Use of the isolated perfused kidney model to assess the acute pharmacologic effects of cyclosporine and its vehicle, cremophor EL

"Nephrotoxicity" secondary to cyclosporine and its clinically used vehicle, Cremophor EL, was examined in the isolated perfused rat kidney model. This model allows the serial determination of renal hemodynamic and tubular functional studies over a 3-hr duration using a normothermic, low hematocrit (13-15%) perfusion system. Initial studies indicated that the addition of small quantities of Cremophor EL resulted in marked renal vasoconstriction with decreased renal blood flow and deterioration in renal tubular function. These effects were highly significant and were of the same magnitude whether or not cyclosporine was present in the system. Cyclosporine was therefore examined after being dissolved in another vehicle, methanol. A 10% (v/v) amount of plasma was necessary in the perfusate to prevent significant adsorption of cyclosporine to the perfusion apparatus. Cyclosporine at concentrations below 100 ng/ml resulted in minor changes in renal hemodynamics. Beginning at 100 ng/ml glomerular filtration rate dropped significantly and renal vascular resistance increased three-fold. Fractional excretion of sodium significantly increased and the urine:plasma inulin ratio significantly decreased. We conclude that the clinically used drug vehicle, Cremophor EL, has significantly adverse effects on renal hemodynamics and tubular function. In addition, CsA causes similar renal toxicity in a dose-dependent fashion. Simultaneous administration of these two nephrotoxic agents could contribute to the high incidence of acute renal failure seen after transplantation. These observations suggest that an alternate vehicle with less renal toxicity might significantly decrease the incidence of this clinical problem.     

Cholestasis in the rat by means of intravenous administration of cyclosporine vehicle, Cremophor EL

The effect of cyclosporine vehicle, Cremophor EL, on bile flow and biliary bile acids and bilirubin output was studied in anesthetized male Wistar rats. Intravenous administration of Cremophor EL or castor oil as a single bolus reduced bile flow and the biliary output of bile acids and bilirubin. The Cremophor EL-induced cholestasis was an immediate and reversible phenomenon, since at 30-35 min after drug injection all parameters evaluated had returned to control values. A slight increase in serum bilirubin concentrations was observed. Our data indicate that the observed cholestasis is related to a reduction in both bile acid-dependent and bile acid-independent bile flow, probably due to a transitory hepatotoxic effect of Cremophor EL. We conclude that the clinically used vehicle for i.v. administration of cyclosporine, Cremophor EL, has adverse effects on hepatobiliary physiology in the rat and suggest that an alternative vehicle should be used.     

The effects of cremophor EL in the anaesthetized dog

The effects of cremophor EL were studied in 13 anaesthetized, paralyzed and ventilated dogs. Twenty per cent cremophor EL in a dose of 4.3 +/- 0.92 ml was infused at a rate of 30 ml X hr-1. In seven dogs, thoracopulmonary compliance, heart rate, systemic arterial pressure (SAP), pulmonary pressures (PAP, PCWP, RAP), cardiac output (CO) and platelet and white cell counts, were measured before the injection of cremophor EL, at the end of infusion and 5, 10, 30 and 150 minutes after the end of infusion. In six dogs, SAP, CO, and blood volume were measured before the injection of cremophor EL, at the end of infusion and 10, 30, 90 and 150 minutes after the end of infusion. Plasma histamine and catecholamines were assayed before the injection of cremophor EL and 2, 5, 10, 30, 90 and 150 minutes after starting the infusion. Cremophor EL induced a marked, sustained and significant decrease in SAP at the end of infusion and at 5, 10 and 30 minutes after the completion of the infusion (-68, -71, -70 and -43 per cent respectively), in PCWP, RAP and CO (-78 per cent at the end of infusion, -32 per cent 150 minutes after the end of infusion). Heart rate and systemic vascular resistance did not vary significantly. Pulmonary vascular resistance increased at the end of infusion, five and ten minutes after the end of infusion (+734, +548 and +439 per cent respectively). Plasma volume decreased 10 and 30 minutes after the end of infusion (-28 and -30.5 per cent respectively).(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of chronic cyclosporine administration on renal blood flow and intrarenal blood flow distribution

Cyclosporine-induced decreases in renal blood flow (RBF) and glomerular function are well documented. Glomerular filtration and tubular function may be affected by changes in both total renal blood flow and cortical blood flow distribution (CBFD). The effect of CsA on RBF, CBFD, glomerular filtration rate, and tubular function was studied in conscious ewes receiving a mean CsA dose of 30 mg/kg/day for 28 days with mean CsA trough levels of 344 +/- 45 ng/ml. RBF and CBFD were determined by the injection of 15 microns radioactive microspheres before and after one month of treatment with CsA or its vehicle, olive oil. RBF decreased by 24% from 7.65 +/- 0.87 to 5.79 +/- 0.42 ml/min/g of kidney in CsA-treated ewes (P = 0.014), while no decrease was noted in the control group (7.92 +/- 1.10 vs. 7.62 +/- 0.71). Intracortical blood flow decreased in proportion to the fall in total renal blood flow--thus CsA treatment did not change the cortical distribution of flow. There was a 25% decrease in GFR, as determined by inulin clearance, in the CsA-treated group (80 +/- 6 vs. 62 +/- 3 ml/min; P = 0.027) while there was a nonsignificant increase in control animals (62 +/- 11 vs. 92 +/- 7 ml/min). There was no evidence of tubular dysfunction in either group. There were also no changes in urinary excretion rates of prostaglandins PGE2, 6-keto-PGF1 alpha or thromboxane B2, nor were there changes in plasma renin activity. CsA induced decreases in RBF occur red without redistribution of cortical blood flow, indicating that altered cortical distribution of blood flow is not responsible for the changes in GFR or tubular function that have been reported. The changes in renal blood flow and glomerular filtration rate are independent of changes in renal prostaglandin production, and are likely not associated with altered plasma renin activity.     

Effect of ischemia on renal blood flow in the rat

The effect of ischemia on renal blood flow (RBF) in the rat has been an unsettled question. This study used a totally extractable indicator, radioactive microspheres, with a reference arterial sample, to study RBF quantitatively 10 and 60 min after 1 h of unilateral renal artery clamping in anesthetized, heparinized rats. The results indicate that after release of the arterial clamp, the kidney is enlarged and exhibits an increase in renal vascular resistance. RBF to the contralateral non-ischemic kidney is not different from that of a time-control series.     

Effects of cyclosporine on cerebral blood flow and metabolism in dogs.

Neurological side effects associated with cyclosporine immunosuppressive therapy are generally believed to occur with CsA blood concentrations above the therapeutic range. The effects of high blood CsA levels on cerebral hemodynamics, metabolism, and electrophysiologic activity were studied in acute (no CsA prior treatment) and chronic (with CsA prior treatment) dogs. In acute animals, when parenteral CsA (10 mg/kg or 25 mg/kg) was administered intravenously (CsA blood level 2000-22,000 ng/ml), slight but significant time-dependent decreases in cerebral blood flow (CBF), prolongation of absolute latencies of somatosensory-evoked potential (SSEP), and brainstem auditory-evoked responses (BAER) were noted. In the CsA chronically administered animals (oral CsA 25 mg/kg/24 hr for 14 days, CsA blood level 1077 ng/ml), baseline cerebral physiologic parameters were normal, and the cerebral responses to further administration of CsA (25 mg/kg, CSA blood level 56,000 ng/ml) intravenously were similar to those of the acute animals. Animals given Cremophor EL, the solvent for parenteral CsA preparation, showed similar cerebral responses to those observed in animals given CsA. Thus this study showed that CsA, regardless of the dose given, whether chronically or acutely administered, or the solvent for CsA all induced similar cerebral physiologic responses. We suggest that the cerebral physiologic and functional changes associated with parenteral CsA administration were small and were likely caused by its solvent, Cremophor EL, rather than CsA itself. Furthermore on the basis of our results, it is unlikely that high blood CsA per se can account for neurological side effects that occur in immunosuppressed patients.     

Mechanism of the decreased renal blood flow in the potassium-depleted conscious rat

Although chronic potassium deficiency is a common clinical problem, the hemodynamic consequences of chronic sustained potassium depletion have not been clearly delineated. In this study, the hemodynamic consequences of chronic potassium depletion were evaluated in the conscious rat. Potassium-depleted rats had a decrease in mean arterial pressure which was caused by a decrease in systemic vascular resistance. In association with these changes in systemic hemodynamics, renal blood flow (RBF) was also decreased. The decreased renal blood flow was caused by an increased renal vascular resistance (RVR). Because plasma renin activity was increased the role of angiotensin II as a renal vasoconstrictor was evaluated by utilizing two angiotensin antagonists. Although the administration of saralasin to potassium-depleted rats did not alter systemic hemodynamics, RVR was decreased and RBF was increased. Similar results were obtained with the converting enzyme inhibitor teprotide. Because products of endoperoxide metabolism may cause renal vasoconstriction, the role of prostaglandins and thromboxanes as renal vasoconstrictors were evaluated by utilizing cyclo-oxygenase and thromboxane synthetase inhibitors. None of these agents altered systemic hemodynamics. Following the administration of indomethacin, RVR was decreased and RBF was increased in potassium-depleted rats. Similar results were obtained with another cyclo-oxygenase inhibitor, meclofenamate, and with imidazole, an inhibitor of thromboxane synthetase. Because neither angiotensin II nor products of endoperoxide metabolism could alone account for the increased renal vascular resistance of potassium depletion, studies were performed in potassium-depleted rats treated with indomethacin plus either saralasin or teprotide. In these potassium-depleted animals, renal blood flow was restored to normal. In conclusion, the decrease in renal blood flow and the increase in renal vascular resistance in potassium depletion is mediated by angiotensin II and a product of prostaglandin endoperoxide metabolism, most likely, thromboxane.     

Partial tolerance in rat renal allograft recipients following multiple blood transfusions and concomitant cyclosporine

Multiple prior administrations of donor-strain blood while under limited cyclosporine cover, consistently induce extensive rat renal allograft survival and transplantation tolerance. Yet it was hypothesized that some chronic rejection mechanisms were nevertheless operative since consistent but nonprogressive minor renal dysfunction was observed long-term. A histopathologic study on these putative tolerant rats was undertaken to test this hypothesis. Twenty long-term LEW recipients of BN renal allografts receiving the blood-CsA regimen were examined histopathologically at day 100 post-transplant. Sixteen control LEW recipients receiving only a BN renal allograft were studied acutely at day 7 posttransplant. The control recipients demonstrated a range of lesions consistent with previous studies on acute renal allograft rejection in the rat. However, tolerant recipients demonstrated mild-to-moderate lesions consistent with chronic mechanisms of rejection including the following: moderate focal interstitial mononuclear inflammatory cellular infiltration, with periglomerular and perivascular accumulation; occasional arteriolar luminal obliteration and glomerular atrophy; focal areas of moderate interstitial fibrosis; mild interstitial hemorrhage; mild-to-moderate tubular atrophy; and focal tubular necrosis. Previously our laboratory has documented that tissue-specific renal basement membrane antigens may be responsible for inciting this pattern of focal chronic interstitial inflammation. However, from the present histopathologic studies, it would appear likely that chronic rejection mechanisms in these recipients, which were defined as tolerant by immunologic criteria, involve both tissue-specific and MHC determinants. Therefore, induction of transplantation tolerance in these indefinite survivors is partial or incomplete.     

The effect of somatostatin on central hemodynamics, renal blood flow, and renal function in dogs

We investigated the effects of infusion of somatostatin (200 and 500 ng/kg/min) on central hemodynamics and renal blood flow measured with radioactive microspheres. At the lower dose infusion rate somatostatin did not alter any hemodynamic parameter, but at 500 ng/kg/min somatostatin caused minor transient bradycardia, lowered cardiac output, and increased peripheral resistance. Both infusions inhibited renal arterial flow, with decreases noted in both cortical and medullary components. The 20% fall in renal perfusion was confirmed by the hippurate clearance technique, and there was a corresponding 17% decrease in the glomerular filtration rate. In contrast, no changes were noted in urine output, urinary concentrations of sodium or potassium, or urine osmolarity. These hemodynamic and renal side effects might limit the therapeutic usefulness of somatostatin infusion.     

Chronic cyclosporine therapy impairs endothelium-dependent relaxation in the renal artery of the rat.

Cyclosporine is an immunosuppressive substance that causes structural and functional alterations in endothelial cells. To examine the effects of chronic cyclosporine therapy on endothelial function, Wistar Kyoto rats received daily s.c. injections of saline, cyclosporine solvent, or cyclosporine (15, 30, or 50 mg/kg) for up to 2 wk. Blood pressure remained unchanged in all groups. Segments of the renal artery were suspended in organ chambers filled with physiological salt solution, and isometric tension was recorded. In rats treated with 30 or 50 mg/kg/day of cyclosporine, endothelium-dependent relaxations to acetylcholine of the renal artery were significantly impaired when compared with vessels obtained from rats injected with saline or solvent. The reduced acetylcholine-induced relaxation of cyclosporine-treated vessels was improved by preincubation of the preparations with the cyclooxygenase inhibitor indomethacin. Endothelium-independent relaxations in response to sodium nitroprusside were unimpaired in renal artery rings after 1 wk of cyclosporine but were reduced after 2 wk of treatment with 30 mg/kg/day. Contractions of the renal artery in response to norepinephrine and serotonin were not altered by cyclosporine. Thus, (1) high-dose cyclosporine therapy impairs endothelium-dependent relaxations in the renal artery of the rat; (2) an endothelium-derived cyclooxygenase product reduces the effects of endothelium-derived relaxing factor in cyclosporine-treated rats; and (3) chronic cyclosporine treatment slightly impairs vascular smooth muscle relaxation, whereas vascular contractility remains unaltered.     

Evaluation of blood flow in free microvascular flaps

Free flap surgery is routine today, yet little is known of its pathophysiology. In this study, the authors evaluated the hemodynamics in different types of free microvascular flaps, by measuring intraoperative transit-time flow. Eighty-six free transplants--21 free TRAM flaps for breast reconstruction, 18 radial forearm flaps for head and neck reconstructions, and 47 muscle flaps for head and neck, trunk and lower extremity reconstructions--were studied. Donor artery flow was highest in the radial artery (mean: 57.5 +/- 50 (SD) ml/min) but dropped (p < 0.001) to one tenth (6.1 +/- 2 ml/min) after anastomosis. The flow was lowest (4.9 +/- 3 ml/min) in the recipient artery of the TRAM flap but, after anastomosis, increased significantly (13.7 +/- 5 ml/min) to the level of the flow in the donor artery. The donor-artery flow in muscle flaps had a mean of 15.9 +/- 11 ml/min, and it significantly increased after anastomosing (23.9 +/- 12 ml/min). Weight-related intake of blood was highest in the radial forearm flap (18.5 +/- 6 ml/ min/100g) and lowest in the TRAM flap (2.5 +/- 1 ml/min/100g). The study showed that blood flow through a free microvascular flap does not depend on recipient artery flow. Even low-flow arteries can be used as recipients, because the flow increases according to free-flap requirements. The blood flow through a free microvascular flap depends on the specific tissue components of the flap.     

The influence of lateral tilt on limb blood flow in advanced pregnancy.

Blood flow of the forearm and calf were measured in 20 expectant mothers by means of strain gauge plethysmography. Measurements were made with the patients initially lying supine, then tilted to the right or left side with a 15 degrees foam rubber wedge and, finally, after returning the subjects to the supine position. Blood flow of the forearm was unaltered by changes in maternal posture. Lower limb perfusion usually increased significantly when the mothers assumed the right or left lateral tilt position, and decreased when the patients again lay supine. The significance of these findings in relationship to aortocaval occlusion by the gravid uterus is discussed. We conclude that lateral tilt of the patient during Caesarean section should contribute significantly to the safety and welfare of both mother and infant.     

The utility of cyclosporine weaning in renal transplantation.

Abrupt conversion of cyclosporine immunosuppression to conventional treatment with azathioprine and prednisone avoids long-term cyclosporine nephrotoxicity, albeit at the cost of a 20% to 40% rejection rate. The authors investigated the benefits and risks of a cyclosporine weaning protocol in 24 cadaveric and 9 live donor kidney recipients treated with a sequential quadruple immunosuppressive protocol. In cadaver kidney recipients, slow tapering of cyclosporine resulted in a 19% (p less than 0.001) improvement in the glomerular filtration rate, as estimated by the inverse ratio of the plasma creatinine concentration. Cadaver kidney recipients were stratified according to graft function (GFR ratio greater than 0.76, less than 0.76) at the of cyclosporine discontinuation. In 12 patients with well-functioning grafts, a 24% improvement was observed, whereas in 12 patients with poor graft function, the gain was limited to 13%. Patients with limited graft function tended to have more acute rejection episodes before cyclosporine weaning (0.92 +/- 0.64 versus 0.42 +/- 0.64, not significant). When the 24 cadaver kidney recipients were stratified according to onset of graft function after transplantation (days to plasma creatinine of 250 mumol/L), need for dialysis, panel reactive antibodies (PRA), and duration of cyclosporine treatment, no significant differences in graft function were observed at the onset or end of cyclosporine weaning. Acute graft rejection before cyclosporine weaning was the only variable associated with a significantly lower estimated glomerular filtration rate ratio at the end of cyclosporine treatment (0.83 +/- 0.11 versus 0.67 +/- 0.16, p less than 0.01). Weaning of cyclosporine was associated with a minimal risk of acute graft rejection. A single patient with stable graft function at the onset of the weaning process experienced an acute but reversible rejection episode 2 months after cyclosporine was discontinued. In summary, gradual weaning of cyclosporine improves graft function, and eliminates the excessive risk of acute graft rejection without the need for additional corticosteroid treatment.     

Effect of monophasic blood flow on the patency of microvascular anastomoses

The effect of monophasic blood flow on the patency of microvascular anastomoses in the rat femoral artery was evaluated by the authors. An experimental model of monophasic blood flow was created in the rat femoral artery to simulate that seen in patients with peripheral vascular disease. Microvascular anastomoses were performed, and the patency at the first hour and 24 hr later was determined. The studies demonstrated that microarterial anastomotic patency was the same when performed in vessels with monophasic or triphasic flow patterns. The authors conclude that monophasic blood flow is not an absolute contraindication for free-tissue transfer.