel nutritional strategy with potential applications in dairy farming for optimizing milk production and maintaining mammary gland health.
基金supported by the National Natural Science Foundation of China(32402710)National Postdoctoral Researchers Funding Program(GZC20232209)+1 种基金Higher Education Institutions Basic Science(Natural Science)Research General Program of Jiangsu Province(24KJB230005)Priority Academic Program Development of Jiangsu Higher Education Institution.
文摘Background Hypoxic stimuli induce follicular atresia by regulating granulosa cell(GC)apoptosis.Notably,mature follicles can still develop and ovulate under hypoxic conditions,highlighting the importance of the hypoxic adaptation in ovarian follicular selection.To date,the role and mechanism of hypoxia-inducible factor 1 subunit alpha(HIF1A)-mediated hypoxic responses in follicular atresia are unclear.This study aimed to investigate whether and how HIF1A regulates follicular atresia via the modulation of O-linked N-acetylglucosamine(O-GlcNAc)protein modification(O-GlcNAcylation).Results Our findings revealed that HIF1A was highly expressed in pig ovaries.Compared with that in healthy follicles,its expression was significantly downregulated in atretic follicles.Under hypoxic conditions,pharmacological inhibition or siRNA-mediated knockdown of HIF1A increased porcine GC apoptosis.Mechanistically,HIF1A knockdown Suppressed O-GlcNAc transferase degradation,leading to increased global O-GlcNAcylation.Using 4D labelfree quantitative proteomics,we identified 53 O-GlcNAcylated proteins.Importantly,O-GlcNAcylation stabilized vascular endothelial zinc finger 1(VEZF1),and HIF1A knockdown upregulated VEZF1 protein levels by promoting O-GlcNAcylation.The HIF1A-VEZF1 axis modulates forkhead box O1(FOXO1)expression by regulating endothelin-1.As a transcription factor,FOXO1 directly binds to the Bcl-2 associated X(BAX)promoter,activating its transcription and ultimately inducing porcine GC apoptosis and follicular atresia.Conclusion Overall,our study elucidates a novel molecular mechanism by which HIF1A deficiency modulates follicular atresia through O-GlcNAcylation-mediated VEZF1 expression.These results not only clarify the molecular mechanism of ovarian follicular development under hypoxic conditions but also offer potential targets for improving follicular selection efficiency in pig breeding.
基金funding from the European Research Council(ERC)under the european union Horizon 2020 research and innovation program(grant agreement ERC co-LS4 ODE(AB)and ERC Synergy Grant 4D Nanoscope(GS))Deutsche Forschungsgemeinschaft DFG-Project number 501752319-TRR369-DIONE-Project No A02 and B05,FOR 2886(TP02),CRC1181(TPA01)DFG funding(450993414)Thunder Imager and the Leibniz Award(GS)。
文摘Osteocytes are the main cells in mineralized bone tissue.Elevated osteocyte apoptosis has been observed in lytic bone lesions of patients with multiple myeloma.However,their precise contribution to bone metastasis remains unclear.Here,we investigated the pathogenic mechanisms driving melanoma-induced osteocyte death.Both in vivo models and in vitro assays were combined with untargeted RNA sequencing approaches to explore the pathways governing melanoma-induced osteocyte death.We could show that ferroptosis is the primary mechanism behind osteocyte death in the context of melanoma bone metastasis.HMOX1 was identified as a crucial regulatory factor in this process,directly involved in inducing ferroptosis and affecting osteocyte viability.We uncover a non-canonical pathway that involves excessive autophagy-mediated ferritin degradation,highlighting the complex relationship between autophagy and ferroptosis in melanoma-induced osteocyte death.In addition,HIF1αpathway was shown as an upstream regulator,providing a potential target for modulating HMOX1 expression and influencing autophagy-dependent ferroptosis.In conclusion,our study provides insight into the pathogenic mechanisms of osteocyte death induced by melanoma bone metastasis,with a specific focus on ferroptosis and its regulation.This would enhance our comprehension of melanoma-induced osteocyte death.
基金supported by the Hebei Provincial Central Guidance Local Science and Technology Development Fund(grant number 236Z7714G).
文摘Cervical cancer is a major malignancy that poses a significant threat to women's health[1].In 2020,an estimated 604,000 new cases and 342,000 deaths were reported globally[2].The most common pathological subtype is squamous cell carcinoma[3,4].However,treatment options for advanced cervical squamous cell carcinoma(CSCC)are limited.Surgery is often not feasible at this stage,resulting in poor prognosis[5,6].Therefore,identifying novel molecular markers and elucidating the mechanisms that drive CSCC growth and metastasis are crucial for improving treatment outcomes.
基金Natural Science Foundation of Hunan Province(2025JJ80078)Open Fund of Hunan University of Chinese Medicine(21PTKF1005)。
文摘Objective To investigate the chemical compositions of Maxing Shigan Decoction(麻杏石甘汤,MXSGD)and elucidate its anti-influenza A virus(IAV)mechanism from prediction to validation.Methods Ultra high-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS)was employed to analyze the chemical compositions of MXSGD.Network pharmacology theories were used to screen and identify shared targets of both the potential targets of active ingredients of MXSGD and IAV.A protein-protein interaction(PPI)network was then constructed,followed by Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analyses.The binding stability between core bioactive compounds and key targets was validated by molecular docking and dynamic simulations.A total of 24 BALB/c mice were infected with IAV to build IAV mouse models.After successful modelling,the mouse models were randomly divided into model,MXSGD high-dose(2.8 g/kg),MXSGD low-dose(1.4 g/kg),and oseltamivir(20.14 mg/kg)groups,with an additional normal mice as control group(n=6 per group).The treatments were administered by gavage daily between 8:00 a.m.and 10:00 a.m.for five consecutive days.Upon completion of the administration,the body weight ratio,lung index,protein content in the bronchoalveolar lavage fluid(BALF),and the levels of inflammatory factors including interleukin(IL)-6 and tumor necrosis factor(TNF)-αin mice were measured to preliminarily analyze the therapeutic efficacy of MXSGD against IAV infection.Furthermore,the expression levels of mechanistic target of rapamycin(mTOR),hypoxia inducible factor(HIF)-1α,and vascular endothelial growth factor(VEGF)proteins in the HIF-1 signaling pathway,which was enriched by network pharmacology,were detected by Western blot.Results A total of 212 chemical components in MXSGD were identified by the UPLC-MS/MS method.These chemical components can be classified into 9 primary categories and 31 secondary categories.After intersecting the chemical component targets with IAV-related targets,a total of 567 potential MXSGD components targeting IAV were identified.The construction of PPI network and the results of both GO and KEGG enrichment analyses revealed that the anti-IAV effects of MXSGD were associated with multiple pathways,including apoptosis,TNF,HIF-1,and IL-17 signaling pathways.The results of molecular docking demonstrated that the binding energies between the core compound 1-methoxyphaseollin and key targets including HIF-1α,mTOR,and VEGF were all lower than–5.0 kcal/mol.Furthermore,molecular dynamics simulations confirmed the structural stability of the resulting complexes.Animal experiments showed that compared with the normal controls,IAV-infected mice showed significantly reduced body weight ratio,markedly increased lung index,protein content in BALF,and the levels of inflammatory factors such as IL-6 and TNF-α(P<0.01),thereby causing damage to the lung tissue;consequently,the expression levels of mTOR,HIF-1α,and VEGF proteins in the lung tissues of these mice were significantly elevated(P<0.01).However,after MXSGD treatment,the mouse models presented a significant increase in body weight ratio,as well as marked decreases in lung index,protein content in BALF,and the levels of inflammatory factors including IL-6 and TNF-α(P<0.01).Furthermore,the therapy alleviated IAV-induced injuries and significantly downregulated the expression levels of mTOR,HIF-1α,and VEGF proteins in lung tissues(P<0.01 or P<0.05).Conclusion MXSGD exerts anti-IAV effects through multi-component,multi-target,and multi-pathway synergism.Among them,1-methoxyphaseollin is identified as a potential key component,which alleviates virus-induced lung injury and inflammatory response via the regulation of HIF-1 signaling pathway,providing experimental evidence for the clinical application of MXSGD.
