Unloaded Liposomes Research Articles

Treatment of experimental autoimmune uveoretinitis with intravitreal injection of infliximab encapsulated in liposomes

AimsTo evaluate the safety and efficacy of intravitreal injection of liposomes encapsulating infliximab in experimental autoimmune uveoretinitis (EAU) rats.MethodsLiposomes containing infliximab were prepared and characterised for mean particle size, entrapment...

Dynamics of unloaded and green tea extract loaded lecithin based liposomal dispersions investigated by nuclear magnetic resonance T2 relaxation

Liposomes are lipid bilayer vesicles that can be used as encapsulation systems for bioactive agents to provide increased protection against environmental stresses (such as pH or temperature extremes). Time Domain Nuclear Magnetic Resonance (TD-NMR) that is based on differentiation of specimen contents with respect to magnetic relaxation rates provides detailed information on amount, state and distribution of water and oil and provide reproducible results on the samples. These make TD-NMR particularly suitable for time-dependent monitoring of emulsion system dynamics. In this study, spin-spin (T2) relaxation times and relaxation spectra were used for characterizing green tea extract loaded and unloaded liposomes prepared with soy (S75) and egg lecithins (E80) by different preparation methods (such as homogenization type, pressure and solvent type). Mean particle sizes of liposomes were found to be the most influential factor in shaping mono-exponential T2 relaxation times. The differences in particle sizes of E80 and S75 samples along with samples with different homogenization pressures could be monitored with T2 relaxation times. Additionally, T2 relaxation times were found to be correlated with particle shape irregularity, and chemical instability of samples due to lipid oxidation. With relaxation spectrum analysis, particular components in the sample could be distinguished (internal/external water and lipid bilayers), which gave more elaborate results on mechanisms of instability.

Distribution of Melaleuca alternifolia essential oil in liposomes with Tween 80 addition and enhancement of in vitro antimicrobial effect

Tea tree oil (TTO) exhibits excellent broad-spectrum antimicrobial activity. In order to preserve it from the degradation in the presence of oxygen, light and temperature, TTO was encapsulated in liposomes (LTTOs) using the thin-membrane hydration and sonication method, and characterised by Zetasizer for size and size distribution, transmission electron microscope for morphology, zeta-potential for surface charge, entrapment efficiency and TTO release from the nanoparticles. The antimicrobial activities of phosphate-buffered saline solution containing TTO, unloaded liposomes and LTTOs suspension were determined by twofold serial broth dilution technique. The size of LTTOs was 75 nm and the encapsulation efficiency of 96.08% was obtained. LTTOs exhibited slow release of TTO and superior broad-spectrum antimicrobial effects compared with free TTO. Liposomes not only effectively encapsulated TTO to form a stable liposome suspension, but also enhanced inhibition and bactericidal effect on the TTO-tolerant strain. The liposomal systems carrying TTO may be a potential alternative for effective antimicrobial agents.

Liposomes loaded with histone deacetylase inhibitors for breast cancer therapy

Histone deacetylase (HDAC) inhibitors (HDACi) of the class I trichostatin A (TSA), CG1521 (CG), and PXD101 (PXD) were incorporated at a high rate (∼1 mM) in liposomes made of egg phosphatidylcholine/cholesterol/distearoylphosphoethanolamine-polyethylenglycol 2000 (64:30:6). Physicochemical parameters (size, zeta potential, loading, stability, release kinetics) of these HDACi-loaded pegylated liposomes were optimized and their cytotoxicity (MTT test) was measured in MCF-7, T47-D, MDA-MB-231 and SkBr3 breast cancer cell lines. In MCF-7 cells, TSA and PXD were efficient inducers of proteasome-mediated estradiol receptor α degradation and they both affected estradiol-induced transcription (TSA > PXD) contrary to CG. Moreover, TSA most efficiently altered breast cancer cell viability as compared to the free drug, CG-liposomes being the weakest, while unloaded liposomes had nearly no cytotoxicity. Pegylated liposomes loaded with TSA or PXD remained stable in size, charge and biological activity for one month when stored at 4 °C. All HDACi-loaded liposomes released slowly the encapsulated drug in vitro, CG-loaded liposomes showed the slowest release kinetic. These formulations could improve the efficacy of HDACi not only in breast cancers but also in other solid tumors because most of these drugs are poor water soluble and unstable in vivo, and their administration remains a challenge.

Treatment of Experimental Autoimmune Uveoretinitis with Intravitreal Injection of Tacrolimus (FK506) Encapsulated in Liposomes

PURPOSE. To evaluate the effects of intravitreal injection of liposomes encapsulating tacrolimus (FK506) on experimental autoimmune uveoretinitis (EAU) in Lewis rats. METHODS. Liposomes containing tacrolimus were prepared by reverse-phase evaporation vesicles. EAU was induced in Lewis rats by subcutaneous injection of interphotoreceptor retinoid-binding protein R16 peptide emulsified in adjuvant. Ten days later, rats were intravitreally injected with saline, tacrolimus, tacrolimus-loaded liposomes, or unloaded liposomes. Clinical signs of inflammation and ocular histologic sections were observed and graded. Retinal function was evaluated by electroretinography (ERG). Tacrolimus concentration was determined in the vitreous body and serum by ELISA. Ocular biodistribution of rhodamine-conjugated liposomes containing tacrolimus (tacrolimus-Rh-lip) was analyzed with a laser scanning confocal microscope. To evaluate the systemic effect of intravitreally injected tacrolimus, delayed-type hypersensitivity (DTH) and lymphocyte proliferation assay (LPA) responses were detected. RESULTS. Treatment of EAU with intravitreal injection of liposomal tacrolimus significantly reduced intraocular inflammation and markedly inhibited the development of EAU, as determined in clinical and histopathologic analyses. No toxic effects could be detected as evaluated by ERG. The concentration of tacrolimus in ocular fluids remained for as long as 14 days after liposomal injection of tacrolimus. Confocal microscopy showed a transretinal distribution of the liposomal particles. DTH and LPA responses were not impaired in liposomal tacrolimus-treated rats. CONCLUSIONS. Intravitreal injection of liposomal tacrolimus was highly effective in suppressing the process of EAU without any side effects on retinal function or systemic cellular immunity. This treatment may represent a new option for the management of intraocular inflammation.

Downregulation of Endotoxin-Induced Uveitis by Intravitreal Injection of Vasoactive Intestinal Peptide Encapsulated in Liposomes

To reestablish the immunosuppressive microenvironment of the eye, disrupted by ocular inflammation during endotoxin-induced uveitis (EIU), by means of intravitreal injection of vasoactive intestinal peptide (VIP) in saline or encapsulated in liposomes, to increase its bioavailability and efficiency. EIU was induced in Lewis rats by subcutaneous injection of lipopolysaccharide (LPS). Simultaneously, animals were intravitreally injected with saline, saline/VIP, VIP-loaded liposomes (VIP-Lip), or unloaded liposomes. EIU severity and cellular infiltration were assessed by clinical examination and specific immunostaining. VIP concentration was determined in ocular fluids by ELISA. Ocular expression of inflammatory cytokine and chemokine mRNAs was detected by semiquantitative RT-PCR. Biodistribution of rhodamine-conjugated liposomes (Rh-Lip) was analyzed by immunohistochemistry in eyes and regional cervical lymph nodes (LNs). Twenty-four hours after intravitreal injection of VIP-Lip, VIP concentration in ocular fluids was 15 times higher than after saline/VIP injection. At that time, EIU clinical severity, ocular infiltrating polymorphonuclear leukocytes (PMNs), and, to a lesser extent, ED1(+) macrophages, as well as inflammatory cytokine and chemokine mRNA expression, were significantly reduced in VIP-Lip-injected rats compared with rats injected with saline/VIP, unloaded liposomes, or saline. Rh-Lip was distributed in vitreous, ciliary body, conjunctiva, retina, and sclera. It was internalized by macrophages and PMNs, and VIP colocalized with liposomes at least up to 14 days after injection. In cervical LNs, resident macrophages internalized VIP-Rh-Lip, and some adjacent lymphocytes showed VIP expression. VIP was efficient at reducing EIU only when formulated in liposomes, which enhanced its immunosuppressive effect and controlled its delivery to all tissues affected by or involved in ocular inflammation.

Neovascular targeting chemotherapy: encapsulation of paclitaxel in cationic liposomes impairs functional tumor microvasculature.

Cationic liposomes have been shown to be internalized selectively by angiogenic tumor endothelial cells after intravenous injection. Therefore, encapsulation of cytotoxic substances in cationic liposomes is a new approach to target tumor vasculature. It was the aim of our study to quantify the effects of paclitaxel encapsulated in cationic liposomes (MBT-0206) on tumor microvasculature and growth in vivo. Experiments were performed in the dorsal skinfold chamber preparation of Syrian Golden hamsters bearing syngeneic A-Mel-3 melanomas. Tumors were treated with intravenous infusion of MBT-0206 (20 mM) resulting in an effective paclitaxel dose of 5 mg/kg body weight (b.w.). Control animals received conventional paclitaxel in Cremophor EL (Taxol(R); 5 mg/kg b.w.), unloaded cationic liposomes (20 mM) or the solvent 5% glucose, respectively. Using intravital microscopy, tumor growth and effects on intratumoral microvasculature were analyzed. Tumor growth was significantly retarded after treatment with MBT-0206 compared to the treatment with paclitaxel. Analysis of intratumoral microcirculation revealed a reduced functional vessel density in tumors after application of liposomal paclitaxel. At the end of the observation time, vessel diameters were significantly smaller in animals treated with paclitaxel encapsulated in cationic liposomes while red blood cell velocity was less affected. This resulted in a significantly reduced blood flow in vessel segments and a reduced microcirculatory perfusion index in these animals. Histochemical TUNEL stain was vessel-associated after treatment with liposomal paclitaxel in contrast to few apoptotic tumor cells in the control groups. Our data demonstrate that encapsulation of paclitaxel in cationic liposomes significantly increased the antitumoral efficacy of the drug. Remarkable microcirculatory changes indicate that encapsulation of paclitaxel in cationic liposomes resulted in a mechanistic switch from tumor cell toxicity to an antivascular therapy.

Pulmonary administration of IgG loaded liposomes for passive immunoprophylaxy

Local passive immunoprophylaxy has been used in pulmonary infectious diseases successfully. However, the short immunoglobulins half-life in the lungs limits the duration of their action. The aim of the present study was to evaluate the efficiency of human polyvalent intravenous immunoglobulins (IVIG) when protected after encapsulation within EPC: DPPG liposomes by dehydration/rehydration. Two IVIG concentrations were chosen: 10 and 1 mg/ml for further studies in mice infected by influenza A. For the highest concentration (10 mg/ml), IVIG loaded liposomes did not significantly differ from IVIG/unloaded liposomes mixture with around 45% association yield. For the lowest concentration (1 mg/ml), two thirds of the IVIG associated were found inside the vesicles. In vivo, IVIG administered intranasally at 10 mg/ml (500 μg per mouse) 4 days before the infection led to 100% survival whatever the formulation. When administered at a lower dose (1 mg/ml—50 μg per mouse) 2 days before the challenge, loaded liposomes were found less efficient than free IVIG while unloaded liposomes showed a slight aspecific immunoprotection. Gastrointestinal clearance must be responsible for a major loss of liposomes compared to IVIG solution because of a higher viscosity of the formulation. Discrepancies with the literature are discussed.

Acceleration of wound healing by topical drug delivery via liposomes.

Despite intensive research, impaired wound healing remains a considerable complication. Therefore, topically applied liposome-encapsulated buflomedil hydrochloride was investigated for its ability to improve wound repair in normal (n=16) and ischemic (n=16) skin tissue. Experiments were performed using the wound healing model of the ear of hairless mice. Standardized skin wounds (4.25 mm(2)) were created by circular excision of the epidermal layer and the subcutaneous tissue. Liposomes were applied daily until complete neovascularization of the wound occurred. Tissue regeneration by complete epithelialization and neovascularization of the wound area, microcirculatory parameters, and leukocyte-endothelium interaction were investigated by means of intravital microscopy. Microvascular perfusion was assessed by laser-Doppler flowmetry. Topical application of buflomedil liposomes led to a significantly (P<0.05) accelerated wound closure in both normal (9.6+/-0.7 days) and ischemic (13.4+/-0.1 days) skin tissue compared with animals that were treated with unloaded liposomes (controls; 13.1+/-0.8 days; 15.3+/-0.6 days). Complete neovascularization of the wound was also enhanced (P<0.05) in buflomedil-treated animals (normal tissue 18.8+/-0.4 days; ischemic tissue 19.6+/-0.7 days) compared with controls (20.6+/-0.6 days; 22. 6+/-1.2 days). These data suggest that buflomedil-loaded liposomes might be of beneficial use for clinical wound care.

Phospholipid/Drug Interactions in Liposomes Studied by Rheological Properties of Monolayers

In order to obtain a good knowledge of the pharmaceutical form, we sought to localize the antiacneic drug RU 58841 in liposomes. The mixed drug/phospholipid monolayer obtained by spreading phospholipids at the air/aqueous surface of drug solutions was studied using the film balance technique to clarify the organization of the film respect to its composition and state of compression. A rheological approach based on Maxwell's model was found to be useful to discriminate between the influence of each constituent in the monolayer. From the study of the isotherms and the rheological behavior of mixed monolayers at low surface pressures it was deduced that the drug could be localized between the phospholipids polar headgroups. In the opposite case, at high surface pressures, the drug molecule was believed to be trapped in the hydrophobic part of the phospholipidic layer and behave like an insoluble one. The surface film formation kinetics of drug-loaded liposomes is faster than the corresponding kinetics of unloaded liposomes.

The use of rifampicin and isoniazid entrapped in liposomes for the treatment of murine tuberculosis

Liposomes loaded with rifampicin and isoniazid were used experimentally to treat mice with severe tuberculosis. The animals were distributed in four groups. The control group and the group treated with unloaded liposomes showed the severest disease. Both groups showed the lowest accumulated survival, about 50 % after 30 days. The numbers of colony-forming-units (CFU) and root specific lung weight (RSLW) were the highest and the histopathology of the lung showed marked diffuse lesions. However, the group treated with unloaded liposomes showed significantly higher growth of M. tuberculosis compared with the control. The group treated with drug and drug loaded liposomes showed a higher survival, about 85 % after 30 days, and the lowest values of CFU and RSLW. The lung histology revealed considerably less inflammation which was focal. The parameters evaluated indicated a significantly better response in the group of animals treated with rifampicin and isoniazid entrapped in liposomes.