Photodynamic therapy (PDT) is an established noninvasive tumor treatment. The hydrophobic natural occurring pigment hypericin shows a lot of attractive properties for the application in PDT. Hence, the administration to biological systems or patients requires the formulation in drug carriers enabling sufficient bioavailability. Therefore, free hypericin was encapsulated by the thin film hydration method or a hypericin-hydroxypropyl-β-cyclodextrin inclusion complex (Hyp-HPβCD) was incorporated by dehydration-rehydration vesicle method in either conventional or ultra-stable tetraether lipid (TEL) liposomes. The hydrodynamic diameter of the prepared nanoformulations ranged between 127 and 212 nm. These results were confirmed by atomic force microscopy. All liposomes showed a good stability under physiological conditions. TEL liposomes which tend to build more rigid bilayers, generate higher encapsulation efficiencies than their conventional counterparts. Furthermore, the suitability for intravenous application was confirmed by hemocompatibility studies resulting in a hemolytic potential less than 20% and a coagulation time less than 50 sec. The uptake of liposomal hypericin into human ovarian carcinoma cells (SK-OV-3) was confirmed using confocal microscopy and further characterized by pathway studies. It was demonstrated that the lipid composition and intraliposomal hypericin localization influenced the anti-vascular effect in the chorioallantoic membrane (CAM). While hypericin TEL liposomes exhibit substantial destruction of the microvasculature drug-in-cyclodextrin TEL liposomes showed no effect. Nevertheless, both formulations yielded severe photocytotoxicity in SK-OV-3 cells in a therapeutic dosage range. Conclusively, hypericin TEL liposomes would be perfectly suited for anti-vascular targeting while Hyp-HPβCD TEL liposomes could deliver the photosensitizer to the tumor site in a more protected manner.
Biomaterials,
which release active compounds after implantation, are an essential
tool for targeted regenerative medicine. In this study, thin multilayer
films loaded with lipid/DNA complexes (lipoplexes) were designed as
surface coatings for in situ transfection applicable in tissue engineering
and regenerative medicine. The film production and embedding of lipoplexes
were based on the layer-by-layer (LbL) deposition technique. Hyaluronic
acid (HA) and chitosan (CHI) were used as the polyelectrolyte components.
The embedded plasmid DNA was complexed using a new designed cationic
lipid formulation, namely, OH4/DOPE 1/1, the advantageous characteristics
of which have been proven already. Three different methods were tested
regarding its efficiency of lipid and DNA deposition. Therefore, several
surface specific analytics were used to characterize the LbL formation,
the lipid DNA embedding, and the surface characteristics of the multilayer
films, such as fluorescence microscopy, surface plasmon resonance
spectroscopy, ellipsometry, zeta potential measurements, atomic force
microscopy, and scanning electron microscopy. Interaction studies
were conducted for optimized lipoplex-loaded polyelectrolyte multilayers
(PEMs) that showed an efficient attachment of C2C12 cells on the surface.
Furthermore, no acute toxic effects were found in cell culture studies,
demonstrating biocompatibility. Cell culture experiments with C2C12
cells, a cell line which is hard to transfect, demonstrated efficient
transfection of the reporter gene encoding for green fluorescent protein.
In vivo experiments using the chicken embryo chorion allantois membrane
animal replacement model showed efficient gene-transferring rates
in living complex tissues, although the DNA-loaded films were stored
over 6 days under wet and dried conditions. Based on these findings,
it can be concluded that OH4/DOPE 1/1 lipoplex-loaded PEMs composed
of HA and CHI can be an efficient tool for in situ transfection in
regenerative medicine.
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