The Mechanism Of DFP Protecting Against Liver And Kidney In Al-Loaded Mice

ObjectivesA1 is one of the most abundant elements in the enviroment.As A1 and A1 compound are widely used as food additive,coagulating agents,drugs,and in kinds of containers and utensils,et al,people contact with A1 badly frequently in daily life. With development of science and technology,reseachers home and abroad investigated the toxicity of A1 deeply and they deemed that A1 intoxication couldn't be disregarded:concentration of A1 in serum and organs will increase if the content of A1 increase in diets.Liver is one of the organs who have most concentrations of A1.A1 is excreted by kidney that may cause A1 accumulation in kidney.A1 accumulation in the organism is associated with a variety of tissue pathologies.There is a consensus that chelation therapy is the procedure to be used to treat A1 intoxication clinically.DFP(1, 2-dimethyl-3-hydroxy-4-pyridone,Deferiprone) is a kind of Fe chelators and was confirmed effective to remove A1 from animal bodies,but was not used to treat A1 intoxication clinically.In the following study,we treated A1-loaded mice with DFP of different concentrations,aim to investigate the mechanism of DFP protecting against liver and kidney in A1-loaded mice to provide foundation for seeking a safe and efficacious chelator for A1 overload.Methods1 The effect of DFP protecting against A1-loaded mice's livers1.1 Animals treatmentHealthy kunming species mice in the weight range of 18～22g were randomly divided into 5 groups by body weight:Negative control group,A1 control group,DFP low-dose group,DFP medium-dose group,DFP high-dose group,20 mice for each group.The other mice were treated with AlCl₃ except negative control group four weeks to prepare for the mice' s model of A1 intoxication.Negative control group were treated with saline four weeks.Negative and A1 control group were treated with saline one week later.At the same time,the low,medium and high-doses groups were intragastrically administered with DFP for 1 week.1.2 The effect of DFP on liver/body weight ratio of Al-loaded miceAfter the last treatment,the mice were given fasting for 24 hours and recorded body weight.Got livers intactly,recorded liver weights,calculated liver/body weight ratio of each group,to study the effect of DFP on liver/body weight ratio of Al-loaded mice.1.3 The effect of DFP on serum proteins and biochemical indicators of Al-loaded miceAfter the last treatment,the mice were given fasting for 24 hours.Blood samples were obtained by picking eyeballs of mice.Then blood samples were centrifugated 10 minites at 3500 rP.m to get serum samples.Determined the concentrations of TP,Alb, Glob,A/G,GOT,GPT,AKP,γ-GT in mice's serum to study the the effect of DFP on serum proteins and biochemical indicators of Al-loaded mice.1.4 The effect of DFP on activity of ATPase in Al-loaded miee's liversThe livers were grinded to make tissue homogenate.The activity of Na⁺ K⁺-ATPase,Ca²⁺ Mg²⁺-ATPase in the homogenate of liver tissues was determined in each group to investigate the effect of DFP to activity of ATPase in Al-loaded mice's livers.1.5 The effect of DFP on concentrations of XOD in Al-loaded miee's liversThe concentrations of XOD in the homogenate of liver tissues were determined in each group to investigate the effect of DFP on concentrations of XOD in Al-loaded mice's livers.1.6 The effect of DFP on contents of aluminum and other essential elements in Al-loaded mice's liversThe tissues of livers were digested by microwave digestion.Aluminum contents were analysised by graphite furnace atomic absorption spectrophotometry and calcium,magnesium,zinc,copper,and iron elements contents were analysised by flame atomic absorption spectrophotometry to investigate the effects of DFP removing aluminum and other essential elements in Al-loaded mice's livers.1.7 Pathomorphological observation of the liver cellsThe livers of the mice were stained by hematoxylin-eosin staining. Pathomorphological observations were carried out to investigate the morphological changes.2 The effect of DFP protecting against Al-loaded mice's kidneys2.1 The effect of DFP on kidney/body weight ratio of Al-loaded miceAfter the last treatment,the mice were given fasting for 24 hours and recorded body weight.Got two kidneys of each mouse intactly,recorded kidneys' weights, calculated kidney/body weight ratios of each group,and investigated the effect of DFP on kidney/body weight ratio of Al-loaded mice.2.2 The effect of DFP on Scr and BUN of AMoaded miceDetermined the concentrations of Scr and BUN in mice's serum to study the effect of DFP protecting against Al-loaded mice's kidney function.2.3 The effect of DFP on activity of ATPase in Al-loaded miee's kidneysThe kidneys were grinded to make tissue homogenate.The activity of Na⁺ K⁺-ATPase,Ca²⁺ Mg²⁺-ATPase in the homogenate of kidney tissues was determined in each group to investigate the effect of DFP on activity of ATPase in Al-loaded mice's kidneys.2.4 The effect of DFP on concentrations of XOD in Al-loaded mice's kidneysThe concentrations of XOD in the homogenate of kidney tissues were determined in each group to investigate the effect of DFP on concentrations of XOD in Al-loaded mice's kidneys.2.5 The effect of DFP on contents of aluminum and calcium,magnesium,zinc, copper and iron elements in Al-loaded mice's kidneysThe tissues of kidneys were digested by microwave digestion.Aluminum concentrations were analysised by graphite furnace atomic absorption spectrophotometry and calcium,magnesium,zinc,copper and iron elements contents were analysised by flame atomic absorption spectrophotometry to investigate the effects of DFP removing A1 and other essential elements in Al-loaded mice's kidneys.2.6 Pathomorphological observation of the kidney cellsThe kidneys of the mice were stained by hematoxylin-eosin staining. Pathomorphological observations were carried out to investigate the morphological changes.Results1 The effect of DFP protecting against Al-loaded mice's livers1.1 The effect of DFP on liver/body weight ratio of Al-loaded miceNo significant differences existed among all groups' liver/body weight ratios.1.2 The effect of DFP on serum proteins and biochemical indicators of Al-loaded miceConcentrations of serum TP in A1 control group was significantly lower than negtive control group(P＜0.01).In comparison with the negtive control group,no significant differences existed in three doses groups.The concentration of serum albumin of mice in A1 control group was lower than negtive control group and middle,high doses groups significantly(P＜0.05,P＜0.01), there was no significant difference between middle,high doses groups and negtive control group.No significant differences existed among all groups' concentrations of Glob and A/G.After the mice were given AlCl₃ for 4 weeks,the concentrations of GOT in mice's serum increased,and decreased after the mice were given high-dose of DFP for lweek.The concentrations of GPT in mice's serum in A1 control group was higher than in negtive control group,and higher than low,middle,high doses groups too.But all the changes had no statistic significance.The activity of serum AKP in A1 control and low,middle,high doses groups were all higher than negtive control group significangly(P＜0.01).No significant differences existed in the low,middle,high doses groups and A1 control group.No significant differences existed among all groups' concentrations of serumγ-GT.1.3 The effect of DFP on activity of ATPase in Al-loaded mice's livers The activity of Ca²⁺Mg²⁺-ATPase in A1 control,low and middle doses groups was lower than that of negtive control group(P＜0.01),and there was no significant difference existed between high and negtive control group.The activity of Na⁺K⁺-ATPase in A1 control group was lower than that of negtive control group,and lower than that of DFP treated groups,but all the difference had no statistic significance.1.4 The effect of DFP on concentrations of XOD in Al-loaded mice's liversThe concentrations of liver XOD in A1 control group and low,middle doses groups were higher than that of negtive control group(P＜0.01),whereas DFP treated goups were lower than A1 control group(P＜0.05,P＜0.01).No significant differences existed between high dose group and negtive control group in the concentrations of liver XOD.1.5 The effect of DFP on contents of aluminum and calcium,magnesium,zinc, copper and iron elements in Al-loaded mice's liversThe results showed that the aluminum contents of A1 control group in livers were significantly higher than that of the negative control group(P＜0.01).Aluminum contents of the three doses group were lower than that of A1 control group(P＜0.01), but no significant difference existed when they were compared with negtive control group.The calcium contents of A1 control group and low,middle doses groups in livers were significantly lower than that of the negative control group(P＜0.05),whereas no significant difference existed between high dose group and negtive control group.No significant difference existed in all groups' liver magnesium,zinc,copper elements contents.The iron contents of A1 control group in livers were higher than that of the negative control group,but there was no significant difference between them.The iron content of all DFP treated groups were lower than A1 control group and negtive control group(P＜0.01).1.6 Pathomorphological observation of the liver cellsThe liver cells of A1 control group presented degeneration,blood sinus were broaden and the hepatic cords arranged disorderly.Slight recovering could be observed in the DFP-treated groups.2 The effect of DFP protecting against Al-loaded mice's kidneys2.1 The effect of DFP on kidney/body weight ratio of Al-loaded miceNo significant differences existed among all groups' kidney/body weight ratios.2.2 The effect of DFP on Scr and BUN of Al-loaded miceNo significant differences existed among all groups' serum Cr concentrations.Compared to negtive group,the BUN concentrations in all A1 treated groups increased(P＜0.01).The BUN concentrations of all DFP treated groups were lower than that of A1 control group,but there was statistic signicance only when high dose group and A1 control group were compared(P＜0.05).2.3 The effect of DFP on activity of ATPase in Al-loaded mice's kidneysThe activity of Na⁺K⁺-ATPase and Ca²⁺Mg²⁺-ATPase in A1 control group was significantly lower than negtive control group(P＜0.01),and was significantly lower than DFP treated groups(P＜0.01).There was no significant difference existed among the three doses groups and negtive group.2.4 The effect of DFP on concentrations of XOD in Al-loaded mice's kidneysThe concentrations of kidney XOD in A1 control group was higher than that of negtive control group(P＜0.01),whereas DFP treated goups were lower than A1 control group(P＜0.01).No significant differences existed between DFP treated group and negtive control group in the concentrations of kidney XOD.2.5 The effect of DFP on contents of aluminum and calcium,magnesium,zinc, copper and iron elements in Al-loaded mice's kidneysThe results showed that the aluminum contents of A1 treated groups in kidneys was significantly higher than that of the negative control group(P＜0.01,P＜0.05). Aluminum contents of the three doses group were lower than that of A1 control group (P＜0.01).The calcium contents of A1 control group and low,middle,high doses groups in kidneys were significantly lower than that of the negative control group(P＜0.05).The calcium contents of low,middle,high doses groups in kidneys were higher than that of the A1 control group,but there were no significant difference existed among them.No significant difference existed in all groups' kidney magnesium,zinc,copper, iron elements concentrations.2.6 Pathomorphological observation of the kidney cellsThe renal glomerulus and tubules of A1 control group presents pathologic changes.Slight recovering could be observed in the DFP-treated groups.Conclusions1.DFP could chelate aluminum and reduce aluminum contents in liver to reduce the damages to mice livers A1 caused.It can remarkably recover the protein production ability of livers,maintain the balance of elements,and inhibit pathological damages of livers.In short,DFP played an roal in protecting the liver.2.DFP could remarkably recover the Al-induced renal function by reducing aluminum contents,protect kidneys away from Al-induced damanges to some extent.