Yoshiyuki Shiroyanagi

Species difference in the inhibitory effect of nonsteroidal anti-inflammatory drugs on the uptake of methotrexate by human kidney slices.

Simultaneous use of nonsteroidal anti-inflammatory drugs (NSAIDs), probenecid, and other drugs has been reported to delay the plasma elimination of methotrexate in patients. Previously, we have reported that inhibition of the uptake process cannot explain such drug-drug interactions using rats. The present study quantitatively evaluated the possible role of the transporters in such drug-drug interactions using human kidney slices and membrane vesicles expressing human ATP-binding cassette (ABC) transporters. The uptake of methotrexate by human kidney slices was saturable with a Km of 45 to 49 μM. Saturable uptake of methotrexate by human kidney slices was markedly inhibited by p-aminohippurate and benzylpenicillin, but only weakly by 5-methyltetrahydrofolate. These transport characteristics are similar to those of a basolateral organic anion transporter (OAT) 3/SLC22A8. NSAIDs and probenecid inhibited the uptake of methotrexate by human kidney slices, and, in particular, salicylate, indomethacin, phenylbutazone, and probenecid were predicted to exhibit significant inhibition at clinically observed plasma concentrations. Among ABC transporters, such as BCRP/ABCG2, multidrug resistance-associated protein (MRP) 2/ABCC2, and MRP4/ABCC4, which are candidates for the luminal efflux of methotrexate, ATP-dependent uptake of methotrexate by MRP4-expressing membrane vesicles was most potently inhibited by NSAIDs. Salicylate and indomethacin were predicted to inhibit MRP4 at clinical plasma concentrations. Diclofenac-glucuronide significantly inhibited MRP2-mediated transport of methotrexate in a concentration-dependent manner, whereas naproxen-glucuronide had no effect. Inhibition of renal uptake (via OAT3) and efflux processes (via MRP2 and MRP4) explains the possible sites of drug-drug interaction for methotrexate with probenecid and some NSAIDs, including their glucuronides.

Multiple Human Isoforms of Drug Transporters Contribute to the Hepatic and Renal Transport of Olmesartan, a Selective Antagonist of the Angiotensin II AT1-Receptor

Olmesartan, a novel angiotensin II AT1-receptor antagonist, is excreted into both bile and urine, with minimal metabolism. Because olmesartan is a hydrophilic anionic compound, some transporters could be involved in its hepatic and renal clearance. In this study, we characterized the role of human drug transporters in the pharmacokinetics of olmesartan and determined the contribution of each transporter to the overall clearance of olmesartan. Olmesartan was significantly taken up into human embryonic kidney 293 cells expressing organic anion-transporting polypeptide (OATP) 1B1, OATP1B3, organic anion transporter (OAT) 1, and OAT3. We also observed its saturable uptake into human hepatocytes and kidney slices. Estimated from the relative activity factor method and application of specific inhibitors, the relative contributions of OATP1B1 and OATP1B3 to the uptake of olmesartan in human hepatocytes were almost the same, whereas OAT3 was predominantly involved in its uptake in kidney slices. The vectorial transport of olmesartan was observed in OATP1B1/multidrug resistance-associated protein (MRP) 2 double transfectants, but not in OATP1B1/multidrug resistance (MDR) 1 and OATP1B1/breast cancer resistance protein (BCRP) transfectants. ATP-dependent transport into membrane vesicles expressing human MRP2 and MRP4 was clearly observed, with Km values of 14.9 and 26.2 μM, respectively, whereas the urinary excretion of olmesartan in Mrp4-knockout mice was not different from that of control mice. We also investigated the transcellular transport of olmesartan medoxomil, a prodrug of olmesartan. Vectorial basal-to-apical transport was observed in OATP1B1/MRP2, OATP1B1/MDR1 double, and OATP1B1/BCRP double transfectants, suggesting the possible involvement of MRP2, MDR1, and BCRP in the limit of intestinal absorption of olmesartan medoxomil. From these results, we suggest that multiple transporters make a significant contribution to the pharmacokinetics of olmesartan and its prodrug.

Urothelium regeneration using viable cultured urothelial cell sheets grafted on demucosalized gastric flaps.

Most of the papers in this section are studies of cellular biology in urological cancer. Each of the four papers assess a particular aspect of four different types of cancer: prostate, bladder, renal, and testicular. The other two reports are on unrelated issues; in the first, the authors write about urothelial regeneration using viable cultured urothelial cell sheets grafted onto demucosalized gastric flaps, and in the second, on the in vivo expression of nitric oxide synthase.

Transplantable Urothelial Cell Sheets Harvested Noninvasively from Temperature-Responsive Culture Surfaces by Reducing Temperature

Augmentation cystoplasty using gastrointestinal flaps may induce severe complications such as lithiasis, urinary tract infection, and electrolyte imbalance. The use of viable, contiguous urothelial cell sheets cultured in vitro should enable us to avoid these complications. Transplantable urothelial cell sheets were obtained by utilizing a temperature-responsive cell culture method, and then examined by immunostaining and electron microscopy. Canine urothelium was produced on the surfaces of temperature-responsive culture dishes covalently bonded with the thermally sensitive polymer, poly(N-isopropylacrylamide). Stratified urothelial cell sheets were cultured and then harvested intact without enzymatic treatment from these dishes by reducing the temperature. Histological structure and cell-to-cell junctions were compared between these urothelial cell sheets and those harvested with dispase. All urothelial cell sheets were harvested from the bonded surfaces by reducing the culture temperature without the need for dispase. Electron microscopy revealed well-developed microridge, microvilli, and cell junction complexes. Conversely, these same cell features were destroyed by dispase treatment. Immunoblotting revealed that dispase fragmented occludin, whereas it remained unchanged in the intact urothelial cell sheets. Novel urothelial cell sheets obtained by culture on temperature-responsive culture surfaces were successfully harvested much less destructively than with dispase. This technology should prove useful in urinary tract tissue engineering in the near future.

Characterization of the Uptake of Organic Anion Transporter (OAT) 1 and OAT3 Substrates by Human Kidney Slices

The activities of renal multispecific organic anion transporters (OATs) 1 and 3 have not been fully evaluated in human kidneys. In the present study, the uptake of some organic anions was characterized in kidney slices from human intact renal cortical tissues: hOAT1 and hOAT3 substrates [p-aminohippurate (PAH) and 2,4-dichlorophenoxyacetate (2,4-D)] and hOAT3 substrates [benzylpenicillin (PCG), dehydroepiandrosterone sulfate (DHEAS), and estrone sulfate (ES)]. Despite large interbatch differences, hOAT1 and hOAT3 mRNA levels correlated well, and there was a good correlation between the uptake of PAH and PCG by kidney slices. The uptake of organic anions by kidney slices was saturable with Km values of 31 to 48 μM for PAH, 0.73 to 4.9 μM for 2,4-D, 14 to 90 μM for PCG, and 9.2 to 11 μM for ES. These parameters were comparable with those for hOAT1 and/or hOAT3. The uptake of DHEAS consists of two saturable components with Km values of 2.2 to 3.9 and 1300 μM, and the Km value of the high-affinity component was close to that for hOAT3. Furthermore, PAH more potently inhibited the uptake of 2,4-D than that of PCG and DHEAS. PCG had a weaker effect on the uptake of PAH and 2,4-D than expected from its Km value. Taken together, it is likely that the uptake of PAH and 2,4-D is due to OAT1, and the uptake of PCG and ES and part of DHEAS uptake are due to OAT3 in human kidney slices. Human kidney slices are useful tools for characterizing the renal uptake of drugs.

Factors associated with failure of extracorporeal shock‐wave lithotripsy for ureteral stones using Dornier lithotripter U/50

Background : In the present series of 170 patients who underwent extracorporeal shock‐wave lithotripsy (SWL) treatment for ureteral stones, the authors determine which patients with ureteral stones had an unsuccessful outcome.

Nephron-sparing tumorectomy for a large benign renal mass : A case of massive bilateral renal angiomyolipomas associated with tuberous sclerosis

Abstract A case of massive bilateral angiomyolipomas (AML) associated with tuberous sclerosis in a 33‐year‐old woman is reported. She was hospitalized because she had been experiencing abdominal fullness and epigastralgia. Several imaging studies revealed massive bilateral renal tumors and she was diagnosed as having renal AML associated with tuberous sclerosis. Left nephrectomy was carried out after renal arterial embolization for intratumor hemorrhage. Two years after left nephrectomy, nephron‐sparing surgery (tumorectomy) for right AML was done because of an increase in the size of the right renal AML and she hoped for a future pregnancy. The left kidney with AML weighed 5700u2003g and the right AML weighed 1700u2003g. Postoperative serous creatinine did not differ from that before operation and an increase in the size of the residual tumor was not observed 8u2003months after operation. We consider that tumorectomy is an effective therapy in patients with a very large tumor involving a solitary kidney.