Nephron number, hypertension, renal disease, and renal failure

Essential hypertension is one of the most common diseases in the Western world, affecting about 26.4% of the adult population, and it is increasing (1). Its causes are heterogeneous and include genetic and environmental factors (2), but several observations point to an important role of the kidney in its genesis (3). In addition to variations in tubular transport mechanisms that could, for example, affect salt handling, structural characteristics of the kidney might also contribute to hypertension. The burden of chronic kidney disease is also increasing worldwide, due to population growth, increasing longevity, and changing risk factors. Although single-cause models of disease are still widely promoted, multideterminant or multihit models that can accommodate multiple risk factors in an individual or in a population are probably more applicable (4,5). In such a framework, nephron endowment is one potential determinant of disease susceptibility. Some time ago, Brenner and colleagues (6,7) proposed that lower nephron numbers predispose both to essential hypertension and to renal disease. They also proposed that hypertension and progressive renal insufficiency might be initiated and accelerated by glomerular hypertrophy and intraglomerular hypertension that develops as nephron number is reduced (8). In this review, we summarize data from recent studies that shed more light on these hypotheses. The data supply a new twist to possible mechanisms of the Barker hypothesis, which proposes that intrauterine growth retardation predisposes to chronic disease in later life (9). The review describes how nephron number is estimated and its range and some determinants and morphologic correlates. It then considers possible causes of low nephron numbers. Finally, associations of hypertension and renal disease with reduced nephron numbers are considered, and some potential clinical implications are discussed.

Does nephron number matter in the development of kidney disease?

The total number of nephrons in normal human kidneys varies over a 10-fold range. This variation in total nephron number leads us to question whether low nephron number increases the risk of renal disease in adulthood. This review considers the available evidence in humans linking low nephron number/reduced nephron endowment and the susceptibility to renal disease. Total nephron number in humans has been directly correlated with birth weight and inversely correlated with age, mean glomerular volume, and hypertension. Low nephron number may be the result of suboptimal nephrogenesis during kidney development and/or loss of nephrons once nephrogenesis has been completed. Low nephron number is frequently, but not always, associated with hypertrophy of remaining glomeruli. This compensatory hypertrophy has also been associated with a greater susceptibility for kidney disease. Three human studies have reported reduced nelphron number in subjects with a history of hypertension. This correlation has been observed in White Europeans, White Americans (but not African Americans) and Australian Aborigines. Studies in additional populations are required, as well as a greater understanding of the fetal environmental and genetic determinants of low nephron number.

Reduced nephron number and glomerulomegaly in Australian Aborigines: A group at high risk for renal disease and hypertension

Aborigines in remote areas of Australia have much higher rates of renal disease, as well as hypertension and cardiovascular disease, than non-Aboriginal Australians. We compared kidney findings in Aboriginal and non-Aboriginal people in one remote region. Glomerular number and mean glomerular volume were estimated with the disector/fractionator combination in the right kidney of 19 Aborigines and 24 non-Aboriginal people undergoing forensic autopsy for sudden or unexpected death in the Top End of the Northern Territory. Aborigines had 30% fewer glomeruli than non-Aborigines-202000 fewer glomeruli per kidney, or an estimated 404000 fewer per person (P=0.036). Their mean glomerular volume was 27% larger (P=0.016). Glomerular number was significantly correlated with adult height, inferring a relationship with birthweight, which, on average, is much lower in Aboriginal than non-Aboriginal people. Aboriginal people with a history of hypertension had 30% fewer glomeruli than those without-250000 fewer per kidney (P=0.03), or 500000 fewer per person, and their mean glomerular volume was about 25% larger. The lower nephron number in Aboriginal people is compatible with their susceptibility to renal failure. The additional nephron deficit associated with hypertension is compatible with other reports. Lower nephron numbers are probably due in part to reduced nephron endowment, which is related to a suboptimal intrauterine environment. Compensatory glomerular hypertrophy in people with fewer nephrons, while minimizing loss of total filtering surface area, might be exacerbating nephron loss. Optimization of fetal growth should ultimately reduce the florid epidemic of renal disease, hypertension, and cardiovascular disease.

Hypertension, glomerular number, and birth weight in African Americans and white subjects in the southeastern United States

Low nephron number has been related to low birth weight and hypertension. In the southeastern United States, the estimated prevalence of chronic kidney disease due to hypertension is five times greater for African Americans than white subjects. This study investigates the relationships between total glomerular number (N-glom), blood pressure, and birth weight in southeastern African Americans and white subjects. Stereological estimates of N-glom were obtained using the physical disector/fractionator technique on autopsy kidneys from 62 African American and 60 white subjects 30-65 years of age. By medical history and recorded blood pressures, 41 African Americans, and 24 white subjects were identified as hypertensive and 21 African Americans and 36 white subjects as normotensive. Mean arterial blood pressure ( MAP) was obtained on 81 and birth weights on 63 subjects. For African Americans, relationships between MAP, N-glom, and birth weight were not significant. For white subjects, they were as follows: MAP and N-glom ( r = -0.4551, P = 0.0047); Nglom and birth weight ( r = 0.5730, P = 0.0022); MAP and birth weight ( r = -0.4228, P = 0.0377). For African Americans, average N-glom of 961 840 +/- 292 750 for normotensive and 867 358 +/- 341 958 for hypertensive patients were not significantly different ( P = 0.285). For white subjects, average N-glom of 923 377 +/- 256 391 for normotensive and 754 319 +/- 329 506 for hypertensive patients were significantly different ( P = 0.03). The data indicate that low nephron number and possibly low birth weight may play a role in the development of hypertension in white subjects but not African Americans.

Le diabète maternel influence la morphogenèse rénale et la programmation périnatale

Le diabète maternel est un facteur de risque majeur pour le développement de malformations congénitales. Dans le syndrome de l’embryopathie diabétique, l’exposition prolongée du fœtus à de hautes concentrations ambientes de glucose induit des dommages qui peuvent affecter plusieurs organes, dont les reins. Les malformations rénales sont la cause de près de 40 pourcent des cas d’insuffisance rénale infantile. L’hyperglycémie constitue un environnement utérin adverse qui nuit à la néphrogenèse et peut causer l’agenèse, la dysplasie (aplasie) ou l’hypoplasie rénale. Les mécanismes moléculaires par lesquels les hautes concentrations ambientes de glucose mènent à la dysmorphogenèse et aux malformations demeurent toutefois mal définis. Le diabète maternel prédispose aussi la progéniture au développement d’autres problèmes à l’âge adulte, tels l’hypertension, l’obésité et le diabète de type 2. Ce phénomène appelé ‘programmation périnatale’ a suscité l’intérêt au cours des dernières décennies, mais les mécanismes responsables demeurent mal compris. Mes études doctorales visaient à élucider les mécanismes moléculaires par lesquels le diabète maternel ou un environnement in utero hyperglycémique affecte la néphrogenèse et programme par la suite la progéniture a développer de l’hypertension par des observations in vitro, ex vivo et in vivo. Nous avons utilisé les cellules MK4, des cellules embryonnaires du mésenchyme métanéphrique de souris, pour nos études in vitro et deux lignées de souris transgéniques (Tg) pour nos études ex vivo et in vivo, soient les souris HoxB7-GFP-Tg et Nephrin-CFP-Tg. Les souris HoxB7-GFP-Tg expriment la protéine fluorescente verte (GFP) dans le bourgeon urétérique (UB), sous le contrôle du promoteur HoxB7. Les souris Nephrin-CFP expriment la protéine fluorescente cyan (CFP) dans les glomérules, sous le contrôle du promoteur nephrin spécifique aux podocytes. Nos études in vitro visaient à déterminer si les hautes concentrations de glucose modulent l’expression du gène Pax2 dans les cellules MK4. Les cellules MK4 ont été traitées pendant 24h avec du milieu contenant soit 5mM D-glucose et 20mM D-mannitol ou 25mM D-glucose et avec ou sans antioxydants ou inhibiteurs de p38 MAPK, p44/42 MAPK, PKC et NF-kB. Nos résultats ont démontré que le D-glucose élevé (25mM) augmente la génération des espèces réactives de l’oxygène (ROS) dans les cellules MK4 et induit spécifiquement l’expression du gène Pax2. Des analogues du glucose tels le D-mannitol, L-glucose ou le 2-Deoxy-D-glucose n’induisent pas cette augmentation dans les cellules MK4. La stimulation de l’expression du gène Pax2 par le D-glucose dans les cellules MK4 peut être bloquée par des inhibiteurs des ROS et de NF-kB, mais pas par des inhibiteurs de p38 MAPK, p44/42 MAPK ou PKC. Ces résultats indiquent que la stimulation de l’expression du gène Pax2 par les concentrations élevées de glucose est due, au moins en partie, à la génération des ROS et l’activation de la voie de signalisation NF-kB, et non pas via les voies PKC, p38 MAPK et p44/42 MAPK. Nos études ex vivo s’intéressaient aux effets d’un milieu hyperglycémique sur la morphogenèse de la ramification du bourgeon urétérique (UB). Des explants de reins embryonnaires (E12 à E18) ont été prélevés par micro-dissection de femelles HoxB7-GFP gestantes. Les explants ont ensuite été cultivés dans un milieu contenant soit 5mM D-glucose et 20mM D-mannitol ou 25mM D-glucose et avec ou sans antioxydants, catalase ou inhibiteur de PI3K/AKT pour diverses durées. Nos résultats ont démontré que le D-glucose stimule la ramification du UB de manière spécifique, et ce via l’expression du gène Pax2. Cette augmentation de la ramification et de l’expression du gène Pax2 peut être bloquée par des inhibiteurs des ROS et de PI3K/AKT. Ces études ont démontré que les hautes concentrations de glucose altèrent la morphogenèse de la ramification du UB via l’expression de Pax2. L’effet stimulant du glucose semble s’effectuer via la génération des ROS et l’activation de la voie de signalisation Akt. Nos études in vivo visaient à déterminer le rôle fondamental du diabète maternel sur les défauts de morphogenèse rénale chez la progéniture. Dans notre modèle animal, le diabète maternel est induit par le streptozotocin (STZ) chez des femelles HoxB7-GFP gestantes (E13). Les souriceaux ont été étudiés à différents âges (naissants et âgés de une, deux ou trois semaines). Nous avons examiné leurs morphologie rénale, nombre de néphrons, expression génique et les événements apoptotiques lors de cette étude à court terme. La progéniture des mères diabétiques avait un plus faible poids, taille et poids des reins, et possédait des glomérules plus petits et moins de néphrons par rapport à la progéniture des mères contrôles. La dysmorphogenèse rénale observée est peut-être causée par l’augmentation de l’apoptose des cellules dans la région du glomérule. Nos résultats ont montré que les souriceaux nés de mères diabétiques possèdent plus de podocytes apoptotiques et plus de marquage contre la caspase-3 active dans leurs tubules rénaux que la progéniture des mères contrôles. Les souriceaux des mères diabétiques montrent une augmentation de l’expression des composants du système rénine angiotensine (RAS) intrarénal comme l’angiotensinogène et la rénine, ainsi qu’une augmentation des isoformes p50 et p65 de NF-kB. Ces résultats indiquent que le diabète maternel active le RAS intrarénal et induit l’apoptose des glomérules, menant à une altération de la morphogenèse rénale de la progéniture. En conclusion, nos études ont permis de démontrer que le glucose élevé ou l’environnement in utero diabétique altère la morphogenèse du UB, qui résulte en un retard dans la néphrogenèse et produit des reins plus petits. Cet effet est dû, au moins en partie, à la génération des ROS, à l’activation du RAS intrarénal et à la voie NF-kB. Nos études futures se concentreront sur les mécanismes par lesquels le diabète maternel induit la programmation périnatale de l’hypertension chez la progéniture adulte. Cette étude à long terme porte sur trois types de progénitures : adultes nés de mères contrôles, diabétiques ou diabétiques traitées avec insuline pendant la gestation. Nous observerons la pression systolique, la morphologie rénale et l’expression de divers gènes et protéines. Nous voulons de plus déterminer si la présence d’un système antioxydant (catalase) peut protéger la progéniture des effets néfastes des ROS causés par l’environnement in utero hyperglycémique. Les souris Catalase-Tg expriment la catalase spécifiquement dans les tubules proximaux et nous permettrons d’explorer notre hypothèse sur le rôle des ROS dans notre modèle expérimental de diabète maternel.

Perfil de expressão dos mIRNAS da família mIR-200 e mIR-192 no rim total e glomérulos isolados de ratos submetidos à restrição protéica in útero

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Efeitos da sobrecarga calórica em ratos machos submetidos ou não a restrição protéica in útero

Pós-graduação em Biologia Geral e Aplicada - IBB

Involvement of Renal Corpuscle microRNA Expression on Epithelial-to-Mesenchymal Transition in Maternal Low Protein Diet in Adult Programmed Rats

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Mutations in Uroplakin IIIA are a rare cause of renal hypodysplasia in humans

BACKGROUND: Renal hypodysplasia, characterized by a decrease in nephron number, small overall kidney size, and maldeveloped renal tissue, is a leading cause of chronic renal failure in young children. Familial clustering and renal hypodysplasia phenotypes observed in transgenic animal models suggest a genetic contribution. Uroplakin IIIa (encoded by UPIIIA) is an integral membrane protein present in urothelial plaques, and the murine UPIIIa knockout is associated with urothelial anomalies and vesicoureteral reflux. De novo UPIIIA mutations recently were identified in 4 of 17 patients with severe bilateral renal adysplasia. METHODS: To evaluate the overall role of UPIIIA in human renal hypodysplasia pathogenesis, we performed UPIIIA mutation analysis in a cohort of 170 pediatric patients affected by severe unilateral or bilateral renal hypodysplasia. Eighty-one patients were affected by bilateral nonobstructive renal hypodysplasia; of these, 61 were without vesicoureteral reflux. Eighty-four patients presented with unilateral nonobstructive renal hypodysplasia, including 24 patients with unilateral multicystic dysplastic kidneys. Family history was positive in 11%. RESULTS: Mutation analysis showed 2 heterozygous mutations not observed in 200 race-matched control chromosomes. In only 1 family was distribution of the UPIIIA mutation consistent with a disease-causing effect. This de novo missense mutation (Gly202Asp) was identified in a patient with unilateral multicystic dysplastic kidneys. The second (intronically located) mutation appeared unlikely to be disease causing because it did not segregate with an obvious disease phenotype in the affected family. CONCLUSION: Our results indicate that de novo mutations in UPIIIA can be involved in defective early kidney development, but probably constitute only a rare cause of human renal hypodysplasia in a minor subset of individuals.

Glomerular and renal vascular structural changes in alpha8 integrin-deficient mice

Integrins are matrix receptors that regulate cell-matrix interactions during development and in adult tissue. In the adult kidney, the alpha8 chain is specifically expressed in glomerular mesangial cells and vascular smooth muscle cells. alpha8-deficient (alpha8-/-) mice demonstrate reductions in renal mass, which can range from complete renal agenesis to the development of kidneys that are only slightly smaller than wild-type kidneys. No histologic abnormalities of these kidneys have been described. However, considering the prominent expression of alpha8 in glomeruli and renal vessels, it seemed unlikely that the kidneys of alpha8-/- mice would be completely normal. Therefore, the renal phenotype of adult alpha8-/- mice was investigated, for assessment of more subtle morphologic alterations in kidney tissue. alpha8-/- mice displayed a significant reduction in nephron number and an increase in glomerular volume, compared with wild-type control animals. Albuminuria was not different in wild-type and alpha8-/- mice. Quantitative morphologic analyses revealed that the glomeruli of alpha8-/- mice were hypercellular, with an increased number of mesangial cells, compared with wild-type mice. Mesangial matrix deposition (as demonstrated for collagen IV and the alpha8 ligand fibronectin) was expanded in alpha8-/- mice, compared with wild-type mice. Collagens I and III, which are not normally present in glomeruli, were detected in the glomeruli of alpha8-/- mice. Staining for other glomerular integrins demonstrated an increased abundance of the collagen receptor alpha2 integrin in alpha8-/- mice. The glomerular capillary length density was significantly greater in alpha8-/- mice than in wild-type mice. Cortical arterial vessel walls were not altered in alpha8-/- mice, but the capillaries of the peritubular network were widened. Despite the strong mesangial and vascular expression of alpha8, glomerular and renal vascular alterations in alpha8-/- mice were relatively mild. Only aged alpha8-/- mice demonstrated increased glomerular capillary widening, compared with control animals. The results suggest that the lack of alpha8 can be largely compensated for, at least in younger alpha8-/- mice. It is not yet clear whether the occurrence of collagens that are not normally present in glomeruli and the increased abundance of the collagen receptor alpha2 contribute to maintaining the glomerular structure in alpha8-/- mice. The compensatory mechanisms involved will be the subject of future research.

Lessons in ethnonephrology

Only recently has the nephrology community moved beyond a fairly singular focus on terminal kidney failure to embrace population-based studies of earlier stages of disease, its markers and risk factors, and of interventions. Observations in developing countries, and in minority, migrant, and disadvantaged groups in westernized countries, have promoted these developments. We are only beginning to interpret renal disease in the context of public health history, social and health transitions, changing population demography, and competing mortality. Its intimate relationships to other health issues are being progressively exposed. Perspectives on the multideterminant etiology of most disease and the pedestrian nature of most risk factors are maturing. We are challenged to reconcile epidemiologic patterns with morphology in diseased renal tissue, and to consider structural markers, such as nephron number and glomerular size, as determinants of disease susceptibility. New work force models are mandated for population-based studies and intervention programs. Intervention programs need to be integrated with other chronic disease initiatives and nested in a matrix of systematic primary care, and although flexible to changing needs, must be sustained over the long term. Cross-disciplinary collaboration is essential in designing those programs, and in promoting them to health-care funders. Substantial expansion and restructuring of the discipline is needed for the nephrology community to participate effectively in those processes.

Low protein diet fed exclusively during mouse oocyte maturation leads to behavioural and cardiovascular abnormalities in offspring

Early embryonic development is known to be susceptible to maternal undernutrition, leading to a disease-related postnatal phenotype. To determine whether this sensitivity extended into oocyte development, we examined the effect of maternal normal protein diet (18% casein; NPD) or isocaloric low protein diet (9% casein; LPD) restricted to one ovulatory cycle (3.5 days) prior to natural mating in female MF-1 mice. After mating, all females received NPD for the remainder of gestation and all offspring were litter size adjusted and fed standard chow. No difference in gestation length, litter size, sex ratio or postnatal growth was observed between treatments. Maternal LPD did, however, induce abnormal anxiety-related behaviour in open field activities in male and female offspring (P <0.05). Maternal LPD offspring also exhibited elevated systolic blood pressure (SBP) in males at 9 and 15 weeks and in both sexes at 21 weeks (P <0.05). Male LPD offspring hypertension was accompanied by attenuated arterial responsiveness in vitro to vasodilators acetylcholine and isoprenaline (P <0.05). LPD female offspring adult kidneys were also smaller, but had increased nephron numbers (P <0.05). Moreover, the relationship between SBP and kidney or heart size or nephron number was altered by diet treatment (P <0.05). These data demonstrate the sensitivity of mouse maturing oocytes in vivo to maternal protein undernutrition and identify both behavioural and cardiovascular postnatal outcomes, indicative of adult disease. These outcomes probably derive from a direct effect of protein restriction, although indirect stress mechanisms may also be contributory. Similar and distinct postnatal outcomes were observed here compared with maternal LPD treatment during post-fertilization preimplantation development which may reflect the relative contribution of the paternal genome. © Journal compilation © 2008 The Physiological Society.

Genetic Abrogation of Adenosine A3 Receptor Prevents Uninephrectomy and High Salt-Induced Hypertension.

BACKGROUND: Early-life reduction in nephron number (uninephrectomy [UNX]) and chronic high salt (HS) intake increase the risk of hypertension and chronic kidney disease. Adenosine signaling via its different receptors has been implicated in modulating renal, cardiovascular, and metabolic functions as well as inflammatory processes; however, the specific role of the A3 receptor in cardiovascular diseases is not clear. In this study, gene-modified mice were used to investigate the hypothesis that lack of A3 signaling prevents the development of hypertension and attenuates renal and cardiovascular injuries following UNX in combination with HS (UNX-HS) in mice. METHODS AND RESULTS: Wild-type (A3 (+/+)) mice subjected to UNX-HS developed hypertension compared with controls (mean arterial pressure 106±3 versus 82±3 mm Hg; P<0.05) and displayed an impaired metabolic phenotype (eg, increased adiposity, reduced glucose tolerance, hyperinsulinemia). These changes were associated with both cardiac hypertrophy and fibrosis together with renal injuries and proteinuria. All of these pathological hallmarks were significantly attenuated in the A3 (-/-) mice. Mechanistically, absence of A3 receptors protected from UNX-HS-associated increase in renal NADPH oxidase activity and Nox2 expression. In addition, circulating cytokines including interleukins 1β, 6, 12, and 10 were increased in A3 (+/+) following UNX-HS, but these cytokines were already elevated in naïve A3 (-/-) mice and did not change following UNX-HS. CONCLUSIONS: Reduction in nephron number combined with chronic HS intake is associated with oxidative stress, chronic inflammation, and development of hypertension in mice. Absence of adenosine A3 receptor signaling was strongly protective in this novel mouse model of renal and cardiovascular disease.

Glomerular number and size in autopsy kidneys: The relationship to birth weight

Background. In the Southeast United States, African Americans have an estimated incidence of hypertension and end-stage renal disease (ESRD) that is five times greater than Caucasians. Higher rates of low birth weight (LBW) among African Americans is suggested to predispose African Americans to the higher risk, possibly by reducing the number of glomeruli that develop in the kidney. This study investigates the relationships between age, race, gender, total glomerular number (N-glom), mean glomerular volume (V-glom), body surface area (BSA), and birth weight. Methods. Stereologic estimates of N-glom and V-glom were obtained using the physical disector/fractionator combination for autopsy kidneys from 37 African Americans and 19 Caucasians. Results. N-glom was normally distributed and ranged from 227,327 to 1,825,380, an 8.0-fold difference. A direct linear relationship was observed between N-glom and birth weight (r=0.423, P=0.0012) with a regression coefficient that predicted an increase of 257,426 glomeruli per kilogram increase in birth weight (alpha=0.050:0.908). Among adults there was a 4.9-fold range in V-glom , and in adults, V-glom was strongly and inversely correlated with N-glom (r=-0.640, P=0.000002). Adult V-glom showed no significant correlation with BSA for males (r=-0.0150, P=0.936), although it did for females (r=0.606, P=0.022). No racial differences in average N-glom or V-glom were observed. Conclusion. Birth weight is a strong determinant of N-glom and thereby of glomerular size in the postnatal kidney. The findings support the hypothesis that LBW by impairing nephron development is a risk factor for hypertension and ESRD in adulthood.