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Objective To observe the effects of salvianolate on blood biochemical indexes, pathological changes of renal tissue and expression of CD2AP and Desmin protein in membranous nephropathy rats induced by cationic bovine serum albumin. And to explore the renal protective effect of salvianolate on membranous nephropathy rats and its possible mechanism. Methods Healthy male SD rats were randomly divided into normal group and modle group. Rat models of membranous nephropathy were reproduced by injection of cationized bovine serum albumin through tail vein. Model successful rats were randomly divided into model group, benazepril group, and salvianolate groups (16.7, 33.3, and 66.7 mg/kg). Each group was given the dose of the corresponding drugs. After treatment, the level of 24 h urine total protein (UTP), serum total cholesterol (TC), triglyceride (TG), total protein (TP), albumin (ALB), urea nitrogen (BUN), and serum creatinine (Scr) were detected. Immunofluorescence, light microscope, and electron microscope were used to observe the pathological changes of rat kidney. Immunohistochemistry and real-time PCR were used to detect the expression of CD2AP and Desmin. Results Compared with the control group, levels of UTP, TC, and TG increased significantly (P < 0.01), levels of serum TP and ALB decreased significantly (P < 0.01) in the model group. Compared with the model group, the UTP, TC, and TG of each treatment group were significantly decreased (P < 0.05, 0.01), while the TP and ALB were significantly increased (P < 0.01). There was no significant difference in the UTP, TC, TG, TP, and ALB among each dosage of salvianolate and benazepril group. And there was no significant difference among each dose group of salvianolate. There was no significant difference among each groups in the level of BUN and Scr. Compared with the normal group, the expression of CD2AP in model group was significantly decreased and Desmin was significantly increased (P < 0.01). Compared with the model group, the expression of CD2AP increased and Desmin decreased in each treatment group (P < 0.01), but there was no difference among the treatment groups. Conclusion Salvianolate has kidney protective effect on membranous nephropathy rats. The mechanism may be related to up-regulating the expression of CD2AP, down-regulating the expression of Desmin, inhibiting podocyte injury and protecting the integrity of the glomerular filtration barrier.
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Objective To observe the expression of mRNA of podocin,nephrin,CD2AP and α - actin - 4 in Doxorubicin - induced nephrotic(ADN)rats,and explore the possible mechanisms of podocyte molecule during the de-velopment of proteinuria. Methods Forty - eight Sprague - Dawley(SD)rats were divided into ADN model group(in-jected with 6. 5 mg/ kg Doxorubicin in tail vein,n = 24)and control group(injected with saline solution in tail vein,n =24). After the nephropathy model was established,6 rats were killed at the end of 1st ,2nd ,4th ,6th week in each group. The changes of the following indicators were observed:(1)24 - hour urinary protein,serum albumin and cholesterol were detected;(2)mRNA expression of nephrin,podocin,CD2AP and α - actin - 4 in cortex of kidney were examined by real time fluorescence quantification PCR. Results The model group came out massive proteinuria(15. 66 ± 1. 50) mg/ 24 h,(45. 98 ± 1. 45)mg/ 24 h,(65. 58 ± 4. 68)mg/ 24 h,(82. 83 ± 8. 43)mg/ 24 h in 1,2,4,6 weeks respec-tively,hypoalbuminemia(27. 4 ±2. 5)g/ L,(23. 6 ±2. 9)g/ L,(20. 6 ±1. 5)g/ L,(6. 9 ± 2. 3)g/ L in 1,2,4,6 weeks respectively and hypercholesterolaemia(2. 00 ± 0. 25)mmol/ L,(2. 16 ± 0. 44)mmol/ L,(4. 02 ± 0. 81)mmol/ L, (7. 54 ± 1. 12)mmol/ L in 1,2,4,6 weeks respectively,and the differences of proteinuria,plasma albumin and total cholesterol compared with control group at each time point had statistical significance(all P ﹤ 0. 01). Compared with the control group,podocin mRNA expression in the model group decreased at the end of 1st week(10. 56 ± 3. 62),de-creased significantly at the end of 2nd week(20. 44 ± 9. 03),and decreased at the end of 4th week(2. 19 ± 0. 18)com-pared with the control group;nephrin mRNA expression decreased at the end of 1st week(2. 41 ± 1. 10)and reached to the peak value,decreased at the end of 4th week(0. 52 ± 0. 18);CD2AP mRNA expression did not change significantly in the 1st week(4. 17 ± 0. 79),increased at the end of 2nd week(6. 74 ± 1. 53),reached to the peak value at the end of 4th week(6. 91 ± 1. 13),but did not change significantly at the end of 6th week(4. 04 ± 0. 82);α - actin - 4 mRNA ex-pression did not change significantly at the end of 1st week(1. 75 ± 0. 48),decreased at the end of 2nd week(2. 01 ± 0. 55),reached to the peak value at the end of 4th week(2. 24 ± 0. 81),but did not change significantly at the end of 6th week(1. 39 ± 0. 18). Compared with the control group,the difference had statistical significance( all P ﹤ 0. 05). Conclusion The abnormal expression of podocyte molecules mRNA in ADN rats may be an important molecular mechanism in the development of proteinuria.
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Objective To study NPHS2 and CD2AP gene mutation with primary steroid-resistant nephroticsyndrome (SRNS) children in Guangdong province,and to investigate the relationship between NPHS2,CD2AP genemutation and SRNS,so as to provide a theoretical basis for the diagnosis and treatment of SRNS in children.Methods Twenty-six SRNS children and 20 cases of the healthy children as controls were chosen randomly in Guangdong province.Genomic DNA was isolated from peripheral blood leucocytes of these patients and the healthy children.Mutational analysis was performed in 8 exons of NPHS2 gene and 18 exons of CD2AP gene after sequencing directly.The results were compared with United States National Center for Biotechnology Information (NCBI) gene database,the detected gene mutation.Results The variation analysis revealed 3 polymorphisms (288C ＞ T,954T ＞ C,1038A ＞ G) in 14 cases out of 26 patients and 4 cases of the healthy children studied,which had been reported before,but there was no significant difference in the genotypic and allelic frequencies of these polymorphisms between the patients and the controls (all P ＞ 0.05).One CD2AP heterozygous mutation (1917 + 20 C ＞ G) was detected in intron in 2 cases of SRNS children.Conclusions NPHS2 gene variation may not be the main mechanism of SRNS in Guangdong province.CD2APgene mutation may increase the possibility of SRNS and focal segmental glomerulosclerosis in children.CD2AP mutation in intron may involve in the pathogenesis of SRNS.
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Mutations in CD2AP, mapped to 6pL2. 3 and encoding CD2 - associated protein ( CD2AP), are responsible for autosomal recessive or autosomal dominant steroid - resistant nephrotic syndrome (SRNS). CD2AP plays a key role in the slitdiaphragm network of the kidney, which is necessary for the structure and ultrafiltration functions of the slitdiaphragm. The CD2AP homozygous mutation results in early - onset SRNS while heterozygous expression of the CD2AP mutation has increased susceptibility to glomerular injury.No recurrence of proteinuria was observed in the patient with SRNS with CD2AP homozygousmutationafter the renal transplantation. Therefore, detection of the CD2AP gene in the patients suffering from SRNS will be beneficial to making therapeutic decisions and predicting prognoses.
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BACKGROUND: Regardless of the underlying diagnosis, the proteinuric condition demonstrates ultrastructural changes in podocytes with retraction and effacement of the foot processes and componental changes in slit diaphragm. We examined the molecular basis for this alteration of the podocyte phenotype, involving quantitative and distributional changes especially on CD2AP as a candidate regulating the modulation of pathogenic changes in the barrier to protein filtration. METHODS: To investigate whether high glucose and AGE induce podocyte cytoskeletal changes, we cultured rat GEpC under normal (5 mM) or high glucose (HG, 30 mM) and AGE- or BSA-added conditions and examined the distribution of CD2AP by confocal microscope and measured the change of CD2AP expression by Western blotting and RT-PCR. RESULTS: We found that CD2AP moved from peripheral to inner cytoplasm in the HG condition by confocal microscopy. In Western blotting, administration of high glucose or AGE decreased the CD2AP productions by 36.9% (p<0.05) and 16.0% (p< 0.05), respectively. Furthermore, both high glucose and AGE decreased the amount of CD2AP more significantly by 64.6% compared to those of control (p<0.01). Such changes was not seen in osmotic control. In RT-PCR, administration of high glucose, AGE or both high glucose and AGE decreased the expression of CD2AP mRNA by 44.9%, 27.9%, and 29.3% (p<0.05), respectively, compared to that of control. CONCLUSION: We could find that HG induce the inward translocation of CD2AP molecule and HG and AGE suppress the production of CD2AP at transcriptional and partly translational level. We suggest that these changes may explain the structural and functional changes of podocytes in diabetic conditions.