Pulsed Electric Fields Induce Extracellular Matrix Remodeling through Matrix Metalloproteinases Activation and Decreased Collagen Production.

Impairment of extracellular matrix remodeling is observed in the tumor microenvironment or fibrosis and results in excessive collagen production and/or decreased degradation by matrix metalloproteinases (MMPs). Thanks to their local application and transient effects, physical stimuli appear as attractive tools to remodel the extracellular matrix. We assessed the potential of pulsed electric field technology, classically applied to drug delivery, to induce collagen remodeling at the tissue scale. A sophisticated in vitro tissue-engineered human dermal substitute was used to show that microsecond and millisecond pulsed electric fields induced (i) a rapid modulation (4 hours after electrostimulation) of mRNA genes composing the matrisome, particularly a downregulation of procollagens and extracellular matrix maturation enzymes such as transglutaminase 2 and lysyl oxidase like; (ii) a transient decrease in procollagens production and hydroxyproline tissue content within a week after electrostimulation; (iii) a long-lasting ROS-dependent overactivation of matrix metalloproteinases for at least 48 hours; and (iv) a downregulation of TGFß1. These observations underpin that pulsed electric fields, a technology already approved for clinical use combined with anticancer agents, are particularly promising to provide local and effective treatment of abnormal extracellular matrix.

Electroporation does not affect human dermal fibroblast proliferation and migration properties directly but indirectly via the secretome.

Aesthetic wound healing is often experienced by patients after electrochemotherapy. We hypothesized that pulsed electric fields applied during electrochemotherapy (ECT) or gene electrotransfer (GET) protocols could stimulate proliferation and migration of human cutaneous cells, as described in protocols for electrostimulation of wound healing. We used videomicroscopy to monitor and quantify in real time primary human dermal fibroblast behavior when exposed in vitro to ECT and GET electric parameters, in terms of survival, proliferation and migration in a calibrated scratch wound assay. Distinct electric field intensities were applied to allow gradient in cell electropermeabilization while maintaining reversible permeabilization conditions, in order to mimic in vivo heterogeneous electric field distribution of complex tissues. Neither galvanotaxis nor statistical modification of fibroblast migration were observed in a calibrated scratch wound assay after application of ECT and GET parameters. The only effect on proliferation was observed under the strongest GET conditions, which drastically reduced the number of fibroblasts through induction of mitochondrial stress and apoptosis. Finally, we found that 24 h-conditioned cell culture medium by electrically stressed fibroblasts tended to increase the migration properties of cells that were not exposed to electric field. RT-qPCR array indicated that several growth factor transcripts were strongly modified after electroporation.

Drug Release by Direct Jump from Poly(ethylene-glycol-b-ε-caprolactone) Nano-Vector to Cell Membrane.

Drug delivery by nanovectors involves numerous processes, one of the most important being its release from the carrier. This point still remains unclear. The current work focuses on this point using poly(ethyleneglycol-b-ε-caprolactone) micelles containing either pheophorbide-a (Pheo-a) as a fluorescent probe and a phototoxic agent or fluorescent copolymers. This study showed that the cellular uptake and the phototoxicity of loaded Pheo-a are ten times higher than those of the free drug and revealed a very low cellular penetration of the fluorescence-labeled micelles. Neither loaded nor free Pheo-a displayed the same cellular localization as the labeled micelles. These results imply that the drug entered the cells without its carrier and probably without a disruption, as suggested by their stability in cell culture medium. These data allowed us to propose that Pheo-a directly migrates from the micelle to the cell without disruption of the vector. This mechanism will be discussed.

trans- and cis-(Cl,Cl)-[RuII(FT)Cl2(NO)](PF6): promising candidates for NO release in the NIR region.

cis- and trans-(Cl,Cl)-[RuII(FT)Cl2(NO)](PF6) complexes show efficient NO photodelivery upon two-photon excitation in the NIR region. Moreover, cytotoxicity and phototoxicity studies provide evidence that these complexes are promising candidates as photoactivatable molecular tools for resection of malignancies.

Chemical composition and antioxidant, anti-inflammatory, and antiproliferation activities of pomegranate (Punica granatum) flowers.

The chemical composition, antioxidant (DPPH and ABTS assays), anti-inflammatory (5-LOX), and cytotoxic (MCF-7) activities from flowers of seven pomegranate varieties (Punica granatum) were investigated. The highest phenolics (330.9±11.3 mg gallic acid equivalent/g dry weight (dw)), flavonoids (29.5±0.8 mg quercetin equivalent/g dw), tannins (30.6±0.6 mg catechin equivalent/g dw), and anthocyanins (0.70±0.03 mg cyanidin-3-glucoside equivalent/g dw) content were determined in the Chetoui (CH) variety. It was found that Garsi (GR) (IC50=4.9±0.2 mg/L by ABTS assay) and Zaghwani (ZG) (IC50=3.9±0.2 mg/L by ABTS assay) varieties exhibited the highest antioxidant activity. For the anti-inflammatory activity, all varieties were active; the ZH variety was the strongest (2.5±0.1 mg/L). The CH, ES, and RA pomegranate varieties were not active against human breast cancer cells MCF-7, whereas inhibition was more evident with extracts from ZH and GR varieties (IC50=33.00±2.64 and 35.00±4.58 mg/L, respectively). Statistical analysis showed that the variety factor influenced significantly (P<.01) the chemical composition and biological activities of pomegranate flowers.

Preparation and evaluation of microporous organogel scaffolds for cell viability and proliferation.

Various porous scaffolds utilizing an organogel were prepared by particulate-leaching method. The porous organogels were made of biodegradable, non-toxic ingredients like soybean oil or caprylic/capric triglyceride as the organic liquids and 12-hydroxystearic acid as the gelator. The scaffolds possessed an effective porosity of 56-65%, and good pore interconnectivity with an average pore size from 220 to 290mum. The biodegradability of such materials was evaluated and lipases were able to totally degrade the scaffolds. The porosity of the material associated with high draining led to suitable scaffolds which were evaluated for CHO cell viability and proliferation using the MTT test. This evaluation was performed over a period of 3 weeks and showed a greater ability to promote cell proliferation for the soybean oil based scaffold than for the caprylic/capric triglyceride one. The histological investigations revealed that this scaffold was able to promote cell colonization and attachment and could induce the production of collagen.

Evaluation of new thiadiazoles and benzothiazoles as potential radioprotectors: free radical scavenging activity in vitro and theoretical studies (QSAR, DFT).

Thiol and aminothiol compounds are among the most efficient chemical radioprotectors. To increase their efficiency, we synthesized two new classes of thiol and aminothiol compounds derived from benzothiazole (T1, T2, AM1, AM2) and thiadiazole (T3, T4, AM3) structures. We examined them for their ability to scavenge free radicals (DPPH*, ABTS(*+), *OH). Thiol derivatives with a thiadiazole structure are the most active compounds scavenging DPPH* and ABTS(*+) free radicals, with an IC(50) of 0.053+/-0.006 and 0.023+/-0.002 mM, respectively, for the derivative T3. Moreover, compounds T1, T2, and T3 at 60 microM gave 83% protection against 2-deoxyribose degradation by *OH. The ability of these compounds to protect DNA against *OH produced by a Fenton reaction and gamma-irradiation (15 Gy)-induced strand breaks was also evaluated on pBR322 plasmid DNA. In both tests thiol derivatives were the most efficient compounds. Derivatives T2 and T3 totally inhibit DNA strand breaks at the concentration of 50 microM. The protection afforded by these derivatives was comparatively higher than that of the radioprotectors WR-2721 and WR-1065. Our data indicate that these two compounds are free radical scavengers and potential antioxidant agents. Finally, DFT and QSAR studies were performed to support the experimental observations.

Polynorbornene polycationic polymers as gene transfer agents. Influence of the counterion for in vitro transfection.

Polycationic derivatives of polynorbornene with different non-cytotoxic counterions, have been prepared by organometallic polymerization of methyleneammonium norbornene and subsequent exchange of the counterion. In this paper the effect of the counterion on the polycationic polymer binding onto plasmid DNA was studied via different ethidium bromide assays, heparin displacement and protection against degradation by DNAse. According to the nature of the counterions and consequently the size of the polymer particles, their complexation with the DNA led to aggregates with variable binding affinity for the plasmid. The relative transfection efficiency of each polyplex was compared, on the basis of reporter gene expression, in cells in culture. The nature of the counterion was seen to affect gene delivery. The order of transfection efficiency of the counterions studied at equivalent charge ratios (NH3+/PO4-) is lactobionate, acetate, chloride. The results obtained with the polynorbornene methyleneammonium lactobionate and acetate are particularly encouraging.