Lina Gubhaju

Accelerated Maturation and Abnormal Morphology in the Preterm Neonatal Kidney

Nephrogenesis is ongoing at the time of birth for the majority of preterm infants, but whether postnatal renal development follows a similar trajectory to normal in utero growth is unknown. Here, we examined tissue collected at autopsy from 28 kidneys from preterm neonates, whose postnatal survival ranged from 2 to 68 days, including 6 that had restricted intrauterine growth. In addition, we examined kidneys from 32 still-born gestational controls. We assessed the width of the nephrogenic zone, number of glomerular generations, cross-sectional area of the renal corpuscle, and glomerular maturity and morphology. Renal maturation accelerated after preterm birth, with an increased number of glomerular generations and a decreased width of the nephrogenic zone in the kidneys of preterm neonates. Of particular concern, compared with gestational controls, preterm kidneys had a greater percentage of morphologically abnormal glomeruli and a significantly larger cross-sectional area of the renal corpuscle, suggestive of renal hyperfiltration. These observations suggest that the preterm kidney may have fewer functional nephrons, thereby increasing vulnerability to impaired renal function in both the early postnatal period and later in life.

Is nephrogenesis affected by preterm birth? Studies in a non-human primate model

Nephrogenesis occurs predominantly in late gestation at a time when preterm infants are already delivered. The aims of this study were to assess the effect of preterm birth and the effect of antenatal glucocorticoid treatment on nephrogenesis. Preterm baboons, which were delivered at 125 days gestation and ventilated for up to 21 days postnatally, were compared with gestational controls. A cohort of preterm baboons that had been exposed to antenatal glucocorticoids were compared with unexposed preterm baboons. The number of glomerular generations was estimated using a medullary ray glomerular-counting method, and glomerular number was estimated using unbiased stereology. CD31 and WT-1 localization was examined using immunohistochemistry and VEGF was localized using in situ hybridization. The number of glomerular generations was not affected by preterm birth, and total glomerular numbers were within the normal range. Kidneys were significantly enlarged in preterm baboons with a significant decrease in glomerular density (number of glomeruli per gram of kidney) in the preterm kidney compared with gestational controls. Neonates exposed to antenatal steroids had an increased kidney-to-body weight ratio and also more developed glomeruli compared with unexposed controls. Abnormal glomeruli, with a cystic Bowmans space and shrunken glomerular tuft, were often present in the superficial renal cortex of both the steroid-exposed and unexposed preterm kidneys; steroid exposure had no significant effect on the proportion of abnormal glomeruli. The proportion of abnormal glomeruli in the preterm kidneys ranged from 0.2 to 18%. In conclusion, although nephrogenesis is ongoing in the extrauterine environment, our findings demonstrate that preterm birth, independent of steroid exposure, is associated with a high proportion of abnormal glomeruli in some, but not all neonatal kidneys. Whether final nephron endowment is affected in those kidneys exhibiting a high proportion of abnormal glomeruli is yet to be confirmed.

Measuring glomerular number and size in perfused kidneys using MRI

The goal of this work was to nondestructively measure glomerular (and thereby nephron) number in the whole kidney. Variations in the number and size of glomeruli have been linked to many renal and systemic diseases. Here, we develop a robust magnetic resonance imaging (MRI) technique based on injection of cationic ferritin (CF) to produce an accurate measurement of number and size of individual glomeruli. High-field (19 Tesla) gradient-echo MR images of perfused rat kidneys after in vivo intravenous injection of CF showed specific labeling of individual glomeruli with CF throughout the kidney. We developed a three-dimensional image-processing algorithm to count every labeled glomerulus. MRI-based counts yielded 33,786 ± 3,753 labeled glomeruli (n = 5 kidneys). Acid maceration counting of contralateral kidneys yielded an estimate of 30,585 ± 2,053 glomeruli (n = 6 kidneys). Disector/fractionator stereology counting yielded an estimate of 34,963 glomeruli (n = 2). MRI-based measurement of apparent glomerular volume of labeled glomeruli was 4.89 × 10(-4) mm(3) (n = 5) compared with the average stereological measurement of 4.99 × 10(-4) mm(3) (n = 2). The MRI-based technique also yielded the intrarenal distribution of apparent glomerular volume, a measurement previously unobtainable in histology. This work makes it possible to nondestructively measure whole-kidney glomerular number and apparent glomerular volumes to study susceptibility to renal diseases and opens the door to similar in vivo measurements in animals and humans.

Quantification of glomerular number and size distribution in normal rat kidneys using magnetic resonance imaging

BACKGROUND Glomerular number and size are important risk factors for chronic kidney disease (CKD) and cardiovascular disease and have traditionally been estimated using invasive techniques. Here, we report a novel technique to count and size every glomerulus in the rat kidney using magnetic resonance imaging (MRI). METHODS The ferromagnetic nature of cationized ferritin allowed visualization of single glomeruli in high-resolution susceptibility-weighted MRI. A segmentation algorithm was used to identify and count all glomeruli within the whole kidney. To prove our concept, we estimated total glomerular number and mean glomerular volume of each kidney using design-based stereology. RESULTS The glomerular counts obtained with MRI agreed well with estimates obtained using traditional methods [MRI, 32 785 (3117); stereology, 35 132 (3123)]. For the first time, the glomerular volume distribution for the entire kidney is shown. Additionally, the method is substantially faster than the current methods. CONCLUSIONS MRI provides a new method for measuring these important microanatomical markers of disease risk and leads the way to in vivo analysis of these parameters, including longitudinal studies of animal models of CKD.

The baboon as a good model for studies of human kidney development.

Because of the improved survival of premature neonates in recent years, it is important to investigate the effects of premature delivery on the kidney, in which nephrogenesis is still ongoing during the third trimester. Hence, an appropriate animal model that is similar to humans is essential. The aim of the current study is to determine the time course of nephrogenesis in the baboon, to establish whether it is a suitable model of human nephrogenesis. At the Southwest Foundation for Biomedical Research (San Antonio, TX), fetal baboons were delivered prematurely by cesarean delivery and at term by natural delivery. Fixed kidneys from 125-, 140-, 175-, and 185-d gestation baboons were assessed morphologically for evidence of a nephrogenic zone. Nephron number, kidney volume, and glomerular and corpuscle volume were also estimated using unbiased stereology. Morphologic assessment confirmed the presence of metanephric mesenchyme and immature glomeruli in the nephrogenic zone of the kidneys from the prematurely delivered fetuses at 125 and 140 d gestation. At 175 d gestation and at term, the nephrons seemed to be mature. Both kidney weight (R2 = 0.918, p = 0.0002) and kidney volume (R2 = 0.837, p = 0.001) were very strongly correlated with nephron number. There was also a direct relationship between gestational age (R2 = 0.589, p = 0.03) and birth weight (R2 = 0.562, p = 0.03) with nephron number. In conclusion, in this study, nephrogenesis in the baboon is complete before term by 175 d gestation, which is similar to humans. Hence, the baboon is a suitable model for future studies to investigate human kidney development.

Preterm Birth and the Kidney: Implications for Long-Term Renal Health:

Although the majority of preterm neonates now survive infancy, there is emerging epidemiological evidence to demonstrate that individuals born preterm exhibit an elevated risk for the development of hypertension and renal impairment later in life, thus supporting the developmental origins of health and disease hypothesis. The increased risk may potentially be attributed to a negative impact of preterm birth on nephron endowment. Indeed, at the time when most preterm neonates are delivered, nephrogenesis in the kidney is still ongoing with the majority of nephrons normally formed during the third trimester of pregnancy. A number of clinical studies have provided evidence of altered renal function during the neonatal period, but to date there have been limited studies describing the consequences of preterm birth on kidney structure. Importantly, studies in the preterm baboon have shown that nephrogenesis is clearly ongoing following preterm birth; however, the presence of abnormal glomeruli (up to 18% in some cases) is of concern. Similar glomerular abnormalities have been described in autopsied preterm infants. Prenatal and postnatal factors such as exposure to certain medications, hyperoxia and intrauterine and/or extrauterine growth restriction are likely to have a significant influence on nephrogenesis and final nephron endowment. Further studies are required to determine the factors contributing to renal maldevelopment and to identify potential interventional strategies to maximize nephron endowment at the start of life, thereby optimizing long-term renal health for preterm individuals.

Effects of ibuprofen treatment on the developing preterm baboon kidney

Preterm neonates are commonly exposed postnatally to pharmacological treatments for a patent ductus arteriosus. Exposure of the developing kidney to nephrotoxic medications may adversely impact renal development. This study aimed to determine the effect of early postnatal ibuprofen treatment, both alone and in combination with a nitric oxide synthase inhibitor (NOSi), on renal development and morphology. Baboon neonates were delivered prematurely at 125-day (125d) gestation (term = 185d) and were euthanized at birth or postnatal day 6. Neonates were divided into four groups: 125d gestational controls (n = 8), Untreated (n = 8), Ibuprofen (n = 6), and ibuprofen (Ibu)+NOSi (n = 4). Animals in the Ibuprofen and Ibu+NOSi groups received five doses of ibuprofen, with the Ibuprofen+NOSi animals additionally administered a NOS inhibitor (N(G)-monomethyl-l-arginine). There was no difference among groups in body weight, kidney weight, or glomerular generation number. Nephrogenic zone width was significantly reduced in the Ibuprofen group (123.5 ± 7.4 μm) compared with the 125d gestational control (176.1 ± 6.9 μm) and Untreated animals (169.7 ± 78.8 μm). In the Ibu+NOSi group, nephrogenic zone width averaged 152.7 ± 3.9 μm, which was not significantly different from any other group. Morphologically abnormal glomeruli were present at a range of 0.0-22.9% in the Untreated group, 0.0-6.1% in the Ibuprofen group, and 0.0-1.4% in the Ibu+NOSi group. In conclusion, early postnatal ibuprofen exposure is associated with a reduced nephrogenic zone width, which may suggest the early cessation of nephrogenesis following treatment. Ultimately, this may impact the number of nephrons formed in the preterm kidney.

Low Birth Weight due to Intrauterine Growth Restriction and/or Preterm Birth: Effects on Nephron Number and Long-Term Renal Health

Epidemiological studies have clearly demonstrated a strong association between low birth weight and long-term renal disease. A potential mediator of this long-term risk is a reduction in nephron endowment in the low birth weight infant at the beginning of life. Importantly, nephrons are only formed early in life; during normal gestation, nephrogenesis is complete by about 32–36 weeks, with no new nephrons formed after this time during the lifetime of the individual. Hence, given that a loss of a critical number of nephrons is the hallmark of renal disease, an increased severity and acceleration of renal disease is likely when the number of nephrons is already reduced prior to disease onset. Low birth weight can result from intrauterine growth restriction (IUGR) or preterm birth; a high proportion of babies born prematurely also exhibit IUGR. In this paper, we describe how IUGR and preterm birth adversely impact on nephrogenesis and how a subsequent reduced nephron endowment at the beginning of life may lead to long-term risk of renal disease, but not necessarily hypertension.

Assessment of renal functional maturation and injury in preterm neonates during the first month of life

Worldwide, approximately 10% of neonates are born preterm. The majority of preterm neonates are born when the kidneys are still developing; therefore, during the early postnatal period renal function is likely reflective of renal immaturity and/or injury. This study evaluated glomerular and tubular function and urinary neutrophil gelatinase-associated lipocalin (NGAL; a marker of renal injury) in preterm neonates during the first month of life. Preterm and term infants were recruited from Monash Newborn (neonatal intensive care unit at Monash Medical Centre) and Jesse McPherson Private Hospital, respectively. Infants were grouped according to gestational age at birth: ≤28 wk (n = 33), 29-31 wk (n = 44), 32-36 wk (n = 32), and term (≥37 wk (n = 22)). Measures of glomerular and tubular function were assessed on postnatal days 3-7, 14, 21, and 28. Glomerular and tubular function was significantly affected by gestational age at birth, as well as by postnatal age. By postnatal day 28, creatinine clearance remained significantly lower among preterm neonates compared with term infants; however, sodium excretion was not significantly different. Pathological proteinuria and high urinary NGAL levels were observed in a number of neonates, which may be indicative of renal injury; however, there was no correlation between the two markers. Findings suggest that neonatal renal function is predominantly influenced by renal maturity, and there was high capacity for postnatal tubular maturation among preterm neonates. There is insufficient evidence to suggest that urinary NGAL is a useful marker of renal injury in the preterm neonate.

The effects of postnatal retinoic acid administration on nephron endowment in the preterm baboon kidney

Administration of retinoic acid (RA), the active metabolite of vitamin A, is linked to the stimulation of nephrogenesis. The aim of this study was to determine whether early postnatal administration of RA could enhance ongoing nephrogenesis in a baboon model of premature birth. Unbiased stereological methods were used to estimate kidney volume, renal corpuscle volume, and nephron number. The percentage of abnormal glomeruli and the number of glomerular generations was also determined in the kidneys of preterm control (n = 6) and preterm +RA (n = 6) animals that received 500 μg/kg/d of all-trans RA after premature delivery. There was no significant difference between the preterm control and the preterm +RA groups in kidney size, nephron number (preterm control: 329,924 ± 41,752; preterm +RA: 354,041 ± 52,095; p = 0.59), renal corpuscle volume, number of glomerular generations, or the percentage of abnormal glomeruli. The proportion of abnormal glomeruli did not appear to be linked to any elements of postnatal care examined. The results of this study indicate that early postnatal administration of RA is unable to stimulate nephrogenesis in the kidney of the preterm baboon. Encouragingly, it does not appear to have any adverse effects on kidney development.