Molybdenum And Cadmium Co-induce Hepatocytes Autophagy Via Nrf2 Signaling Pathway And Endoplasmic Reticulum Stress In Duck

To investigate whether Nrf2 signaling pathway and endoplasmic reticulum stress mediated autophagy are involved in hepatotoxicity co-induced by molybdenum and cadmium in ducks.160 healthy 8-day-old ducks were used as the experimental animals to establish Mo or/and Cd sub-chronic poisoning models.Hexaammonium molybdate[（NH₄）₆Mo₇O₂₄·4H₂O]and cadmium sulfate（3Cd SO₄·8H₂O）were resources of Mo and Cd were added to the per kilogram（kg）basic diet,respectively.Ducks were divided into four groups:Control group（0 mg Mo+0 mg Cd）;Mo group（100 mg Mo）;Cd group（4mg Cd）;Mo+Cd group（100 mg Mo+4 mg Cd）.On the 16^thweek in the experiment,10ducks were randomly selected from each group to collect liver samples.The ultrastructure of hepatocytes,LC3II immunofluorescence,oxidative stress indices,Nrf2 signaling pathway,endoplasmic reticulum stress and autophagy levels were detected by using transmission electron microscope,laser scanning confocal microscope,RT-q PCR,western blotting and so on.The results showed that:1.Mo or/and Cd remarkably downregulated GSH-Px,CAT and T-SOD activities,upregulated MDA and H₂O₂concentrations,the changes in joint group were most obvious,showing that Mo or/and Cd exposure caused oxidative stress in hepatocytes.2.Mo or/and Cd dramatically decreased the Nrf2 signaling pathway related genes（Nrf2,HO-1,NQO1,GST,CAT,SOD1,GCLC and GCLM）m RNA and protein（Nrf2）expression levels,and the changes in joint group were most obvious,proving that Mo or/and Cd exposure inhibited the Nrf2 signaling pathway in hepatocytes.3.Mo or/and Cd obviously enhanced the endoplasmic reticulum stress related genes（GRP78,PERK,e IF2a,ATF4,IRE1 and ATF6）m RNA and proteins（PERK,IRE1 and ATF6）expression levels,and the changes in joint group were most obvious,certifying that Mo or/and Cd exposure activated the endoplasmic reticulum stress signaling pathway in hepatocytes.4.Mo or/and Cd notably increased the number of autophagosomes and LC3II fluorescence aggregation points,upregulated the autophagy related genes（Atg5,LC3A,LC3B and Beclin-1）m RNA and proteins（LC3II/LC3I and Beclin-1）expression levels,downregulated the autophagy related genes（p62,m TOR and Dynein）m RNA and protein（p62）expression levels,and the changes in joint group were most obvious,verifying that Mo or/and Cd exposure induced autophagy in hepatocytes.In conclusion,Mo or/and Cd induced oxidative stress,inhibited Nrf2 signaling pathway,activated endoplasmic reticulum stress and induced autophagy.Therefore,autophagy induced by Mo or/and Cd may be associated with the activation of crosstalk between Nrf2-mediated antioxidant defense response and endoplasmic reticulum stress in duck hepatocytes.Mo and Cd may aggravate toxic damage to the hepatocytes.