Immune monitoring in kidney transplant recipients could predict acute rejection by a new method: flow cytometric microcarrier assay.

BACKGROUND: Peripheral blood lymphocytes (PBL) of kidney transplant recipients stimulated in vitro release tumor necrosis factor (TNF)-α and interferon (IFN)-γ into the supernate as detected by a flow cytometric microcarrier assay (FCMA) that we used to predict acute rejection episodes. METHODS: Fifty-two kidney transplant recipients were divided into 2 groups; stable function (STA; n = 30) and acute rejection (ARG; n = 22) for comparison with healthy volunteers (n = 10). PBL were stimulated for 8 hours with phorbol myphnistate acetate and ionomycin, thereafter detecting TNF-α and IFN-γ in culture supernates by FCMA. Receiver operating characteristics (ROC) procedures were used to assess the sensitivity and specificity to predict acute rejection. RESULTS: The fluorescence intensity of TNF-α and IFN-γ in culture supernates was significant higher among healthy controls than STA: 68.38 ± 28.59 vs 51.08 ± 34.05, respectively (P < .05). The intensity of TNF-α and IFN-γ in ARG (144.47 ± 81.21 and 116.61 ± 53.89, respectively) was significant higher than STA (P < .001). The sensitivity and specificity to predict acute rejection were 86.4% and 86.7%, respectively, when analyzed by ROC curves combining TNF-α and IFN-γ. The intensity in noncultured plasma from ARG or STA was significant lower than that in culture supernates from ARG and STA with sensitivity and specificity to predict acute rejection episodes of 63.6% and 73.3%, respectively, when combining TNF-α and IFN-γ. CONCLUSIONS: Monitoring the expression of TNF-α and IFN-γ in cell culture supernates after stimulation of kidney transplant recipient PBL in vitro using FCMA predicted acute rejection episodes.

Establishment of an animal model of biliary ischemic stenosis with clamping in mice.

OBJECTIVE: The objective of this study was to explore a method to establish biliary ischemic stenosis in mice. METHODS: After the optimal time of biliary ischemia was determined, 20 Kunming mice were equally divided into 2 groups. In the experimental group a 0.4-cm length of common bile duct was clamped for 90 minutes with 2 micro-vessel clamps (width = 0.1 cm). The common bile duct was not clamped in the control group. Twenty-one days later, biliary tract visualization was performed in all mice. Blood samples were collected from the inferior vena cava to determine the serum levels of total bilirubin (TBIL) and alanine aminotransferase (ALT). Meanwhile, samples of the common bile duct and liver tissue were extracted for microscopic examination to observe morphological changes. RESULTS: In the experimental group, obvious dilatation of the common bile duct appeared over the clamp site. There was no dilatation of the common bile duct in the control group. Twenty-one days later, serum levels of TBIL and ALT were significantly higher among the experimental compared with the control group. Microscopic examination showed that the part of common bile duct at the clamp site was significantly expanded, with a smaller or occluded bile duct lumen necrotic mucosa with determination, and tubular wall with fibrosis and excrustation. A few dead liver cells and many inflammatory cells were observed in liver tissue samples. CONCLUSIONS: A biliary ischemic stenosis model was established using a clamping method in mice, which may provide a reliable technique for basic and clinical research into mechanisms of biliary ischemic stenosis after liver transplantation.

Sirolimus-based immunosuppressive therapy in liver transplant recipient with tacrolimus-related chronic renal insufficiency.

BACKGROUND: While providing potent immunosuppression for liver transplant recipients, calcineurin inhibitors (CNI) exhibit nephrotoxicity as a major side effect. The purpose of this study was to evaluate the safety and efficacy of conversion from CNI to sirolimus (SRL) among liver transplant recipients with CNI-induced chronic nephrotoxicity. METHODS: Between January 2004 and June 2005, we performed conversion in 16 recipients after a median period of 8.5 months after liver transplantation. The indication for conversion was CNI-related nephrotoxicity with a serum creatinine (sCr) value >132.6 umol/L. Renal function was measured before and after conversion to SRL. Clinical and laboratory data related to the clinical course of the patients were recorded to investigate the safety and efficacy of conversion. RESULTS: Sixteen patients were converted to SRL after developing nephrotoxicity. Their renal function improved gradually after conversion. The levels of sCr decreased significantly within the first 30 days (164.1 +/- 12.48 micromol/L to 130.1 +/- 5.573 micromol/L), and over the next 60 days after conversion (97.86 +/- 11.69 micromol/L to 90.7 +/- 8.95 micromol/L) (P < .01). Similarly, the mean glomerular filtration rate (GFR) increased significantly during the same period. Four recipients experienced hypercholesterolemia, 1 with ankle edema, and 1 with acute rejection. The median follow-up was 2.4 years. No patient discontinued SRL due to side effects. No patient needed dialysis or kidney transplantation during the study period. CONCLUSIONS: SRL is a safe, effective replacement agent as primary immunosuppressive therapy following withdrawal of CNIs in liver transplant recipients with CNI-induced chronic nephrotoxicity.

[Effects of gentamycin on the fetal kidneys].

Gentamycin was given in 24 women during termination of mid-trimester pregnancy. The concentration of gentamycin in the blood of pregnant women, umbilical cords and in tissue of placenta, fetal kidneys were determined by fluorescence polarization immunoassays, and the morphological changes of fetal kidneys were investigated as well. The results indicated that the concentration of gentamycin in circulation of pregnant women was not significantly different from that of the umbilical cords (P greater than 0.05), but in tissue of fetal kidneys gentamycin concentration was remarkably higher than that in placenta (P less than 0.05). The following pathological findings of fetal kidneys were seen: marked congestion in glomeruli, cloudy swelling in the proximal tubules, reduced cilial cells and granular degeneration in epithelium. All these have shown that the placenta did not act as a barrier on gentamycin. It should be noted that while gentamycin in therapeutic dosage was given to pregnant women, it would be accumulated in fetal kidneys and caused damage.

The route of incorporation of nitrogen atom into rifamycin during its biosynthesis.

In this paper, it was first shown that under various conditions of nitrogen supply the rifamycin yield was positively correlated with the mycelial specific activity of glutamine synthetase (GS), then the enhancing effect of glutamine, the product of GS, on rifamycin biosynthesis was demonstrated with resting cell system. The stimulatory effect of glutamine was more pronounced than that of glutamate, and not reduced by a GS specific inhibitor, DON. However, the increase in yield brought about by glutamate, and by asparagine was strongly inhibited by this inhibitor. Glutamine-CO15NH2 amd glutamate-alpha-15NH2 were synthesized and compared for the incorporation of 15N into rifamycin. It was found that the amide nitrogen was incorporated to a much greater extent than the alpha-amino nitrogen, showing that glutamine was a direct precursor of the nitrogen atom in rifamycin. In addition, synthesis of A-32 (C7N), an intermediate secreted by an inactive mutant rif 1, was also greatly stimulated by glutamine, and the synthetic C7N was found to be able to stimulate the biosynthesis of rifamycin. Based upon the above results, the route of incorporation of nitrogen atom into rifamycin is summarized as follows: (Formula: see text).