Jean-Bernard Gouyon

In utero exposure to immunosuppressive drugs: experimental and clinical studies

Abstract Over the last few decades, the number of pregnant women under immunosuppressive (IS) therapy following transplantation or autoimmune diseases has increased. At first, IS drugs, including prednisone, azathioprine, and cyclosporine A were used, but now new molecules such as tacrolimus and mycophenolate mofetil have appeared. These IS drugs cross the placental barrier and enter into the fetal circulation, which poses a risk for fetal development. Experimental data have shown that IS drugs often have deleterious effects on fetuses, while human data have reported an increased rate of abortion, prematurity, intrauterine growth retardation (IUGR), and low birth weight, without significant increases in malformation rates. However, only limited information is available about the newly used molecules. Although fetal and neonatal data are reassuring, long-term effects of IS drugs on fertility, immune response and renal function, as well as the consequences of prematurity and IUGR, should be monitored.

Long-Term Effects of In Utero Exposure to Cyclosporin A on Renal Function in the Rabbit

The number of pregnant women who receive cyclosporin A (CsA) after transplantation or for autoimmune disease has increased. CsA and its metabolites can cross the placental barrier and thus interfere with fetal development. It was shown previously that rabbits that were exposed in utero to 10 mg/kg per d CsA from the 14th to the 18th day of gestation presented a 25% nephron reduction. Thus, this study was conducted to assess the long-term systemic and renal effects of a CsA-induced nephron reduction. Twenty-two pregnant New Zealand white rabbits were randomly divided into two groups: Twelve received 10 mg/kg per d CsA from day 14 to day 18 of gestation, and 10 were used as controls. Rabbits that were born to these animals were evaluated at 4, 11, 18, and 35 wk of life. Pups that were exposed antenatally to CsA presented first a permanent nephron deficit; second, glomerular, tubular, and intrarenal hemodynamics dysfunction; third, enlarged kidneys with numerous tubular and glomerular lesions; and, fourth, an endothelin-dependent systemic hypertension that worsened with age. In utero exposure to CsA induced a nephron reduction that led to systemic hypertension and progressive chronic renal insufficiency in adulthood. A long-term clinical survey is mandatory in infants who are born to mothers who were treated with cyclosporin during pregnancy.

Recurrence of SOX2 anophthalmia syndrome with gonosomal mosaicism in a phenotypically normal mother.

Laurence Faivre,* Kathleen A. Williamson, Valérie Faber, Nicole Laurent, Marianne Grimaldi, Christel Thauvin-Robinet, Christine Durand, Francine Mugneret, Jean-Bernard Gouyon, Alain Bron, Frédéric Huet, Caroline Hayward, Veronica van Heyningen, and David R. FitzPatrick Centre de Génétique, Hôpital d’Enfants, Dijon, France MRC Human Genetics Unit, Western General Hospital, Edinburgh, United Kingdom Service de Pédiatrie, Hôpital d’enfants, Dijon, France Service d’Anatomopathologie, CHU le Bocage, Dijon, France Service de Radiologie, Hôpital d’Enfants, Dijon, France Service de Cytogénétique, CHU Le Bocage, Dijon, France Service d’Opthalmologie, CHU Le Bocage, Dijon, France

Cyclosporin A Administration during Pregnancy Induces a Permanent Nephron Deficit in Young Rabbits

Cyclosporin A (CsA) is an immunosuppressive agent used to prevent graft rejection and to treat autoimmune disorders. Successful pregnancies can be achieved among CsA-treated women, although it is known that CsA is nephrotoxic and crosses the human placenta. The aim of this study was to evaluate the harmlessness of CsA toward the embryonic kidney. Twenty-one pregnant rabbits were divided into four groups. Groups of six and four female animals were subjected to daily injections of 10 mg/kg per d CsA (administered subcutaneously) for 5 d, from day 14 to day 18 of gestation or from day 20 to day 24 of gestation, respectively. In the third group, five female animals received the CsA diluent (Cremophor) from day 14 to day 18 of gestation. The fourth group consisted of six untreated female animals. Pregnancy outcomes among CsA-treated does demonstrated a reduced number of living pups, which were also growth-retarded, with exposure to CsA from day 20 to day 24 of gestation. However, pups exposed to CsA from day 14 to day 18 of gestation exhibited normal fetal growth, and blood concentrations of CsA matched human data. Examinations of kidneys at birth demonstrated vacuolation of proximal and collecting tubules and ureteric bud ends. Increased glomerular volumes and decreased nephron densities suggested nephron mass reduction, which was quantitatively evaluated in 1-mo-old animals. The nephron numbers were reduced by 25 and 33% in day 14 to 18 CsA-treated and day 20 to 24 CsA-treated animals, respectively, which displayed compensatory adaptation of the existing nephrons. However, foci of segmental glomerular sclerosis were already present, which would possibly jeopardize renal function later in life.

Use of peritoneal dialysis, continuous arteriovenous hemofiltration, and continuous arteriovenous hemodiafiltration for removal of ammonium chloride and glutamine in rabbits

OBJECTIVE We compared the ability of peritoneal dialysis, hemofiltration, and continuous hemodiafiltration to remove infused ammonium chloride. STUDY DESIGN Anesthetized adult rabbits received an intravenous infusion of ammonium chloride. Two methods of removal of ammonium chloride were performed in each animal and compared. In group 1 (n = 6), peritoneal dialysis (dialysate = 75 ml.kg-1) and continuous arteriovenous hemofiltration (CAVH) with a polysulfone 800 cm2 hemofilter (Minifilter Plus; Amicon Division, W. R. Grace & Co., Danvers, Mass.) were simultaneously performed for 40 minutes. In group 2 (n = 6), peritoneal dialysis and continuous arteriovenous hemodiafiltration (CAVHD) (dialysate flow = 1000 ml.hr-1) were simultaneous performed for 40 minutes. In group 3 (n = 6), CAVH and CAVHD were performed successively in random order for 30 minutes each. RESULTS Animals had high and stable ammonium chloride and glutamine plasma levels during the experimental procedure. No significant difference in ammonium chloride clearance was observed between PD and CAVH (group 1). In comparison with PD or CAVH, CAVHD resulted in significantly higher clearances of ammonium chloride (40% +/- 10% vs 96% +/- 34%, respectively) and of glutamine (195% +/- 17% vs 77% +/- 25%, respectively). CONCLUSION The overall results indicate that CAVHD should be considered for hyperammonemia when peritoneal dialysis is indicated but unfeasible or inefficient.

Protective Effect of Perindoprilat in the Hypoxemia-Induced Renal Dysfunction in the Newborn Rabbit

The renal effects of acute hypoxemia and the ability of perindoprilat, a potent angiotensin-converting enzyme inhibitor, to prevent these effects were assessed in 31 anesthesized and mechanically ventilated newborn (5 to 8 d of age) rabbits. Renal blood flow (RBF) and GFR were determined by the clearances of para-aminohippuric acid and inulin, respectively. Each animal acted as its own control. In eight normoxemic rabbits (group 1), the i.v. infusion of saline did not change renal and hemodynamic functions. In eight additional rabbits, acute hypoxemia (Pao2 = 40 mm Hg) induced a significant decrease in mean blood pressure (-8 ± 2%), RBF (-36 ± 3%), and GFR (-31 ± 3%) and an increase in renal vascular resistance (+50 ± 12%). A third group of newborn animals (n = 7) was used to determine the renal effects of perindoprilat administration (20 µg/kg) under normoxemic conditions. RBF significantly increased (+15 ± 2%) and renal vascular resistance significantly decreased (-15 ± 3%), whereas GFR, mean blood pressure, and filtration fraction did not change significantly. In group 4 (n = 7), perindoprilat infusion completely prevented the hypoxemia-induced alterations in GFR and renal vascular resistance and partially prevented the fall in RBF. These results demonstrate that angiotensin II modulates the renal immature microcirculation and that inhibition of its formation effectively prevents the hypoxemia-induced decrease in GFR.

Prevention of hypoxemia-induced renal dysfunction by perindoprilat in the rabbit

The role of angiotensin II, a potent postglomerular vasoconstrictor, in the hypoxemia-induced renal changes is still controversial. The ability of perindoprilat, an angiotensin converting-enzyme inhibitor, to prevent the acute renal effects of hypoxemia was assessed in 22 anesthetized-ventilated rabbits. In 8 untreated rabbits, hypoxemia induced a significant drop in mean blood pressure (MBP) (-12 +/- 2%), glomerular filtration rate (GFR) (-16 +/- 3%) and renal blood flow (RBF) (-12 +/- 3%) with a concomittant increase in renal vascular resistance (RVR) (+18 +/- 5%) and urine flow rate (+33 +/- 14%), and without any changes in filtration fraction (FF) (-4 +/- 2%). This suggests the occurrence of glomerular vasoconstriction during the hypoxemic stress. In 7 normoxemic rabbits, intravenous perindoprilat (20 microg/kg) induced an increase in urine flow rate (+17 +/- 4%) and RBF (+17 +/- 4%), and a decrease in MBP (-6 +/- 1%), RVR (-14 +/- 3%) and FF (-11 +/- 2%) without a significant change in GFR. The drop in FF and the increase in RBF suggests preferential postglomerular vasodilatation. In 7 rabbits, perindoprilat prevented the occurence of the hypoxemia-induced changes in RBF and RVR without improving MBP. FF decreased significantly (-18 +/- 2%), while the drop in GFR (-7 +/- 2%) was partially blunted and the increase in urine flow rate (+25 +/- 9%) was confirmed. These results could be explained by the inhibition of the angiotensin-mediated efferent vasoconstriction and by the inhibition of bradykinin degradation by perindoprilat. These data confirm the ability of converting-enzyme inhibitors to prevent the renal hypoperfusion induced by acute hypoxemia.

Urea removal by hemofiltration and hemodiafiltration.

Continuous hemofiltration is usually regarded as a safe method for controlling fluid overload in neonates presenting acute renal failure. We considered that continuous hemodiafiltration with a hemofilter type especially designed for use in the neonatal period could improve urea removal as compared with hemofiltration. Continuous arteriovenous hemofiltration (CAVH) and hemodiafiltration (CAVHD) were consecutively performed on 7 anesthetized adult rabbits which were given an urea infusion. The hemofilter was a 800-cm2 polysulfone model (Amicon Minifilter Plus). Mean values for physiological and operational parameters were comparable during CAVH and CAVHD, i.e., mean blood pressure (69.6 +/- 14.9 and 70 +/- 13.1 mm Hg, respectively), arterial oncotic pressure (16.0 +/- 2.4 and 16.0 +/- 1.7 mm Hg), hematocrit (36.2 +/- 5.8 and 36.7 +/- 4.4%), protein plasma level (38.9 +/- 7.0 and 39.0 +/- 6.4 g/l), urea plasma level (20.6 +/- 6.5 and 19.9 +/- 7.0 mmol/l), plasma flow in the hemofilter (7.89 +/- 8.48 and 7.12 +/- 3.08 ml/min), and ultrafiltrate rate (1.17 +/- 0.49 and 1.73 +/- 0.78 ml/min). CAVHD allowed a significant increase in urea clearance (4.74 +/- 4.51 ml/min) as compared with CAVH (1.23 +/- 0.81 ml/min). These experimental results suggest that CAVHD usefulness should be assessed in neonates presenting acute renal failure.

Calcium, Sodium and Potassium Urinary Excretion during the First Five Days of Life in Very Preterm Infants

Background: Hypercalciuria has been associated with the risk of nephrocalcinosis and renal stones in both adults and children. Renal calcifications are frequently encountered in preterm infants because this particular population presents most of the risk factors of increased urinary calcium excretion. Urinary calcium excretion has been shown to correlate with sodium and potassium excretions in adult patients, but these correlations have not been demonstrated in the early neonatal period yet. Objective: To define the relationship between calcium urinary excretion and sodium or potassium excretions in the first 5 days of life in preterm babies. Methods: A prospective study was conducted in 16 preterm infants born before 32 weeks of gestation (body weight 1,373 ± 310 g; gestational age 29.1 ± 1.6 weeks). Fifteen consecutive 8-hour urine collections were performed for each infant from the 8th hour of life. A plasma sample was obtained at the end of each urine collection. Sodium, potassium, calcium and creatinine were measured in urine and blood samples as often as possible. Results: (1) Urine sodium excretion was 6.56 ± 4.35 mmol/kg per day. (2) Urinary calcium excretion was 5.9 ± 5.4 mg/kg per day and the urinary calcium/creatinine ratio was 0.48 ± 0.39 mg/mg. (3) Urinary calcium and sodium excretion were positively correlated (r = 0.65, p = 0.0001), while an inverse correlation was found between calcium and potassium excretion (r = 0.31, p = 0.004). Conclusion: The mean values of urinary calcium excretion and calcium/creatinine ratio observed in our population were higher than 4 mg/kg per day and 0.4 mg/mg, respectively, i.e. boundary values previously associated with an increased risk of nephrocalcinosis. We hypothesize that an increase in urinary sodium excretion in this population may facilitate calcium excretion.

Disparate effects of chronic and acute theophylline on cyclosporine A nephrotoxicity

Abstract We previously developed a model of acute cyclosporine A (CsA)-induced vasomotor nephrotoxicity in rabbits. As exogenous adenosine infusion mimics the haemodynamic changes that characterize acute renal failure (ARF), we wanted to know whether adenosine was a mediator in this model and whether an adenosine receptor blocker could prevent the CsA-induced ARF. Group 1 were untreated controls. Group 2 received CsA (25 mg/kg per day) for 5 days. Renal function parameters were measured, showing ARF in all animals compared to controls. Theophylline (1 mg/kg i.v. bolus) was then administered and renal function was reassessed. Theophylline significantly reduced renal vascular resistance (–8%) and increased renal blood flow (RBF) (+20%), glomerular filtration rate (GFR) (+50%), filtration fraction (+24%) and diuresis (+73%), suggesting that adenosine was involved in the CsA-induced ARF. In group 3, theophylline (30 mg/kg per day) was given concomitantly with CsA for 5 days. GFR was normalized, but theophylline did not hinder the drop in RBF seen with CsA alone in group 2. Microscopy observation of the kidneys showed that chronic theophylline administration aggravated the morphological changes induced by CsA alone. We conclude that CsA administration for 5 days induced a vasomotor nephropathy with an adenosine-mediated afferent arteriolar constriction which cannot be prevented by concomitant theophylline administration.