Surface specific peptide immobilization on radiografted polymers as potential screening assays for antiangiogenic immunotherapy

International audienceAngiogenesis is a key process of cancer development and metastasis. It's inhibition is an important and promising strategy to block tumor growth and invasion. One of these approaches, based on antiangiogenic immunotherapy, is the recognition of a specific region of an angiogenic growth factor, called VEGF-A, by monoclonal antibodies. Thus, we aimed to design a novel assay to screen potential monoclonal antibodies directed against VEGF-A. In a first approach, we chose to perform covalent coupling of angiogenesis active cyclopeptides onto biocompatible thermoplastic transparent PVDF films and to fully characterize the chemical structure, the surface state and the biochemical properties of the synthesized devices. Electron beam radiation created radical sites on PVDF films without adding any toxic chemicals. These primary radicals and some induced peroxides were used as initiators for acrylic acid polymerization. Under our experimental conditions, surface grafting was favoured. Functionalization of PVDF-g-PAA films with peptides via a spacer arm was possible by performing two subsequent coupling reactions. EDC was used as coupling agent. Spacer arm saturation of the film surface was achieved for 25 mol% yield meaning that one spacer arm on four carboxylic acids were covalently bound. Peptide immobilization resulted in binding 10 times less leading to a final 3 mol% yield. Binding densities are governed by their individual space requirements. Each chemical step has been followed by FTIR in ATR mode, NMR using HR MAS technique and XPS. From XPS results, a layer of peptide covered PVDF-g-PAA film surface. The amounts of covalently immobilized peptide were determined using indirect UV spectroscopy on supernatant reaction solution. Yields were correlated with high resolution NMR results. The peptide/antibody recognition validated our system showing the conservation of peptide tridimensional structure with a positive response to specific antibodies. Because of the covalent protein linkage to PVDF films, a simple cleaning with immunoaffinity chromatography buffer allows the films to be reused

Serine residue 115 of MAPK-activated protein kinase MK5 is crucial for its PKA-regulated nuclear export and biological function

The mitogen-activated protein kinase-activated protein kinase-5 (MK5) resides predominantly in the nucleus of resting cells, but p38MAPK, extracellular signal-regulated kinases-3 and -4 (ERK3 and ERK4), and protein kinase A (PKA) induce nucleocytoplasmic redistribution of MK5. The mechanism by which PKA causes nuclear export remains unsolved. In the study reported here we demonstrated that Ser-115 is an in vitro PKA phosphoacceptor site, and that PKA, but not p38MAPK, ERK3 or ERK4, is unable to redistribute MK5 S115A to the cytoplasm. However, the phosphomimicking MK5 S115D mutant resides in the cytoplasm in untreated cells. While p38MAPK, ERK3 and ERK4 fail to trigger nuclear export of the kinase dead T182A and K51E MK5 mutants, S115D/T182A and K51E/S115D mutants were able to enter the cytoplasm of resting cells. Finally, we demonstrated that mutations in Ser-115 affect the biological properties of MK5. Taken together, our results suggest that Ser-115 plays an essential role in PKA-regulated nuclear export of MK5, and that it also may regulate the biological functions of MK5

Biochemical aspects of overtraining in endurance sports: A review

SCOPUS: re.jinfo:eu-repo/semantics/publishe

Current Trends in the Development of FTIR Imaging for the Quantitative Analysis of Biological Samples

Fourier transform infrared imaging instrumentation has come of age for the rapid acquisition of biosample IR images, and thus now allows developing analytical methods based upon the molecular information of samples contents. For the biomedicine field, Fourier transform infrared imaging should be able to play a role in the molecular characterization of cells and tissues where no other analytical method provides quantitative information. Now, a compromise may be obtained between an acceptable acquisition time of IR images, which should never exceed a few tens of minutes, and the necessary spatial resolution (down to the diffraction limit, although limited for biology), spectral resolution (2 to 8 $cm^{-1}$ for biosample analyses), and signal-to-noise ratio level, to provide a diagnostic answer to clinicians during the surgery time. Here, we will discuss the potential of Fourier transform infrared imaging in face to major pathologies (myopathies, brain tumors, metabolic diseases) for which current imaging methods remain unable to provide sufficient information for a precise diagnosis. Quantitative molecular information may be extracted from infrared images of samples as soon as samples volume/thickness is controlled as well as absorption and absorptivity of the molecules to analyze may be isolated from other absorbing compounds. In this context, metabolic parameters appear as the main targets for providing critical information about the physiological status of a biosample due to their characteristic infrared spectra. As examples, it will be shown how to isolate and quantify glucose, lactic-acid, urea, etc. absorptions from complex biosample infrared images

Current Trends in the Development of FTIR Imaging for the Quantitative Analysis of Biological Samples

Fourier transform infrared imaging instrumentation has come of age for the rapid acquisition of biosample IR images, and thus now allows developing analytical methods based upon the molecular information of samples contents. For the biomedicine field, Fourier transform infrared imaging should be able to play a role in the molecular characterization of cells and tissues where no other analytical method provides quantitative information. Now, a compromise may be obtained between an acceptable acquisition time of IR images, which should never exceed a few tens of minutes, and the necessary spatial resolution (down to the diffraction limit, although limited for biology), spectral resolution (2 to 8 $cm^{-1}$ for biosample analyses), and signal-to-noise ratio level, to provide a diagnostic answer to clinicians during the surgery time. Here, we will discuss the potential of Fourier transform infrared imaging in face to major pathologies (myopathies, brain tumors, metabolic diseases) for which current imaging methods remain unable to provide sufficient information for a precise diagnosis. Quantitative molecular information may be extracted from infrared images of samples as soon as samples volume/thickness is controlled as well as absorption and absorptivity of the molecules to analyze may be isolated from other absorbing compounds. In this context, metabolic parameters appear as the main targets for providing critical information about the physiological status of a biosample due to their characteristic infrared spectra. As examples, it will be shown how to isolate and quantify glucose, lactic-acid, urea, etc. absorptions from complex biosample infrared images

Modifying PTR-MS operating conditions for quantitative headspace analysis of hydro-alcoholic beverages. 2. Brandy characterization and discrimination by PTR-MS

Abstract not availableGuillaume Fiches, Isabelle Déléris, Anne Saint-Eve, Pascal Brunerie, Isabelle Soucho

Modifying PTR-MS operating conditions for quantitative headspace analysis of hydro-alcoholic beverages. 1. Variation of the mean collision energy to control ionization processes occurring during PTR-MS analyses of 10-40% (v/v) ethanol-water solutions

Abstract not availableGuillaume Fiches, Isabelle Déléris, Anne Saint-Eve, Brigitte Pollet, Pascal Brunerie, Isabelle Soucho

Phosphorus limitation strategy to increase propionic acid flux towards 3-hydroxyvaleric acid monomers in [i]Cupriavidus necator[/i]

Properties of polyhydroxybutyrate-co-hydroxyvalerate (P(3HB-co-3HV)) depend on their 3HV content. 3HV can be produced by [i]Cupriavidus necator[/i] from propionic acid. Few studies explored carbon distribution and dynamics of 3HV and 3HB monomers production, and none of them have been done with phosphorus as limiting nutrient. In this study, fed-batch cultures of [i]C. necator[/i] with propionic acid, as sole carbon source or mixed with butyric acid, were performed. Phosphorus deficiency allowed sustaining 3HV production rate and decreasing 3HB production rate, leading to an instant production of up to 100% of 3HV. When a residual growth is sustained by a phosphorus feeding, the maximum 3HV percentage produced from propionic acid is limited to 33% (Mole. Mole (1)). The association of a second carbon source like butyric acid lead to higher conversion of propionic acid into 3HV. This study showed the importance of the limiting nutrient and of the culture strategy to get the appropriate product