Shiro Oguma

The UW solution for canine kidney preservation. Its specific effect on renal hemodynamics and microvasculature.

The preservation effects of UW solution on renal hemodynamics and microvascular systems were studied in canine kidney autografts. In 72-hr UW-preserved kidneys, the microvessels of both cortex and medulla were completely visualized with silicon rubber compound 1 hr after reperfusion. Histology also showed extremely well-preserved arterioglomerular and tubular systems. These results were correlated with good renal blood flow, prompt recovery of posttransplant graft function, and 100% two-week survival of dogs. In contrast, kidneys preserved for 72 hr with Euro-Collins solution showed necrotic and obstructive changes of the microvasculature and deterioration of renal hemodynamics. In 120-hr UW-preserved kidneys, the microcirculation of the medullary region became poor after reflow when there was fairly intact perfusion of the cortical region, indicating an ischemia-related intrarenal blood flow maldistribution. The 120-hr kidneys subsequently failed in spite of having a good blood flow and morphologically well-maintained microvasculature after reperfusion. These data demonstrated that much, but not all, of the beneficial effect of UW solution in kidney preservation might be attributed to its remarkable protection of renal microvasculature. Correction of intrarenal blood maldistribution caused by a discrepancy in tolerance to ischemia of the vascular and tubular systems might be important in successfully preserving the kidney for 120 hr.

Metabolic responses during hemodialysis determined by quantitative 1H NMR spectroscopy

A large proportion of patients with end-stage renal disease have lifelong hemodialysis (HD) treatment. HD rapidly and indiscriminately removes necessary small metabolites together with uremic toxins from plasma into dialysate. To investigate metabolic responses to HD, we determined the levels of metabolites through time-course monitoring of (1)H NMR spectroscopy of dialysate during HD. The dialysate sample is stable for analysis because it contains only small metabolites without proteins. It was collected non-invasively from 9 HD patients with chronic glomerular nephropathy, at 6 time points during 4h of HD in 5 sessions. Creatinine, alanine, lactate, pyruvate and valine were simultaneously quantified on a one-dimensional single-pulse spectrum with a single standard compound. The concentration of creatinine exhibited monotonous decay with time, while that of valine decreased slowly and then maintained its levels throughout an HD. Lactate, alanine and pyruvate increased at 2-3h after the initiation of HD. They exhibited remarkable responses to HD with production from the body. The time-course of change in the 4 metabolites of lactate, pyruvate, alanine, and valine had reproducible behavior unique to each patient during the HD. This finding may be applied to distinguish metabolic status in HD patients.

Plasma Levels of Biotin Metabolites Are Elevated in Hemodialysis Patients with Cramps.

Patients with renal failure undergoing hemodialysis (HD) are susceptible to muscle cramps during and after HD. Muscle cramps are defined as the sudden onset of a prolonged involuntary muscle contraction accompanied by severe pain. Through HD, water-soluble vitamins are drawn out with water. Since biotin, a water-soluble vitamin, plays an essential role as one of the coenzymes in producing energy, we have hypothesized that deficiency of biotin may be responsible for HD-associated cramps. We previously reported that biotin administration ameliorated the muscle cramps, despite the elevated plasma biotin levels before HD and biotin administration, as judged by an enzyme-linked immunosorbent assay (ELISA). However, the ELISA measures not only biotin but also total avidin-binding substances (TABS) including biotin metabolites. In the present study, we determined biotin in HD patients as well as healthy controls, using a newly developed method with ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). The plasma samples were collected from 28 HD patients (16 patients with cramps and 12 patients without cramps) before HD and biotin administration and from 11 controls. The results showed that the accumulation of biotin and TABS in plasma of HD patients compared to controls. Importantly, the levels of biotin metabolites, i.e. TABS subtracted by biotin, increased significantly in patients with cramps over those without cramps. Moreover, the levels of biotin metabolites were significantly higher in patients with a poor response to administered biotin, compared to those with a good response. We propose that accumulated biotin metabolites impair biotins functions as a coenzyme.

A simple and rapid ultra-high-performance liquid chromatography-tandem mass spectrometry method to determine plasma biotin in hemodialysis patients.

A simple, rapid, and selective method for determination of plasma biotin was developed using ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). After single-step protein precipitation with methanol, biotin and stable isotope-labeled biotin as an internal standard (IS) were chromatographed on a pentafluorophenyl stationary-phase column (2.1 × 100 mm, 2.7 μm) under isocratic conditions using 10 mm ammonium formate-acetonitrile (93:7, v/v) at a flow rate of 0.6 mL/min. The total chromatographic runtime was 5 min for each injection. Detection was performed in a positive electrospray ionization mode by monitoring selected ion transitions at m/z 245.1/227.0 and 249.1/231.0 for biotin and the IS, respectively. The calibration curve was linear in the range of 0.05-2 ng/mL using 300 μL of plasma. The intra- and inter-day precisions were all <7.1%. The accuracy varied from -0.7 to 8.2%. The developed UHPLC-MS/MS method was successfully applied to determine plasma biotin concentrations in hemodialysis patients. Copyright