Objective Tyro-3 and Axl receptors are expressed in brain in a region-specific manner and their bioactivities in the central nervous system remain still elusive.The aim of the present study was to investigate their fu...Objective Tyro-3 and Axl receptors are expressed in brain in a region-specific manner and their bioactivities in the central nervous system remain still elusive.The aim of the present study was to investigate their functions in neuronal differentiation.Methods PC12 cells overexpressing Tyro-3 or Axl were established by transfection with full-length CMV-Tyro3-eCFP or CMV-Axl-eGFP plasmid,respectively.CMV-eGFP plasmid served as a control vector.After that,the fluorescence intensity and distributions of green fluorescent protein （GFP） and cyan fluorescent protein （CFP） in the cells with or without nerve growth factor （NGF） treatment were real-time monitored.Results Expressions of Tyro-3 and Axl receptors were under the regulation of NGF and associated with neuronal differentiation.This was not observed in CMV-eGFP-transfected PC12 cells.Besides,confocal microscopy revealed that NGF affected intracellular localization of full-length Axl-eGFP and Tyro-3eCFP in PC12 cells.Moreover,the development of outgrowth of differentiated PC12 cells under stimulation of NGF was promoted by overexpression of Tyro-3 or Axl.Conclusion Expressions of Tyro-3 and Axl receptors are under the regulation of NGF and are involved in NGF-induced neuronal differentiation of PC12 cells.展开更多
In this study, we investigated the role of the TYRO3/Akt signaling pathway in hypoxic injury to hippocampal neurons. 3-（4,5-Dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide assay showed that hypoxia inhibited t...In this study, we investigated the role of the TYRO3/Akt signaling pathway in hypoxic injury to hippocampal neurons. 3-（4,5-Dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide assay showed that hypoxia inhibited the proliferation and viability of hippocampal neurons. Terminal deoxynucleotidyl transferase-mediated d UTP nick end labeling assay demonstrated that hypoxia induced neuronal apoptosis in a time-dependent manner, with a greater number of apoptotic cells with longer hypoxic exposure. Immunofluorescence labeling revealed that hypoxia suppressed TYRO3 expression. Western blot assay showed that hypoxia decreased Akt phosphorylation levels in a time-dependent manner. Taken together, these findings suggest that hypoxia inhibits the proliferation of hippocampal neurons and promotes apoptosis, and that the inhibition of the TYRO3/Akt signaling pathway plays an important role in hypoxia-induced neuronal injury.展开更多
Objective To study the effects of TYRO protein kinase-binding protein(TYROBP)deficiency on learning behavior,glia activation and pro-inflammatory cycokines,and Tau phosphorylation of a new Alzheimer’s disease(AD)mous...Objective To study the effects of TYRO protein kinase-binding protein(TYROBP)deficiency on learning behavior,glia activation and pro-inflammatory cycokines,and Tau phosphorylation of a new Alzheimer’s disease(AD)mouse model carrying a PSEN1 p.G378E mutation.Methods A new AD mouse model carrying PSEN1 p.G378E mutation was built based on our previously found AD family which might be ascribed to the PSEN1 mutation,and then crossed with TYROBP deficient mice to produce the heterozygous hybrid mice(PSEN1^(G378E)/WT;Tyrobp^(+/-))and the homozygous hybrid mice(PSEN1^(G378E/G378E);Tyrobp^(-/-)).Water maze test was used to detect spatial learning and memory ability of mice.After the mice were sacrificed,the hippocampus was excised for further analysis.Immunofluorescence was used to identify the cell that expresses TYROBP and the number of microglia and astrocyte.Western blot was used to detect the expression levels of Tau and phosphorylated Tau(p-Tau),and ELISA to measure the levels of pro-inflammatory cytokines.Results Our results showed that TYROBP specifically expressed in the microglia of mouse hippocampus.Absence of TYROBP in PSEN1^(G378E) mutation mouse model prevented the deterioration of learning behavior,decreased the numbers of microglia and astrocytes,and the levels of interleukin-6,interleukin-1βand tumor necrosis factor-αin the hippocampus(all P<0.05).The ratios of AT8/Tau5,PHF1/Tau5,pT181/Tau5,pT231/Tau5 and p-ERK/ERK were all higher in homozygous hybrid mice(PSEN1^(G378E/G378E);Tyrobp^(-/-) mice)compared with PSEN1^(G378E/G378E) mice(all P<0.05).Conclusions TYROBP deficiency might play a protective role in the modulation of neuroinflammation of AD.However,the relationship between neuroinflammation processes involving microglia and astrocyte activation,and release of pro-inflammatory cytokines,and p-Tau pathology needs further study.展开更多
The Tyro3, Axl and Mer (TAM) receptor tyrosine kinases are activated by endogenous ligands, protein S1 (PROS1) and growth arrest-specific gene 6 (Gas6), and those have important effects on cell biology. These receptor...The Tyro3, Axl and Mer (TAM) receptor tyrosine kinases are activated by endogenous ligands, protein S1 (PROS1) and growth arrest-specific gene 6 (Gas6), and those have important effects on cell biology. These receptors (Rs) can be shad from the cell membrane and their soluble(s) forms can be found in plasma. We investigated the fluctuation and interactive role of sTAMRs and its ligands in patients with hepatocellular carcinoma (HCC), hepatitis groups, and healthy normal adult controls (NC). The measurement cases were 45 patients with HCC group (stage 1 in 4, stage 2 in 8, stage 3 in 16, and stage 4 in 17), 4 patients with fulminant hepatitis (FH), 14 patients with acute hepatitis (AH), 10 patients with chronic hepatitis (CH), 16 patients with liver cirrhosis (LC), and 20 NCs matched by age. Plasma levels of three sTAMRs and their ligands were measured by ELISA. In comparison with NCs, Gas6, des-γ-carboxy Gas6, and sTAMRs levels were significantly higher in HCC patients, but free PROS1 levels were significantly lower. The sTyro3 and sAxl levels peaked HCC stages 2 and 3 respectively, and gradually decreased afterwards while maintaining high levels. sMer levels increased with the progression of HCC. Gas6 and des-γ-carboxy Gas6 levels gradually increased, and PROS1 levels decreased with the progression of HCC. Gas6 levels were positively correlated with sAxl levels, whereas sMer levels were negatively correlated with free PROS1 levels. sTAMRs and Gas6 levels increased in parallel to the progression of HCC fibrosis. Through the progression of HCC, Axl played the major role in TAMRs activation. However, sTYro3 continued increasing rapidly from the early stage, and that of Mer increased throughout the progression. Roles of Axl may be changed in Mer, because des-γ-carboxy Gas6 levels increasing with Gas6 in the advanced stage of HCC cannot send a signal to Axl.展开更多
Water deficit-induced ABA accumulation is an ideal model or 搒timulus-response?system to investigate cellular stress signaling in plant cells, using such a model the cellular stress signaling triggered by water defici...Water deficit-induced ABA accumulation is an ideal model or 搒timulus-response?system to investigate cellular stress signaling in plant cells, using such a model the cellular stress signaling triggered by water deficit was inves-tigated in Maize L. coleoptile. Water deficit-induced ABA accumulation was sensitively blocked by NaVO3, a potent inhibitor both to plasma membrane H+-ATPase (PM-H+- ATPase) and protein tyrosine phosphatase (PTPase). How-ever, while PM- H+-ATPase activity was unaffected under water deficit and PM- H+-ATPase activator did not induce an ABA accumulation instead of water deficit, water deficit induced an increase in the protein phosphatase activity, and furthermore, ABA accumulation was inhibited by PAO, a specific inhibitor of PTPase. These results indicate that pro-tein phosphtases may be involved in the cellular signaling in response to water deficit. Further studies identified at least four species of protein phosphtase as assayed by using pNPP as substrate, among which one component was especially sensitive to NaVO3. The NaVO3-sensitive enzyme was puri-fied and finally showed a protein band about 66 kD on SDS/PAGE. The purified enzyme showed a great activity to some specific PTPase substrates at pH 6.0. In addition to NaVO3, the enzyme was also sensitive to some other PTPase inhibitors such as Zn2+ and MO33+, but not to Ca2+ and Mg2+, indicating that it might be a protein tyrosine phosphatase. Interestingly, the purified enzyme could be deactivated by some reducing agent DTT, which was previously proved to be an inhibitor of water deficit-induced ABA accumulation. This result further proved that PTPase might be involved in the cellular signaling of ABA accumulation in response to water deficit.展开更多
基金supported by the National Basic Research Development Program of China(No.2006CB500700)the National Natural Science Foundation of China(No. 30900421/c090201)
文摘Objective Tyro-3 and Axl receptors are expressed in brain in a region-specific manner and their bioactivities in the central nervous system remain still elusive.The aim of the present study was to investigate their functions in neuronal differentiation.Methods PC12 cells overexpressing Tyro-3 or Axl were established by transfection with full-length CMV-Tyro3-eCFP or CMV-Axl-eGFP plasmid,respectively.CMV-eGFP plasmid served as a control vector.After that,the fluorescence intensity and distributions of green fluorescent protein （GFP） and cyan fluorescent protein （CFP） in the cells with or without nerve growth factor （NGF） treatment were real-time monitored.Results Expressions of Tyro-3 and Axl receptors were under the regulation of NGF and associated with neuronal differentiation.This was not observed in CMV-eGFP-transfected PC12 cells.Besides,confocal microscopy revealed that NGF affected intracellular localization of full-length Axl-eGFP and Tyro-3eCFP in PC12 cells.Moreover,the development of outgrowth of differentiated PC12 cells under stimulation of NGF was promoted by overexpression of Tyro-3 or Axl.Conclusion Expressions of Tyro-3 and Axl receptors are under the regulation of NGF and are involved in NGF-induced neuronal differentiation of PC12 cells.
基金supported by the National Natural Science Foundation of China,No.81001541the Natural Science Foundation of Fujian Province of China,No.2013J01331
文摘In this study, we investigated the role of the TYRO3/Akt signaling pathway in hypoxic injury to hippocampal neurons. 3-（4,5-Dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide assay showed that hypoxia inhibited the proliferation and viability of hippocampal neurons. Terminal deoxynucleotidyl transferase-mediated d UTP nick end labeling assay demonstrated that hypoxia induced neuronal apoptosis in a time-dependent manner, with a greater number of apoptotic cells with longer hypoxic exposure. Immunofluorescence labeling revealed that hypoxia suppressed TYRO3 expression. Western blot assay showed that hypoxia decreased Akt phosphorylation levels in a time-dependent manner. Taken together, these findings suggest that hypoxia inhibits the proliferation of hippocampal neurons and promotes apoptosis, and that the inhibition of the TYRO3/Akt signaling pathway plays an important role in hypoxia-induced neuronal injury.
基金This study was partly supported by the National Natural Science Foundation of China(81771360).
文摘Objective To study the effects of TYRO protein kinase-binding protein(TYROBP)deficiency on learning behavior,glia activation and pro-inflammatory cycokines,and Tau phosphorylation of a new Alzheimer’s disease(AD)mouse model carrying a PSEN1 p.G378E mutation.Methods A new AD mouse model carrying PSEN1 p.G378E mutation was built based on our previously found AD family which might be ascribed to the PSEN1 mutation,and then crossed with TYROBP deficient mice to produce the heterozygous hybrid mice(PSEN1^(G378E)/WT;Tyrobp^(+/-))and the homozygous hybrid mice(PSEN1^(G378E/G378E);Tyrobp^(-/-)).Water maze test was used to detect spatial learning and memory ability of mice.After the mice were sacrificed,the hippocampus was excised for further analysis.Immunofluorescence was used to identify the cell that expresses TYROBP and the number of microglia and astrocyte.Western blot was used to detect the expression levels of Tau and phosphorylated Tau(p-Tau),and ELISA to measure the levels of pro-inflammatory cytokines.Results Our results showed that TYROBP specifically expressed in the microglia of mouse hippocampus.Absence of TYROBP in PSEN1^(G378E) mutation mouse model prevented the deterioration of learning behavior,decreased the numbers of microglia and astrocytes,and the levels of interleukin-6,interleukin-1βand tumor necrosis factor-αin the hippocampus(all P<0.05).The ratios of AT8/Tau5,PHF1/Tau5,pT181/Tau5,pT231/Tau5 and p-ERK/ERK were all higher in homozygous hybrid mice(PSEN1^(G378E/G378E);Tyrobp^(-/-) mice)compared with PSEN1^(G378E/G378E) mice(all P<0.05).Conclusions TYROBP deficiency might play a protective role in the modulation of neuroinflammation of AD.However,the relationship between neuroinflammation processes involving microglia and astrocyte activation,and release of pro-inflammatory cytokines,and p-Tau pathology needs further study.
文摘The Tyro3, Axl and Mer (TAM) receptor tyrosine kinases are activated by endogenous ligands, protein S1 (PROS1) and growth arrest-specific gene 6 (Gas6), and those have important effects on cell biology. These receptors (Rs) can be shad from the cell membrane and their soluble(s) forms can be found in plasma. We investigated the fluctuation and interactive role of sTAMRs and its ligands in patients with hepatocellular carcinoma (HCC), hepatitis groups, and healthy normal adult controls (NC). The measurement cases were 45 patients with HCC group (stage 1 in 4, stage 2 in 8, stage 3 in 16, and stage 4 in 17), 4 patients with fulminant hepatitis (FH), 14 patients with acute hepatitis (AH), 10 patients with chronic hepatitis (CH), 16 patients with liver cirrhosis (LC), and 20 NCs matched by age. Plasma levels of three sTAMRs and their ligands were measured by ELISA. In comparison with NCs, Gas6, des-γ-carboxy Gas6, and sTAMRs levels were significantly higher in HCC patients, but free PROS1 levels were significantly lower. The sTyro3 and sAxl levels peaked HCC stages 2 and 3 respectively, and gradually decreased afterwards while maintaining high levels. sMer levels increased with the progression of HCC. Gas6 and des-γ-carboxy Gas6 levels gradually increased, and PROS1 levels decreased with the progression of HCC. Gas6 levels were positively correlated with sAxl levels, whereas sMer levels were negatively correlated with free PROS1 levels. sTAMRs and Gas6 levels increased in parallel to the progression of HCC fibrosis. Through the progression of HCC, Axl played the major role in TAMRs activation. However, sTYro3 continued increasing rapidly from the early stage, and that of Mer increased throughout the progression. Roles of Axl may be changed in Mer, because des-γ-carboxy Gas6 levels increasing with Gas6 in the advanced stage of HCC cannot send a signal to Axl.
基金the National Natural Science Foundation of China (Grant Nos.30070071& 30270135)and the National Key Basic Research Project (Grant No. G1999011700)
文摘Water deficit-induced ABA accumulation is an ideal model or 搒timulus-response?system to investigate cellular stress signaling in plant cells, using such a model the cellular stress signaling triggered by water deficit was inves-tigated in Maize L. coleoptile. Water deficit-induced ABA accumulation was sensitively blocked by NaVO3, a potent inhibitor both to plasma membrane H+-ATPase (PM-H+- ATPase) and protein tyrosine phosphatase (PTPase). How-ever, while PM- H+-ATPase activity was unaffected under water deficit and PM- H+-ATPase activator did not induce an ABA accumulation instead of water deficit, water deficit induced an increase in the protein phosphatase activity, and furthermore, ABA accumulation was inhibited by PAO, a specific inhibitor of PTPase. These results indicate that pro-tein phosphtases may be involved in the cellular signaling in response to water deficit. Further studies identified at least four species of protein phosphtase as assayed by using pNPP as substrate, among which one component was especially sensitive to NaVO3. The NaVO3-sensitive enzyme was puri-fied and finally showed a protein band about 66 kD on SDS/PAGE. The purified enzyme showed a great activity to some specific PTPase substrates at pH 6.0. In addition to NaVO3, the enzyme was also sensitive to some other PTPase inhibitors such as Zn2+ and MO33+, but not to Ca2+ and Mg2+, indicating that it might be a protein tyrosine phosphatase. Interestingly, the purified enzyme could be deactivated by some reducing agent DTT, which was previously proved to be an inhibitor of water deficit-induced ABA accumulation. This result further proved that PTPase might be involved in the cellular signaling of ABA accumulation in response to water deficit.
