Electropermeabilization is one of the nonviral methods successfully used to transfer genes into living cells in vitro and in vivo . Although this approach shows promise in the field of gene therapy, very little is known about the basic processes supporting DNA transfer. The present investigation studies this process at the single-cell level by using digitized fluorescence microscopy. Permeabilization is a prerequisite for gene transfer. Its assay by propidium-iodide (PI) penetration shows that it occurs at the sides of the cell membrane facing the two electrodes, whereas fluorescently labeled plasmids only interact with the electropermeabilized side of the cell facing the cathode. The plasmid interaction with the electropermeabilized part of the cell surface results in the formation of localized aggregates. These membrane-associated spots are formed only when pulses with a longer duration than a critical value are applied. These complexes are formed within 1 s after the pulses and cannot be destroyed by pulses of reversed polarities. They remain at the membrane level up to 10 min after pulsing. Although freely accessible to DNA dye (TOTO-1) 1 min after the pulses, they are fully protected when the addition takes place 10 min after. They diffuse in the cytoplasm 30 min after pulses and are present around the nucleus 24 h later.

Electrochemotherapy (ECT) enhances the effectiveness of chemotherapeutic agents by administering the drug in combination with short intense electric pulses. ECT is effective because electric pulses permeabilize tumour cell membranes and allow non-permeant drugs, such as bleomycin, to enter the cells. The aim of this study was to demonstrate the anti-tumour effectiveness of ECT with bleomycin on cutaneous and subcutaneous tumours. This article summarizes results obtained in independent clinical trials performed by five cancer centres. A total of 291 cutaneous or subcutaneous tumours of basal cell carcinoma (32), malignant melanoma (142), adenocarcinoma (30) and head and neck squamous cell carcinoma (87) were treated in 50 patients. Short and intense electric pulses were applied to tumours percutaneously after intravenous or intratumour administration of bleomycin. The tumours were measured and the response to the treatment evaluated 30 days after the treatment. Objective responses were obtained in 233 (85.3%) of the 273 evaluable tumours that were treated with ECT. Clinical complete responses were achieved in 154 (56.4%) tumours, and partial responses were observed in 79 (28.9%) tumours. The application of electric pulses to the patients was safe and well tolerated. An instantaneous contraction of the underlying muscles was noticed. Minimal adverse side-effects were observed. ECT was shown to be an effective local treatment. ECT was effective regardless of the histological type of the tumour. Therefore, ECT offers an approach to the treatment of cutaneous and subcutaneous tumours in patients with minimal adverse side-effects and with a high response rate.

The cellular action of growth factors, among them basic fibroblast growth factor (bFGF), is mediated by their interaction with a cell surface receptor, but the mechanism of transfer of mitogenic (or other) signals to the nucleus has not been identified. In this work, we show that bFGF is translocated to and accumulated in the nucleolus. Furthermore, the nucleolar localization of bFGF is correlated with a stimulation of transcription of ribosomal genes during G0----G1 transition induced by bFGF alone in adult bovine aortic endothelial cells (ABAE cells). Stimulation of ribosomal gene transcription is preceded by a significant increase of the major nonhistone nucleolar protein, nucleolin. In vitro, the growth factor has a direct effect on the enhancement of RNA polymerase I activity in isolated nuclei from quiescent sparse (G0) ABAE cells. The direct action of bFGF on the level of ribosomal gene transcription could correspond to an additional growth-signaling pathway, mediated by this growth factor.