Biomedical subjects

Sally-Ann Cryan

Publications and source records attributed to Sally-Ann Cryan.

5 recordsLinked to original sources

PubMed2005

Secretory leucoprotease inhibitor binds to NF-kappaB binding sites in monocytes and inhibits p65 binding.

Secretory leucoprotease inhibitor (SLPI) is a nonglycosylated protein produced by epithelial cells. In addition to its antiprotease activity, SLPI has been shown to exhibit antiinflammatory properties, including down-regulation of tumor necrosis factor alpha expression by lipopolysaccharide (LPS) in macrophages and inhibition of nuclear factor (NF)-kappaB activation in a rat model of acute lung injury. We have previously shown that SLPI can inhibit LPS-induced NF-kappaB activation in monocytic cells by inhibiting degradation of IkappaBalpha without affecting the LPS-induced phosphorylation and ubiquitination of IkappaBalpha. Here, we present evidence to show that upon incubation with peripheral blood monocytes (PBMs) and the U937 monocytic cell line, SLPI enters the cells, becoming rapidly localized to the cytoplasm and nucleus, and affects NF-kappaB activation by binding directly to NF-kappaB binding sites in a site-specific manner. SLPI can also prevent p65 interaction with the NF-kappaB consensus region at concentrations commensurate with the physiological nuclear levels of SLPI and p65. We also demonstrate the presence of SLPI in nuclear fractions of PBMs and alveolar macrophages from individuals with cystic fibrosis and community-acquired pneumonia. Therefore, SLPI inhibition of NF-kappaB activation is mediated, in part, by competitive binding to the NF-kappaB consensus-binding site.

Animals↗

PubMed2005

Carrier-based strategies for targeting protein and peptide drugs to the lungs.

With greater interest in delivery of protein and peptide-based drugs to the lungs for topical and systemic activity, a range of new devices and formulations are being investigated. While a great deal of recent research has focused on the development of novel devices, attention must now be paid to the formulation of these macromolecular drugs. The emphasis in this review will be on targeting of protein/peptide drugs by inhalation using carriers and ligands.

Administration, Inhalation↗

PubMed2004

Cell transfection with polycationic cyclodextrin vectors.

Polycationic cyclodextrins (CDs) were complexed with plasmid DNA and their effectiveness as vectors was tested on COS-7 cells. These CDs were modified with pyridylamino, alkylimidazole, methoxyethylamino or primary amine groups at 6-positions of the glucose units. Uncharged CDs, beta-CD, hydroxypropyl-beta-CD, and dimethyl-beta-CD were also tested, but these did not form stable complexes with the DNA and produced only a slight improvement in transfection level over DNA alone. The polycationic CDs neutralised DNA to form stable nanoparticulate complexes. The transfection efficiency of these CDs was dependent on the substituents present, with the most efficient having either an amino, pyridylamino or butylimidazole group at the 6-positions and unmodified 2- and 3-hydroxyls. One of the most effective vectors, heptakispyridylamino CD, produced a 4000-fold increase in transfection level over DNA alone. Levels were improved 10-fold by use of the endosomolytic agent, chloroquine. The transfection efficiency of the best of these systems in serum equals that of DOTAP in serum. Studies with (32)P-labelled plasmid DNA indicate that the polycationic CDs are exceptional promoters of DNA cellular-uptake, the most efficient surpassing DOTAP. Uptake is dependent on proteoglycan-mediated binding to cells. The data imply that intracellular trafficking but not cellular uptake, may be the rate-limiting step in the transfection process. These initial results indicate that CDs are useful templates for further modification to produce molecular constructs capable of enhanced gene delivery.

Animals↗

PubMed2003

Mechanistic studies on nonviral gene delivery to the intestine using in vitro differentiated cell culture models and an in vivo rat intestinal loop.

PURPOSE: To identify factors influencing nonviral vector transfection in differentiated CaCo-2 and mucus-secreting coculture, CaCo-2: Ht29GlucH, cell culture models and to compare these in vitro results with in vivo transfection efficiency in rat intestine. METHODS: A range of nonviral vectors including DOTAP, Lipofectin, Superfect, PEI, and polylysine were investigated. CaCo-2 and a mucus-secreting coculture were used at 21 days. Transfection efficiency was assessed using pCMVluc (firefly luciferase) plasmid, and radio-labeled plasmid was used to determine the binding and internalization of plasmid DNA. The in vivo model used was a ligated rat intestinal loop. RESULTS: Transfection levels decreased by over 1000-fold in differentiated models relative to nondifferentiated COS-7 cells and were related to reductions in luciferase production by individual cells. Active internalization of DNA by the differentiated cells decreased. Removal of mucus by the mucolytic agent N-acetylcysteine, from the coculture system significantly reduced (p < 0.05) transfection efficiency. In vivo the transfection efficiency of PEI proved superior to DOTAP. CONCLUSIONS: Nonviral gene delivery to the hostile environment of the intestine is possible. Mechanistic studies using differentiated intestinal cell models aid identification of the rate-limiting steps to transfection and represent a more physiologically relevant approach to predict gene delivery to the intestine.

Animals↗

PubMedDate not supplied

Increased intracellular targeting to airway cells using octaarginine-coated liposomes: in vitro assessment of their suitability for inhalation.

Delivery of macromolecular drugs to airway cells after inhalation can be limited by rapid clearance, in vivo degradation, and poor intracellular targeting. Liposome carriers offer an effective method of improving drug stability, but conventional liposomes have limited intracellular targeting capacity and are cleared rapidly by the lungs. Further modification is required to improve liposome-cell interaction and intracellular targeting. Therefore, we proposed conjugating three arginine-rich membrane translocating peptides, namely, HIV-TAT, Antennapedia, and octaarginine, to neutral liposomes as a biocompatible alternative to cationic lipids for intracellular delivery of macromolecules to airway cells. Conjugation did not significantly affect liposome stability, and each system was nebulized to produce aerosols of mean aerodynamic diameter < 1.5 microm. The peptides caused a significant (p < 0.05) increase in liposome-airway cell association compared to untagged liposomes and to DOTAP liposomes. Up to 30% of the peptide-conjugated liposomes added were bound and internalized (via a temperature-dependent, endocytic process) after just 2 h. The novel carriers all delivered encapsulated dextrans rapidly and efficiently to the cytoplasm of Calu-3 cells. Once internalized by the cells, the modified carriers localize for the most part in the cytoplasm with only a small amount of nuclear localization. These peptide-conjugated liposomes were significantly (p < 0.05) less toxic than DOTAP liposomes with octaarginine-coated liposomes the least toxic. These systems, particularly octaarginine-coated liposomes, offer many advantages for drug delivery to airway epithelial cells including increased stability, improved cell binding, and cell uptake with an improved toxicity profile.

Cell Adhesion↗