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ABSTRACT: In this work, degradable cationic shell cross-linked knedel-like (deg-cSCK) nanoparticles were developed as an alternative platform to replace similar nondegradable cSCK nanoparticles that have been utilized for nucleic acids delivery. An amphiphilic diblock copolymer poly(acrylamidoethylamine)90-block-poly(dl-lactide)40 (PAEA90-b-PDLLA40) was synthesized, self-assembled in aqueous solution, and shell cross-linked using a hydrolyzable cross-linker to afford deg-cSCKs with an average core diameter of 45 ± 7 nm. These nanoparticles were fluorescently labeled for in vitro tracking. The enzymatic- and hydrolytic-degradability, siRNA binding affinity, cell uptake and cytotoxicity of the deg-cSCKs were evaluated. Esterase-catalyzed hydrolysis of the nanoparticles resulted in the degradation of ca. 24% of the PDLLA core into lactic acid within 5 d, as opposed to only ca. 9% degradation from aqueous solutions of the deg-cSCK nanoparticles in the absence of enzyme. Cellular uptake of deg-cSCKs was efficient, while exhibiting low cytotoxicity with LD50 values of ca. 90 and 30 μg/mL in RAW 264.7 mouse macrophages and MLE 12 cell lines, respectively, ca. 5- to 6-fold lower than the cytotoxicity observed for nondegradable cSCK analogs. Additionally, deg-cSCKs were able to complex siRNA at an N/P ratio as low as 2, and were efficiently able to facilitate cellular uptake of the complexed nucleic acids.
Biomacromolecules 03/2013; · 5.48 Impact Factor
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ABSTRACT: Cilia are traditionally classified as motile or primary. Motile cilia are restricted to specific populations of well-differentiated epithelial cells, including those in the airway, brain ventricles, and oviducts. Primary cilia are nonmotile, solitary structures that are present in many cell types, and often have sensory functions such as in the retina and renal tubules. Primary cilia were also implicated in the regulation of fundamental processes in development. Rare depictions of primary cilia in embryonic airways led us to hypothesize that primary cilia in airway cells are temporally related to motile ciliogenesis. We identified primary cilia in undifferentiated, cultured airway epithelial cells from mice and humans and in developing lungs. The solitary cilia in the airways express proteins considered unique to primary cilia, including polycystin-1 and polycystin-2. A temporal analysis of airway epithelial cell differentiation showed that cells with primary cilia acquire markers of motile ciliogenesis, suggesting that motile ciliated cells originate from primary ciliated cells. Whereas motile ciliogenesis requires Foxj1, primary ciliogenesis does not, and the expression of Foxj1 was associated with a loss of primary cilia, just before the appearance of motile cilia. Primary cilia were not found in well-differentiated airway epithelial cells. However, after injury, they appear in the luminal layer of epithelium and in basal cells. The transient nature of primary cilia, together with the temporal and spatial patterns of expression in the development and repair of airway epithelium, suggests a critical role of primary cilia in determining outcomes during airway epithelial cell differentiation.
American Journal of Respiratory Cell and Molecular Biology 12/2010; 43(6):731-9. · 5.13 Impact Factor
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ABSTRACT: Viral bronchiolitis is the leading cause of hospitalization in young infants. It is associated with the development of childhood asthma and contributes to morbidity and mortality in the elderly. Currently no therapies effectively attenuate inflammation during the acute viral infection, or prevent the risk of post-viral asthma. We hypothesized that early treatment of a paramyxoviral bronchiolitis with azithromycin would attenuate acute and chronic airway inflammation.
Mice were inoculated with parainfluenza type 1, Sendai Virus (SeV), and treated daily with PBS or azithromycin for 7 days post-inoculation. On day 8 and 21 we assessed airway inflammation in lung tissue, and quantified immune cells and inflammatory mediators in bronchoalveolar lavage (BAL).
Compared to treatment with PBS, azithromycin significantly attenuated post-viral weight loss. During the peak of acute inflammation (day 8), azithromycin decreased total leukocyte accumulation in the lung tissue and BAL, with the largest fold-reduction in BAL neutrophils. This decreased inflammation was independent of changes in viral load. Azithromycin significantly attenuated the concentration of BAL inflammatory mediators and enhanced resolution of chronic airway inflammation evident by decreased BAL inflammatory mediators on day 21.
In this mouse model of paramyxoviral bronchiolitis, azithromycin attenuated acute and chronic airway inflammation. These findings demonstrate anti-inflammatory effects of azithromycin that are not related to anti-viral activity. Our findings support the rationale for future prospective randomized clinical trials that will evaluate the effects of macrolides on acute viral bronchiolitis and their long-term consequences.
Respiratory research 01/2010; 11:90. · 3.36 Impact Factor
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ABSTRACT: Polymer chemistry offers the possibility of synthesizing multifunctional nanoparticles which incorporate moieties that enhance diagnostic and therapeutic targeting of cargo delivery to the lung. However, since rules for predicting particle behavior following modification are not well-defined, it is essential that probes for tracking fate in vivo are also included. Accordingly, we designed polyacrylamide-based hydrogel particles of differing sizes, functionalized with a nona-arginine cell-penetrating peptide (Arg(9)), and labeled with imaging components to assess lung retention and cellular uptake after intratracheal administration. Radiolabeled microparticles (1-5 microm diameter) and nanoparticles (20-40 nm diameter) without and with Arg(9) showed diffuse airspace distribution by positron emission tomography imaging. Biodistribution studies revealed that particle clearance and extrapulmonary distribution was, in part, size dependent. Microparticles were rapidly cleared by mucociliary routes but, unexpectedly, also through the circulation. In contrast, nanoparticles had prolonged lung retention enhanced by Arg(9) and were significantly restricted to the lung. For all particle types, uptake was predominant in alveolar macrophages and, to a lesser extent, lung epithelial cells. In general, particles did not induce local inflammatory responses, with the exception of microparticles bearing Arg(9). Whereas microparticles may be advantageous for short-term applications, nanosized particles constitute an efficient high-retention and non-inflammatory vehicle for the delivery of diagnostic imaging agents and therapeutics to lung airspaces and alveolar macrophages that can be enhanced by Arg(9). Importantly, our results show that minor particle modifications may significantly impact in vivo behavior within the complex environments of the lung, underscoring the need for animal modeling.
Molecular Pharmaceutics 10/2009; 6(6):1891-902. · 4.78 Impact Factor
