Measuring protein structure and stability of protein-nanoparticle systems with synchrotron radiation circular dichroism.

We measure the structural and stability changes of proteins at nanomolar concentration upon interaction with nanoparticles. Using synchrotron radiation circular dichroism (SRCD), we measure a decrease of 6 °C in the thermal unfolding of human serum albumin upon interaction with silver nanoparticles while this does not happen with gold. The use of SRCD allows measuring critical parameters on protein-nanoparticle interactions, and it will provide experimental data on the relative stability of key biological proteins for nanotoxicology.

Effect of native and NH3 plasma-functionalized polymeric membranes on the gene expression profiles of primary hepatocytes.

Little is known about how cells respond to different biomaterials at the molecular level. Biomaterials could stimulate specific cellular responses at the molecular level, such as activation of signalling pathways that control gene activity involved in the maintenance, growth and functional regeneration of liver tissue in vitro. This aspect is an important step in liver tissue engineering. Currently, there are no data available concerning the modulation of cellular genomic response by using synthetic membranes in a bioartificial system. For the first time we investigated gene expression profiles of primary hepatocytes cultured on different substrates: collagen sandwich, native and NH(3) plasma-grafted PEEK-WC-PU membranes. Gene expression in cell suspension prepared after cell isolation was used as a control. Generally, microarray data revealed that the expression of the majority of genes remained unchanged compared to the control. Among 31 000 genes, 52 were significantly changed: 20 were upregulated and 32 downregulated. There were similar changes in gene expression of hepatocytes cultured in the membranes and collagen sandwich. However, some genes involved in the cell proliferation and functional metabolic pathways are more expressed in cells cultured on the membranes and especially on the functionalized ones. Both membranes sustained liver functions at the molecular level, demonstrating their suitability for the reconstruction of liver and as a toxicogenomic tool to predict the liver response to novel drugs.

Improved functions of human hepatocytes on NH3 plasma-grafted PEEK-WC-PU membranes.

PEEK-WC-PU membranes were modified with an NH(3) glow discharge process to graft N-containing functional groups at their surface in order to improve the maintenance of human hepatocytes. Native and modified membrane surfaces were characterized with XPS, ToF-SIMS and WCA measurements. We have investigated morphological behaviour and specific functions of primary human hepatocytes on native and modified PEEK-WC-PU membranes in a small-scale gas-permeable bioreactor. N-containing groups grafted at the surface of the membranes improved the initial steps of adhesion and the maintenance of phenotype and differentiated functions of cells. Confocal microscopy of cell morphology evidenced human hepatocytes exhibiting a polygonal shape and organizing a 3D structure. The presence of CK19 positive cells, a marker of biliary duct epithelium, was also found on native and modified membranes. Liver specific functions, investigated in terms of urea production, albumin synthesis and diazepam biotransformation, were maintained at high levels up to 19 days, particularly on surface-modified membranes.

Rat embryonic liver cell expansion and differentiation on NH3 plasma-grafted PEEK-WC-PU membranes.

Biomaterials can potentially influence stem and progenitor cell proliferation and differentiation in both a positive and a negative way. Herein, we report on the expansion and differentiation of rat embryonic (E17) liver (RLC-18) cells on new bioactive membrane made of PEEK-WC-PU, whose surface was grafted with nitrogen functionalities by means of NH(3) glow discharges. The performance of the developed membrane was evaluated by analyzing the expression of the liver specific functions of cells cultured in a 6-well gas-permeable bioreactor. It was found that native and NH(3) plasma-grafted PEEK-WC-PU membranes enabled expansion of liver cells in the bioreactor. Liver embryonic cells on the membranes exhibited higher functional activities compared to those cultured on conventional culture dishes as demonstrated by higher albumin and urea production. They showed gene expression of alpha-fetoprotein and albumin in a time-dependent manner of the hepatic differentiation process. LDH assay and SEM analyses revealed that a high number of viable liver stem cells attached to the membranes. Unexpectedly, liver progenitors cultured on membranes had higher telomerase activity than ones in the plates, preventing cell senescence. Thus, membranes are able to sustain in vitro the same in vivo liver functions and to allow the expansion of progenitor cells.

Pd nanoparticles catalyzed stereospecific synthesis of beta-aryl cinnamic esters in ionic liquids.

Reaction of the catalyst 1 or Pd(OAc)(2) with tetrabutylammonium acetate, dissolved in tetrabutylammonium bromide, leads to a fast formation of Pd nanoparticles which efficiently catalyze the stereospecific reaction of cinnamates with aryl halides to give beta-aryl-substituted cinnamic esters. The role of tetrabutylammonium acetate is crucial in determining the formation of nanoparticles and stereospecificity of the C-C coupling process.