Masatoshi Sakurada

Analysis of urinary donor-derived DNA in renal transplant recipients with acute rejection.

Abstract: In renal transplantation we usually diagnose an acute rejection by based on the results of a needle biopsy; however, this takes time and findings in some cases are not definite. We analysed the urine of renal recipients for the presence of donor DNA in an attempt to establish a diagnostic means of acute rejection. Sixty‐four renal transplant recipients were examined. Thirty‐seven patients had no trouble after transplantation and 22 patients developed acute rejection, diagnosed based on serum creatinine levels and/or needle biopsy findings of the graft. Five patients had drug‐induced renal dysfunction. In female recipients with a male graft we examined urine for the presence of Y‐chromosome (SRY and DYZ‐1) and in recipients receiving a HLA mismatched graft we investigated the HLA‐DR gene (DRB1) by the polymerase chain reaction (PCR) method. Among female recipients with a male graft there were 14 patients with stable renal function and SRY and DYZ‐1 on Y‐chromosome were negative in 13 (93%) and positive in one, whereas SRY and DYZ‐1 of urine were positive in the four female patients with acute rejection and these DNA fragments disappeared in three after rejection therapy. One patient was subjected to haemodialysis. Among 23 recipients of a graft from HLA mismatched donors with stable renal function, DRB1 was negative in 21(91%). Among 18 patients with acute rejection DRB1 was positive in 16 (93%) and negative in two. These DNA fragments disappeared in 13 patients after rejection therapy. In all patients with drug‐induced renal dysfunction donor‐derived DNA was negative. Presence of donor‐specific DNA in the urine of the recipient is associated strongly with acute rejection and analysis of DNA derived from donor cells in urine might be an effective and accurate method for the diagnosis of acute rejection of a renal transplant.

Isolation of dermatan sulfate with high heparin cofactor II-mediated thrombin-inhibitory activity from porcine spleen

We prepared dermatan sulfate specimens from various porcine tissues, and compared their heparin cofactor II-mediated thrombin-inhibitory activities and chemical natures, including disaccharide composition. Electrophoresis of the specimens on cellulose acetate membrane indicated that spleen dermatan sulfate was the most acidic of the dermatan sulfates prepared from the various porcine tissues. Analysis of the disaccharide units of the dermatan sulfate specimens by high-performance liquid chromatography revealed that spleen dermatan sulfate was rich in 4,6-di-O-sulfated N-acetylgalactosamine residues as compared with those of the other tissues. Spleen dermatan sulfate exhibited the highest thrombin-inhibitory activity, which may be related to its high content of the disulfated N-acetylgalactosamine residue.

Electron leakage from mitochondria and lipid peroxidation after cold ischemia of liver

Abstract In liver transplantation primary graft nonfunction (PGN) occurs in 10–15% of all cases. The cause of PGN is likely associated with free radical and lipid peroxidation at reperfusion, however, the mechanism of PGN is still unknown. We investigated in rats the electron leakage of mitochondria, i.e., free radical generation, and lipid peroxidation of mitochondria after cold ischemia to clarify the mechanism of PGN. After cold ischemia, generation of free radical was recognized, but decreased in proportion to cold ischemic time. The chemiluminescence value increased slightly after 8 h of cold ischemia, however, thereafter gradually decreased. Mitochondrial ATP synthesis also decreased in proportion to cold ischemic time. From these results we concluded that free radical generation occurs while the mitochondria maintain energy synthesis and the generation of free radical decreases with cold preservation. Although mitochondria with cold ischemia suffer lipid peroxidation with ease, leaked electrons from the mitochondria are not a main cause of the lipid peroxidation of mitochondria at reperfusion.

THE BLOOD LEVEL OF ENDOTOXIN AFTER LIVER TRANSPLANTATION IN THE PIG

ブタ肝移植において移植後のエンドトキシン (Et) の変動について検討した. 3時間 (n=5) および12時間保存 (n=2) の移植モデルで全例において血中Etが検出され, 移植後24時間の間に8～380pg/mlのピ-ク値を示し以後漸減した. このEtは従来の合成基質法では陽性に検出されたがβ-glucan感受性因子を選択的に除去したエンドトキシン特異テストでは全く検出されなかった, また移植前後での血液培養は全例が陰性であった. この現象はヒトの肝不全時にみられる, 矢島らのいうところのnon-septic endotoxemiaと同一のものであると考えられた. 移植肝の状態が悪く, 数時間で死亡した例では血中Etが低値を示す傾向が認められ, この意味で生着予後の1つの指標となりうると考えられたが, この現象は従来のspill-over学説では説明困難であった.