No abstract

The function and regulation of keratin 8 (K8) and 18 (K18), intermediate filament (IF) proteins of the liver, are not fully understood. We employed the liver damage induced by microcystin-LR (MC-LR), a liver-specific inhibitor of type-1 and type-2A protein phosphatases, in normal and in keratin assembly-incompetent mouse strains as a model to elucidate the roles of IF phosphorylation in situ. The mouse strains used were wild-type (wt) mice and mice with abnormal filament assembly, caused by a targeted null mutation of the K8 gene or caused by expression of a point-mutated dominant negative human K18. In vivo 32 P-labeled wt mice, subsequently injected with a lethal dose of MC-LR, showed hyperphosphorylation, disassembly, and reorganization of K8/K18, in particular K18, indicating high phosphate turnover on liver keratins in situ. At lethal doses, the keratin assembly-incompetent mice displayed liver lesions faster than wt mice, as indicated histopathologically and by liver-specific plasma enzyme elevations. The histological changes included centrilobular hemorrhage in all mouse strains. The assembly-incompetent mice showed a marked vacuolization of periportal hepatocytes. Indistinguishable MC-LR-induced reorganization of microfilaments was observed in all mice, indicating that this effect on microfilaments is not dependent on the presence of functional K8/K18 networks. At sublethal doses of MC-LR, all animals had the same potential to recover from the liver damage. Our study shows that K8/K18 filament assembly is regulated in vivo by serine phosphorylation. The absence or occurrence of defective K8/K18 filaments render animals more prone to liver damage, which supports the previously suggested roles of keratin IFs in maintenance of structural integrity. (HEPATOLOGY 1998;28: 116-128.)The intermediate filament (IF) proteins of hepatocytes consist of the simple epithelial IFs, keratin 8 (K8; type II) and keratin 18 (K18; type I). K8 and K18 form obligate heteropolymers assembling into complex filamentous networks spanning the hepatocyte cytoplasm.

Propofol, currently available as a lipid emulsion (Intralipid loo/), is widely used for induction and maintenance of general anaesthesia [ 11. We have recently evaluated the effects of total intravenous anaesthesia with propofol on the immune response during minor surgery [2] and found increased percentages of T helper lymphocytes in the blood after propofol anaesthesia but not after conventional balanced anaesthesia with thiopentone and nitrous oxide. T helper cells are a subset of T lymphocytes pivotal in augmenting both cell-mediated [3] and humoral immunity [4]. Major surgery causes changes in the immune response which are related to the extent of surgical trauma [5] and the neuroendocrine stress response [6]. In this study we compared the effects of propofol infusion anaesthesia and combined isoflurane anaesthesia on the immune response to abdominal hysterectomy. Patients and methodsWe studied 30 women (median age 47 years; range 36-65 years), ASA grades 1 or 2 scheduled for elective abdominal hysterectomy. The patients were allocated randomly to two groups. Patients with immunosuppressive treatment, malignant disease or marked obesity (weight exceeding the ideal body weight by 30% or more) were not studied.

The natural folic acid activity (FAA) in the serum, on Sephadex® G-200 gel filtration, is divided into 3 fractions: the first in the band of largemolecular proteins; the second in that of small-molecular proteins; and the third outside the protein band. The second fraction, in the present study, was purified by hydroxyapatite chromatography, with the result that FAA emerged over a narrow area of fractions. In immunoelectrophoresis anddisc electrophoresis this area contained only albumin

Background:Microalbuminuria is an established early marker of diabetic nephropathy and an important cardiovascular risk factor in diabetes and hypertension. We aimed to develop a rapid point-of-care assay for the measurement of urine albumin. Methods: The competitive homogeneous assay used an albumin-specific monoclonal antibody labeled with a stable fluorescent europium chelate as donor and an albumin labeled with cyanine 5 (Cy5) as acceptor. The assay was performed at room temperature in single microtitration wells that contained all the required dryform reagents. The close proximity between the two labels in the immune complex allowed fluorescence resonance energy to be transferred from the pulseexcited europium chelate to the acceptor Cy5. The emission of long-lived energy transfer signal from the sensitized Cy5 was measured at 665 nm with time-resolved fluorometry that eliminated short-lived background. Results: The assay procedure required 12 min for a 10-L urine sample. The working range was from 10 to ϳ320 mg/L, and the lower limit of detection was 5.5 mg/L. The within-and between-run CVs were 6.9 -10% and 7.5-13%, respectively. Recovery was 103-122%. The assay correlated well (r 2 ‫؍‬ 0.98; n ‫؍‬ 37) with a laboratory-based immunoassay, although mean (SD) results were 7 (29)% lower. Conclusions: The speed and ease of performance of this assay recommend it for near-patient use. The assay is the first to combine a fluorescence resonance energy transfer-type rapid competitive assay with an all-in-one dry reagent.