Effects of extrusion, lipid concentration and purity on physico-chemical and biological properties of cationic liposomes for gene vaccine applications

We developed cationic liposomes containing DNA through a conventional process involving steps of (i) preformation of liposomes, (ii) extrusion, (iii) drying and rehydration and (iv) DNA complexation. Owing to its high prophylactic potentiality against tuberculosis, which had already been demonstrated in preclinical assays, we introduced modifications into the conventional process towards getting a simpler and more economical process for further scale-up. Elimination of the extrusion step, increasing the lipid concentration (from 16 to 64 mM) of the preformed liposomes and using good manufacturing practice bulk lipids (96-98% purity) instead of analytical grade purity lipids (99.9-100%) were the modifications studied. The differences in the physico-chemical properties, such as average diameter, zeta potential, melting point and morphology of the liposomes prepared through the modified process, were not as significant for the biological properties, such as DNA loading on the cationic liposomes, and effective immune response in mice after immunisation as the control liposomes prepared through the conventional process. Beneficially, the modified process increased productivity by 22% and reduced the cost of raw material by 75%.

Bacterial leakage along the implant-abutment interface: culture and DNA Checkerboard hybridization analyses

Objective Bacterial species have been found harboring the internal surface of dental implants as consequence of their failed connections. The aim of the present study was to compare the detection frequency of bacterial leakage from human saliva through the implantabutment interface, under non-loading conditions, using either DNA Checkerboard or culture method. Materials and methods Thirty dental implants with hexagonal platforms were connected to pre-machined abutments according to the manufacturers specifications. The assemblies were individually incubated in human saliva under anaerobic conditions for 7 similar to days at 37 degrees C. Afterward, contents from the inner parts of the implants were collected and evaluated with either DNA Checkerboard (s similar to=similar to 15) or culture (n similar to=similar to 15). Subsequently, identification and quantitation of bacterial species from saliva and implants were carried out for the group evaluated with the DNA Checkerboard method. Results Both DNA Checkerboard and culture showed positive signals of bacterial leakage in 6 of the 15 evaluated samples. Capnocytophaga gingivalis and Streptococcus mutans were the most frequently detected species harboring the internal surface of the implants followed by Veillonella parvula. Conclusion Occurrence of bacterial leakage along the implantabutment interface is comparably detected with both DNA Checkerboard hybridization and conventional culture methods.

Mechanical loading induces the expression of a Pol I regulon at the onset of skeletal muscle hypertrophy

von Walden F, Casagrande V, Ostlund Farrants AK, Nader GA. Mechanical loading induces the expression of a Pol I regulon at the onset of skeletal muscle hypertrophy. Am J Physiol Cell Physiol 302: C1523-C1530, 2012. First published March 7, 2012; doi:10.1152/ajpcell.00460.2011.-The main goal of the present study was to investigate the regulation of ribosomal DNA (rDNA) gene transcription at the onset of skeletal muscle hypertrophy. Mice were subjected to functional overload of the plantaris by bilateral removal of the synergist muscles. Mechanical loading resulted in muscle hypertrophy with an increase in rRNA content. rDNA transcription, as determined by 45S pre-rRNA abundance, paralleled the increase in rRNA content and was consistent with the onset of the hypertrophic response. Increased transcription and protein expression of c-Myc and its downstream polymerase I (Pol I) regulon (POL1RB, TIF-1A, PAF53, TTF1, TAF1C) was also consistent with the increase in rRNA. Similarly, factors involved in rDNA transcription, such as the upstream binding factor and the Williams syndrome transcription factor, were induced by mechanical loading in a corresponding temporal fashion. Chromatin immunoprecipitation revealed that these factors, together with Pol I, were enriched at the rDNA promoter. This, in addition to an increase in histone H3 lysine 9 acetylation, demonstrates that mechanical loading regulates rRNA synthesis by inducing a gene expression program consisting of a Pol I regulon, together with accessory factors involved in transcription and chromatin remodeling at the rDNA promoter. Altogether, these data indicate that transcriptional and epigenetic mechanisms take place in the regulation of ribosome production at the onset of muscle hypertrophy.