The Effect Of PINK1/Parkin-Mediated Mitophagy On Kidney Damage Induced By Aflatoxin B₁ In Mice

Aflatoxin B₁（AFB₁）is a highly toxic contaminant that accumulates in the kidneys when it enters the body,causing kidney damage.Oxidative stress is the pathological basis of AFB₁-induced kidney injury and can lead to mitochondrial damage,while damaged mitochondria can trigger apoptosis and aggravate kidney injury.PINK1/Parkin-mediated mitophagy participates in the clearance of damaged mitochondria from kidney tissue cells and plays an important role in maintaining kidney function.However,whether it is activated in AFB₁-induced kidney injury and the role it plays in it are inconclusive.Thus,this study proposes the theoretical hypothesis that"PINK1/Parkin-mediated mitophagy is involved in protecting AFB₁ against kidney damage"and proposes to conduct two studies:（1）AFB₁ toxicity test:40 wild-type（WT）C57BL/6 male mice split into control group（CG,0 mg/kg B.W.AFB₁）,low-dose group（LG,0.5 mg/kg B.W.AFB₁）,medium-dose group（MG,0.75 mg/kg B.W.AFB₁）and high-dose group（HG,1 mg/kg B.W.AFB₁）and exposed to AFB₁ for 28 d.The growth and development,kidney state,oxidation and antioxidant levels,mitochondrial state,apoptosis level,and mitophagy related indicators in mice were evaluated,aiming to clarify the effect of AFB₁ on kidney injury and mitophagy in mice,as well as to screen for the optimal dose of AFB₁ for use in Parkin gene knockout(Parkin^-/-)intervention trial.（2）Parkin^-/-intervention trial:20 WT C57BL/6 male mice and 20 Parkin^-/-C57BL/6 male mice split into WT control group（WCG,0 mg/kg B.W.AFB₁）,WT poisoning group（WAG,1 mg/kg B.W.AFB₁）,Parkin^-/-control group（PCG,0 mg/kg B.W.AFB₁）and Parkin^-/-poisoning group（PAG,1 mg/kg B.W.AFB₁）and exposed to AFB₁ for 28 d.The growth and development,kidney state,oxidation and antioxidant levels,mitochondrial state,apoptosis level,and mitophagy related indicators in mice were evaluated,aiming to investigate the effect of PINK1/Parkin-mediated mitophagy on AFB₁-cuased kidney lesion.（1）AFB₁ toxicity test results1)Compared with the CG,the MG and HG mice showed significantly lower body weight and kidney coefficient;significant microscopic and ultrastructural kidney damage;and significantly higher serum BUN and CREA levels,indicating that AFB₁ could cause structural and functional damage to the kidney in mice.2)Compared with the CG,the kidney ROS content was significantly higher and SOD activity was significantly lower in each AFB₁-treated group,indicating that AFB₁ could induce oxidative stress in mice kidney.3)Compared with the CG,the MG and HG mice showed significant structural damage to renal mitochondria,and significantly lower MMP level and ATP content,indicating that AFB₁could induce mitochondrial structure and function damage in mice kidney.4)Compared with the CG,the MG and HG mice showed significantly higher Bax protein level,and significantly lower Bcl-2 protein level,indicating that AFB₁ could induce apoptosis in mice kidney.5)Compared with the CG,the MG and HG mice showed significantly higher PINK1,Parkin,and LC3 m RNA and protein levels,and significantly lower p62 m RNA and protein levels,indicating that AFB₁ could activate PINK1/Parkin-mediated mitophagy in mice kidney.In summary,the dose point with significant kidney injury and the highest intensity of mitophagy,namely 1 mg/kg,was used as the AFB₁ dose for Parkin^-/-intervention trial.（2）Parkin^-/-intervention trial results1)Compared with the WAG,the PINK1 and LC3 m RNA and protein levels of PAG mice were reduced,while the p62 m RNA and protein levels were raised,indicating that Parkin^-/-could inhibit PINK1/Parkin-mediated mitophagy induced by AFB₁ in mice kidney.2)Compared with the WAG,the body weight and kidney coefficient of PAG mice were declined;the kidney microscopic and ultrastructural damage were aggravated;the contents of BUN and CREA were raised,indicating that Parkin^-/-could exacerbate AFB₁-induced renal structure and function damage in mice.3)Compared with the WAG,the contents of ROS and MDA of PAG mice were raised,while the activities of GSH-Px and SOD were decreased,indicating that Parkin^-/-could aggravate AFB₁-induced oxidative stress in mice kidney.4)Compared with the WAG,the kidney mitochondrial structural lesion of PAG mice was aggravated,the level of MMP and the content of ATP were declined,indicating that Parkin^-/-could aggravate AFB₁-induced mitochondrial structural and functional damage in mice kidney.5)TUNEL staining showed that compared with the WAG,the Bax protein level and the Caspase-3 activity of PAG mice were raised,while the Bcl-2 protein level was reduced,indicating that Parkin^-/-could aggravate AFB₁-induced apoptosis in mice kidney.Conclusion:AFB₁ exposure can activate renal PINK1/Parkin-mediated mitophagy,and PINK1/Parkin-mediated mitophagy plays a protective role in AFB₁-induced kidney damage,and its protective effect is related to alleviating oxidative stress,mitochondrial damage,and apoptosis.