Effect Of Intrauterine Growth Retardation And Postnatal Nutritional Intervention On Renal Damages In Rats

PARTⅠInfluence of intrauterine growth retardation on blood pressure and renalfunction in ratsBackgroundEpidemiological studies have suggested that intrauterine growth retardation (IUGR) is a risk factor for adult deseases, including hypertension, type 2 diabetes mellitus, coronary artery disease, and chronic kidney diseases. This indicates that factors in the prenatal and early postnatal environment can "programming" the individual for increased risk of diseases later in life. But the mechanism is unknown. The goal of present study is to establish an IUGR rat model by maternal protein restriction during pregnancy and analyze the effect of IUGR on blood pressure and renal function, and in order to determine the influence of prenatal environment on hypertension and kidney diseases.MethodsA rat model of IUGR was employed. Pregnant Sprague-Dawley rats were kept on 6% protein isocaloric diet through gestation. We chose males as our study objects. Systolic blood pressure was monitored by the tail cuff method, and 24hr urinary protein was measured by routine biochemical method at 4 weeks, 8weeks and 12weeks respectively. At 12 weeks old, the total number of glomeruli was determined by the maceration method, renal plasma flow (RPF) was calculated by the renal clearance of para-aminohippuricacid, and glomerular filtration rate (GFR) was calculated by inulin clearance.ResultsThe offspring from the low-protein pregnancies had significantly lower birth weights than those in controls (6．92±0．33g vs. 4．81±0．27g, P<0．001) . The total number of glomeruli per kidney was decreased by 21% (22900±926 glomeruli vs. 28861±1044 glomeruli, P＜0．01) . They developed increased blood pressure levels by 8 weeks of age (125．2±2．3mmHg vs. 109．4±3．1mmHg, P＜0．01) and the hypertension progressed at 12 weeks (132．1±2．9 mmHg vs. 115．2±2．3mmHg, P<0．01) . By 12 weeks of age, they had increased kidney weight /body weight (0．378±0．019% vs. 0．334±0．007%,P＜0．05) and mild proteinuria (118．46±21．85 mg·kg^-1·d^-1 vs. 69．72±10．35 mg·kg^-1·d^-1 ,P＜0．001) , but GFR and RPF showed no significant differences (P>0．05, respectively).ConclusionThis study suggests that adult hypertension is susceptible to IUGR associated withmaternal low-protein diet, and reduced nephron endowment might play a role in thethe aetiology of hypertension. PARTⅡStudy on mechanism of nephron deficit in rat model of intrauterine growthretardationBackgroundBoth human and animal model studies demonstrate that intrauterine growth retardation (IUGR) lead to a decrease in glomeruli; however, the molecular mechanisms are still unknown. Nephrogenesis involves the rapid remodeling of structures, which requires massive proliferation and apoptosis, and is regulated by expression of lots of genes, such as WT1, Bcl-2, Bax, and p53．Therefore, We hypothesized that such nephron deficits in IUGR are associated with altered cell turnover and expression of related genes in fetal kidney.MethodsA rat model of IUGR was built by maternal low-protein (6%) diet throughout pregnancy. We chose newborn male pups as our study objects. The proliferation and apoptosis in kidney was showed by Ki-67 detection and TUNEL method. Renal expressions for WT1, Bcl-2, Bax, and p53 mRNAs was examined by real-time PCR. Western blot was used to examine the expression of WT1 and Bcl-2 gene products in renal tissue. The final number of glomeruli was determined at 2 weeks of age when nephrogenesis has finished.ResultsAt two weeks postnatally, IUGR offspring had fewer glomeruli per kidney than those in controls (22891±1242 glomeruli vs. 28466±919 glomeruli, P＜0．001) . Histopathologic analysis showed not only the cortex appeared thinner in 1-day-old IUGR offsprings compared to controls (391．94±24．73μm vs. 425．50±24．01μm, P＜0．01) , but also the proportion of cortex that was in the nephrogenic zone was increased (38．1±3．4% vs. 24．3±6．7%, P＜0．01) . In IUGR newborns, TUNEL positive cells were more numerous in the nephrogenic zone (21．26±1．94% vs. 12．14±1．96%, P＜0．001) . Renal WT1 and Bcl-2 mRNA levels were significantly reduced in newborn IUGR pups (4．96±0．17 vs. 5．33±0．22, 4．28±0．14 vs. 4．46±0．11, P＜0．01, respectively), and the Bcl-2 mRNA/Bax mRNA ratio was also decreased (0．739±0．022 vs. 0．770±0．030, P＜0．05) , but there was no change in the expression of p53 mRNA (P>0．05) . The WT1 and Bcl-2 protein expressions were also significantly decreased (0．364±0．015 vs 0．554±0．053, 0．573±0．022 vs. 0．914± 0．059, P＜0．001, respectively) in IUGR newborns.ConclusionThese results suggest that reduction of nephron number in IUGR rat may beassociated with enhanced apoptosis in kidney development. Decreased WT1 andBcl-2 expressions and reduction of the Bcl-2/Bax ratio may contribute to themolecular mechanisms behind these findings. partⅢEffect of postnatal high protein diet on renal damages of rats with intrauterinegrowth retardationBackgroundIt was believed that postnatal high protein diet was helpful for catch-up growth in intrauterine growth retardation (IUGR) children. But recently, the "predictive adaptive response(PAS)"theory proposed that the mismatching of prenatal and postnatal enviorment can increase the risk of adult diseases in later life. And high protein in itself may lead to glomerular hyperfusion and hyperfiltration, resulting in more serious damage to the kidney. Based on those concern, we designed the disparity between the prenatal and postnatal enviorment, and tried to investigate the influence of high protein diet on renal development and renal function in IUGR rats.MethodsA rat model of IUGR was built by maternal low-protein (6%) diet throughout pregnancy. Male pups were divided randomly into 2 groups, fed either a control diet containing 22% protein (IUGR group) or high protein diet containing 30% protein (HP group), and normal male pups as control (Con group). Systolic blood pressure was monitored by the tail cuff method, and 24hr urinary protein was measured by routine biochemical method at 4 weeks, 8weeks and 12weeks respectively. At 12 weeks old, the total number of glomeruli was determined by the maceration method, renal pathologic morphology was observed under light and electronic microscope, renal expressions for desmin mRNA was examined by real-time PCR.ResultsAt the 12^th week, the blood pressure and proteinuria in HP group were more server than those of IUGR group(138．6±2．8mmHg vs. 132．1±2．9 mmHg, P＜0．01; 202．61±62．55 mg·kg^-1·d^-1 vs. 118．46±21．85, P＜0．001) , but the total number of glomeruli per kidney in HP group was not significantly different from IUGR group(23043±595 glomeruli vs. 22900±926 glomeruli, P>0．05) . The morphometric analysis revealed that the extracellular matrix area was significantly inceased in HP group compared with that in controls (31．6±2．3% vs. 21．3±2．1%, P＜0．01) . Electron microscopy showed that fusion of foot process was scarcely observed in IUGR rats, otherwise there was partial fusion of foot process and hyperplasia of mesangial cells in HP rats. Renal desmin mRNA levels was greatly increased in both IUGR and HP groups,(IUGR:3．33±0．41, HP:4．06±0．27 vs. Con:2．77±0．37, P＜0．05, respectively), withmore significant in HP group.ConclusionFeeding postnatal high protein diet to IUGR rats leads to no increased glomerularnumber, but more severe hypertension and proteinuria. Damage to the podocytes maybe one of the mechanisms in which IUGR and postnatal high protein diet lead toproteinuria.