Background This study aimed to determine the effects of tumor necrosis factor(TNF-a) on endothelial cytoskeleton morphology and permeability,and to detect the underlying signaling mechanisms involved in these response...Background This study aimed to determine the effects of tumor necrosis factor(TNF-a) on endothelial cytoskeleton morphology and permeability,and to detect the underlying signaling mechanisms involved in these responses. Methods Cultured endothelial cells(ECs) were exposed to TNF-a,and EC cytoskeletal changes were evaluated by observing fluorescence of F-actin following ligation with labeled antibodies.Endothelial permeability was detected by measuring the flux of HRP-albumin across the EC monolayers.To explore the signaling pathways behind TNF-a-induced EC alteration, ECs were treated with either the RhoGTPase inhibitor Y27632 or the MAPK inhibitors PD98059 and SB203580 before TNF-a administration.To further elucidate possible involvement of the RhoA and ERK pathways in TNF-induced EC changes,retrovirus-carried recombinant dominant-negative forms and constitutive-activative forms of RhoA,namely T19NRhoA and Q63LRhoA,were pre-infect-ed into ECs prior to TNF-a exposure.Results TNF-a induced F-actin cytoskeleton rearrangement,as well as EC hyperpermeability in a dose and time-dependent manner.The effects were attenuated in cells pretreated with Y27632 or PD98059,respectively.EC pre-infection with T19NRhoA also alleviated the effects of TNF-a.Furthermore,retrovirus-mediated administration of activated forms of Q63LRhoA alone induced rearrangement of F-actin and hyperpermeability as well as induced the activation of pERK.Conclusions These results indicate that RhoA-ERK/MAPK signal pathway play important roles in the mediation of TNF-a induced EC barrier dysfunction associated with morphological changes of the Factin.展开更多
Disruption of the blood-spinal cord barrier(BSCB)is a critical event in the secondary injury following spinal cord injury(SCI).Mertk has been reported to play an important role in regulating inflammation and cytoskele...Disruption of the blood-spinal cord barrier(BSCB)is a critical event in the secondary injury following spinal cord injury(SCI).Mertk has been reported to play an important role in regulating inflammation and cytoskeletal dynamics.However,the specific involvement of Mertk in BSCB remains elusive.Here,we demonstrated a distinct role of Mertk in the repair of BSCB.Mertk expression is decreased in endothelial cells following SCI.Overexpression of Mertk upregulated tight junction proteins(TJs),reducing BSCB permeability and subsequently inhibiting inflammation and apoptosis.Ultimately,this led to enhanced neural regeneration and functional recovery.Further experiments revealed that the RhoA/Rock1/P-MLC pathway plays a key role in the effects of Mertk.These findings highlight the role of Mertk in promoting SCI recovery through its ability to mitigate BSCB permeability and may provide potential targets for SCI repair.展开更多
Osteosarcoma is the most common malignant bone tumour,and the metastasis of osteosarcoma is an important cause of death.Evidence has shown that the mevalonate pathway is highly activated and is expected to be a new ta...Osteosarcoma is the most common malignant bone tumour,and the metastasis of osteosarcoma is an important cause of death.Evidence has shown that the mevalonate pathway is highly activated and is expected to be a new target for tumour therapy.In this study,we investigated the effect of mevalonate signalling on osteosarcoma metastasis and its molecular mechanism.First,we found that the key rate-limiting enzyme of mevalonate signalling,3-hydroxy-3-methylglutaryl-CoA reductase(HMGCR),was highly expressed in osteosarcoma cells,and inhibition of HMGCR with simvastatin significantly inhibited the motility of 143B cells.Next,we found that YAP1 activity was significantly upregulated in osteosarcoma cells and that YAP1 knockdown inhibited the motility of 143B cells.We also found that the mevalonate pathway regulated the motility of 143B cells by modulating YAP1 phosphorylation and cellular localization.Moreover,we found that the activity of YAP1 was regulated by the mevalonate pathway by modulating the cell membrane localization of RhoA.Finally,we demonstrated that inhibition of the mevalonate pathway notably reduced the lung metastasis of 143B cells,as reflected by the decreased incidence and number of metastatic nodules and the increased survival time of the nude mice.Taken together,our findings suggest that the mevalonate pathway can promote the metastasis of osteosarcoma by activating YAP1 via RhoA.Inhibition of the mevalonate pathway may be a promising therapeutic strategy for osteosarcoma metastasis.展开更多
文摘Background This study aimed to determine the effects of tumor necrosis factor(TNF-a) on endothelial cytoskeleton morphology and permeability,and to detect the underlying signaling mechanisms involved in these responses. Methods Cultured endothelial cells(ECs) were exposed to TNF-a,and EC cytoskeletal changes were evaluated by observing fluorescence of F-actin following ligation with labeled antibodies.Endothelial permeability was detected by measuring the flux of HRP-albumin across the EC monolayers.To explore the signaling pathways behind TNF-a-induced EC alteration, ECs were treated with either the RhoGTPase inhibitor Y27632 or the MAPK inhibitors PD98059 and SB203580 before TNF-a administration.To further elucidate possible involvement of the RhoA and ERK pathways in TNF-induced EC changes,retrovirus-carried recombinant dominant-negative forms and constitutive-activative forms of RhoA,namely T19NRhoA and Q63LRhoA,were pre-infect-ed into ECs prior to TNF-a exposure.Results TNF-a induced F-actin cytoskeleton rearrangement,as well as EC hyperpermeability in a dose and time-dependent manner.The effects were attenuated in cells pretreated with Y27632 or PD98059,respectively.EC pre-infection with T19NRhoA also alleviated the effects of TNF-a.Furthermore,retrovirus-mediated administration of activated forms of Q63LRhoA alone induced rearrangement of F-actin and hyperpermeability as well as induced the activation of pERK.Conclusions These results indicate that RhoA-ERK/MAPK signal pathway play important roles in the mediation of TNF-a induced EC barrier dysfunction associated with morphological changes of the Factin.
基金Natural Science Foundation of Guangdong Province(2017A030313111)National Natural Science Foundation of China(81974329).
文摘Disruption of the blood-spinal cord barrier(BSCB)is a critical event in the secondary injury following spinal cord injury(SCI).Mertk has been reported to play an important role in regulating inflammation and cytoskeletal dynamics.However,the specific involvement of Mertk in BSCB remains elusive.Here,we demonstrated a distinct role of Mertk in the repair of BSCB.Mertk expression is decreased in endothelial cells following SCI.Overexpression of Mertk upregulated tight junction proteins(TJs),reducing BSCB permeability and subsequently inhibiting inflammation and apoptosis.Ultimately,this led to enhanced neural regeneration and functional recovery.Further experiments revealed that the RhoA/Rock1/P-MLC pathway plays a key role in the effects of Mertk.These findings highlight the role of Mertk in promoting SCI recovery through its ability to mitigate BSCB permeability and may provide potential targets for SCI repair.
基金This work was supported by the Natural Science Foundation of Chongqing(No.cstc2019jcyj-msxmX0358).
文摘Osteosarcoma is the most common malignant bone tumour,and the metastasis of osteosarcoma is an important cause of death.Evidence has shown that the mevalonate pathway is highly activated and is expected to be a new target for tumour therapy.In this study,we investigated the effect of mevalonate signalling on osteosarcoma metastasis and its molecular mechanism.First,we found that the key rate-limiting enzyme of mevalonate signalling,3-hydroxy-3-methylglutaryl-CoA reductase(HMGCR),was highly expressed in osteosarcoma cells,and inhibition of HMGCR with simvastatin significantly inhibited the motility of 143B cells.Next,we found that YAP1 activity was significantly upregulated in osteosarcoma cells and that YAP1 knockdown inhibited the motility of 143B cells.We also found that the mevalonate pathway regulated the motility of 143B cells by modulating YAP1 phosphorylation and cellular localization.Moreover,we found that the activity of YAP1 was regulated by the mevalonate pathway by modulating the cell membrane localization of RhoA.Finally,we demonstrated that inhibition of the mevalonate pathway notably reduced the lung metastasis of 143B cells,as reflected by the decreased incidence and number of metastatic nodules and the increased survival time of the nude mice.Taken together,our findings suggest that the mevalonate pathway can promote the metastasis of osteosarcoma by activating YAP1 via RhoA.Inhibition of the mevalonate pathway may be a promising therapeutic strategy for osteosarcoma metastasis.
