Real-time in vivo imaging of surface-modified liposomes to evaluate their behavior after pulmonary administration

Our previous study demonstrated that surface modification of liposomes using polyvinyl alcohol with a hydrophobic anchor (PVA-R) achieved sustained absorption from the lung after pulmonary administration and prolonged the pharmacological effects of the model peptide drug. In the present study, the behavior of PVA-R-modified liposomes in the lung and whole body was monitored using a real-time in vivo imaging system. Subsequently, the influence of surface modification with PVA-R on liposomal behavior in lung tissue was examined. Indocyanine green (ICG) was used as a near-infrared label of PVA-R-modified liposomes and was used to observe their dynamic behavior using non-invasive in vivo imaging (IVIS® imaging system) after pulmonary administration to rats. PVA-R-modified submicron-sized liposomes (ssLips) induced long-term retention in the lung compared with unmodified liposomes. Moreover, liposome association with alveolar macrophages (NR8383) was decreased by PVA-R modification in vitro. Therefore, PVA-R modification may prevent rapid elimination of ssLips by macrophages, thereby increasing retention in the lung.     

Vitamin E, membrane order, and antioxidant behavior in lung microsomes and reconstituted lipid vesicles

Vitamin E, a dietary antioxidant, is known to inhibit peroxidation of membrane lipids and to protect the lungs of vitamin E-deficient animals and to a lesser extent vitamin E-sufficient animals from oxidant injury. Since the protective interaction between vitamin E and biological membranes may be related to alterations in composition and physical state of membrane lipids, we evaluated the effect of vitamin E deficiency on lung microsomal lipids and membrane fluidity. Both intact microsomes and lipid vesicles prepared from the total lipid extracts of these microsomes were used. The percentage incorporation of vitamin E and cholesterol, membrane fluidity, and lipid peroxidation were measured in microsomes as well as their lipid vesicles. Fluidity was measured by monitoring changes in fluorescence anisotropy for 1,6-diphenyl-1,3,5-hexatriene (DPH). Lipid peroxidation was measured by thiobarbituric acid reaction. There were significant increases in the phospholipid (p less than 0.01), the total cholesterol (p less than 0.05), and the total saturated fatty acids (p less than 0.05) and decreases in total polyunsaturated fatty acid (p less than 0.01) content of vitamin E-deficient microsomes. There were no detectable peroxidative products in freshly isolated microsomes from either vitamin E-sufficient or -deficient lungs. However, lipids from vitamin E-deficient microsomal membranes were more susceptible to free radical initiated peroxidation than lipids from vitamin E-sufficient microsomes. Fluidity in vitamin E-deficient microsomes or in their lipid vesicles was significantly (p less than 0.05) decreased compared to the respective controls. In vitamin E-deficient microsomes or their lipid vesicles, the incorporation rate of vitamin E was two- to three-fold greater than in vesicles of vitamin E-sufficient microsomes or their lipid vesicles. However, the percentage incorporation of cholesterol was identical in both vitamin E-deficient and vitamin E-sufficient microsomes or in their respective lipid vesicles. As a result of vitamin E incorporation, fluidity was significantly decreased (p less than 0.05) in vitamin E-sufficient vesicles and was further decreased (p less than 0.001) in vitamin E-deficient vesicles. Incorporation of cholesterol also decreased fluidity in both vitamin E-deficient and vitamin E-sufficient vesicles but to the same extent (p less than 0.001). Lipid peroxide formation was two-fold greater in the vitamin E-deficient than in the vitamin E-sufficient vesicles.(ABSTRACT TRUNCATED AT 400 WORDS)     

Formulation and characterization of azidothymidine-loaded liposomes

Entrapment efficiency (EE%) and in vitro stability of azidothymidine (AZT)-loaded hand-shaken multilamellar vesicles (MLVs), freeze and thaw vesicles (FATMLVs), and reverse phase evaporation vesicles (REVs) were compared. AZT entrapment in FATMLVs was further studied by varying initial lipid concentrations, drug concentration, and lipid composition. The results suggest that AZT entrapment is dependent on the aqueous volume entrapped within liposomes, and the interaction between the drug and liposomal bilayer may not be significant. Increasing the lipid concentration increases the liposomal entrapment of AZT but the encapsulation yield decreases above a lipid concentration of 30 μmol/mL. No significant difference was observed in EE% when the AZT concentration was varied from 5 to 20 mg/mL. The entrapment efficiency was highest (43.2%) for DSPC/CHOL/PS (molar ratio 6:3:3) vesicles but DSPC/CHOL/PS liposome formulations in a molar ratio of 4:3:3 or 4:5:1 and DSPC/CHOL/SA liposome formulations in a molar ratio of 4:5:1 were found to be more stable in vitro. In vitro drug release from liposomes was dependent on bilayer composition and the method of preparation.     

Pharmacokinetics of oligodeoxynucleotides encapsulated in liposomes: effect of lipid composition and preparation method

1. The effect of the method employed to prepare liposomes and their lipid composition were evaluated in terms of the encapsulation efficiency and pharmacokinetic features of two oligodeoxynucleotides of a 21 mer: the normal (N-Odn) and the phosphorothioate (S-Odn) oligodeoxynucleotide. 2. Liposomes were prepared by the classical method of multilamellar vesicles (MV) and by the dehydration-rehydration method (DR). Two lipid mixtures were used to prepare liposomes - the predominant lipid being phosphatidylcholine (PC) and sphingomyelin (SM) respectively. 3. The DR method for liposome preparation provided the highest encapsulation e ciency, regardless of liposome lipid composition and the type of oligodeoxynucleotide involved (N-Odn or S-Odn). 4. The pharmacokinetics of free and liposome encapsulated oligodeoxynucleotides was studied in mouse following i.v. administration. Liposome encapsulated oligodeoxynucleotides exhibited a significantly lower plasma clearance and longer half-life and residence time than free oligodeoxynucleotides. The method used to obtain the liposomes affected plasma clearance, which was lower for liposomes elaborated by the DR method than for liposomes prepared with the MV method. The use of S-Odn in place of N-Odn decreased the plasma clearance of oligodeoxynucleotide when administered encapsulated in liposomes, regardless of the lipid composition and method used to obtain the liposomes.     

Pharmacokinetics of oligodeoxynucleotides encapsulated in liposomes: effect of lipid composition and preparation method

1. The effect of the method employed to prepare liposomes and their lipid composition were evaluated in terms of the encapsulation efficiency and pharmacokinetic features of two oligodeoxynucleotides of a 21 mer: the normal (N-Odn) and the phosphorothioate (S-Odn) oligodeoxynucleotide. 2. Liposomes were prepared by the classical method of multilamellar vesicles (MV) and by the dehydration-rehydration method (DR). Two lipid mixtures were used to prepare liposomes--the predominant lipid being phosphatidylcholine (PC) and sphingomyelin (SM) respectively. 3. The DR method for liposome preparation provided the highest encapsulation efficiency, regardless of liposome lipid composition and the type of oligodeoxynucleotide involved (N-Odn or S-Odn). 4. The pharmacokinetics of free and liposome encapsulated oligodeoxynucleotides was studied in mouse following i.v. administration. Liposome encapsulated oligodeoxynucleotides exhibited a significantly lower plasma clearance and longer half-life and residence time than free oligodeoxynucleotides. The method used to obtain the liposomes affected plasma clearance, which was lower for liposomes elaborated by the DR method than for liposomes prepared with the MV method. The use of S-Odn in place of N-Odn decreased the plasma clearance of oligodeoxynucleotide when administered encapsulated in liposomes, regardless of the lipid composition and method used to obtain the liposomes.     

Preparation and characterization of liposomes incorporating cucurbitacin E,a natural cytotoxic triterpene

Cucurbitacin E (Cuc E), highly oxygenated triterpene from cucurbitaceae family, was demonstrated to possess anti-proliferative, anti-inflammatory and anti-oxidant activities. Here, we studied the effect of Cuc E on the properties of the phospholipid membrane. Large unilamellar vesicles, with and without sulforhodamine B (SRB), were prepared in the absence and presence of Cuc E. The fluorescence increase, resulting from SRB release from vesicles due to inhibition of auto-quenching, was used to assess the permeability of liposome at 4° and 37 °C. At 4 °C, blank liposomes and those containing Cuc E were stable; while at 37 °C Cuc E loaded liposomes showed less stability than blank ones. The loading efficiency of Cuc E into the vesicles was demonstrated to be 85% by HPLC. Dynamic light scattering measurements showed that liposomes embedding Cuc E were smaller than those which did not. Multilamellar vesicles were manufactured from dipalmitoylphosphatidylcholine in the presence and absence of Cuc E (molar ratio 7%). Results obtained by differential scanning calorimetry suggest that Cuc E binds mainly at the polar/apolar interfacial region of lipid bilayers and may incorporate into the lipid bilayer. Cuc E does not produce phase separation and thus might be included in liposome formulation.     

Aerosolized liposome-based delivery of amphotericin B to alveolar macrophages

The present study was aimed at preparation, characterization, and performance evaluation of amphotericin B (Amp B)-loaded aerosolized liposomes for their selective presentation to lungs (alveolar macrophages), that being the densest site of Aspergillosis infection. Egg phosphatidylcholine (PC)- and cholesterol (Chol)-based liposomes were modified by coating them with alveolar macrophage-specific ligands (O-palmitoyl mannan, OPM, and O-polmitoyl pullulan, OPP). The prepared formulations were characterized in vitro for vesicle morphology, mean vesicle size, vesicle size distribution and percent drug entrapment. Pressurized packed systems based on preformed liposomal formulations in chlorofluorocarbon aerosol propellants were prepared. In vitro airways penetration efficiency of the liposomal aerosols was determined by percent dose reaching the peripheral airways, it was recorded 1.4-1.6 times lower as compared to plain drug solution-based aerosol. In vivo tissue distribution studies on albino rats suggested the preferential accumulation of OPM- and OPP-coated formulations in the lung macrophages. Higher lung drug concentration was recorded in case of ligand-anchored liposomal aerosols as compared to plain drug solution and plain liposome-based aerosols. The drug was estimated in the lung in high concentration even after 24 h. The drug-localization index calculated after 6 h was nearly 1.42-, 4.47-, and 4.16-fold, respectively, for plain, OPM-, and OPP-coated liposomal aerosols as compared to plain drug solution-based aerosols. These results suggest that the ligand anchored liposomal aerosols are not only effective in rapid attainment of high-drug concentration in lungs with high population of alveolar macrophages but also maintain the same over prolonged period of time. The significance of targeting potential of the developed systems was established.     

Evaluation of Percutaneous Absorption of Naproxen from Different Liposomal Formulations

The present study concerns the percutaneous absorption of naproxen (NPX), as model anti-inflammatory drug, included in liposome formulations constituted of different lipids: stratum corneum lipids (SCL) and phosphatidylcholine/cholesterol (PC/CHOL). Liposome dispersions were produced using two different methods: reverse-phase evaporation (REV) and thin layer evaporation (TLE). Morphology and dimensions of the disperse phase were characterized by cryo-transmission electron microscopy (cryo-TEM) and photon correlation spectroscopy, respectively. X-ray diffraction was employed to determine the structural organization of the vesicles. In vitro diffusion was studied by Franz cell on liposome dispersions viscosized by carbomer. Tape stripping was performed to investigate in vivo the performance of differently composed liposomes as NPX delivery system. Cryo-TEM showed spherical vesicles and bigger irregular elongated nanoparticles for TLE SCL liposomes. REV resulted in spherical and elongated multilamellar vesicles. Also X-ray diffraction evidenced La or Lp multilamellar vesicles for PC/CHOL and SCL liposome respectively. The in vitro study showed a lower NPX flux for SCL with respect to PC/CHOL liposome. Tape stripping corroborate the in vitro findings regarding SCL, suggesting that liposomes create a drug reservoir mixing with SC lipids, whilst PC/CHOL liposome promoted NPX permeation through the skin. Liposome lipid composition seems to affect NPX permeation through the skin. © 2009 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 99:2819-2829, 2010     

Physicochemical characterization and skin permeation of liposome formulations containing clindamycin phosphate

This study was undertaken to evaluate the physicochemical properties and skin permeation of liposome formulations containing clindamycin phosphate (CP), especially when charge was imparted to the liposome. Five different liposome formulations were prepared using Phospholipon 85G (PL) and cholesterol (CH) by conventional lipid film hydration technique. Molar ratio of CH to PL was varied in the range of 0.16–1.0. Charged liposomes were prepared in the same way with addition of 1,2-dioleoyl-3-trimethylammonium-propane chloride salt (DOTAP) and 1,2-dimyristoyl-sn-glycero-3-phosphate monosodium salt (DMPA) as charge carrier lipid for cationic or anionic charge of the liposome, respectively. Fresh full-thickness mice skin was taken and used for skin permeation study using Keshary-Chien diffusion cell with 1.77 cm² diffusion area at 37°C. All liposome formulations prepared showed homogeneous size distribution with mean particle size of about 1 μm or less. Among the five liposome formulations prepared, formulation with the molar ratio of 0.5 showed the best result in the physicochemical properties such as polydispersity index, entrapment efficiency, size evolution, and ability of the liposome to retain CP as of entrapped in the vesicles. Charge-impartation of the formulation with cationic charge carrier lipid resulted in additional benefit in terms of inhibition of size evolution, the ability of the liposome to retain CP in the vesicles, and skin permeation. Steady state flux of the drug through the mice skin in the cationic liposome vesicles was 0.75 ± 0.01 μg/cm²h while that in the control (dissolved into mixed alcohol solution) was 0.17 μg/cm²h. One half molar ratio of CH to PL was optimal in terms of physicochemical properties of the liposome formulation containing CP, and incorporation of cationic charge carrier lipid appeared to provide additional benefits for the stability of the liposome formulation and skin permeation of the drug.     

超氧化物歧化酶脂质体在大鼠体内的药代动力学和组织分布

目的考察3种超氧化物歧化酶(SOD)脂质体静脉给药后在大鼠体内的药代动力学和组织分布。方法 用反相蒸发法制备SOD脂质体，采用黄嘌呤氧化酶法检测SOD活力，静脉注射给药后，测定大鼠血中SOD含量变化和不同组织中SOD含量变化。结果在血浆中，SOD水溶液、SOD普通脂质体、用DSPE-PEG2000修饰的SOD脂质体、用Tween 80修饰的SOD脂质体的半衰期分别为0.25，0.34，0.66和0.41 h；AUC分别为12.48，24.66，41.16和33.02 μg·h·mL^-1。与普通脂质体比较，经过DSPE-PEG和Tween 80修饰后的脂质体，使肝、脾中SOD的含量有不同程度的降低，脑中含量有所提高。结论3种SOD脂质体均可不同程度地延长SOD的血浆半衰期，并以用DSPE-PEG2000修饰的SOD脂质体效果最好。与普通脂质体相比，用Tween 80修饰的SOD脂质体可以提高进入脑中的SOD量，用DSPE-PEG2000修饰的SOD脂质体可以减少肝脾对SOD的摄取。     

Phospholipid vesicles as carriers in aquaculture: preparation and stability study of thiamine hydrochloride-loaded liposomes

The aim of this work is to study liposomes as carriers of nutrients and therapeutic agents in aquaculture with Venerupis decussatus and Venerupis pullastra larvae. Multilamellar (MLVs) and large unilamellar (LUVs) vesicles were prepared from a commercial mixture of soy phosphatidylcholine, rich in unsaturated and polyunsaturated fatty acids, cholesterol, and hydrated with a solution of vitamin B1 both in distilled and sea water. Carboxyfluorescein-loaded liposomes were also prepared in order to test the uptake of vesicles by larvae. The stability of formulations was checked by monitoring the size of vesicles and their drug leakage. In order to limit the vitamin loss, liposome freeze-drying was studied. Dried formulations were also prepared by using different amounts of trehalose as cryoprotectant. We found that freeze-dried vesicles, rehydrated after two weeks, had a vitamin retention (R%) equal to 95%, while their diameter significantly increased. By contrast, liposomes freeze-dried in the presence of trehalose displayed a lower R%, but higher bilayer stability. Finally, when CF-loaded vesicles were added to Venerupis decussatus and Venerupis pullastra larvae incubated in filtered sea water, a bright and diffused fluorescence was present in most of the larvae, a fact which can be regarded as evidence of liposome uptake by Venerupis larvae.     

The effects of desferrioxamine on cisplatin-induced lipid peroxidation and the activities of antioxidant enzymes in rat kidneys

Cisplatin-induced nephrotoxicity is associated with an increase in lipid peroxidation and oxygen free radicals in rat kidneys. In this study, the effects of desferrioxamine were compared to vitamin C and E on cisplatin-induced lipid peroxidation and antioxidant enzyme activities in rat kidneys. Rats were divided into five groups, with 15 Wistar rats in each group. In the control group, rats received 1 mL/100 g isotonic saline solution intraperitoneally (i.p.). In Group II, 10 mg/kg cisplatin i.p. was injected to rats. Thirty minutes before the same dosage of cisplatin administration, 100 mg/kg i.p. vitamin C or E was given to rats in groups III and IV, respectively. Rats in Group V received 250 mg/kg desferrioxamine i.p., before the same dose of cisplatin administration. All rats were killed by cervical dislocation after 72 hours. The kidneys were immediately removed and washed in cold saline. Spectrophotometric method was used for all analyses. While catalase, glutathione reductase (GR), and superoxide dismutase (SOD) levels were found to be significantly decreased (P < 0.001), malondialdehyde (MDA) (P < 0.05) and hydrogen peroxide (H2O2) (P < 0.001) levels were significantly increased in the cisplatin group when compared to the controls. MDA levels were decreased by desferrioxamine (P < 0.005) as well as vitamin C and E (P < 0.05 and P < 0.001, respectively). These three compounds induced a significant increase in SOD levels (P < 0.05), but only in the vitamin C group, were SOD levels not significantly different than the levels of the controls (P > 0.05). In the desferrioxamine (P < 0.05), vitamin C and E groups (P < 0.001 for both), the cisplatin elevated H2O2 levels were decreased. None of these drugs had any effect on GR and catalase levels (P > 0.05). Desferrioxamine is useful to prevent cisplatin-induced lipid peroxidation, however, vitamin C and E are more effective on antioxidant enzymes than desferrioxamine.     

Pulmonary disposition of budesonide from liposomal dry powder inhaler

The Purpose of this study was to establish the use of a developed dry powder inhaler of budesonide liposomes in pulmonary drug delivery. Budesonide liposomes composed of egg phosphatidyl choline (EPC) and cholesterol were prepared using a lipid-film hydration technique. The liposomal dispersion was freeze dried and formulated to a dry powder inhaler. The entrapped drug values (91.79% to 78.99%) of freeze dried liposomes were estimated in prepared batches after purification from the free drug by centrifugation of the rehydrated vesicles. In vitro drug retention was evaluated using methanolic phosphate buffer saline and bronchoalveolar lavage, following incubation at 37 degrees C. All batches were found to retain more than 63.54% of budesonide within liposomes at the end of 24 h. Rehydrated budesonide liposomes or nonencapsulated budesonide was delivered to rat lungs by intratracheal administration. The pulmonary drug disposition was assessed by simultaneous monitoring of drug levels in the bronchoalveolar lavage and lung tissue. After intratracheal administration, cumulative drug levels in the lung tissue indicated that the targeting factor was at least 1.66 times higher in liposomes. The maximal drug concentration in the lung homogenate for the liposomal dry powder inhaler was 36.64 micrograms as compared to 78.56 micrograms with the plain drug. Similarly, the time for maximum drug concentration in the lung homogenate for the liposomal dry powder inhaler was 9-12 h as compared to 3 h for that of the plain drug. Hence, the use of a developed liposomal budesonide dry powder inhaler was found to provide desired drug levels in the lungs for a prolonged period of time, which is expected to enhance the therapeutic index of the drug and probably reduce the dose and cost of therapy as well.     

Enhanced anti-inflammation of inhaled dexamethasone palmitate using mannosylated liposomes in an endotoxin-induced lung inflammation model

Inhalation of bacterial endotoxin induces pulmonary inflammation by activation of nuclear factor kappaB (NFkappaB), production of cytokines and chemokines, and neutrophil activation. Although glucocorticoids are the drugs of choice, administration of free drugs results in adverse effects as a result of a lack of selectivity for the inflammatory effector cells. Because alveolar macrophages play a key role in the inflammatory response in the lung, selective targeting of glucocorticoids to alveolar macrophages offers efficacious pharmacological intervention with minimal side effects. We have demonstrated previously the selective targeting of mannosylated liposomes to alveolar macrophages via mannose receptor-mediated endocytosis after intratracheal administration. In this study, the anti-inflammatory effects of dexamethasone palmitate incorporated in mannosylated liposomes (DPML) at 0.5 mg/kg via intratracheal administration were investigated in lipopolysaccharide-induced lung inflammation in rats. DPML significantly inhibited tumor necrosis factor alpha, interleukin-1beta, and cytokine-induced neutrophil chemoattractant-1 levels, suppressed neutrophil infiltration and myeloperoxidase activity, and inhibited NFkappaB and p38 mitogen-activated protein kinase activation in the lung. These results prove the value of inhaled mannosylated liposomes as powerful targeting systems for the delivery of anti-inflammatory drugs to alveolar macrophages to improve their efficacy against lung inflammation.     

Enhanced systemic exposure of fexofenadine via the intranasal administration of chitosan-coated liposome

The present study aimed to develop the intranasal delivery system of fexofenadine for the prolonged drug release via the preparation of mucoadhesive liposome. By using thin layer film hydration method, liposome of fexofenadine was prepared with DPPC/DPPG, resulting in the small lipid vesicles (359 ± 5.5 nm) with narrow size distribution (PI<0.1). Subsequently, the surface of anionic liposome was coated by chitosan and in vitro characteristics of liposomes were evaluated along with the pharmacokinetic studies in rats. Chitosan coated liposomes were stable for 6-month storage at 4 °C without any significant size change and drug leakage. Furthermore, it exhibited strong mucoadhesive properties in mucin adsorption test, which was 3-fold higher than uncoated liposomes. Compared to the oral delivery of powder formulation, the intranasal delivery of fexofenadine significantly (p<0.05) increased systemic exposure of fexofenadine in rats. Particularly, the intranasal administration of chitosan coated liposome exhibited approximately 5 fold enhancement of AUC with more sustained drug release in rats compared to the oral delivery. In conclusion, intranasal administration of chitosan coated liposome appeared to be effective to enhance the bioavailability as well as prolonged exposure of fexofenadine in rats.     

Experimental pulmonary delivery of cyclosporin A by liposome aerosol

The utilization of CsA–liposome for aerosol delivery by jet nebulizers has potential advantages for clinical development including: aqueous compatibility, sustained pulmonary release to maintain therapeutic drug levels and facilitated delivery to alveolar macrophages and pulmonary lymphocytes. Inhalation of cyclosporin A (CsA)–dilauroylphosphatidylcholine (DLPC) liposome aerosols will theoretically result in localized and sustained delivery of therapeutic CsA concentrations within the lung as an alternative to local immunotherapy for pulmonary diseases. In the lung, targeted delivery of therapeutic CsA concentrations would require lower dosages than via conventional intravenous or oral routes of administration. Potential benefits from targeted lung delivery could include reduced systemic toxicity and prolonged immunosuppressive activity. Aerosol delivery systems have been developed to deposit drugs directly onto pulmonary surfaces at the sites of disease within the lung. A novel HPLC method for tissue analysis of CsA–liposomes is developed and utilized with a solid-phase extraction method to measure CsA recovered from Balb/c mouse lung tissues. A concentrated formulation containing 5 mg CsA–37.5 mg DLPC/ml was nebulized with an Aerotech II nebulizer generating an aerosol particle size distribution (mass median aerodynamic diameter (MMAD)) of 1.7 μm and geometric standard deviation (GSD) of 2.0. After a 15-min aerosol exposure, little of no CsA was detected in the blood, liver, kidney or spleen. The lung contained the highest organ CsA levels with high immunosuppressive activity demonstrating effective pulmonary targeting of the CsA–DLPC liposome aerosol. The results of this system will be utilized as the experimental basis for future pharmacokinetic, toxicological, immunosuppression and other biological studies.     

肺靶向阿奇霉素脂质体的制备及其在小鼠体内的分布

目的研究肺靶向阿奇霉素阳离子脂质体的制备方法并考察其在小鼠体内的分布。方法利用旋转薄膜-冻融法制备肺靶向阿奇霉素脂质体。用高效液相色谱法测定给药后小鼠体内各组织中的药物浓度。结果制得的脂质体平均粒径为6.582 μm，表面电荷为+19.5 mV，包封率大于75%，稳定性好。药物体外释药符合Higuchi方程。小鼠尾静脉给药后，阳离子脂质体主要被肺摄取，在肺部的滞留时间明显延长，AUC值约为阿奇霉素溶液的8.4倍。结论采用薄膜-冻融法，添加十八胺可制得具有较高包封率及稳定性的阿奇霉素阳离子脂质体，在小鼠肺部的分布优于注射液，能达到肺靶向目的。     

Preparation and properties of poly(vinyl alcohol)-stabilized liposomes

The purpose of this work is to evaluate the improvement in physical stability of poly(vinyl alcohol) (PVA) modified liposomes. Liposomes composed of soya phosphatidylcholile (SPC) and cholesterol (1:1 molar ratio) were prepared by reverse phase evaporation method. Two types of interaction between liposome and PVA were investigated: PVA addition into lipid bilayer during liposome preparation and coating of already formed liposomes with PVA. The microparticles system was morphologically characterized by transmission electron microscopy (TEM) and particles analysis. Changes in particles size and zeta potential confirmed the existence of a thick polymer layer on the surface of liposomes. The amount of PVA adsorbing to liposomes and the encapsulation efficiency increased with increasing polymer concentration. The physical stability was evaluated by measuring the release rate of contents at 20 and 37 degrees C, the PVA modified liposomes were more stable than the conventional liposomes. Comparing with PVA-coated liposomes, the liposomes with PVA addition to the bilayer were more stable, and had higher entrapment efficiency.     

Optimization of cationic lipid/DNA complexes for systemic gene transfer to tumor lesions

Intravenous (i.v.) administration of cationic lipid N-[( 1-(2-3-dioleyloxy)propyl)]-N-N-N-trimethylammonium chloride (DOTMA)-based transfection complexes in mice with subcutaneous squamous cell tumors yielded plasmid delivery and expression in tumor lesions. The efficiency of gene transfer in tumors was significantly lower than in the lung. This was consistent with low plasmid levels associated with the tumor, suggesting that plasmid delivery to the tumor site was a limiting factor. Lowering the lipid/DNA charge ratio from 5:1 to 0.8:1 (+/-) did not change DNA levels in tumor but significantly reduced DNA levels in lung. However, expression levels were significantly reduced in both tissues at lower lipid/DNA charge ratios. Complexes prepared from small unilamellar liposomes gave significantly lower expression levels in the lungs but similar expression levels in tumors when compared to complexes prepared from larger unilamellar liposomes. The small liposome complexes were better tolerated than large liposome complexes. Varying the cationic lipid to colipid (cholesterol or DOPE) molar ratio from 4: 1 to 1: 1 significantly reduced expression levels in both tumor and lung. Cationic lipid substitution, using a cholesterol cationic lipid, diethyldiamino-carbamyl-cholesterol instead of DOTMA, produced reduced expression in all other tissues except tumor. Incorporation of PEG into preformed transfection complexes reduced DNA delivery to lung, increased circulation half-life, and enhanced DNA delivery to tumor. In a lung metastatic mouse tumor model, where the accessibility of the i.v. administered transfection complexes to tumor lesions should be less challenging, DOTMA: CHOL complexes (4: 1 lipid to colipid molar ratio, 3: 1 +/- lipid to plasmid charge ratio) were preferentially localized in tumor lesions. These data demonstrate that systemic gene transfer to distal tumor sites by lipid/ DNA complexes may be limited by low plasmid delivery. Modifying the chemical surface properties of transfection complexes enhanced both DNA delivery and expression in tumor and is one approach that may overcome limitations.     

Quantitative Histology and Cytochrome P-450 Immunocytochemistry of the Lung Parenchyma Following 6 Months of Exposure of Strain A/J Mice to Cigarette Sidestream Smoke

Abstract

Male strain A/J mice were exposed for 6 h/day, 5 days/wk to aged and diluted cigarette sidestream smoke (ADSS) at a chamber concentration of 4 mg/m³ of total suspended particulate matter (TSP). After 6 mo, the lungs were examined for altered expression of cytochrome P-450 isozymes and for differences in total alveolar tissue volume or surface area, as well as changes in the numbers of epithelial type II cells and alveolar macrophages. Morphologic measurements showed no statistically significant differences for the air, alveolar tissue, or capillary volumes of the lungs or changes in the total number of epithelial type II cells or alveolar macrophages. In contrast, cytochrome P-4501A1 was elevated in the lungs of ADSS-exposed animals and localized in capillary endothelial cells. CYP2B1 was present in airway epithelial cells as well as in epithelial cells throughout the lung parenchyma, but its distribution was not changed by ADSS exposure. Isozyme CYP2E1 was also found in airway epithelial cells, but not in the lung parenchyma, with no differences noted between ADSS exposed animals and controls. CYP2F2 was found in the bronchiolar Clara cells and in type II cells located within the alveolar parenchyma, but was also unchanged. It is concluded that chronic exposure to cigarette ADSS at 4 mg/m³ of TSP produces no changes in alveolar macrophages or epithelial type II cells in mouse lung but increases the expression of cytochrome P4501A1.