Mechanism Of HIF-1? Regulating Notch1 Signaling Pathway On Pulmonary Fibrosis Induced By Paraquat Poisoning

Objective: Paraquat(PQ)is a widely used herbicide.It is highly toxic to humans and animals and has no antidote.Late PQ poisoning can lead to lung fibrosis and death from respiratory failure.According to the physiological process of lung epithelial cells transformed into interstitial cells after poisoning.We believe that hypoxia-inducible factor(HIF)plays an important role in PQ poisoning leading to pulmonary fibrosis.Under the condition that PQ poisoning causes severe cell hypoxia,HIF activates many target genes downstream by combining hypoxia response elements,thereby participating in the angiogenesis and epithelialmesenchymal transition(EMT)of lung epithelial cells.HIF can also directly react with cell signaling pathways.This article aims to explore the mechanism by which HIF-1? regulates pulmonary EMT caused by PQ poisoning through the Notch1 signaling pathway,thereby controlling the occurrence and progression of fibrosis,and finding potential targets for the treatment of PQ poisoning.Method:1.Establishment of a mouse model of PQ poisoning: Exposure by intraperitoneal injection to construct a model of pulmonary fibrosis in mice after PQ exposure.Methods: Group A saline control group,Group B 10 mg / kg(single exposure group),Group C: 20 mg / kg(single exposure group),Group D(10 mg / kg group once every other day,three times in total)observe the anesthesia and kill the mice on the 21 st day after exposure Tissue,microscopic observation of pathological changes,Masson staining,Ashcroft score and other methods to assess the degree of fibrosis,extraction of mouse lung tissue protein and Western Blot to detect the expression of HIF-1? and Notch1.2.Establishment of a cell model for PQ poisoning: A549 cell lines cultured in vitro were infected with PQ at final concentrations of 0,10,30,and 60 ?g / ml.After 6 days of exposure,MTT method to detect cell viability,Transwell experiment observed the migration ability of fibrotic cells,Western Blot and Realtime RT-PCR detect ?-SMA and E-cadherin and other fibrosis indicators,and focus on the detection of protein and gene expression of HIF-1? and Notch1 in cell poisoning models.3.siRNA interference verification experiment: on the basis of the above PQ poisoning cell model,knock down the expression of HIF-1? and Notch1 with specific iRNA,analyze the expression of ?-SMA and E-cadherin and the change of EMT degree,verify HIF-1?-Down-regulation of Notch1 pathway alleviates EMT after PQ exposure.4.Co-IP experiment: Based on the above PQ poisoning cell model,Co-IP was used to analyze the direct interaction between HIF-1? and Notch1 protein.Results:1.In a mouse model of fibrosis caused by PQ poisoning,HIF-1? and Notch1 are highly expressed in lung tissue.2.In the PQ poisoning cell model,HIF-1? and Notch1 are highly expressed in the infected cells.3.In the model of PQ poisoning cells,knocking down the expression of HIF-1? and Notch1 with specific iRNA can reduce the expression of EMT marker ?-SMA and increase the expression of epithelial cell marker E-cadherin,suggesting that The chemical process has a certain relief effect.4.In the model of PQ poisoning cells,there may be a direct interaction between HIF-1? and Notch1 molecules.Conclusion:The process of PQ-induced lung fibrosis induced by chronic PQ poisoning was simulated at the in vivo and in vitro levels in mice.This process can activate the HIF-1?-Notch1 signaling pathway,and the expression of HIF-1? and Notch1 is up-regulated,and both There is a direct interaction between the molecules.Interference with the HIF-1? / Notch1 pathway can down-regulate the characteristics of lung cell EMT caused by PQ poisoning.