Intrauterine growth retardation aggravates the course of acute mesangioproliferative glomerulonephritis in the rat
2006; Elsevier BV; Volume: 70; Issue: 11 Linguagem: Inglês
10.1038/sj.ki.5001966
ISSN1523-1755
AutoresChristian Plank, Iris Östreicher, Andrea Hartner, Ines Marek, Friedrich G. Struwe, Kerstin Amann, Karl F. Hilgers, Wolfgang Rascher, Jörg Dötsch,
Tópico(s)Neonatal Respiratory Health Research
ResumoIntrauterine growth retardation (IUGR) aggravates the course of acute mesangioproliferative glomerulonephritis (GN) in the rat. Observational studies in children suggest that IUGR may be associated with a severe course of kidney diseases such as IgA nephropathy. We tested the hypothesis that IUGR leads to aggravation of acute mesangioproliferative GN in former IUGR rats. IUGR was induced in Wistar rats by isocaloric protein restriction in pregnant dams. Litter size was reduced to six male neonates in low protein animals (LP) and normal protein animals (NP). At 8 weeks GN was induced by injection of an anti-Thy-1.1 antibody. Rats were killed on days 4 and 14 after induction of GN and kidneys were investigated for inflammation and sclerosis using real-time polymerase chain reaction and histological methods. On day 4 after induction of GN, LP animals showed more glomerulosclerosis and tubulointerstitial lesions. On day 14, inflammatory markers (expression of monocyte chemoattractant protein 1, osteopontin, tumor necrosis factor and interleukin-6), extracellular matrix accumulation and markers of sclerosis (plasminogen activator inhibitor-1 expression, transforming growth factor-β1 expression, score for glomerulosclerosis, glomerular deposition of collagen I and collagen IV) were more severe in LP animals. Some degree of induction of inflammatory and profibrotic markers was also present in non-nephritic LP animals. However, these rats did not display marked glomerulosclerosis or interstitial fibrosis. We conclude that after IUGR inflammatory damage is aggravated and the reparation of the kidney is impaired during the course of acute mesangioproliferative GN, leading to more sclerotic lesions. Intrauterine growth retardation (IUGR) aggravates the course of acute mesangioproliferative glomerulonephritis (GN) in the rat. Observational studies in children suggest that IUGR may be associated with a severe course of kidney diseases such as IgA nephropathy. We tested the hypothesis that IUGR leads to aggravation of acute mesangioproliferative GN in former IUGR rats. IUGR was induced in Wistar rats by isocaloric protein restriction in pregnant dams. Litter size was reduced to six male neonates in low protein animals (LP) and normal protein animals (NP). At 8 weeks GN was induced by injection of an anti-Thy-1.1 antibody. Rats were killed on days 4 and 14 after induction of GN and kidneys were investigated for inflammation and sclerosis using real-time polymerase chain reaction and histological methods. On day 4 after induction of GN, LP animals showed more glomerulosclerosis and tubulointerstitial lesions. On day 14, inflammatory markers (expression of monocyte chemoattractant protein 1, osteopontin, tumor necrosis factor and interleukin-6), extracellular matrix accumulation and markers of sclerosis (plasminogen activator inhibitor-1 expression, transforming growth factor-β1 expression, score for glomerulosclerosis, glomerular deposition of collagen I and collagen IV) were more severe in LP animals. Some degree of induction of inflammatory and profibrotic markers was also present in non-nephritic LP animals. However, these rats did not display marked glomerulosclerosis or interstitial fibrosis. We conclude that after IUGR inflammatory damage is aggravated and the reparation of the kidney is impaired during the course of acute mesangioproliferative GN, leading to more sclerotic lesions. Intrauterine growth retardation (IUGR) is a risk factor for the development of a metabolic syndrome later in life.1.Hales C.N. Early programming of glucose metabolism, insulin action and longevity.Adv Exp Med Biol. 2000; 478: 57-64Crossref PubMed Google Scholar, 2.Barker D.J. Intrauterine programming of adult disease.Mol Med Today. 1995; 1: 418-423Abstract Full Text PDF PubMed Scopus (323) Google Scholar, 3.Reyes L. Manalich R. Long-term consequences of low birth weight.Kidney Int Suppl. 2005; 97: S107-S111Abstract Full Text Full Text PDF PubMed Scopus (68) Google Scholar In former IUGR individuals, type II diabetes, hypertension, hyperlipidemia, and cardiovascular disease occur more frequently than in individuals with normal birth weight.4.Barker D.J. In utero programming of chronic disease.Clin Sci (London). 1998; 95: 115-128Crossref PubMed Google Scholar,5.McMillen I.C. Robinson J.S. Developmental origins of the metabolic syndrome: prediction, plasticity, and programming.Physiol Rev. 2005; 85: 571-633Crossref PubMed Scopus (1440) Google Scholar Moreover, a higher incidence of end-stage renal disease at adulthood was described in patients with former IUGR,6.Lackland D.T. Bendall H.E. Osmond C. et al.Low birth weights contribute to high rates of early-onset chronic renal failure in the southeastern United States.Arch Intern Med. 2000; 160: 1472-1476Crossref PubMed Scopus (296) Google Scholar,7.Lackland D.T. Egan B.M. Fan Z.J. et al.Low birth weight contributes to the excess prevalence of end-stage renal disease in African Americans.J Clin Hypertens (Greenwich). 2001; 3: 29-31Crossref PubMed Scopus (53) Google Scholar which, at least in part, can be attributed to the higher incidence of metabolic diseases, like diabetes type II or hypertension. On the other hand, there is increasing evidence for an association of IUGR and glomerular disease in children independent of a pre-existing metabolic disease. Three retrospective analyses found a greater incidence of complications and an unfavorable progression of nephrotic syndrome in children with IUGR.8.Zidar N. Avgustin Cavic M. Kenda R.B. et al.Unfavorable course of minimal change nephrotic syndrome in children with intrauterine growth retardation.Kidney Int. 1998; 54: 1320-1323Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar, 9.Sheu J.N. Chen J.H. Minimal change nephrotic syndrome in children with intrauterine growth retardation.Am J Kidney Dis. 2001; 37: 909-914Abstract Full Text PDF PubMed Scopus (36) Google Scholar, 10.Na Y.W. Yang H.J. Choi J.H. et al.Effect of intrauterine growth retardation on the progression of nephrotic syndrome.Am J Nephrol. 2002; 22: 463-467Crossref PubMed Scopus (25) Google Scholar IgA nephropathy is one of the most frequent renal diseases during childhood. Children with IUGR showing birth weight below the 10th percentile for the respective gestational age were shown to be at higher risk to develop progressive glomerulosclerosis during this disease compared to children without IUGR.11.Zidar N. Cavic M.A. Kenda R.B. et al.Effect of intrauterine growth retardation on the clinical course and prognosis of IgA glomerulonephritis in children.Nephron. 1998; 79: 28-32Crossref PubMed Scopus (79) Google Scholar Little is known about potential pathways linking IUGR and the later progression of inflammatory renal diseases. Therefore, we tested the hypothesis that IUGR leads to a more severe course of glomerular disease. Of the different animal models of IUGR,12.Bertram C.E. Hanson M.A. Animal models and programming of the metabolic syndrome.Br Med Bull. 2001; 60: 103-121Crossref PubMed Scopus (223) Google Scholar,13.Vehaskari V.M. Woods L.L. Prenatal programming of hypertension: lessons from experimental models.J Am Soc Nephrol. 2005; 16: 2545-2556Crossref PubMed Scopus (96) Google Scholar we chose the low protein (LP) diet model in the rat, which is widely used.14.Langley-Evans S.C. Welham S.J. Sherman R.C. et al.Weanling rats exposed to maternal low-protein diets during discrete periods of gestation exhibit differing severity of hypertension.Clin Sci (London). 1996; 91: 607-615Crossref PubMed Scopus (222) Google Scholar, 15.Langley-Evans S.C. Hypertension induced by foetal exposure to a maternal low-protein diet, in the rat, is prevented by pharmacological blockade of maternal glucocorticoid synthesis.J Hypertens. 1997; 15: 537-544Crossref PubMed Scopus (181) Google Scholar, 16.Vehaskari V.M. Aviles D.H. Manning J. Prenatal programming of adult hypertension in the rat.Kidney Int. 2001; 59: 238-245Abstract Full Text Full Text PDF PubMed Scopus (327) Google Scholar, 17.Bertram C. Trowern A.R. Copin N. et al.The maternal diet during pregnancy programs altered expression of the glucocorticoid receptor and type 2 11 beta-hydroxysteroid dehydrogenase: potential molecular mechanisms underlying the programming of hypertension in utero.Endocrinology. 2001; 142: 2841-2853Crossref PubMed Scopus (286) Google Scholar, 18.Woods L.L. Ingelfinger J.R. Nyengaard J.R. et al.Maternal protein restriction suppresses the newborn renin-angiotensin system and programs adult hypertension in rats.Pediatr Res. 2001; 49: 460-467Crossref PubMed Scopus (471) Google Scholar IUGR is induced in the offspring by protein restriction to half or less of the normal protein (NP) content to the pregnant dams. Most often this prenatal protein restriction is followed by NP content in post-natal diet modeling the change between prenatal restriction of nutrition supply and adequate post-natal nutrition.13.Vehaskari V.M. Woods L.L. Prenatal programming of hypertension: lessons from experimental models.J Am Soc Nephrol. 2005; 16: 2545-2556Crossref PubMed Scopus (96) Google Scholar Typical aspects of this model are reduced birth weight, development of arterial hypertension in later life, and a reduced glomerular number.13.Vehaskari V.M. Woods L.L. Prenatal programming of hypertension: lessons from experimental models.J Am Soc Nephrol. 2005; 16: 2545-2556Crossref PubMed Scopus (96) Google Scholar As a model for renal inflammation, we selected acute mesangioproliferative glomerulonephritis (GN) in rats. This form of GN is initiated by injection of an anti-Thy-1.1 antibody causing a self limiting GN with features reminiscent of human IgA nephropathy.19.Bagchus W.M. Hoedemaeker P.J. Rozing J. et al.Glomerulonephritis induced by monoclonal anti-Thy 1.1 antibodies. A sequential histological and ultrastructural study in the rat.Lab Invest. 1986; 55: 680-687PubMed Google Scholar,20.Hartner A. Hilgers K.F. Bitzer M. et al.Dynamic expression patterns of transforming growth factor-beta(2) and transforming growth factor-beta receptors in experimental glomerulonephritis.J Mol Med. 2003; 81: 32-42Crossref PubMed Scopus (29) Google Scholar We hypothesized that IUGR influences the course of GN in later life owing to a reduced regeneration capacity. LP diet of the pregnant dams led to a reduction of birth weight and body length of their offspring at birth in comparison to litters of NP fed dams (Table 1). The total number of animals per litter was not different (16.0±0.8 pups in NP mothers versus 15.0±1.3 pups in LP mothers, P=0.61). Until day 12 after birth, however, the offspring of the mothers fed LP diet caught up with the offspring of mothers with normal 17% protein diet (NP). At the age of 8 weeks, the body weight of LP and NP was comparable (Table 1). At day 70, mean arterial blood pressure was not different in LP and NP animals (Table 1). There was no development of proteinuria and no significant difference in endogenous creatinine clearance (Table 1). Relative kidney weight was not different on day 70 (Table 1).Table 1Physiological and stereological data of control animals and animals with IUGRNPLPBirth weight (g)6.4±0.15.6±0.12*P<0.05 versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy).Body length at birth (cm)5.0±0.034.8±0.04*P<0.05 versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy).Body weight at the time of induction of GN=week 8 (g)316.2±3.89317.7±6.62Urinary protein excretion (mg/24 h/100 g BWT)4.05±0.585.08±6.43Creatinine clearance (ml/h/100 g BWT)37.77±4.8231.90±2.05Creatinine clearance (ml/h/1000 glomeruli)2.83±0.552.91±0.31Mean arterial pressure (mm Hg)105.3±1.64101.3±2.69Relative kidney weight (mg/g BWT)4.04±0.093.98±0.17Number of glomeruli/kidney54 960±223140 370±2638*P<0.05 versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy).Total glomerular volume (mm3)44.87±3.1637.37±5.47Mean glomerular volume (× 103 μm3)817.2±54.92907.6±89.23Glomerular filtration area per kidney (mm2)172.1±20.0104.5±13.5*P<0.05 versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy).IUGR, intrauterine growth retardation; LP, low protein; NP, normal protein.Values are given as means±s.e.m.Data for body weight at birth, body length at birth, and body weight at the time of induction of GN are given from animals of all groups (NP, n=22, LP, n=21).All other data are given from the day 70 animals (NP d70, n=12, LP d70, n=11).Six NP d70 and five LP d70 animals were randomly selected for stereological analysis.* P<0.05 versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy). Open table in a new tab IUGR, intrauterine growth retardation; LP, low protein; NP, normal protein. Values are given as means±s.e.m. Data for body weight at birth, body length at birth, and body weight at the time of induction of GN are given from animals of all groups (NP, n=22, LP, n=21). All other data are given from the day 70 animals (NP d70, n=12, LP d70, n=11). Six NP d70 and five LP d70 animals were randomly selected for stereological analysis. The total number of glomeruli per kidney was about 27% lower in LP than in NP (Table 1). Total glomerular volume and mean glomerular volume were not different. Thus, total filtration area per kidney was lower in LP animals than in NP animals (Table 1). Looking for early profibrotic and inflammatory cytokine expression at day 70, we detected a significant induction in the mRNA expression of transforming growth factor-β1(TGF-β1) (3.68±0.56 fold induction in LP, P<0.05) and plasminogen activator inhibitor-1 (PAI-1) (2.54±0.58 fold induction in LP, P<0.05), as well as osteopontin (OPN) (2.58±0.50 fold induction in LP, P<0.05), monocyte chemoattractant protein 1 (MCP-1) (2.82±0.34 fold induction in LP, P<0.05) and tumor necrosis factor (TNF) (3.75±1.48 fold induction in LP, P<0.05) mRNA expression in LP. At day 70 (Figure 1, upper panels) minimal signs of glomerulosclerosis could be detected by semiquantitative scoring in LP (glomerulosclerosis index (GSI): 0.11±0.03 in NP versus 0.39±0.03 in LP; P<0.01). Scoring for tubulointerstitial lesions showed no differences at day 70 (data not shown). To exclude cell proliferation in the glomerulus, we analyzed the number of proliferating cell nuclear antigen (PCNA)-positive glomerular cells, which was similar in LP and NP (0.16±0.03 PCNA-positive cells/glomerulus in non-nephritic NP and 0.16±0.05 in non-nephritic LP). OPN and MCP-1 induction at day 70 was neither followed by an increased glomerular (0.85±0.11 cells/glomerulus in NP versus 0.77±0.06 cells/glomerulus in LP) nor by an increased interstitial (2.53±0.12 cells/interstitial cross-section in NP versus 2.76±2.20 cells/interstitial cross-section in LP) macrophage infiltration cells staining positive for mouse monoclonal anti-rat monocyte-like macrophage antibody (ED-1-positive cells) in LP animals. No differences in the deposition of glomerular collagen IV could be detected between LP and NP animals at day 70 (14.3±1.9% per glomerular cross-section in NP versus 17.8±2.0% in LP). The induction of mesangioproliferative GN was followed by proteinuria, microaneurysm formation, inflammation, macrophage infiltration, cell proliferation, mild glomerulosclerosis, and tubulointerstitial damage (Figure 1, middle panels). In response to anti-Thy-1.1 injection NP and LP animals developed proteinuria. Nevertheless, proteinuria was not different between LP and NP (Table 2). Endogenous creatinine clearance per 1000 glomeruli as surrogate for renal function was significantly lower in nephritic LP animals (Table 2). Mean arterial blood pressure was not increased in nephritic LP animals (Table 2) compared to NP.Table 2Proteinuria, creatinine clearance, mean arterial blood pressure, relative kidney weight, glomerular cell proliferation, macrophage infiltration, and stereological data of rats at days 4 and 14 of GNNP+GN d4LP+GN d4NP+GN d14LP+GN d14Urinary protein excretion (mg/24 h/100 g BWT)5.35±0.588.82±2.8520.18±6.9217.14±2.30Endogen creatinine clearance (ml/h/100 g BWT)85.36±4.8270.78±3.83*P<0.05 LP versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy).53.19±1.8963.23±1.90*P<0.05 LP versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy).Endogen creatinine clearance (mg/h/1000 glomeruli)7.69±0.8010.98±0.60*P<0.05 LP versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy).5.90±0.5910.25±7.48*P<0.05 LP versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy).Mean arterial blood pressure (mm Hg)100.3±4.65115.0±2.8599.2±7.02121.3±7.48Relative kidney weight (mg/g BWT)4.03±0.134.14±0.153.72±0.084.67±0.25*P<0.05 LP versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy).Glomerular cell proliferation (PCNA-positive cells/glomerular cross section)5.66±0.674.85±0.910.85±0.181.24±0.26Glomerular macrophage infiltration (ED-1-positive cells/glomerular cross-section)5.72±0.606.23±1.811.68±0.111.07±0.07*P<0.05 LP versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy).Interstitial macrophage infiltration/ED-1-positive cells/cross-section)4.31±0.674.68±1.943.26±0.842.86±0.34Number of glomeruli/kidney36 960±218323 400±1066*P<0.05 LP versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy).33 450±270522 380±1292*P<0.05 LP versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy).Total glomerular volume (mm3)37.77±3.7822.78±1.12*P<0.05 LP versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy).26.06±4.1830.37±1.41Mean glomerular volume (× 103 μm3)1016±71.98982±80.63807.1±138.11366±61.94*P<0.05 LP versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy).Glomerular filtration area per kidney (mm2)155.3±35.47165.7±12.3779.23±15.068.38±8.12BWT, body weight; d4, day 4; d14, day 14; GN, glomerulonephritis; LP, low protein; NP, normal protein; PCNA, proliferating cell nuclear antigen.Values are given as means ±s.e.m. LP animals were always compared to their corresponding NP group.* P<0.05 LP versus NP. LP, growth-retarded rats (mothers fed LP diet during pregnancy). NP, control rats (mothers fed NP diet during pregnancy). Open table in a new tab BWT, body weight; d4, day 4; d14, day 14; GN, glomerulonephritis; LP, low protein; NP, normal protein; PCNA, proliferating cell nuclear antigen. Values are given as means ±s.e.m. LP animals were always compared to their corresponding NP group. To quantify the extent of complement-mediated mesangiolysis after anti-Thy-1.1 injection, we scored microaneurysm formation. As expected, glomerular microaneurysms were detected in both LP and NP at day 4 of disease (see Figure 1, middle panels), being equally distributed in both groups (score for microaneurysms 0.77±0.09 in NP and 0.94±0.24 in LP animals, not significant). As an important feature in the course of Anti-Thy-1.1 nephritis, we analyzed the early proinflammatory cytokine pattern in renal cortex. The mRNA expression of MCP-1, the inflammatory cytokine interleukin-6, TNF, and OPN (Figure 2) in LP was not different to NP. As a consequence, glomerular and interstitial infiltration by macrophages measured by ED-1-positive staining was similar in LP and NP (Table 2). As a marker for glomerular cell proliferation we analyzed the number of PCNA-positive cells per glomerulus at day 4 of GN showing similar cell proliferation in NP and LP (Table 2). For the detection of early changes in the induction of fibrotic lesions, we analyzed the cortical mRNA expression of TGF-β1, PAI-1, and collagen I. At this early stage of GN expression levels were comparable in NP and LP (Figure 3). Nevertheless, glomerulosclerosis, measured by GSI and tubulointerstitial damage (score for tubulointerstitial lesions) were more severe in nephritic LP animals compared to the nephritic NP group (GSI: 0.12±0.03 in NP and 0.71±0.12 in LP, P<0.01, tubulointerstitial lesion score: 0.32±0.03 in NP and 0.75±0.13 in LP) (see Figure 1, middle panels). In this early phase of GN, glomerular collagen I and collagen IV deposition was not different between nephritic LP and NP animals (Figure 4a and b). Interstitial collagen I deposition was lower in NP compared to LP animals (Figure 4c).Figure 3Expression of profibrotic markers and collagen I in the renal cortex of rats at days 4 (d4) and 14 (d14) of GN. LP, growth-retarded rats (mothers fed LP diet during pregnancy). All values are normalized to the corresponding NP group (control rats with GN, mothers fed NP diet during pregnancy). Expression of (a) TGF-β1, (b) collagen I, and (c) PAI-1. *P<0.05 LP versus NP.View Large Image Figure ViewerDownload (PPT)Figure 4Glomerular and interstitial collagen deposition in rats at days 4 (d4) and 14 (d14) of GN. NP, control rats (mothers fed NP diet during pregnancy), LP, growth-retarded rats (mothers fed LP diet during pregnancy). Positive area for (a) glomerular collagen I per glomerular cross-section (%), (b) glomerular collagen IV per glomerular cross-section (%), (c) interstitial collagen I per glomerular cross-section. *P<0.05, **P<0.01.View Large Image Figure ViewerDownload (PPT) The total number of glomeruli was significantly lower in LP animals compared to NP animals on day 4 (Table 3). Total glomerular volume was reduced in nephritic LP animals, whereas the mean glomerular volume was similar in both groups. Glomerular filtration area per kidney was not different at day 4 of GN.Table 3Designed primer pairs and TaqMan probes used in this studyMCP-1Forward: 5′-CCTCCACCACTATGCAGGTCTC-3′Reverse: 5′-GCACGTGGATGCTACAGGC-3′Probe: 5′(FAM)-TCACGCTTCTGGGCCTGTTGTTCA-(TAMRA)3′OPNForward: 5′-AAAGTGGCTGAGTTTGGCAG-3′Reverse: 5′-AAGTGGCTACAGCATCTGAGTGT-3′Probe: 5′(FAM)-TCAGAGGAGAAGGCGCATTACAGCA- (TAMRA)3′PAI-1Forward: 5′-TCCGCCATCACCAACATTTT-3′Reverse: 5′-GTCAGTCATGCCCAGCTTCTC-3′Probe: 5′(FAM)-CCGCCTCCTCATCCTGCCTAAGTTCTCT- (TAMRA)3′IL-6Forward: 5′-TCCAAACTGGATATAACCAGGAAAT-3′Reverse: 5′-TTGTCTTTCTTGTTATCTTGTAAGTTGTTCTT-3′Probe: 5′(FAM)-AATCTGCTCTGGTCTTCTGGAGTTCGGTTTCTA- (TAMRA)3′TNFForward: 5′-GACCCTCACACTCAGATCATCTTCT-3′Reverse: 5′-TTGTCTTTGAGATCCATGCCATT-3′Probe: 5′(FAM)-ACGTCGTAGCAAACCACCAAGCGGA- (TAMRA)3′TGF-β1Forward: 5′-CACCCGCGTGCTAATGGT-3′Reverse: 5′-GGCACTGCTTCCCGAATG-3′Probe: 5′(FAM)-ACCGCAACAACGCAATCTATGACA-(TAMRA)3′GAPDHForward: 5′-ACGGGAAACCCATCACCAT-3′Reverse: 5′-CCAGCATCACCCCATTTGA-3′Probe: 5′(FAM)-TTCCAGGAGCGAGATCCCGTCAAG-TAMRA)3′PBGDForward: 5′-AGACCATGCAGGCCACCA-3′Reverse: 5′-CAACCAACTGTGGGTCATCCT-3′Probe: 5′(FAM)-AGGTCCCTGTTCAGCAAGAAGATGGTCC- (TAMRA)3′β-ActinForward: 5′-TGAGCTGCCTGACGGTCAG-3′Reverse: 5′-TGCCACAGGATTCCATACCC-3′Probe: 5′(FAM)-CACTATCGGCAATGAGCGGTTCCG-(TAMRA)3′Collagen-IaCollagen I was detected by SYBR Green analysis.Forward: 5′-GACGCAGAAGTCATAGGAGTC-3′Reverse: 5′-GGAAGTCCAGGCTGTCCA G-3′GAPDH, glyceraldehyde-3-phosphate dehydrogenase; IL-6, interleukin-6; MCP-1, monocyte chemoattractant protein 1; OPN, osteopontin; PAI-1, plasminogen activator inhibitor-1; PBGD, porphobilinogen-deaminase; TGF-β1, transforming growth factor-β1; TNF, tumor necrosis factor.a Collagen I was detected by SYBR Green analysis. Open table in a new tab GAPDH, glyceraldehyde-3-phosphate dehydrogenase; IL-6, interleukin-6; MCP-1, monocyte chemoattractant protein 1; OPN, osteopontin; PAI-1, plasminogen activator inhibitor-1; PBGD, porphobilinogen-deaminase; TGF-β1, transforming growth factor-β1; TNF, tumor necrosis factor. After 14 days of GN, urinary protein excretion was higher compared to the animals with GN at day 4. LP animals showed no higher protein excretion than NP animals (Table 2). Renal function according to endogenous creatinine clearance was slightly higher in LP than in NP animals at day 14 of GN (Table 2). Mean arterial blood pressure was slightly higher in LP animals than in NP animals without reaching statistical significance. Relative kidney weight was significantly higher in LP animals on day 14 of GN (Table 3). Glomeruli of both NP and LP were still hypercellular, showed matrix expansion and formation of extracapillary proliferates (Figure 1, lower panels). The number of microaneurysms had decreased in all animals compared to day 4. There was still a significant higher microaneurysm score index in nephritic LP animals (0.33±0.03 in LP versus 0.06±0.01 in NP, P<0.01), showing a slower reparation of microaneurysms. A higher mRNA expression of inflammatory mediators was detected in nephritic LP day 14 as compared to nephritic NP day 14. Expression of MCP-1 (Figure 2a) and OPN (Figure 2b) as well as expression of the inflammatory cytokines TNF (Figure 2c) and interleukin-6 (Figure 2d) was significantly higher at day 14 of disease. This prolongation of the inflammatory response in LP corresponded with a prolonged expression of fibrotic markers. Expression of the profibrotic cytokine TGF-β (Figure 3a), the PAI-1 (Figure 3c), and collagen I (Figure 3b) was significantly higher in LP than in NP. Assessment of the GSI revealed a significantly higher score in LP compared to NP (1.7±0.1 in LP versus 1.2±0.1 in NP, P<0.05). Moreover, LP rats tended to have a higher index for tubulointerstitial damage (1.2±0.04 in LP versus 0.6±1.7 in NP, P=0.056) and more extracapillary proliferates (6.9±1.4% of glomeruli affected in LP versus 2.9±1.7% in NP, P=0.15). The increased mRNA expression of collagen I resulted in an enhanced deposition of collagen I in the glomeruli of LP rats (Figure 4a). There was no difference of interstitial collagen I and glomerular collagen IV deposition in both groups (Figure 4c and b). The number of glomerular PCNA-positive cells was not different in LP and NP animals at day 14 of GN. Glomerular macrophage infiltration was significantly lower in LP animals (Table 2). No difference was found for interstitial macrophage infiltration at day 14 of GN. The total number of glomeruli was significantly lower in LP animals, total glomerular volume was similar in both groups, and mean glomerular volume in LP animals was significantly higher compared to NP animals at day 14 of GN (Table 2). Glomerular filtration area per kidney was not different in LP animals compared to NP animals on day 14 (Table 2). IUGR is discussed as an independent risk factor for the later development of metabolic, cardiovascular, and renal disease.21.Luyckx V.A. Brenner B.M. Low birth weight, nephron number, and kidney disease.Kidney Int Suppl. 2005; 97: S68-S77Abstract Full Text Full Text PDF PubMed Scopus (261) Google Scholar Until now only a few observational studies analyzed the course of glomerular diseases like idiopathic nephrotic syndrome and IgA nephropathy in children in relation to their birth weight.8.Zidar N. Avgustin Cavic M. Kenda R.B. et al.Unfavorable course of minimal change nephrotic syndrome in children with intrauterine growth retardation.Kidney Int. 1998; 54: 1320-1323Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar, 9.Sheu J.N. Chen J.H. Minimal change nephrotic syndrome in children with intrauterine growth retardation.Am J Kidney Dis. 2001; 37: 909-914Abstract Full Text PDF PubMed Scopus (36) Google Scholar, 10.Na Y.W. Yang H.J. Choi J.H. et al.Effect of intrauterine growth retardation on the progression of nephrotic syndrome.Am J Nephrol. 2002; 22: 463-467Crossref PubMed Scopus (25) Google Scholar, 11.Zidar N. Cavic M.A. Kenda R.B. et al.Effect of intrauterine growth retardation on the clinical course and prognosis of IgA glomerulonephritis in children.Nephron. 1998; 79: 28-32Crossref PubMed Scopus (79) Google Scholar We studied the course of acute anti-Thy-1.1 GN in rats with IUGR. As a model of IUGR, we chose maternal protein restriction during gestation. In contrast to other IUGR models like bilateral uterine artery ligation, no disturbed glucose tolerance or development of diabetes in young animals can be detected.12.Bertram C.E. Hanson M.A. Animal models and programming of the metabolic syndrome.Br Med Bull. 2001; 60: 103-121Crossref PubMed Scopus (223) Google Scholar Because placenta insufficiency is an important cause for IUGR in humans, bilateral uterine artery ligation was considered to more closely resemble human IUGR in Western countries.22.Fernandez-Twinn D.S. Ozanne S.E. Ekizoglou S. et al.The maternal endocrine environment in the low-protein model of intra-uterine growth restriction.Br J Nutr. 2003; 90: 815-822Crossref PubMed Scopus (130) Google Scholar Protein malnutrition of dams may be seen as an equivalent for maternal protein malnutrition in developing countries.13.Vehaskari V.M. Woods L.L. Prenatal programming of hypertension: lessons from experimental models.J Am Soc Nephrol. 2005; 16: 2545-2556Crossref PubMed Scopus (96) Google Schol
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