Enlarged cell-associated proteoglycans abolish E-cadherin functionality in invasive tumor cells.

Mouse and dog epithelial cell lines, expressing high levels of the Ca(2+)-dependent cell-cell adhesion molecule E-cadherin in vitro, generated invasive and metastatic tumors in athymic mice. From these tumors, neoplastic cell lines were isolated. All ex vivo isolates retained high expression levels of E-cadherin at their surface. Nevertheless, some showed a fusiform morphotype, were defective in Ca(2+)-dependent cell aggregation, and were invasive in vitro, indicating that E-cadherin was not functional. Cell-associated proteoglycans were found to be enlarged in these variants as compared to their counterparts with functional E-cadherin. Treatment of the cells with the drug 4-methylumbelliferyl beta-D-xyloside specifically reduced the amount and size of cell-associated proteoglycans. This same drug induced an epithelial morphotype, increased Ca(2+)- and E-cadherin-dependent cell aggregation, and abrogated invasiveness without influencing E-cadherin expression levels. Our results indicate that enlarged proteoglycans can prevent the homophilic binding of E-cadherin, probably by steric hindrance. This is one more mechanism by which carcinomas may counteract invasion-suppressor genes and acquire malignancy.

Performance of a modified starch hydrophilic matrix for the sustained release of theophylline in healthy volunteers.

Two experimental formulations of theophylline with a hydrophilic starch matrix were evaluated for their sustained-release characteristics after single administration in healthy human volunteers. Theo-dur was chosen as a reference sustained-release formulation. In a first study, the extent of absorption was similar for a syrup, for Theo-dur, and for the experimental formulation of theophylline with 70% drum-dried corn starch as the sustained-release agent (DDCS-70). The maximal plasma concentration (Cmax) was significantly lower, and the time to reach Cmax as well as the time span during which the plasma concentration was at least 75% of the Cmax were significantly higher for Theo-dur than for the DDCS-70 formulation. A sustained-release profile, as for Theo-dur, was not reached for DDCS-70. In a second study the influence of the starch:drug ratio on the bioavailability was investigated. The decrease in starch content from 70 to 50% of the formulation did not improve the plasma concentration-time profile towards a sustained-release profile.

Influence of the sterilization process on alginate dispersions.

The influence of different sterilization procedures on alginate dispersions was studied by measuring viscosity and molecular weight changes. Autoclaving caused a 64% decrease in viscosity. Heating at a low temperature over several cycles was less efficient in sterilizing alginates and there was a progressive breakdown of the alginate chain over the succeeding cycles. Heating during ethylene oxide sterilization also resulted in reduced viscosity and breakdown. Membrane filtration yielded a sterile product with no significant reduction in viscosity or mol. wt.

A fluorescence method for the determination of the molecular weight cut-off of alginate-polylysine microcapsules.

Several applications of microcapsules for the encapsulation of living cells or macromolecules require well defined pore sizes. The molecular weight cut-off of alginate-polylysine microcapsules has been determined using a range of fluorescent labelled dextran molecules. The diffusion of the fluorescein isothiocyanate labelled (FITC)-dextrans into the microcapsules was followed by fluorescence and confocal laser scanning microscopy. The permeability of microcapsules for FITC-dextrans with a molecular weight of 4,700 daltons and the impermeability for FITC-dextrans with a molecular weight of 40,500 daltons was confirmed with both techniques. Determination of the molecular weight cut-off, using confocal laser scanning microscopy was more reliable and required a smaller sample than fluorescence measurements.

Light stability of molsidomine in infusion fluids.

The influence of artificial light, daylight or stimulated sunlight on the stability of molsidomine was investigated. In static experiments an 80 micrograms mL-1 solution of molsidomine in saline was stored in an unprotected infusion bag. During dynamic experiments the molsidomine solution (80 micrograms mL-1) was dropped at 12.5 mL h-1 from an unprotected infusion bag, from an infusion bag covered with aluminium-foil, or from an infusion bag protected with a UV-cover. Either unprotected infusion tubing or infusion tubing with a UV-filter were connected to the infusion bags. Static as well as dynamic experiments showed a half-life of about 20 min for the unprotected molsidomine solutions, when placed behind a window during a sunny day. Protection from light of the infusion bag but not of the infusion tubing had only a minor influence on the drug half-life. Protection of the infusion bag and the infusion tubing with a UV-filter increased the half-life to several days. These results confirm that both the infusion bag and the infusion tubing need adequate light protection during molsidomine administration.

Sorption of isosorbide dinitrate to central venous catheters.

As several studies demonstrated the sorption of isosorbide dinitrate (ISDN) to intravenous delivery systems, a study on the sorption of ISDN to several central venous catheters was performed. Fourteen different catheters were perfused during a 2 h period, under simulated infusion conditions, with ISDN (250 micrograms mL-1) in 0.9% (w/v) sodium chloride. The drug loss to a polyethylene and a silicone catheter was 0.2 and 6.1%, respectively. The sorption to a polyvinylchloride heparin-coated thermodilution catheter was 1-6.9% depending on the length of the catheter. The drug loss to polyurethane catheters of different composition varied between 3.8 and 28.9%. For polyurethane catheters composed of cycloaliphatic polyurethanes an inverse relation was shown between Shore A hardness and sorption.

Host reaction against empty alginate-polylysine microcapsules. Influence of preparation procedure.

Microcapsules, prepared with alginate and polylysine, were injected intraperitoneally into mice and the number of peritoneal leucocytes as well as the cells sticking to the capsule wall were counted after 4-28 days. A significant increase in host reaction was observed when the microcapsules contained an outer layer of polylysine as compared with calcium alginate beads without polylysine or microcapsules coated with an outer layer of alginate. The alginate sources influenced the host reaction significantly. After an intraperitoneal residence of 4 days, the microcapsules were mainly surrounded by macrophages. After 28 days, several cell layers surrounded the microcapsules; macrophages, multinucleate giant cells, fibroblasts and mesothelial cells.

Host reaction against alginate-polylysine microcapsules containing living cells.

Syngeneic and nude mice were injected intraperitoneally with saline, empty microcapsules, aggregates of tumourogenic MO4 cells, encapsulated non-tumourogenic MO cells and encapsulated MO4 cells. The host reaction 4 days after injection, was evaluated by counting the leucocytes in the peritoneal cavity and the cells sticking to recovered microcapsules. A significantly lower number of peritoneal leucocytes was found in mice injected with empty microcapsules or encapsulated MO cells as compared with encapsulated MO4 cells. Histological evaluation showed a significantly higher number of cells sticking to microcapsules containing cells as compared with empty microcapsules. These observations showed that the cellular host reaction against intraperitoneal implants can be evaluated by counting the leucocytes in the peritoneal lavage and histology of recovered capsules.

Bioavailability of two ibuprofen oral paste formulations in fed or nonfed ponies.

The bioavailability and pharmacokinetics of ibuprofen, a nonsteroidal antiinflammatory drug, was studied in healthy Shetland ponies. Ibuprofen was administered IV, as a suspension, and as a solid solution oral paste to ponies from which food was withheld. The suspension paste was also administered to ponies that received hay and water ad libitum. Both formulations had an absolute bioavailability of about 80%. Bioavailability was not influenced by feeding.

Effect of adjuvant administration on weight gain, infectious diseases, and mortality of veal calves.

Young weaned calves are susceptible to viral and bacterial infections. In two studies 279 and 500 weaned calves were injected at random with an adjuvant, with saline or were left untreated in order to evaluate their weight gain or illness and mortality during the first 10 weeks of fattening on a fat-stock farm. However, in contrast to studies reported in the literature, no significantly higher increase in weight gain and no significant improvement of illness and mortality were found with adjuvant treatment.

Near-infrared (NIR) monitoring of H2O2 vapor concentration during vapor hydrogen peroxide (VHP) sterilisation.

PURPOSE: There is an increasing use in the pharmaceutical industry of barrier systems such as transfer isolators, sterilisation tunnels and work station isolators. As Vapor Hydrogen Peroxide (VHP) sterilisation of isolators and lyophilizers becomes an important sterilisation method, there is an acute need for a VHP monitoring system to be used for in-process control and validation. In this study, near infrared (NIR) spectrofotometry was evaluated as a potential technique to monitor hydrogen peroxide. Additionally the H2O2 vapor permeability of different packaging materials, commonly used in steam and ethylene oxide sterilisation, was evaluated. METHODS: NIR spectrofotometry, using a gas cell connected with optic fibres, was evaluated as a potential technique to monitor hydrogen peroxide vapor and water vapor during VHP sterilisation of an isolator. A NIR spectrum was taken every 30 s during VHP sterilisation of an isolator. The influence of injection rate, air flow rate, working temperature and gas distribution was investigated. The H2O2 vapor permeability of different packaging materials was determined by placing the gas cell in the sterilisation bags and sealing the bags hermetically. The sterilisation bag was then subjected to VHP sterilisation. RESULTS: The NIR spectra taken at steady state sterilization conditions showed 4 absorption peaks: at 1364, 1378 and 1400 nm attributed to water and at 1420 nm attributed to H2O2 vapor. By measuring the absorbance level at these wavelengths, the actual concentration of H2O and H2O2 vapor in the isolator was calculated. The water vapor permeation of the sterilisation bags, measured with NIR, appeared to be equal for all materials tested. Whereas Tyvek was the most permeable material for hydrogen peroxide vapor (82.7% of the reference concentration outside the bag), only 30% was found in bags made of medical paper. Sterilisation bags consisting of laminate films and PVC sealed to medical paper showed intermediate permeability. CONCLUSIONS: Near-infrared (NIR) spectroscopy using a gas cell with optic fibres is a useful technique to monitor VHP sterilisation cycles. There was a difference in H2O2 vapor permeability of different packaging materials, commonly used in steam and ethylene oxide sterilisation.

The molecular weight cut-off of microcapsules is determined by the reaction between alginate and polylysine.

Mammalian cells encapsulated in alginate-polylysine microcapsules are used as artificial organs in cancer research and in biotechnology. These applications require microcapsules with a reproducible mol. wt. cut-off. The high cost of the polycation, polylysine, requires an efficient preparation procedure. This article shows that the overall reported contact time of 5 minutes at ambient conditions should be increased several times in order to reach a maximal binding between the calcium alginate beads and 0.1% (w/v) polylysine solutions. An increase of the polylysine concentration from 0.0125% to 0.8% (w/v) resulted in a faster maximal binding, but the amount of polylysine bound increased also. Immersion of calcium alginate beads with a diameter of 750 mum, prepared from 1 mL alginate, in 30 mL of a 0.8% (w/v) polylysine solution, resulted in a polylysine spill of more than 89%. The time required to reach a maximal binding was related to the reaction temperature. The interaction zone between calcium alginate beads and fluorescein isothiocyanate-labeled polylysine solutions was visualized with a confocal laser scanning microscope as a function of time. Microcapsules, prepared at 40 degrees C with 0.1% (w/v) polylysine solutions with mol. wts. between 12 and 249.2 kD, were permeable for fluorescein isothiocyanate-labeled dextran, mol. wt. 4.7, but not for 40.5 kD. Higher polylysine concentrations resulted in a membrane with a mol. wt. cut-off lower than 4.7 kD.

Micro-encapsulation of MDCK-ras-e cells prevents loss of E-cadherin invasion-suppressor function in vivo.

The invasion-suppressor molecule E-cadherin mediates Ca(2+)-dependent cell aggregation and prevents invasion. E-cadherin-positive Madin-Darby canine kidney (MDCK) cells that were non-invasive in vitro formed, upon i.p. injection, tumors that were invasive. Differentiated tubular tumor areas showed an intense immuno-signal for E-cadherin at intercellular contacts, whereas undifferentiated structures did not. Cell lines derived from such tumors turned out to be invasive in vitro and showed decreased Ca(2+)-dependent cell aggregation but no change in E-cadherin immunopositivity. This combination of phenotypes indicated a loss of the E-cadherin invasion-suppressor function. Micro-encapsulation of i.p.-injected cells prevented the loss of the E-cadherin invasion-suppressor function. We concluded that this loss in vivo was dependent upon immediate contacts between tumor cells and host cells or upon host factors that could not cross the capsule membrane.