The importance, prevalence and determination of vitamins B6 and B12 in food matrices: A review.

Vitamins are a vast group of fundamental organic compounds, which are not produced by the human body but are essential for the living organisms' good health. Vitamins B6 and B12 belong to the same group of hydrophilic vitamins. Structurally unrelated, they share the same purpose as essential components for normal cellular operation, growth and development. Vitamin B6 is an enzymatic co-factor that is vital for countless biochemical reactions, and is also important in sugar and fatty acid metabolization. It encompasses three natural and inter-convertible pyridine-derivatives: pyridoxine, pyridoxal and pyridoxamine. Vitamin B12 is a cobalt organometallic complex also indispensable in numerous human physiological functions. It has four bioactive forms: cyanocobalamin, methylcobalamin, hydroxocobalamin and 5'-deoxyadenosylcobalamin, and only a few prokaryotes have the ability to biosynthesize cobalamin. This work reviews the significant aspects of vitamins B6 and B12: their vital roles, consequences of deficit; food sources; and methods of determination and respective matrices, with heavy emphasis on chromatographic techniques developed within the last two decades.

Survivin silencing as a promising strategy to enhance the sensitivity of cancer cells to chemotherapeutic agents.

Since clinical application of conventional cancer therapies is usually limited by drug resistance and toxic side effects, combination of chemotherapeutic agents with gene therapy appears as an attractive therapeutic strategy to overcome these issues. Being selectively expressed in tumor tissues, survivin is a promising target for the development of anticancer strategies aimed at eliminating tumor cells while sparing normal tissues. In this work, we achieved substantial protein knockdown in a number of human cell lines, namely, A549, HeLa and MCF-7 cells which overexpress survivin, after treatment with anti-survivin siRNAs, which was associated with a significant reduction of cell viability, when compared to treatment with control siRNAs. Interestingly, when the survivin-silencing approach was combined with a chemotherapeutic agent, an enhancement of the therapeutic effect was achieved. Treatment with anti-survivin siRNAs resulted in high levels of caspase 3/7 activation, and an enhancement of this effect was observed when survivin silencing was combined with vinblastine. In addition, we showed that for A549 and HeLa cells survivin silencing contributes to the reversion of cell resistance to doxorubicin. Overall, we demonstrate that the combination of a survivin-directed silencing strategy with chemotherapeutic agents constitutes a valuable approach for cancer treatment.

Hydrological contraction patterns and duration of drying period shape microbial-mediated litter decomposition.

The length and number of streams experiencing intermittency is expected to increase in response to human population growth, associated water use, and climate change. In these streams, habitat contraction may occur at distinct rates giving rise to drying periods of distinct duration. To date, the impact of drought installation rate and duration have been mostly overlooked. In this microcosm study, stream conditioned oak leaf litter was subjected to either a short (5 weeks) or a long (8 weeks) drying period, originating from a very slow, slow, or abrupt contraction. The effects of these treatments were compared at the end of the drying period in terms of microbial-mediated litter mass loss, fungal biomass, respiration, and sporulation rates. A very slow contraction pattern led to 1.3 times higher mass loss than both slow or abrupt contraction. Fungal biomass, respiration and sporulation rates were up to 2.3 times lower under slow than abrupt contraction. Both drying period durations inhibited leaf decomposition, suggesting an early, critical effect of drying on microbial-mediated processing, regardless of contraction pattern. This seems to be related to an impoverishment of leaf associated fungal communities and resultant lower functional efficacy - species richness decreased by up to 75% in response to a long (vs. short) drying period, despite the maintenance of mycelial biomass. Our results show the relevance of aquatic hyphomycetes to litter decomposition in dry streambeds, particularly following slower habitat contraction patterns. Faster wet-to-dry transitions and longer drying periods strongly impaired microbial functioning, with potential impacts on global processing rates and cascading effects through changes of detritus quality. If confirmed in field tests, such impacts on stream functioning may be mitigated by preserving riparian forests, which may protect against extreme drying events by buffering temperature changes.

In situ TEM study of grain growth in nanocrystalline copper thin films.

Nanocrystalline metals demonstrate a range of fascinating properties, including high levels of mechanical strength. However, as these materials are exposed to high temperatures, it is critical to determine the grain size evolution, as this process can drastically change the mechanical properties. In this work, nanocrystalline sputtered Cu thin films with 43 +/- 2 nm grain size were produced by dc-magnetron sputtering. Specimens were subsequently annealed in situ in a transmission electron microscope at 100, 300 and 500 degrees C. Not only was grain growth more evident at 500 degrees C but also the fraction of twins found. An analysis of grain growth kinetics revealed a time exponent of 3 and activation energy of 35 kJ mol(-1). This value is explained by the high energy stored in the form of dislocation, grain boundaries and twin boundaries existing in nanocrystalline copper, as well as the high probability for atoms to move across grains in nanocrystalline materials.

TEM characterization of As-deposited and annealed Ni/Al multilayer thin film.

Reactive multilayer thin films that undergo highly exothermic reactions are attractive choices for applications in ignition, propulsion, and joining systems. Ni/Al reactive multilayer thin films were deposited by dc magnetron sputtering with a period of 14 nm. The microstructure of the as-deposited and heat-treated Ni/Al multilayers was studied by transmission electron microscopy (TEM) and scanning transmission electron microscopy (STEM) in plan view and in cross section. The cross-section samples for TEM and STEM were prepared by focused ion beam lift-out technique. TEM analysis indicates that the as-deposited samples were composed of Ni and Al. High-resolution TEM images reveal the presence of NiAl in small localized regions. Microstructural characterization shows that heat treating at 450 and 700°C transforms the Ni/Al multilayered structure into equiaxed NiAl fine grains.

Real-world data from the Portuguese Nivolumab Expanded Access Program (EAP) in previously treated Non Small Cell Lung Cancer (NSCLC).

OBJECTIVE: The main aim of the study was to evaluate the efficacy and safety profile of Nivolumab, an immune-checkpoint-inhibitor antibody, in advanced, previously treated, Non-Small Cell Lung Cancer (NSCLC) patients, in a real world setting. METHODS: We performed a retrospective, multicentre data analysis of patients who were included in the Portuguese Nivolumab Expanded Access Program (EAP). Eligibility criteria included histologically or citologically confirmed NSCLC, stage IIIB and IV, evaluable disease, sufficient organ function and at least one prior line of chemotherapy. The endpoints included Overall Response Rate (ORR), Disease Control Rate (DCR), Progression Free Survival (PFS) and Overall Survival (OS). Safety analysis was performed with the National Cancer Institute Common Terminology Criteria for Adverse Events (CTCAE), version 4.0, and immune-related Adverse Events (irAEs) were treated according to protocol treatment guidelines. Tumour response was assessed using the Response Evaluation Criteria in Solid Tumours (RECIST) version 1.1. Data was analysed using SPSS, version 21.0 (IBM Statistics). RESULTS: From June 2015 to December 2016, a total of 229 patients with advanced NSCLC were enrolled at 30 Portuguese centres. Clinical data were collected up to the end of July 2018. The baseline median age was 64 years (range 37-83) and the majority of patients were males (70.3%) and former/current smokers (69.4%). Patients with non-squamous histology predominated (88.1%), and 67.6% of the patients had received 2 or more prior lines of chemotherapy. Out of 229 patients, data was available for 219 patients (3 patients did not start treatment, while data was unavailable in 7 patients); of the 219 patients, 15.5% were not evaluated for radiological tumour assessment, 1.4% had complete response (CR), 21% partial response (PR), 31% stable disease (SD) and 31.1% progressive disease (PD). Thus, the ORR was 22.4% and DCR was 53.4% in this population. At the time of survival analysis the median PFS was 4.91 months (95% CI, 3.89-6.11) and median OS was 13.21 months (95% CI, 9.89-16.53). The safety profile was in line with clinical trial data. CONCLUSIONS: Efficacy and safety results observed in this retrospective analysis were consistent with observations reported in clinical trials and from other centres.

Evaluation of the antitumoral effect mediated by IL-12 and HSV-tk genes when delivered by a novel lipid-based system.

In the present work, we used a novel albumin-associated lipoplex formulation, containing the cationic lipid 1-palmitoyl-2-oleoyl-sn-glycero-3-ethylphosphocholine (EPOPC) and cholesterol (Chol), to evaluate the antitumoral efficacy of two gene therapy strategies: immuno-gene therapy, mediated by IL-12 gene expression, and "suicide" gene therapy, mediated by HSV-tk gene expression followed by ganciclovir (GCV) treatment. Our data show that, in an animal model bearing a subcutaneous TSA (mouse mammary adenocarcinoma) tumor, intratumoral administration of the albumin-associated complexes containing the plasmid encoding IL-12 results in a strong antitumoral effect, as demonstrated by the smaller tumor size, the higher T-lymphocyte tumor infiltration and the more extensive tumor necrotic and hemorrhagic areas, as compared to that observed in animals treated with control complexes. On the other hand, the application of the "suicide" gene therapy strategy results in a significant antitumoral activity, which is similar to that achieved with the immuno-gene therapy strategy, although involving different antineoplastic mechanisms. For the tested model, albumin-associated complexes were shown to efficiently mediate intratumoral delivery of therapeutic genes, thus leading to a significant antitumoral effect. This finding is particularly relevant since TSA tumors are characterized for being poorly immunogenic, aggressive and exhibiting high proliferation capacity.

Early infection with Trichuris trichiura and allergen skin test reactivity in later childhood.

BACKGROUND: Allergic diseases cause a large and increasing burden in developed countries and in urban centres in middle-income countries. The causes of this increase are unknown and, currently, there are no interventions to prevent the development of allergic diseases. The 'hygiene hypothesis' has tried to explain the increase through a reduction in the frequency of childhood infections causing a failure to program the immune system for adequate immune regulation. Intestinal helminth parasites are prevalent in childhood in developing countries and are associated with a lower prevalence of allergen skin test reactivity and asthma. OBJECTIVES: To investigate whether children who had intestinal helminth infections during early childhood have a lower prevalence of allergen skin test reactivity later in childhood. METHODS: We re-visited a population of 1055 children from whom stool samples had been collected for detection of intestinal helminth infections for another study, and collected new stool samples and performed allergen skin prick testing. Information on potential confounding variables was collected. RESULTS: Children with heavy infections with Trichuris trichiura in early childhood had a significantly reduced prevalence of allergen skin test reactivity in later childhood, even in the absence of T. trichiura infection at the time of skin testing in later childhood. CONCLUSION: Early heavy infections with T. trichiura may protect against the development of allergen skin test reactivity in later childhood. Novel treatments to program immune-regulation in early childhood in a way that mimics the effects of early infections with T. trichiura may offer new strategies for the prevention of allergic disease.

Simultaneous evaluation of viability and Bcl-2 in small-cell lung cancer.

When designing molecular targeted therapeutic strategies against cancer, it is important to correlate protein expression and cell viability. However, such goal can be difficult if performed in separate assays, especially when only a fraction of cells has been efficiently transfected. Therefore, the aim of the present study was to establish a flow cytometry procedure to assess simultaneously Bcl-2 protein level and viability in small-cell lung cancer (SCLC) cells. Viability assessment was performed by staining cells with Annexin V-fluorescein isothiocyanate (FITC) and 7-aminoactinomycin D (7-AAD). Intracellular detection of Bcl-2 was carried out by immunodetection with monoclonal antibodies. Regarding viability determination, the FSC/7-AAD plot identifies the same percentage of viable cells as the FSC/Annexin V-FITC plot, although with greater sensitivity. The procedures involving cells' fixation with 1% paraformaldehyde and permeabilization with digitonin, required for intracellular Bcl-2 immunostaining did not compromise the association of 7-ADD (nor Annexin V-FITC) previously incubated with SCLC cells. It was therefore possible to simultaneously assess cell viability and Bcl-2 protein in SCLC cells. A simple, sensitive, and versatile procedure was established for the first time for the simultaneous evaluation of cell viability and intracellular detection of Bcl-2 in SCLC.

Non-viral lipid-based nanoparticles for targeted cancer systemic gene silencing.

New molecular biology techniques have uncovered the hidden role of genes in cancer. Identification of activated oncogenes, as fundamental genetic differences relative to normal cells, has made it possible to consider such genes as targets for antitumor therapy, namely by applying gene silencing strategies. In this regard, antisense oligonucleotides or small interfering RNAs, constitute promising therapeutic tools. The widespread clinical application of such molecules as modulators of gene expression, is still dependent on several aspects that limit their bioavailability, including: enhanced biological stability, favourable pharmacokinetics, enhanced tumor cell uptake and, consequently, efficient targeted delivery. One of the most promising strategies to overcome the barriers faced by gene silencing molecules, upon systemic administration, involves the use of lipid-based nanoparticles. The first part of this review aims at providing the reader with the molecular mechanism of action of the most important gene silencing molecules used in anticancer therapy. The primary obstacle for translating gene silencing technology from an effective research tool into a feasible therapeutic strategy remains its efficient delivery to the targeted cell type in vivo. Therefore, an overview of different lipid-based strategies for nucleic acid delivery will be presented on the second part. As we learn more about the pharmacokinetics and pharmacodynamics of the carrier and/or of the gene silencing molecules, it will be possible to further improve the delivery strategy that likely in the future will lead to the ideal non-viral particle for targeted cancer systemic gene silencing.

Starch nanocapsules containing a novel neutrophil elastase inhibitor with improved pharmaceutical performance.

Psoriasis and atopic dermatitis patients show an excessive amount of elastase in peripheral blood neutrophils due to an imbalance between this proteolytic enzyme and its endogenous inhibitors, the search for new human neutrophil elastase (HNE) inhibitors are required. The HNE is an attractive therapeutic target and inhibitors with new molecular architectures have been extensively investigated. In this context a promising novel synthetic human neutrophil elastase inhibitor (ER143) was associated to a starch-based nanoparticulate system (StNC) with improved pharmaceutical performance, using a quality by design approach to support product development and optimization. The resulting formulation was characterized in terms of and in vitro release, permeation and retention studies in newborn pig skin, using Franz diffusion cells revealing the StNC have the ability to control the drug release rate and contribute to a high skin retention and/or permeation profiles. The anti-inflammatory activity accessed in vivo using the croton oil-induced ear inflammation model in mice showed that erythema and edema were attenuated in 98% following local application. These observations suggest the association of ER143 to the StNC promotes a deeper skin penetration and retention, also confirming StNC as a potential topical delivery system.

Cellular and molecular events associated with the antitumor response induced by the cytosine deaminase/5-fluorocytosine suicide gene therapy system in a rat liver metastasis model.

The bacterial cytosine deaminase (CD) gene converts the non-toxic prodrug 5-fluorocytosine (5-FC) into 5-fluorouracil. We have previously shown, in a rat liver metastasis model from colon carcinoma, that intratumoral injection of a CD-expressing plasmid into the animals followed by 5-FC treatment results in the regression of the treated tumor as well as distant uninjected tumors. The aim of this study was to further analyze the mechanisms associated with tumor regression induced upon application of suicide CD/5-FC strategy. Tumor regression was associated with an increased apoptosis, the recruitment of natural killer cells, CD4- and CD8 T lymphocytes within the tumors and an increased expression of several cytokines/chemokines mRNAs. These data indicate that the CD/5-FC suicide strategy is associated with the triggering of cellular and molecular events leading to an efficient antitumor immune response involving both innate and acquired immunity.

Trypanosomiasis by Trypanosoma vivax in cattle in the Brazilian semiarid: Description of an outbreak and lesions in the nervous system.

An outbreak of trypanosomiasis by Trypanosoma vivax is reported in the semiarid of Paraíba, Northeastern Brazil from May to August 2002. Sixty-four cows out of 130 were affected; 11 died and the other recovered after treatment with diminazene aceturate. Affected animals had fever, anemia, weight loss, hypoglycemia, increased serum levels of aspartate aminotransferase and, in nine cows, nervous signs. All cows with nervous signs died; six of them recovered after treatment, but the disease relapsed. Six cows aborted and one delivered a calf that died immediately after parturition. Thirty-two out of 100 calves were affected and five died. Nervous signs were not observed in the calves. Gross lesions were thickening of the meninges, enlarged lymph nodes and prominent white pulp of the spleen. The main histological lesion was meningoencephalitis and malacia in the brain of cows with nervous signs. No antibodies against trypanosomes were found in 33 blood samples collected before the outbreak in the affected farm and in 29 samples collected at the same time in two other neighbor farms. Until January 2003, all 89 animals tested had antibodies against T. vivax, suggesting the occurrence of sub clinical infections in cattle without clinical signs. Only two out of 85 serum samples collected on April 2004 were positive for T. vivax antibodies. Data obtained suggested that the semiarid region is non-endemic for trypanosomiasis and that disease occurred due to introduction of the parasite in a susceptible population after an apparent rise in the Tabanus spp. population.

Relief influence on the spatial distribution of the Atlantic Forest cover on the Ibiúna Plateau, SP.

Several studies suggest that, on a large scale, relief conditions influence the Atlantic Forest cover. The aim of this work was to explore these relationships on a local scale, in Caucaia do Alto, on the Ibiúna Plateau. Within an area of about 78 km(2), the distribution of forest cover, divided into two successional stages, was associated with relief attribute data (slope, slope orientation and altitude). The mapping of the vegetation was based on the interpretation of stereoscopic pairs of aerial photographs, from April 2000, on a scale of 1:10,000, while the relief attributes were obtained by geoprocessing from digitalized topographic maps on a scale of 1:10,000. Statistical analyses, based on qui-square tests, revealed that there was a more extensive forest cover, irrespective of the successional stage, in steeper areas (>10 degrees) located at higher altitudes (>923 m), but no influence of the slope orientation. There was no sign of direct influence of relief on the forest cover through environmental gradients that might have contributed to the forest regeneration. Likewise, there was no evidence that these results could have been influenced by the distance from roads or urban areas or with respect to permanent preservation areas. Relief seems to influence the forest cover indirectly, since agricultural land use is preferably made in flatter and lower areas. These results suggest a general distribution pattern of the forest remnants, independent of the scale of study, on which relief indirectly has a strong influence, since it determines human occupation.

Clinical Presentation Resembling Mucosal Disease Associated with 'HoBi'-like Pestivirus in a Field Outbreak.

The genus Pestivirus of the family Flaviviridae consists of four recognized species: Bovine viral diarrhoea virus 1 (BVDV-1), Bovine viral diarrhoea virus 2 (BVDV-2), Classical swine fever virus (CSFV) and Border disease virus (BDV). Recently, atypical pestiviruses ('HoBi'-like pestiviruses) were identified in batches of contaminated foetal calf serum and in naturally infected cattle with and without clinical symptoms. Here, we describe the first report of a mucosal disease-like clinical presentation (MD) associated with a 'HoBi'-like pestivirus occurring in a cattle herd. The outbreak was investigated using immunohistochemistry, antibody detection, viral isolation and RT-PCR. The sequence and phylogenetic analysis of 5'NCR, N(pro) and E2 regions of the RT-PCR positive samples showed that four different 'HoBi'-like strains were circulating in the herd. The main clinical signs and lesions were observed in the respiratory and digestive systems, but skin lesions and corneal opacity were also observed. MD characteristic lesions and a pestivirus with cytopathic biotype were detected in one calf. The present study is the first report of a MD like presentation associated with natural infection with 'HoBi'-like pestivirus. This report describes the clinical signs and provides a pathologic framework of an outbreak associated with at least two different 'HoBi'-like strains. Based on these observations, it appears that these atypical pestiviruses are most likely underdiagnosed in Brazilian cattle.

On the mechanisms of internalization and intracellular delivery mediated by pH-sensitive liposomes.

We investigated the molecular mechanisms by which pH-sensitive liposomes surpass the cytoplasmic and endosomal membranes to deliver their aqueous contents into the cytoplasm. Various liposome formulations were evaluated for their efficacy to mediate intracellular delivery of encapsulated material, including a novel sterically stabilized pH-sensitive formulation ((DOPE:CHEMS:DSPE-PEG(2000) (6:4:0.3)) that was previously developed in our laboratories. In an attempt to fully characterize the nature of liposome-cell interactions different approaches based on a dual-labeling fluorescence assay were used. Our results indicate that the efficacy of interaction of pH-sensitive liposomes, both plain and sterically stabilized, with cells is strongly determined by the inclusion of DOPE in their composition, independently of the type of the amphiphilic stabilizer used. In fact, DOPE-containing liposomes shown to be non-pH sensitive by biophysical assays, mediated cytoplasmic delivery of their contents as efficiently as well known pH-sensitive formulations (e.g. DOPE:CHEMS). However, among the different formulations studied, DOPE:CHEMS liposomes were those exhibiting the highest extent of cell association. Moreover, our results with cells pretreated with metabolic inhibitors or lysosomotropic agents clearly indicate that DOPE-containing liposomes are internalized essentially by endocytosis and that acidification of the endosomes is not the only mechanism involved in the destabilization of the liposomes inside the cell.

Evaluation of lipid-based reagents to mediate intracellular gene delivery.

We characterized different cationic lipid-based gene delivery systems consisting of both liposomes and nonliposomal structures, in terms of their in vitro transfection activity, resistance to the presence of serum, protective effect against nuclease degradation and stability under different storage conditions. The effect of lipid/DNA charge ratio of the resulting complexes on these properties was also evaluated. Our results indicate that the highest levels of transfection activity were observed for complexes prepared from nonliposomal structures composed of FuGENE 6. However, their DNA protective effect was shown to be lower than that observed for cationic liposome formulations when prepared at the optimal (+/-) charge ratio. Our results suggest that lipoplexes are resistant to serum up to 30% when prepared at a 2:1 lipid/DNA charge ratio. However, when they were prepared at higher (+/-) charge ratios, they become sensitive to serum for even lower concentrations (10%). Replacement of dioleoyl-phosphatidylethanolamine (DOPE) by cholesterol enhanced the resistance of the complexes to the inhibitory effect of serum. This different biological activity in the presence of serum was attributed to different extents of binding of serum proteins to the complexes, as evaluated by the immunoblotting assay. Studies on the stability under storage show that lipoplexes maintain most of their biological activity when stored at -80 degrees C, following their fast freezing in liquid nitrogen.

Human serum albumin enhances DNA transfection by lipoplexes and confers resistance to inhibition by serum.

Cationic liposome-DNA complexes ('lipoplexes') are used as gene delivery vehicles and may overcome some of the limitations of viral vectors for gene therapy applications. The interaction of highly positively charged lipoplexes with biological macromolecules in blood and tissues is one of the drawbacks of this system. We examined whether coating cationic liposomes with human serum albumin (HSA) could generate complexes that maintained transfection activity. The association of HSA with liposomes composed of 1, 2-dioleoyl-3-(trimethylammonium) propane and dioleoylphosphatidylethanolamine, and subsequent complexation with the plasmid pCMVluc greatly increased luciferase expression in epithelial and lymphocytic cell lines above that obtained with plain lipoplexes. The percentage of cells transfected also increased by an order of magnitude. The zeta potential of the ternary complexes was lower than that of the lipoplexes. Transfection activity by HSA-lipoplexes was not inhibited by up to 30% serum. The combined use of HSA and a pH-sensitive peptide resulted in significant gene expression in human primary macrophages. HSA-lipoplexes mediated significantly higher gene expression than plain lipoplexes or naked DNA in the lungs and spleen of mice. Our results indicate that negatively charged HSA-lipoplexes can facilitate efficient transfection of cultured cells, and that they may overcome some of the problems associated with the use of highly positively charged complexes for gene delivery in vivo.

Interaction of cationic liposomes and their DNA complexes with monocytic leukemia cells.

Cationic liposomes complexed with DNA have been used extensively as non-viral vectors for the intracellular delivery of reporter or therapeutic genes in culture and in vivo. We examined the relationship between the characteristics of the lipoplexes, their mode of interaction with monocytic THP-1 cells and their ability to transfect these cells. We determined the size and zeta potential of cationic liposomes (composed of 1,2-dioleoyl-3-(trimethylammonium) propane (DOTAP) and its mixtures with neutral lipids), and lipoplexes at different (+/-) charge ratios. As the (+/-) charge ratio of the lipoplexes decreased to (1/1), a significant reduction in zeta potential and an increase in size was observed. The increase in size resulted from fusion between liposomes promoted by DNA, as demonstrated by a lipid mixing assay, and from aggregation of the complexes. Interaction of liposomes and lipoplexes with THP-1 cells was assessed by monitoring lipid mixing ('fusion') as well as binding and cell association. While no lipid mixing was observed with the 1/2 (+/-) lipid/DNA complexes, lipoplexes with higher (+/-) charge ratios underwent significant fusion in conjunction with extensive cell binding. Liposome binding to cells was dependent on the positive charge of the liposomes, and their fusion could be modulated by the co-lipid. DOTAP/phosphatidylethanolamine (1:1) liposomes fused with THP-1 cells, unlike DOTAP/phosphatidylcholine (1:1) liposomes, although both liposome types bound to the cells to a similar extent. The use of inhibitors of endocytosis indicated that fusion of the cationic liposomes with cells occurred mainly at the plasma membrane level. The presence of serum increased the size of the cationic liposomes, but not that of the lipoplexes. Low concentrations of serum (3%) completely inhibited the fusion of cationic liposomes with cells, while inhibiting binding by only 20%. Our results suggest that binding of cationic liposomes and lipoplexes to cells is governed primarily by electrostatic interactions, whereas their fusion is regulated by the lipid composition and sterically favorable interactions with cell surface molecules. In addition our results indicate no correlation between fusion of the lipoplexes with the plasma membrane and the levels of transfection.