Effect of postischemic reperfusion on microcirculation and lipid metabolism of skeletal muscle

To study the effect of ischemia reperfusion injury on microvascular reactivity and tissue metabolism in skeletal muscle, a Sprague-Dawley rat cremaster muscle was prepared as a tourniquet ischemia model and subjected to 2 hr ischemia followed by 1 hr reperfusion to simulate the timing of ischemia during microvascular surgery. The dose-response curve of arteriolar reactivity to norepinephrine, lipid peroxidation, and ultrastructure of capillaries was determined in both the control and postischemic reperfusion stages. Judging from the results, we summarize our observations as follows: (1) Postischemic reperfusion significantly increased arteriolar reactivity to norepinephrine, in which the EC50 for vasoconstriction decreased in all three orders of arterioles. These results suggest that reperfusion could have impaired the vasodilation control mechanism, possibly being endothelium dependent. (2) Lipid peroxidation increased sixfold in the reperfusion group, suggesting that oxygen free radicals have produced significant tissue damage under the created conditions. (3) Significant endothelial damage in the capillaries shown by electron microscope observation supports these studies, indicating that ischemia/reperfusion in clinically transplanted skeletal muscles could cause significant damage to the tissue microcirculation both physiologically and metabolically.     

Validation of 19F-magnetic resonance determination of myocardial blood volume

Site-specific regeneration of the liver after 70% partial hepatectomy was investigated noninvasively in terms of protein synthesis using PET with L-[methyl-11C]methionine in a living animal. Protein synthesis in rat liver at 24 hr after partial hepatectomy did not occur uniformly in the whole liver but intensely in the middle part of the regenerating organ in comparison with the other parts. The activity was significantly low at the posterior aspect of the liver. On the other hand, site-specific protein synthesis was not observed in normal liver. These results were confirmed by bioimaging analyzer system (BAS) analysis, an invasive method that indicates radioactivities of precise intrahepatic sites. DNA synthesis in regenerating liver was also monitored with [2-14C]thymidine and analyzed by BAS 24 hr after 70% hepatectomy. Site-specific DNA synthesis in regenerating liver corresponding to the protein synthesis was also observed by BAS analysis. These results indicate that liver regeneration occurs intensely in the middle part of the liver and that PET enables noninvasive in vivo biochemical analysis.     

The effect of the cyclin-dependent kinase inhibitor olomoucine on cell cycle kinetics

The effect of the cyclin-dependent (CDK) inhibitors olomoucine and roscovitine on cell kinetics was studied. To this end, nonsmall cell lung cancer (NSCLC) cell line MR65 and neuroblastoma cell line CHP-212 were pulse labeled with bromodeoxyuridine (BrdUrd) and chased in culture medium, to which various concentrations of olomoucine or roscovitine were added. A dose-dependent inhibition of the G1/S-phase and G2/ M-/G1 transitions was observed. Furthermore, S-phase progression was also inhibited in a dose-dependent manner. Similarly, roscovitine, another CDK inhibitor with a 10-fold higher efficiency for both CDK1 and CDK2 as compared to olomoucine, showed the same effects at a 10-fold lower concentration. At the highest tested doses both olomoucine (200 microM) and roscovitine (40 microM) induced a complete cell cycle block in both cell lines, paralleled by the appearance of apoptotic figures. In these cultures a decrease in CDK1 protein level was found as shown by Western blotting. Bivariate CDK1/DNA analysis confirmed these observations and showed that a subpopulation of cells with characteristics of apoptosis became CDK1 negative. The presented data suggest that cyclins and CDKs are involved at an important nodal point shared by pathways regulating cellular proliferation and apoptosis.     

Double sodium and potassium sulphates revealed by microprobe analysis in dried cervical mucus: a mid-cycle crystallographic index

The use of physical techniques such as scanning electron microscopy and microprobe analysis permits the analysis of valuable details of the structure of dried samples of mid-cycle cervical mucus during the period of maximum ferning. From a crystallographic point of view, particular attention was paid to the location, morphology and chemical composition of anisotropic organic sulphate microstructures. As distinct from the isotropic sodium chloride pattern described by Papanicolaou, these appear in a narrow period of time corresponding to the middle of the period of maximum ferning. Anisotropic microstructures were revealed to be tiny isolated crystals or polycrystalline aggregates composed of double salts of potassium and sodium. The potassium/sodium cation ratio, equal to 3/1, was constant in the best specimens. Using a simple polarizing microscope, the detection of anisotropic microstructures may be useful to determine the period of maximum fertility more accurately. In addition, experiments have been carried out in order to determine the relative importance of sulphate anisotropic structures as compared with the whole crystallized surface and to investigate precisely their transience after they appear.     

Neural network analysis of combined conventional and experimental prognostic markers in prostate cancer: a pilot study

Radiolabeling permits the detection of trace amounts of zwitterionic detergent remaining in extracted hydrophobic or membrane proteins. To develop a sensitive and specific assay for its presence, the commonly used zwitterionic membrane protein detergent 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (Chaps) was synthesized in a tritiated form. Synthesis via 7-ketodeoxycholic acid gave [7-3H]Chaps in 53% yield with a specific activity of 0.85 mCi/mmol. A novel solvent extraction system for cholic acid obviated the need for chromatographic isolation of this intermediate. The protocol can be readily modified to yield [7-3H]Chaps of higher specific activity. [7-3H]Chaps was used to monitor the efficiency of various strategies for detergent removal from concentrated bacterial culture supernatants containing 0.2% (w/v) Chaps. Dialysis removed 95% of Chaps and the addition of detergent-affinity beads to the dialysis buffer resulted in 97% removal of Chaps. Gel-filtration chromatography removed 99.9% of Chaps, while a detergent-affinity bead chromatography column removed 99.99%. Overall, gel-filtration chromatography was the most convenient and economical method for the one-step removal of the zwittergent from complex biological mixtures.