To study the effects of tumor necrosis factor （TNF）-α on matrix metalloproteinase （MMP）-9 expression and activity in alveolar macrophages （AM） and to investigate the role of NF-κB in the induction, AM were col...To study the effects of tumor necrosis factor （TNF）-α on matrix metalloproteinase （MMP）-9 expression and activity in alveolar macrophages （AM） and to investigate the role of NF-κB in the induction, AM were collected from bronchoalveolar lavage fluid （BALF） of healthy subjects and patients with chronic obstructive pulmonary disease （COPD）. MMP-9 expression and activity were detected by semi-quantitative reverse transcription-polymerase chain reaction （RT-PCR）, Western blotting and zymography. NF-κB activity was detected by electrophoretic mobility shift assay （EMSA）. MMP-9 expression and activity induced by TNF-α in AM from healthy subjects or patients with COPD were significantly increased in a dose-dependent manner （P〈0.05）. NF-κB activity induced by TNF-α was significantly increased in AM from patients with COPD, and pyrrolidine dithiocarbamate （PDTC） and N-acetyl-L-cysteine （NAC） significantly inhibited the activation of NF-κB induced by TNF-α （P〈0.05）. The presents study suggested that the expression and activity of MMP-9 from AM can be induced by TNF-α, and TNF-α/NF-κB signal pathway may play an important role in the induction.展开更多
Ni34.1 Fe27.9B18 Si18 Nb2 coating was deposited on mild steel substrate using high power laser cladding followed by laser remelting process. The laser processing was conducted by the powder feeding method using low pu...Ni34.1 Fe27.9B18 Si18 Nb2 coating was deposited on mild steel substrate using high power laser cladding followed by laser remelting process. The laser processing was conducted by the powder feeding method using low purity materials without shielding box. To learn the surface amorphous matrix coating forming mechanism, the coating without remelting process was also studied. The phases and microstructures were analyzed by X-ray diffraction （XRD）, scanning- and transmission-electron microscopy（ SEM, TEM）. The microhardness and corrosion resistance property of the coating were also measured. The results of SEM, XRD and TEM analysis show that the remelted coating has an amorphous matrix layer embedded with some crystals due to high cooling rate during remelting process. The crystals phases are identified as Fe2 B phase, γ （Fe, Ni ） phase and α- Fe phase. No oxidation phases are found in the coating surface. Hardness profiles reveal microhardness more than 1 100 HVo.5 over the full depth of the amorphous matrix layer, while the unremtled coating and the substrate show relatively lower hardness than the remelted layer. Corrosion resistance tests exhibit that the remelted coating is nobler than the unremelted coating and the substrate material.展开更多
To ascertain the molecular basis of Ca2+-mediated activation of matrix metalloproteinase-9 (MMP-9), we determined the accessibility of tryptophan residues to externally added acrylamide as quencher in the absence and ...To ascertain the molecular basis of Ca2+-mediated activation of matrix metalloproteinase-9 (MMP-9), we determined the accessibility of tryptophan residues to externally added acrylamide as quencher in the absence and presence of the metal ion. The steady-state and time resolved fluorescence data revealed that MMP-9 possesses two classes of tryptophan residues, “exposed” and “buried” which are quenched by the collisional rate constants (kq) of 3.2′ 109M-1.s-1 and 7.5′ 108M-1.s-1, respectively. These values are impaired by approximately two and three-fold, respectively, in the presence of 10 mM Ca2+. The Stern-Volmer constants (Ksv values) predicted from the time resolved fluorescence data (in the absence of Ca2+ ) satisfied the dynamic quenching model of the enzyme’s tryptophan residues. This was not the case in the presence of Ca2+;the steady-state acrylamide quenching data could only be explained by a combination of “dynamic” and “static” quenching models. A cumulative account of these data led to the suggestion that the binding of Ca2+ modulated the tertiary structure of the protein by decreasing the dynamic flexibility of the enzyme, which is manifested in further structuring of the enzyme’s active site pocket toward facilitating catalysis. Arguments are presented that the binding of Ca2+ at distal sites “dynamically” communicates with the active site residues of MMP-9 during catalysis.展开更多
文摘To study the effects of tumor necrosis factor （TNF）-α on matrix metalloproteinase （MMP）-9 expression and activity in alveolar macrophages （AM） and to investigate the role of NF-κB in the induction, AM were collected from bronchoalveolar lavage fluid （BALF） of healthy subjects and patients with chronic obstructive pulmonary disease （COPD）. MMP-9 expression and activity were detected by semi-quantitative reverse transcription-polymerase chain reaction （RT-PCR）, Western blotting and zymography. NF-κB activity was detected by electrophoretic mobility shift assay （EMSA）. MMP-9 expression and activity induced by TNF-α in AM from healthy subjects or patients with COPD were significantly increased in a dose-dependent manner （P〈0.05）. NF-κB activity induced by TNF-α was significantly increased in AM from patients with COPD, and pyrrolidine dithiocarbamate （PDTC） and N-acetyl-L-cysteine （NAC） significantly inhibited the activation of NF-κB induced by TNF-α （P〈0.05）. The presents study suggested that the expression and activity of MMP-9 from AM can be induced by TNF-α, and TNF-α/NF-κB signal pathway may play an important role in the induction.
基金Acknowledgements The authors would like to thank the financial support provided by the National Natural Science Foundation of China （No. 50971091 ）, the Ministry of the Science and Technology of the People＇s Republic of China （No. 2009DFB50350） , the Science and Technology Commission of Shanghai Municipality （ No. 08520704900） and the Economy and Information Commission of Shanghai Municipality （ No. zx08089）.
文摘Ni34.1 Fe27.9B18 Si18 Nb2 coating was deposited on mild steel substrate using high power laser cladding followed by laser remelting process. The laser processing was conducted by the powder feeding method using low purity materials without shielding box. To learn the surface amorphous matrix coating forming mechanism, the coating without remelting process was also studied. The phases and microstructures were analyzed by X-ray diffraction （XRD）, scanning- and transmission-electron microscopy（ SEM, TEM）. The microhardness and corrosion resistance property of the coating were also measured. The results of SEM, XRD and TEM analysis show that the remelted coating has an amorphous matrix layer embedded with some crystals due to high cooling rate during remelting process. The crystals phases are identified as Fe2 B phase, γ （Fe, Ni ） phase and α- Fe phase. No oxidation phases are found in the coating surface. Hardness profiles reveal microhardness more than 1 100 HVo.5 over the full depth of the amorphous matrix layer, while the unremtled coating and the substrate show relatively lower hardness than the remelted layer. Corrosion resistance tests exhibit that the remelted coating is nobler than the unremelted coating and the substrate material.
文摘To ascertain the molecular basis of Ca2+-mediated activation of matrix metalloproteinase-9 (MMP-9), we determined the accessibility of tryptophan residues to externally added acrylamide as quencher in the absence and presence of the metal ion. The steady-state and time resolved fluorescence data revealed that MMP-9 possesses two classes of tryptophan residues, “exposed” and “buried” which are quenched by the collisional rate constants (kq) of 3.2′ 109M-1.s-1 and 7.5′ 108M-1.s-1, respectively. These values are impaired by approximately two and three-fold, respectively, in the presence of 10 mM Ca2+. The Stern-Volmer constants (Ksv values) predicted from the time resolved fluorescence data (in the absence of Ca2+ ) satisfied the dynamic quenching model of the enzyme’s tryptophan residues. This was not the case in the presence of Ca2+;the steady-state acrylamide quenching data could only be explained by a combination of “dynamic” and “static” quenching models. A cumulative account of these data led to the suggestion that the binding of Ca2+ modulated the tertiary structure of the protein by decreasing the dynamic flexibility of the enzyme, which is manifested in further structuring of the enzyme’s active site pocket toward facilitating catalysis. Arguments are presented that the binding of Ca2+ at distal sites “dynamically” communicates with the active site residues of MMP-9 during catalysis.
