Henrik Starklint

Nephrotoxicity of cyclosporin A in humans: effects on glomerular filtration and tubular reabsorption rates

Abstract. The contention that cyclosporin A (CyA) nephrotoxicity may be due to renal afferent arteriolar constriction was inferred from rat studies showing CyA to increase renal vascular resistance, to reduce glomerular filtration rate (GFR) and delivery of tubular fluid from the end of the proximal tubule to the loop of Henle (Vprox), and to increase proximal fractional reabsorption. In order to test whether the mechanism of human CyA nephrotoxicity is similar to its rat analogue, and whether CyA treatment causes prolonged renal malfunction after drug withdrawal, renal function was investigated with clearance techniques including lithium clearance (CLi) as a measure of Vprox. The subjects were patients (n= 11) with previously normal renal function, given CyA in the treatment of ocular manifestation of extrarenal disease, or bone‐marrow transplant recipients. Nine out of these eleven patients were investigated before and during CyA treatment: GFR (P<0·05) and Vprox (P<0·005) decreased while proximal fractional reabsorption increased (P<0·01). In six patients investigated before CyA was given, and re‐examined a mean of 273 days (range 84–384 days) after CyA withdrawal, CLi was reduced (P<0·05) while mean GFR was not significantly lowered (0·5>P > 0·2). In one of these six patients GFR was reduced to a subnormal value of 26 ml min‐1 (1·73m2 body surface)‐1. In conclusion, human and rat CyA nephrotoxicity have the same pattern of renal functional deterioration. Cyclosporin A nephrotoxicity was evident in patients investigated a mean of 9 months after CyA withdrawal.

Effect of cooling and warming rate on glycerolized rabbit kidneys

Cooling and warming rates are known to be important determinants of viability for cryopreserved cells, but optimal rates have not previously been determined for any whole organ. In this study, rabbit kidneys, permeated with 2 M glycerol were cooled to -80 degrees C at four rates varying from 1 degrees C/hr to 3.1 degrees C/min and then rewarmed at four rates from 1 degrees C/hr to 4.2 degrees C/min, giving 16 experimental treatments. After gradual deglycerolization at 10 degrees C, each kidney was autografted and observed for 30 min. Assessment was by measurement of vascular resistance, immediate post-thaw lactate dehydrogenase (LDH) release, gross appearance, light- and electron microscopy, and tissue K+/Na+ ratio 30 min after transplantation. The best results were obtained after cooling at 1 degrees C/hr; warming rate had little apparent influence on the criteria used to assess function with the exception of LDH release, which indicated a preferred warming rate around 1 degrees C/min. Histological studies revealed extensive vascular damage, notably to the glomerular capillaries, that was minimized by very slow cooling. Freeze substitution, carried out on samples removed at -80 degrees C, demonstrated extensive ice formation in the interstitial space and, at the faster cooling rates, in the glomerular capillaries. Intracapillary ice formation was reduced in the kidneys cooled at 1 degrees C/hr.

Nephrotoxicity of cyclosporin A. A lithium clearance and micropuncture study in rats

Abstract. Renal function was studied in rats treated with cyclosporin A (CyA). Peroral CyA 25 mg kg‐1 day‐1 depressed glomerular filtration rate (GFR) from 1284 ± 429 to 500 ± 228 μl min‐1 g‐1 kidney weight (KW) (P < 0·01). Absolute rate of proximal tubular reabsorption (APR) decreased from 1075 ± 437 to 468 ± 203 μl min gKW‐1 (P < 0·01). Proximal tubular fractional reabsorption (PFR) was 67·7 and 68·5% measured with the TT/OT and fractional lithium‐clearance methods, respectively. Amiloride had no effect on lithium‐clearance in CyA treated rats. Acute isotonic volume expansion increased GFR and APR towards normal, while PFR remained increased. Increased sodium clearance did not normalize renal function. CyA intravenously (12·5 mg kg‐1) depressed GFR and APR acutely, while PFR increased. Proximal intratubular pressures were low normal (mean 11·6 mmHg). Proximal transit times were prolonged (mean 25·2 s, P < 0·01). Renal morphology was normal. The data are evidence against a primary tubular damage of CyA, and makes it less likely that the major lesion is located to the glomerular membrane. The results suggest that CyA nephrotoxicity mainly is due to a haemodynamic effect.

Antagonist capacities of nifedipine, captopril, phenoxybenzamine, prostacyclin and indomethacin on cyclosporin A induced impairment of rat renal function

Abstract. Experimental evidence indicates that cyclosporin A (CyA) nephrotoxicity is due to renal arteriolar constriction, reducing renal blood flow, glomerular filtration rate (GFR), and delivery of tubular fluid from the end of the proximal tubule to the loop of Henle. The proximal tubular fractional reabsorption (PFR) is increased. Therefore, the impact on renal function of vasodilating agents was studied in rats given CyA. Conscious catheterized rats and clearance techniques were used. In acute experiments a preexisting CyA‐nephrotoxicity was resistant to infusion of phenoxybenzamine, prostacyclin, captopril, nifedipine and indomethacin. Concomitant treatment with captopril and CyA did not improve renal function, while concomitant treatment with CyA and nifedipine improved GFR to 1·13±0·34 ml min‐1 g‐1 kidney weight (gKW) (n=19, P<0·05), as compared to CyA and placebo treated controls (n=12, 0·83±0·32 ml min‐1 g‐1 KW). Nifedipine also reduced FPR (88·6±5·1% vs. 83·2±5·6%. P<0·01), and increased lithium clearance from 99±54 to 184±64 μl min‐1 g‐1 KW (P<0·001). The results are further evidence that CyA nephrotoxicity includes renal vasoconstriction, and indicates that calcium entry blockade is nephroprotective in the case of CyA toxicity.

Introduction and removal of cryoprotective agents with rabbit kidneys: Assessment by transplantation

Rabbit kidneys were perfused with up to 4 M glycerol or propane-1,2-diol (propylene glycol, PG) in three vehicle solutions: one normokalemic and made hypertonic with mannitol (HP5), one hyperkalemic but without mannitol (HP6), and one hyperkalemic and with mannitol (HP7). Subsequent function was assessed by autotransplantation. Up to 3 M glycerol in HP5 was well tolerated but not in HP6 or HP7. Conversely, up to 3 M PG in HP7 was compatible with excellent post-transplant function, but the same concentration in HP5 was severely damaging. PG (4 M) in either solution was severely injurious and no kidneys survived perfusion with this concentration. Vascular resistance was well controlled by the vehicle solutions with mannitol, but it was generally higher during perfusion with the hyperkalemic HP7 compared with the normokalemic HP5. No kidneys perfused with 3 M solutions of either of the cryoprotective agents and cooled briefly to -6 degrees C without freezing had any post-transplant function, and neither did kidneys perfused with 3 M PG or 4 M glycerol tolerate slow cooling to -80 degrees C and warming. The need to optimize perfusate composition for the CPA being used is clear, and the dramatic increase in toxicity of PG when the concentration exceeds 3 M supports the suggestion that mixtures of PG and glycerol should be considered. The observation of damage at high subzero temperatures, before freezing has occurred, requires further detailed study.

The survival of pig to rabbit renal xenografts during inhibition of thromboxane synthesis

Five rabbits treated with the thromboxane synthetase inhibitor Dazoxiben and five control rabbits received pig renal xenografts. Plasma albumin, complement factor C3, TXB2, 6-keto-PGF1 alpha, and serum TXB2 and 6-keto-PGF1 alpha were determined before and 1/2 hour after transplantation. The xenograft survival was significantly decreased in the Dazoxiben treated animals compared to the placebo treated animals determined as time to total cyanosis of the graft, total urine production, and time to stop of urine production. Lack of TXB2 production during blood coagulation confirmed inhibition of platelet thromboxane synthesis. The other determined variables showed no significant differences between the treated and the placebo animals.

Impaired glomerular and tubular function as a short-term effect of sirolimus treatment in the rat.

Aims: To investigate acute and short-term effects of sirolimus (SRL) on glomerulo-tubular function, blood pressure (BP), and renal morphology in the rat. Methods: Male Sprague-Dawley rats, weighing initially 140–180 g were treated with SRL in three series: SRL 0.2, 0.4, or 0.8 mg/kg/day intraperitoneally for up to 28 days after skin allo-transplantation from Lewis donors (to establish a dosage with significant immunosuppressive effect). SRL 0.4 mg/kg intravenously (acute effects). SRL 0.4 mg/kg/day intraperitoneally for 7 days (short-term effects). Inulin, lithium (CLi) and sodium clearance, and intra-arterial BP were measured in conscious catheterized rats. Morphological kidney studies were completed after post-mortem fixation. Results: Maximum immunosuppressive effect was achieved with SRL 0.4 mg/kg/day. SRL acutely increased GFR and CLi, whereas fractional proximal reabsorption (PFR) declined. In the short-term study SRL had opposite effects on GFR and CLi, unaffected proximal tubular reabsorption and PFR, raised BP, diminished food consumption, and slower increase in body weight. Morphological changes were non-specific. Conclusion: In a dosage giving maximum immunosuppressive effect, SRL revealed acute effects on glomerular and proximal tubular function thus indicating increased outflow from the proximal tubules whereas one week of SRL treatment produced a change resembling the known nephrotoxic effects of the calcineurine inhibitors.

Platelet aggregation is not essential for xenograft rejection

Five rabbit kidneys were perfused at 80 mm Hg with heparinized human blood, five with platelet poor human blood, and five with human blood added prostacyclin. Time until blood flow decreased to 2 ml/min was significantly delayed by removal of platelets or addition of prostacyclin. Histological examination did not reveal any difference between the groups. It is concluded that platelets play an enhancing role, but they are not essential for xenograft rejection.

Glycerol used as a cryoprotectant in subzero preservation of rabbit kidneys

Summary Perfusion fluids containing different concentrations of glycerol were examined using autotransplantation of kidneys in rabbits. The isolated left kidney was flushed with the perfusion fluid in question for 10 min, stored for 45 min in a refrigerator at +4°C, and finally reperfused for 5 min with the same fluid immediately before transplantation. Graft function was estimated by measuring serum creatinine and creatinine clearance in the first 24 hr after transplantation. Three perfusion fluids containing 5, 10, and 20% of glycerol were examined. The function of kidney grafts flushed with fluids containing up to 10% glycerol did not differ significantly from the function of kidneys preserved with a modified Collins solution using the same procedure; whereas perfusion with fluids containing 20% glycerol resulted in a large increase in serum creatinine and a smaller creatinine clearance. Following these results, a group of kidneys were perfused continuously with a fluid containing 10% glycerol for 2 hr, stored by simple hypothermia at −1°C for 19–22 hr, and then transplanted. Five out of 20 animals survived. Serum creatinine showed a temporary increase after transplantation but returned subsequently to almost normal.

An Association of Hypothalamic Hamartoma, Central Precocious Puberty and Juvenile Granulosa Cell Tumour in Early Childhood

A case of central precocious puberty from infancy due to a hypothalamic hamartoma and associated with an ovarian juvenile granulosa cell tumour is presented. Central precocious puberty was diagnosed by gonadotropin stimulation tests and LHRH agonist therapy was successful. A MR scan, but not a CT scan, demonstrated the hypothalamic hamartoma. The possible influence of early LH stimulation for the development of the granulosa cell tumour is discussed.