Ultrastructural localization of adhalin in normal murine skeletal myofiber

The ultrastructural localization of adhalin and its relations to dystrophin, β-dystroglycan, and β-spectrin were studied in normal murine skeletal myofibers. The C-terminal peptides of adhalin and β-dystroglycan were synthesized based on their cDNAs, and the affinity-purified antibodies against these peptides were produced. Single-immunolabeling electron microscopy showed that the adhalin was located just inside the muscle plasma membrane or inside the myofiber a short distance from the plasma membrane. The adhalin signal was also noted at the sarcoplasmic side of plasmalemmd invaginations or at vesicular structures in subsarcolemmal areas. Double-immunogold-labeling electron microscopy disclosed a similar localization of dystrophin, β-dystroglycan, and β-spectrin. The close association of adhalin with dystrophin or β-dystroglycan was demonstrated by formation of doublets by signals of antibodies of adhalin with those of dystrophin or β-dystroglycan and was confirmed by statistical analyses. This study demonstrated that the location of adhalin is close to that of dystrophin and β-dystroglycan at the muscle plasma membrane.     

Effectiveness of short-contact topical tretinoin in promoting wound healing in db/db mice.

We have investigated the effects of short-contact topical application of tretinoin using 16 male db/db mice by creating two full-thickness wounds on the skin of the back. A 0.1% tretinoin aqueous gel was applied to one wound for five minutes daily for five successive days while only aqueous gel was applied to the other (control). The mean (SD) percentage surface area unhealed before and after treatment were 0.88 (0.3) and 0.64 (0.15). In tretinoin-treated mice and non-treated mice (controls), the mean (SD) thicknesses of granulation were 1.383 (697) micro m and 683 (413) micro m, the density of capillary vessels in granulation was 12.2 (5.5)% and 5.7 (3.9)%, respectively. Differences between the two groups were significant for each variable. Wound healing was accelerated with short-contact topical application of tretinoin in db/db mice.     

Intracellular free magnesium of red blood cells in patients with renal disease.

While serum magnesium (Mg) level is increased in patients with end-stage renal disease (ESRD), it is decreased in renal transplant recipients (TR) receiving ciclosporin. This study was performed to examine the cation metabolism of red blood cells (RBC) in these patients. Intracellular free Mg was measured with 31P-nuclear magnetic resonance spectrometry, and ouabain-sensitive sodium (Na) efflux rate (Eos) was measured from the increase in RBC-Na concentration when RBC were incubated in the presence of ouabain. The ouabain-sensitive Na efflux rate constant (ERCos) was obtained by dividing Eos by RBC-Na concentration. RBC free Mg and ERCos were significantly higher in the TR group than in the ESRD group. There was a significant correlation between RBC free Mg and ERCos (r = 0.474, p less than 0.01). These results support the views that the regulation mechanism for intracellular free Mg is different from that for extracellular Mg in patients with renal disease, and intracellular free Mg modulates Na pump activity of RBC.     

The influence of reflux of bile and pancreatic juice on gastric carcinogenesis in rats

The effect of reflux of the duodenal contents on the development of gastric stump carcinoma without carcinogen in rats was studied. At the first experiment, seventy male Wistar rats were subjected to gastrojejunostomy for the diversion of duodenal contents into the resected stomach through the afferent and the efferent loops. The incidence of adenocarcinoma in the afferent anastomosis were 0/8 (0%) at the 10th week, 3/16 (18.8%) at the 20th week and 7/20 (35%) at the 40th week, so the incidence was apt to rise in parallel to the time. In the second experiment, ninety rats were divided into 4 groups: Group I received the same operation as the first experiment, group II received the operation with Braun's anastomosis, group III received the operation for reflux of bile alone to the remnant stomach, and group IV received the operation for reflux of pancreatic juice alone. Animals surviving after surgery were sacrificed at the 40th week. The incidence were 33.3% in group I, 0% in group II, 0% in group III, and 8.3% in group IV, respectively. These findings suggest that reflux of duodenal contents (both bile and pancreatic juice) is an important factor in the gastric stump carcinogenesis.     

Disposition Characteristics of Macromolecules in Tumor-Bearing Mice

As part of the strategy for the design of macromolecular carriers for drug targeting, the disposition characteristics of macromolecules were studied in mice bearing tumors that served as target tissues. Eight kinds of macromolecules including four polysaccharides and four proteins with different molecular weights and electric charges were used; tissue distribution and tumor localization after intravenous injection were studied. Pharmacokinetic analysis revealed that the tissue radioactivity uptake rate index calculated in terms of clearance was different among the tested compounds; especially, the urinary radioactivity excretion clearances and the total hepatic radioactivity uptake clearances varied widely. Compounds with low molecular weights (approximately 10 kD) or positive charges showed lower tumor radioactivity accumulation; radioactivity was rapidly eliminated from the plasma via rapid urinary excretion or extensive hepatic uptake, respectively. On the other hand, large and negatively charged compounds, carboxymethyl dextran, bovine serum albumin, and mouse immunoglobulin G, showed higher radioactivity accumulation in the tumor (calculated total amounts were 15.6, 10.8, and 20.8% of the dose, respectively) and prolonged retention in the circulation. These results demonstrated that the total systemic exposure rather than the uptake rate index was correlated with total tumor uptake. Molecular weight and electric charge of the macromolecules significantly affected their disposition characteristics and, consequently, determined radioactivity accumulation in the tumor. It was concluded that a drug–carrier complex designed for systemic tumor targeting should be polyanionic in nature and larger than 70,000 in molecular weight.