AIM: To identify and characterize the function of non-mu-scle myosin Ⅱ (NMM Ⅱ) isoforms in primary rat hepatic stellate cells (HSCs).METHODS: Primary HSCs were isolated from male Spra-gue-Dawley rats by pronase/coll...AIM: To identify and characterize the function of non-mu-scle myosin Ⅱ (NMM Ⅱ) isoforms in primary rat hepatic stellate cells (HSCs).METHODS: Primary HSCs were isolated from male Spra-gue-Dawley rats by pronase/collagenase digestion. Total RNA and protein were harvested from quiescent and culture-activated HSCs. NMM Ⅱ isoform (Ⅱ-A, Ⅱ-B and Ⅱ-C) gene and protein expression were measured by RealTime polymerase chain reaction and Western blot analyses respectively. NMM Ⅱ protein localization was visualized in vitro using immunocytochemical analysis. For in vivo assessment, liver tissue was harvested from bile duct-ligated (BDL) rats and NMM Ⅱisoform expression determined by immunohistochemistry. Using a selective myosin Ⅱ inhibitor and siRNA-mediated knockdown of each isoform, NMM Ⅱ functionality inprimary rat HSCs was determined by contraction and migration assays.RESULTS: NMM Ⅱ-A and Ⅱ-B mRNA expression was increased in culture-activated HSCs (Day 14) with sig-niflicant increases seen in all pairwise comparisons (Ⅱ-A: 12.67 ± 0.99 (quiescent) vs 17.36 ± 0.78 (Day 14), P < 0.05; Ⅱ-B: 4.94 ± 0.62 (quiescent) vs 13.90 ±0.85 (Day 14), P < 0.001). Protein expression exhibited similar expression patterns (Ⅱ-A: 1.87 ± 2.50 (quiescent) vs 58.64 ± 8.76 (Day 14), P < 0.05; Ⅱ-B: 1.17 ± 1.93 (quiescent) vs 103.71 ± 21.73 (Day 14), P < 0.05). No signif icant differences were observed in NMM Ⅱ-C mRNA and protein expression between quiescent and activated HSCs. In culture-activated HSCs, NMM Ⅱ-A and Ⅱ-B merged with F-actin at the cellular periphery and throughout cytoplasm respectively. In vitro stud-ies showed increased expression of NMM Ⅱ-B in HSCs activated by BDL compared to sham-operated animals. There were no apparent increases of NMM Ⅱ-A and Ⅱ-C protein expression in HSCs during hepatic BDL injury. To determine the contribution of NMM Ⅱ-A and Ⅱ-B to migration and contraction, NMM Ⅱ-A and Ⅱ-B expres-sion were downregulated with siRNA. NMM Ⅱ-A and/or Ⅱ-B siRNA inhibited HSC migration by approximately 25% compared to scramble siRNA-treated cells. Conversely, siRNA-mediated NMM Ⅱ-A and Ⅱ-B inhibition had no signif icant effect on HSC contraction; however, contraction was inhibited with the myosin Ⅱ inhibitor, blebbistatin (38.7% ± 1.9%).CONCLUSION: Increased expression of NMM Ⅱ-A and Ⅱ-B regulates HSC migration, while other myosin Ⅱclasses likely modulate contraction, contributing to development and severity of liver f ibrosis.展开更多
A novel copper（II） complex with the reduced Schiff base, [Cu（L）2]·H2O （I, HL = N-（4-hydroxybenzyl）-L-serine）, was prepared in aqueous solution and characterized by elemental analysis, FT-IR, electrospray ...A novel copper（II） complex with the reduced Schiff base, [Cu（L）2]·H2O （I, HL = N-（4-hydroxybenzyl）-L-serine）, was prepared in aqueous solution and characterized by elemental analysis, FT-IR, electrospray ionization mass spectrometry and single-crystal X-ray diffraction. Complex I crystallizes in the orthorhombic system, space group P212121, with a = 8.9040（18）, b = 9.1530（18）, c = 24.891（5）A^°, V = 2028.6（7） A^°3, Z = 4, C20H26CuN2O9, Mr = 501.97, Dc = 1.644g·cm^3, μ = 1.135 mm^-1, F（000） = 1044, GOOF = 1.194, the final R = 0.0484 and wR = 0.1420 for 6186 observed reflections （I 〉 2σ（I））. In I, two L^- anions are coordinated to the copper ion in tridentate and bidentate chelating modes, respectively, resulting in the coordinated geometry of copper ion to be a distorted square pyramid. The intermolecular hydrogen bonds between the complexes, complexes and lattice water molecules lead to a 2D supramolecular network. The bioactivity of the complex as a potential PTPs inhibitory agent in vitro was investigated, displaying potent inhibition against PTP1B （IC50, 0.27 μM） and TCPTP （IC50, 0.57 μM） with a moderate selectivity.展开更多
Thrombosis formation on disrupted atherosclerotic plaque is the most common acuse of cardiovascular diseases, in the pathophysiology, increased platelet reactivity is a descriptor of the risk of cardiovascular events ...Thrombosis formation on disrupted atherosclerotic plaque is the most common acuse of cardiovascular diseases, in the pathophysiology, increased platelet reactivity is a descriptor of the risk of cardiovascular events in healthy persons and in patients with overt coronary artery disease. Regardless of the stimulus for activation platelet-platelet interation and thrombus formation is ultimately regulated through the GP Ⅱ b/Ⅲ a receptor展开更多
文摘AIM: To identify and characterize the function of non-mu-scle myosin Ⅱ (NMM Ⅱ) isoforms in primary rat hepatic stellate cells (HSCs).METHODS: Primary HSCs were isolated from male Spra-gue-Dawley rats by pronase/collagenase digestion. Total RNA and protein were harvested from quiescent and culture-activated HSCs. NMM Ⅱ isoform (Ⅱ-A, Ⅱ-B and Ⅱ-C) gene and protein expression were measured by RealTime polymerase chain reaction and Western blot analyses respectively. NMM Ⅱ protein localization was visualized in vitro using immunocytochemical analysis. For in vivo assessment, liver tissue was harvested from bile duct-ligated (BDL) rats and NMM Ⅱisoform expression determined by immunohistochemistry. Using a selective myosin Ⅱ inhibitor and siRNA-mediated knockdown of each isoform, NMM Ⅱ functionality inprimary rat HSCs was determined by contraction and migration assays.RESULTS: NMM Ⅱ-A and Ⅱ-B mRNA expression was increased in culture-activated HSCs (Day 14) with sig-niflicant increases seen in all pairwise comparisons (Ⅱ-A: 12.67 ± 0.99 (quiescent) vs 17.36 ± 0.78 (Day 14), P < 0.05; Ⅱ-B: 4.94 ± 0.62 (quiescent) vs 13.90 ±0.85 (Day 14), P < 0.001). Protein expression exhibited similar expression patterns (Ⅱ-A: 1.87 ± 2.50 (quiescent) vs 58.64 ± 8.76 (Day 14), P < 0.05; Ⅱ-B: 1.17 ± 1.93 (quiescent) vs 103.71 ± 21.73 (Day 14), P < 0.05). No signif icant differences were observed in NMM Ⅱ-C mRNA and protein expression between quiescent and activated HSCs. In culture-activated HSCs, NMM Ⅱ-A and Ⅱ-B merged with F-actin at the cellular periphery and throughout cytoplasm respectively. In vitro stud-ies showed increased expression of NMM Ⅱ-B in HSCs activated by BDL compared to sham-operated animals. There were no apparent increases of NMM Ⅱ-A and Ⅱ-C protein expression in HSCs during hepatic BDL injury. To determine the contribution of NMM Ⅱ-A and Ⅱ-B to migration and contraction, NMM Ⅱ-A and Ⅱ-B expres-sion were downregulated with siRNA. NMM Ⅱ-A and/or Ⅱ-B siRNA inhibited HSC migration by approximately 25% compared to scramble siRNA-treated cells. Conversely, siRNA-mediated NMM Ⅱ-A and Ⅱ-B inhibition had no signif icant effect on HSC contraction; however, contraction was inhibited with the myosin Ⅱ inhibitor, blebbistatin (38.7% ± 1.9%).CONCLUSION: Increased expression of NMM Ⅱ-A and Ⅱ-B regulates HSC migration, while other myosin Ⅱclasses likely modulate contraction, contributing to development and severity of liver f ibrosis.
基金Supported by NNSFC(Nos.21271121,21471092,21571118)
文摘A novel copper（II） complex with the reduced Schiff base, [Cu（L）2]·H2O （I, HL = N-（4-hydroxybenzyl）-L-serine）, was prepared in aqueous solution and characterized by elemental analysis, FT-IR, electrospray ionization mass spectrometry and single-crystal X-ray diffraction. Complex I crystallizes in the orthorhombic system, space group P212121, with a = 8.9040（18）, b = 9.1530（18）, c = 24.891（5）A^°, V = 2028.6（7） A^°3, Z = 4, C20H26CuN2O9, Mr = 501.97, Dc = 1.644g·cm^3, μ = 1.135 mm^-1, F（000） = 1044, GOOF = 1.194, the final R = 0.0484 and wR = 0.1420 for 6186 observed reflections （I 〉 2σ（I））. In I, two L^- anions are coordinated to the copper ion in tridentate and bidentate chelating modes, respectively, resulting in the coordinated geometry of copper ion to be a distorted square pyramid. The intermolecular hydrogen bonds between the complexes, complexes and lattice water molecules lead to a 2D supramolecular network. The bioactivity of the complex as a potential PTPs inhibitory agent in vitro was investigated, displaying potent inhibition against PTP1B （IC50, 0.27 μM） and TCPTP （IC50, 0.57 μM） with a moderate selectivity.
文摘Thrombosis formation on disrupted atherosclerotic plaque is the most common acuse of cardiovascular diseases, in the pathophysiology, increased platelet reactivity is a descriptor of the risk of cardiovascular events in healthy persons and in patients with overt coronary artery disease. Regardless of the stimulus for activation platelet-platelet interation and thrombus formation is ultimately regulated through the GP Ⅱ b/Ⅲ a receptor
