Jan Egil Melvik

Increase in cis-dichlorodiammineplatinum(II) cytotoxicity upon reversible electropermeabilization of the plasma membrane in cultured human NHIK 3025 cells

A series of brief electrical high-voltage discharges were given to cultured NHIK 3025 cells to render the plasma membrane transiently permeable to drugs. Using [14C]sucrose as an inert marker which normally does not cross plasma membranes, increased permeability could be demonstrated for no longer than 10 min following electrical treatment, indicating that the permeabilization was entirely reversible. The reversibility of the treatment was further demonstrated by a lack of effect on cell growth and colony-forming ability. When cells were given electrical discharges immediately before or during exposure to cis-dichlorodiammineplatinum(II)(cis-DDP) the cytotoxic drug effect increased. By using electrical discharges during a 2 hr drug treatment period the cytotoxicity was enhanced to an extent corresponding to at least a 3-fold increase in drug uptake relative to unpermeabilized cells. This increase in drug uptake was confirmed by direct measurements of the amount of cell-associated Pt by atomic absorbtion spectroscopy. The results suggest that uptake across the plasma membrane may be the rate-limiting factor in the cytotoxic effect of cis-DDP. Furthermore, the methodology applied in the present study may prove useful in assessing the influence of membrane permeability on the effect of other cytotoxic drugs.

Formulation and bacterial phototoxicity of curcumin loaded alginate foams for wound treatment applications: Studies on curcumin and curcuminoides XLII

Curcumin loaded alginate foams are proposed for application in antimicrobial photodynamic therapy of infected wounds. The drug loaded foams were formulated to provide a burst release of the photosensitizer when hydrated. The foams remained intact after hydration and would be possible to remove from the wound prior to irradiation without causing any tissue damage. The characterization of the prepared foams showed that both curcumin loaded and unloaded foams hydrated within 1  min and absorbed from 12 to 16 times their dry weight of a model physiological fluid. Curcumin, the model photosensitizer, has an extremely low solubility in water and may aggregate in aqueous environment. Cyclodextrins (CDs) and polyethylene glycol 400 (PEG 400) were therefore selected as solubilizers of curcumin in the foams to provide a burst release of the photosensitizer. Exposure to the prepared foams in combination with visible light irradiation (∼9.7  J/cm(2)) resulted in >6  log reduction of Entrococcus faecalis cells. However, curcumin mediated photokilling of Escherichia coli was ineffective when CDs were selected as solubilizer of curcumin in the foams. An 81% reduction in viable E. coli cells was detected after treatment with the foam containing PEG 400 as the only solubilizer of curcumin combined with visible light irradiation (∼29  J/cm(2)).

Evaluation of Novel Alginate Foams as Drug Delivery Systems in Antimicrobial Photodynamic Therapy (aPDT) of Infected Wounds—An In Vitro Study: Studies on Curcumin and Curcuminoides XL

The aim of the present study was to incorporate a model water-insoluble photosensitizer, curcumin, in novel alginate foams, further to evaluate the suitability of the curcumin loaded foams in antimicrobial photodynamic therapy of infected wounds. Six foam formulations were prepared and characterized with respect to physical characteristics, in vitro release and storage- and photo-stability of curcumin. One formulation was sterilized (gamma-sterilization). The foams contained hydroxypropyl-beta-cyclodextrins or hydroxypropyl-gamma-cyclodextrins as solubilizers of curcumin. A reference foam without cyclodextrins was prepared with PEG 400 as the solubilizer. At a curcumin load of 0.153% (w/w), the water insoluble photosensitizer was uniformly distributed in the hydrophilic foams matrix. All foams were easy to handle, flexible and hydrated rapidly in a model physiological fluid. Release of curcumin in its monomeric form was demonstrated in vitro and found to be dependent on the type and amount of cyclodextrins in the formulation. Curcumin was stable during storage, but susceptible to photodegradation in the foams, especially when the formulations contain PEG 400 or hydroxypropyl-gamma-cyclodextrins. Curcumin did not degrade after gamma-sterilization, however a decrease in the in vitro release rate of curcumin and changes in the foams physical characteristics were detected.

DNA flow cytometry in human testicular cancer.

DNA flow cytometry revealed aneuploid tumour stemlines in 19 of 20 primary testicular cancers without significant difference of the ploidy values between seminomas and non-seminomas. In 7 of 8 analyzable histograms the S-phase activity was 22-51%. A metastatic mature teratoma had 6% cells in S-phase. These results support the clinical observation that testicular cancer is usually a rapidly growing human tumour. The high percentage of aneuploidy in testicular cancer may be of clinical value in the diagnosis of this malignancy.

Reduction of cis-dichlorodiammineplatinum-induced cell inactivation by methionine

Abstract The effect of cis-dichlorodiammineplatinum(II) (cisplatin) in combination with methionine was investigated on human NHIK 3025 cells cultivated in vitro. Simultaneous treatment with cisplatin and high dose methionine was less lethal by means of colony-forming ability than treatment with cisplatin alone. This reduction in cell inactivation was seen only when methionine was present during the exposure with cisplatin and was strongly dependent upon the concentration of methionine. Cells pretreated with methionine before treatment with cisplatin were found to be more sensitive than cells treated with cisplatin alone. The protective effect of methionine was also compared with benzaldehyde since we have previously found that this drug may inhibit the uptake of cisplatin through the cell membrane. The data suggest that methionine and benzeldehyde protect against cisplatin cytotoxicity independently of each other and by entirely different mechanisms.

Flow cytometric DNA measurements in paraffin-embedded bladder carcinoma tissue before and after precystectomy radiotherapy

From 1976 through 1983, 48 patients with T2/T3 bladder carcinomas were treated with pre-operative radiotherapy (46 Gy) and total cystectomy. DNA histograms were recorded by flow cytometry (FCM) from all pre-treatment paraffin-embedded tumour biopsies and, if still present after radiotherapy, from tumour tissue from the cystectomy specimen. Five patients were excluded from the study because no DNA histograms could be recorded due to extensive destruction of the cell nuclei during preparation. Thus, 43 patients are completely evaluable. Before the start of radiotherapy, 32 bladder carcinomas were interpreted as non-diploid, whereas 11 were diploid. Non-diploidy of the pre-treatment biopsy was associated with radiotherapy-induced stage reduction (p = 0.13). The survival rates in patients with diploid and non-diploid tumours were 65% and 45% respectively at 4 years (p = 0.13). Although not statistically significant, our results suggest that DNA-FCM may provide clinically relevant information about the tumour biology and response to radiotherapy concerning bladder carcinomas.

Rheological characterization of an injectable alginate gel system

BackgroundThis work investigates a general method for producing alginate gel matrices using an internal mode of gelation that depends solely on soluble alginate and alginate/gelling ion particles. The method involves the formulation of two-component kits comprised of soluble alginate and insoluble alginate/gelling ion particles. Gelling kinetics, elastic and Young’s moduli were investigated for selected parameters with regard to soluble alginate guluronate content, molecular weight, calcium or strontium gelling ions and alginate gelling ion particle sizes in the range between 25 and 125 micrometers.ResultsBy mixing the two components and varying the parameters mentioned above, alginate gel matrices with tailor-made viscoelastic properties and gelling kinetics were obtained. Final gel elasticity depended on alginate type, concentration and gelling ion. The gelling rate could be manipulated, e.g. through selection of the alginate type and molecular weight, particle sizes and the concentration of non-gelling ions.ConclusionsFormulations of the injectable and moldable alginate system presented have recently been used within specific medical applications and may have potential within regenerative medicine or other fields.

Oxygen- and temperature-dependent cytotoxic and radiosensitizing effects of cis-dichlorodiammineplatinum(II) on human NHIK 3025 cells in vitro

The radiosensitizing effect of the chemotherapeutic drug cis-dichlorodiammineplatinum(II) (cis-DDP) was tested on human NHIK 3025 cells cultivated in vitro. cis-DDP was found to exert a radiomodifying effect under hypoxic but not under aerobic conditions. These results confirm that cis-DDP may act as a radiosensitizer of hypoxic cells; however, the radiosensitizing effect was seen only at concentrations of cis-DDP having a considerable cytotoxic activity, and for practical reasons concerning survival level the highest drug concentration that was investigated was 15 microM at 37 degrees C. The radiosensitizing effect was of a dose-modifying type and with a dose-modifying factor (DMF) of 1.2 at 15 microM in hypoxic cells. The radiosensitizing as well as the cytotoxic effect of cis-DDP was found to be strongly temperature dependent. Isoeffect doses of cis-DDP was reduced with a factor of 3 at 22 as compared to 37 degrees C. We also found that hypoxic cells were less sensitive to cis-DDP than cells treated in the presence of oxygen. To test the correlation between cytotoxicity and radiosensitization on the one hand and cellular uptake of cis-DDP on the other, cell-associated Pt was measured by atomic absorption spectroscopy. From these studies the cytotoxicity of cis-DDP at 22 and 37 degrees C under aerobic conditions was found to be the same as long as the amount of cell-associated Pt (i.e., the cellular uptake) was the same. However, whether the cells were treated under hypoxic or aerobic conditions, the cellular uptake of Pt was the same. While the radiosensitizing effect was present at 37 and at 40 degrees C, no such effect could be found at 22 degrees C. Since the cytotoxicity of cis-DDP as well as the drug uptake was reduced about three times at 22 as compared to 37 degrees C, we increased the concentration threefold, to 50 microM at 22 degrees C. Still no radiosensitizing effect was found at this temperature.

Oxygen consumption in T‐47D cells immobilized in alginate

Encapsulation or entrapment of cells is increasingly being used in a wide variety of scientific studies for tissue engineering and development of novel medical devices. The effect on cell metabolism of such systems is, in general, not well characterized. In this work, a simple system for monitoring respiration of cells embedded in 3‐D alginate cultures was characterized.

Microenvironment‐dependent respiration of T‐47D cells cultured in alginate biostructures

The main objective of this paper was to investigate whether the oxygen consumption rate (OCR) of cells entrapped in alginate hydrogels depends on presence of soluble factors present in foetal bovine serum (FBS).