Takafumi Yura

Cytochrome P450 4A expression and arachidonic acid omega-hydroxylation in the kidney of the spontaneously hypertensive rat.

20-Hydroxyeicosatetraenoic acid (20-HETE) is a major arachidonate metabolite in the kidney of the spontaneously hypertensive rat (SHR). The increase in its synthesis has been associated with the elevation of blood pressure in the SHR. The omega-hydroxylation of arachidonic acid is an activity associated with members of the CYP4A gene family which, in the rat, comprises three major isoforms: 4A1, 4A2 and 4A3. 20-HETE displays potent and diverse biological activities which can affect pro- and anti-hypertensive mechanisms dependent upon where, when and by which isoform it has been produced. Therefore, it is important to identify and characterize its biosynthetic system. We compared CYP4A mRNA and protein expression to patterns of 20-HETE synthesis in the SHR kidney. The reverse transcription/polymerase chain reaction (RT/PCR) technique was used to amplify CYP4A mRNA in microdissected nephron segments. Southern blot hybridization of PCR products obtained from nephron segments with the CYP4A1 cDNA probe demonstrated strong signals in S2 and S3 segments of the proximal tubule. Immunoblots of nephron segments using a polyclonal anti-rat liver CYP4A1 antibody which cross-reacts with CYP4A2 and CYP4A3, and 14C-arachidonic acid metabolism, confirmed that arachidonic acid omega-hydroxylation, i.e., 14C-20HETE formation, and CYP4A proteins were also localized mainly in the S2 and S3 segments. Correlation also existed between the age-dependent increase in arachidonate omega-hydroxylation in the kidney and CYP4A mRNA levels as measured by Northern hybridization of total RNA using the CYP4A1 cDNA probe. Immunoblot analysis revealed that at 7 weeks, where 20-HETE production is at its maximum, all three proteins are expressed. CYP4A3 and 4A1 immunoreactive proteins appeared at 3 weeks, showed maximum levels at 5 and 7 weeks, respectively, and gradually decreased to lower levels at 13 and 20 weeks, whereas CYP4A2 levels were undetectable at 3, 5 and 7 weeks but appeared at 13-20 weeks. Additional immunoblots indicated that renal cortical CYP4A1 protein levels were higher in SHR compared to Sprague-Dawley and Wistar-Kyoto rats. The increased levels of CYP4A1-immunoreactive band at 7 weeks corresponded to the maximal activity of arachidonate omega-hydroxylation. Thus, CYP4A1 might play a significant role in contributing to the increased cortical/proximal production of 20-HETE seen in 7-week-old SHR. However, given the high homology among members of the CYP4A gene family and the lack of specific tools to discern among these isoforms, additional studies have to be carried out to substantiate our findings.

Stimulatory Effect of 8-Epi-PGF2α, An F2-Isoprostane, on Endothelin-1 Release

Summary: 8-Epi-prostaglandin F2α (8-epi-PGF2α) is an F2-isoprostane produced in vivo by a cyclooxygenase-independent, free radical-catalyzed lipid peroxidation mechanism. It exhibits renal vasoconstrictor effects by binding to a receptor related to, but distinct from, that of thromboxane A2 (TxA2). In cultured bovine aortic endothelial cells (BAECs), competitive binding assays using [3H]-8-epi-PGF2α indicated the existence of two distinct binding sites. The Kd values were similar to those of cultured rat aortic smooth-muscle cells, suggesting that the high- and the low-affinity binding sites represent isoprostane and TxA2 receptors, respectively. 8-Epi-PGF2α dose-dependently stimulated endothelin-1 (ET-1) secretion from BAECs. These increases were partially inhibited by a TxA2 receptor antagonist, consistent with the premise that isoprostanes and TxA2 recognize closely related receptors. The presence of specific binding sites for F2-isoprostanes on endothelial cells widens the scope of the possible pathophysiologic significance of these eicosanoids, released during oxidant injury, to include alteration of endothelial cell biology. The release of ET-1 by 8-epi-PGF2α may help to explain the large increases in plasma and urinary concentrations for both ET-1 and 8-epi-PGF2α in patients with hepatorenal syndrome.

Antineutrophil Cytoplasmic Antibodies (ANCA)-Associated Crescentic Glomerulonephritis and Propylthiouracil Therapy

Cutaneous vasculitis is an uncommon complication of propylthiouracil therapy. Although its pathogenesis remains to be established, detection of antineutrophil cytoplasmic antibodies (ANCA) in association with this type of vasculitis has recently been described. We report here 2 patients who developed ANCA-associated crescentic glomerulonephritis without evidence of cutaneous vasculitis during propylthiouracil treatment of hyperthyroidism. Improvement of the renal function and the disappearance of ANCA were correlately found after discontinuation of propylthiouracil and by corticosteroid therapy in both patients.

Quenching the Thirst in Dialysis Patients

In a double-blind cross-over trial, 22 stable end-stage renal failure patients on maintenance haemodialysis were subjected to conventional dialysis with dialysate containing 137 mEq/l sodium and constant ultrafiltration (UF) and to a different dialysis therapy, in which, by linear sodium modelling, the dialysate sodium was reduced from 137 to 128 mEq/l. A computerized UF program was used to gradually reduce the UF to a minimum towards the end of the session. Severity of thirst, interdialytic weight gain and intradialytic complications were less with low sodium dialysate. It allowed adequate UF with absolute hemodynamic stability. The reduced incidence of complication with low sodium dialysate therapy was probably because they required less UF.

Angiography with Nonionic X-Ray Contrast Media in Severe Chronic Renal Failure: Renal Function and Contrast Retention

The effects of contrast media on renal function and the cortical retention of contrast media after abdominal angiography were investigated. Sixteen nondiabetic patients with predialytic chronic renal failure received either the nonionic dimeric contrast medium iodixanol or the monomeric contrast medium iohexol in a double-blind randomized study. All patients were well hydrated before, during and after angiography. Mean 99mTc-DTPA clearance was 14.0 ml/min/1.73 m2 in the iodixanol group, and 9.3 ml/min/1.73 m2 in the iohexol group at baseline. No statistically significant changes were seen after angiography. Serum creatinine increased significantly 48 and 72 h after angiography in both groups, and then normalized. Creatinine clearance was reduced only in the iohexol group, at 72-96 h. The urinary excretion of renal enzymes and of total protein did not change significantly. No patients developed oliguria or required dialysis during the 7-day observation period. Increased attenuation in the renal cortex, measured by computed tomography and probably reflecting intracellular retention of contrast medium, peaked at 24 h, and was observed in both groups during the follow-up period. Thus, although transient and minor changes in glomerular filtration rate were noted, both iodixanol and iohexol were safe for use in angiography in nondiabetic patients with severe chronic failure when the patients were well hydrated.

Role for Doppler Ultrasound in the Assessment of Renal Circulation: Effects of Dopamine and Dobutamine on Renal Hemodynamics in Humans

To examine the utility of Doppler ultrasound in assessing renal hemodynamics, we investigated the effects of dopamine and dobutamine on renal blood flow using Doppler ultrasound technique and conventional clearance tests in 7 healthy volunteers. After visualization of arterial blood flow in the right renal hilus by two-dimensional color flow mapping, the phasic blood flow velocity in the vessel was obtained by a pulsed Doppler method. Intravenous infusion of dopamine at a low dose increased the velocity and decreased the waveform pulsatility of renal artery blood flow without causing any significant changes in blood pressure, heart rate, or cardiac index. In contrast, dobutamine infusion increased the peak systolic velocity in a dose-dependent manner, but did not increase the mean velocity or decrease the waveform pulsatility. Percent changes of renal blood flow during infusions of both agents correlated well with those of the mean velocity. Furthermore, the degrees of changes of the waveform pulsatility were consistent with those of renal vascular resistance obtained from clearance tests and blood pressure. Our results suggest that mean velocity reflects renal blood flow and the pulsatility of blood flow waveform represents renal vascular resistance. We conclude that the effects of vasoactive agents on renal blood flow and renal vascular resistance can be estimated noninvasively, directly, and repeatedly using Doppler ultrasound.

Total and Split Renal Function Assessed by Ultrasound Doppler Techniques

We evaluated total and split renal functions from the pattern for renal arterial blood flow detected by ultrasound Doppler in healthy subjects and patients with varying degrees of renal function and disorders other than renovascular hypertension or severe aortic valvular disease. A renal-time pulsed ultrasonic echo-Doppler device at 2.5 MHz was used with a translumbar approach. The ratio of peak diastolic (D) to systolic (S) velocity correlated well with both p-aminohippurate clearance and creatinine clearance. Acceleration time was correlated with the clearance of neither compound. To evaluate the clinical usefulness of ultrasound Doppler in the assessment of split renal function, we compared the D/S ratio with the renal function obtained by radionuclide methods for individuals. The split renal glomerular filtration rate, calculated by a method which makes use of the early renal uptake of 99mTc-diethylenetriam-inepentaacetic acid, correlated well with the D/S ratio. These results indicate that the ultrasonic measurement of renal arterial blood flow by the pulsed Doppler method should be useful for assessment of total and split renal functions.

Oxyphil Cell Function in Secondary Parathyroid Hyperplasia

Oxyphil cell function in secondary parathyroid hyperplasia due to chronic renal failure was evaluated using in situ hybridization and heterotransplantation of parathyroid tissue. In situ hybridization and histologic analysis were performed on continuous frozen sections using 22 parathyroid tissues. A restricted area composed exclusively of oxyphil cells was observed in 10 specimens, and an area of only chief cells was found in 12 specimens. Silver grains demonstrating the existence of parathyroid hormone (PTH) mRNA were 18.8 +/- 7.8 (mean +/- SD) in oxyphil cells while those in chief cells were 17.2 +/- 7.5. PTH mRNA was abundant in both the oxyphil and chief cells. Further analysis of oxyphil cell function was assessed by the heterotransplantation of parathyroid nodules, consisting exclusively of oxyphil or chief cells, into nude mice. The function of these implants was assessed by measuring the concentration of human intact PTH which did not cross-react with mouse PTH. Serum PTH concentrations were correlated with the volume of implanted tissue. Elevations of PTH concentrations were similar in the mice transplanted with oxyphil or chief cells, indicating that both cell types had similar PTH secretory activity. The basic histologic characteristics of both cell types were not altered following transplantation. These results demonstrate that oxyphil cells in secondary parathyroid hyperplasia synthesize and secrete PTH, and that this secretion contributes to the pathophysiology of hyperparathyroidism.

Hypophosphatemia in End Stage Renal Disease

A case of hypophosphatemia in a 55-year-old black female on maintenance hemodialysis is described. She developed multiple bone fractures and congestive heart failure during her 10-year period on hemod

Successful Resection of Pheochromocytoma in a Hemodialysis Patient

A 45-year-old woman with a pheochromocytoma who had been on regular hemodialysis for 4 years and underwent successful surgery is described. Careful preoperative management, including the use of prazosin and weight control, was carried out to prevent severe intraoperative and postoperative cardiovascular complications. Prazosin was given at an initial dose of 0.5 mg/day, and the dosage was increased to 20 mg/day prior to surgery. The increase in intravascular volume led to a gain of 3 kg in body weight. No deterioration of the cardiovascular or respiratory function was caused by these maneuvers, and surgery was performed without significant complications except for a rapid rise of blood pressure during tumor resection. To our knowledge, only one similar case report could be found in the English literature.