Mechanism of antagonism of paclitaxel-bivalirudin balloon coating complex on proliferation and migration of human coronary artery smooth muscle cells through regulating pathways of nuclear factor-κB and nuclear factor E2 related factor 2/antioxidant response element

Li Hongmei; Li Haiyan

doi:10.3969/j.issn.1674-4055.2023.08.09

Abstract：Objective To investigate the microscopic mechanism of paclitaxel-bivalirudin balloon coating complex(LFN)in inhibiting proliferation and migration of human coronary artery smooth muscle cells(HCASMC)for preventing restenosis after PCI based on dual pathway activation model of nuclear factor-κB(NF-κB)and nuclear factor E2 related factor 2(Nrf2)/antioxidant response element(ARE)of HCASMC.Methods HCASMC of 4-6 generations was selected to activate dual pathways of NF-κB and Nrf2/ARE by taken lipopolysaccharide(LPS)as an inducer for establishing a dual pathway activated cell model of HCASMC inflammation and oxidative stress.HCASMC,cultured in vitro,were divided into blank control group,LPS group,LPS+LFN group.The influence of LFN on proliferation rate of HCASMC was detected by using CCK-8 method,and expression of proliferating cell nuclear antigen(PCNA)was detected by using immunofluorescence for verifying HCASMC proliferation status.The influence of LFN on migration and invasion ability of HCASMC was detected by using Transwell method,and expression changes of α-smooth muscle actin(α-SMA),osteopontin(OPN),NF-κB p65 and Nrf2.The influence of LFN on phenotype transformation of HCASMC was discussed,and regulation effect of LFN on key proteins in activated NF-κB and Nrf2/ARE pathways in HCASMC was studied.The changes of key site genes and protein expressions after LFN intervention were identified in dual pathways by using fluorescence quantitative polymerase chain reaction(Q-PCR)and Western blotting assay for exploring further the potential mechanism of LFN regulating HCASMC proliferation and migration activated by inflammatory and oxidative stress.Results Paclitaxel(1 μmol/L)combined with bivalirudin(0.2 mg/ml)inhibited significantly HCASMC proliferation after model activation,weakened HCASMC migration and invasion,and reversed activated HCASMC from secreting type to the contractile type.Furthermore,compared with LPS group,LFN inhibited expression of IκBα,blocked activation of NF-κB into the nucleus,At the same time,LFN inhibited expression of Keap-1,improved Nrf2 nuclear translocation,activated gene and protein expressions of downstream NQO-1 and HO-1(P<0.05),and significantly relief intracellular oxidative stress state.Conclusion LFN can significantly inhibit inflammatory and oxidative stress states of HCASMC activated by dual pathways of NF-κB and Nrf2/ARE.This inhibitory effect is achieved through regulating expressions of nuclear transcription factors and their ligands in NF-κB and Nrf2/ARE pathways,as well as influencing activation of downstream effector molecules in the pathways.It provides a cytological basis for molecular mechanism of LFN coated balloon against restenosis after PCI.