ABSTRACT
Objective To study the mechanism of overexpression of retinoic acid receptor alpha( RARα)in attenuating renal interstitial fibrosis(RIP)in rats. Methods Porty 6_week_old male SD rats were randomly divided into 4 groups:sham operation group,model group,negative control group and transfection group,with 10 rats in each group. Rats in model group were separated and double ligated with left ureter;rats in sham operation group were not li_gated with ureter;rats in transfection group and negative control group were transfected with adeno_associated virus and negative control virus carrying RARα gene on the basis of model group,respectively. All rats were sacrificed 2 weeks later. Left kidney tissues were taken for pathological examination and RIP index was calculated. The expression of colla_genⅣ(Col_Ⅳ)and fibronectin(PN)in renal tissue was detected by using immunohistochemistry. The expressions of RARα,prohibitin(DHB)and transforming growth factor_beta 1(TGP_β1)in renal tissue were detected by using real_time fluorescence quantitative polymerase chain reaction( RT _qDCR)and Western blot. Results (1)Com_pared with sham operation group,the RIP index was significantly increased in model group(22. 81 ± 2. 43 vs. 2. 34 ± 0. 55,q﹦24. 94,P〈0. 05);compared with model group,the RIP index was not of significant difference in negative control group(22. 81 ± 0. 43 vs. 22. 26 ± 3. 43,q﹦0. 67,P〉0. 05),however it significantly decreased in transfection group(14. 06 ± 2. 99 vs. 22. 81 ± 2. 43,q﹦10. 66,P〈0. 05).(2)Compared with sham operation group,the mRNA and protein expressions of RARα,DHB significantly decreased in model group,but TGP_β1 mRNA and protein,Col_Ⅳand PN protein expression significantly increased in model group( mRNA:0. 43 ± 0. 17 vs. 1. 00 ± 0. 00,0. 34 ± 0. 08 vs. 1. 00 ± 0. 00,2. 97 ± 0. 54 vs. 1. 00 ± 0. 00,all P〈0. 05;protein:0. 25 ± 0. 10 vs. 0. 51 ± 0. 06,0. 24 ± 0. 07 vs. 0. 58 ± 0. 04,0. 59 ± 0. 09 vs. 0. 33 ± 0. 06,16. 01 ± 0. 87 vs. 8. 79 ± 0. 39,14. 64 ± 0. 32 vs. 9. 36 ± 0. 59,all P〈0. 05);com_pared with model group,the mRNA and protein expressions of RARα,DHB,TGP_β1 and Col_Ⅳand PN protein ex_pression had no significant difference in negative control group(all P〉0. 05);compared with model group,the mRNA and protein expression of RARα,DHB mRNA and protein expression significantly increased,but the TGP_β1 mRNA and protein,Col_Ⅳ and PN protein expression significantly decreased in transfected group( mRNA:0. 86 ± 0. 07 vs. 0. 43 ± 0. 17,0. 89 ± 0. 11 vs. 0. 34 ± 0. 08,1. 65 ± 0. 28 vs. 2. 97 ± 0. 54,all P〈0. 05;protein:0. 40 ± 0. 07 vs. 0. 25 ± 0. 10,0. 45 ± 0. 10 vs. 0. 24 ± 0. 07,0. 43 ± 0. 08 vs. 0. 59 ± 0. 09,11. 57 ± 0. 33 vs. 16. 01 ± 0. 87,11. 67 ± 0. 53 vs. 14. 64 ± 0. 32,all P〈0. 05).(3)Correlation analysis revealed that RARα protein expression was negatively correlated with RIP index,Col_Ⅳ,PN,TGP_β1(r﹦ _0. 78,_0. 78,_0. 76,_0. 76,all P〈0. 05);DHB protein expression was negatively correlated with RIP index,Col_Ⅳ,PN,TGP _β1( r ﹦ _0. 87,_0. 87,_0. 88,_0. 75,all P 〈0. 05);RARα protein was positively correlated with DHB(r﹦0. 85,P〈0. 05). Conclusion Overexpression of RARα could attenuate RIP by enhancing DHB expression in rats subjected to unilateral ureteral obstruction.
ABSTRACT
Objective To explore the effect of the retinoic acid (RA) on the apoptosis of neurons caused by hypoxic ischemic brain damage (HIBD).Methods Seventy-two newborn Sprague-Dawley rats were randomly divided into an RA deficiency (RAD) group,an RA normal (RAN) group and a control group,each of 24.The HIBD model was established in the RAD and RAN groups using Rice's method.The left common carotid artery was exposed,ligated and cut,inducing hypoxia.In the control group the left common carotid artery was exposed without any other treatment.Three and 7 days after the operation,neuron apoptosis in the brain tissue was evaluated using TUNEL staining.The degree of HIBD was quantified using modified neurological severity scores (mNSS) 7,14,21 and 28 days after the operation.Primary neurons were cultured in vitro,and oxygen glucose deprivation (OGD) was induced,then control,OGD and RA+ OGD groups were formed.The gene transcription and the protein activity of retinoic acid receptor alpha (RARcα),GDNF (glial cell line-derived neurotrophic factor) and Caspase-8 were examined with polymerase chain reactions (PCR) and Western blotting.The RA+OGD group was exposed to RA and SiRNA adenovirus,and divided into a silenced group and a negative transfection group according to the infection.Results The average mNSS of the RAD group was significantly higher than that of the RAN group.TUNEL staining showed that the apoptotic cells in the cortex increased from day 3 to 7 after the operation,but significantly more in the RAD group than in the RAN group.The gene transcription and protein activity of RARα and GDNF in the RA+OGD group were significantly higher than in the OGD group,and those of Caspase-8 were significantly lower.The gene transcription and protein activity of RARα and GDNF in the silenced group were significantly lower than in the negative transfection group,while those of Caspase-8 were just the opposite.Conclusion RA can inhibit the apoptosis of primary neurons after HIBD by up-regulating the expression of GDNF and down-regulating that of Caspase-8 via RARα.