Identification of novel non-myelin biomarkers in multiple sclerosis using an improved phage-display approach

Although the etiology of multiple sclerosis is not yet understood, it is accepted that its pathogenesis involves both autoimmune and neurodegenerative processes, in which the role of autoreactive T-cells has been elucidated. Instead, the contribution of humoral response is still unclear, even if the presence of intrathecal antibodies and B-cells follicle-like structures in meninges of patients has been demonstrated. Several myelin and non-myelin antigens have been identified, but none has been validated as humoral biomarker. In particular autoantibodies against myelin proteins have been found also in healthy individuals, whereas non-myelin antigens have been implicated in neurodegenerative phase of the disease. To provide further putative autoantigens of multiple sclerosis, we investigated the antigen specificity of immunoglobulins present both in sera and in cerebrospinal fluid of patients using phage display technology in a new improved format. A human brain cDNA phage display library was constructed and enriched for open-read-frame fragments. This library was selected against pooled and purified immunoglobulins from cerebrospinal fluid and sera of multiple sclerosis patients. The antigen library was also screened against an antibody scFv library obtained from RNA of B cells purified from the cerebrospinal fluid of two relapsing remitting patients. From all biopanning a complex of 14 antigens were identified; in particular, one of these antigens, corresponding to DDX24 protein, was present in all selections. The ability of more frequently isolated antigens to discriminate between sera from patients with multiple sclerosis or other neurological diseases was investigated. The more promising novel candidate autoantigens were DDX24 and TCERG1. Both are implicated in RNA modification and regulation which can be altered in neurodegenerative processes. Therefore, we propose that they could be a marker of a particular disease activity state

Novel Protocol for the Chemical Synthesis of Crustacean Hyperglycemic Hormone Analogues — An Efficient Experimental Tool for Studying Their Functions

The crustacean Hyperglycemic Hormone (cHH) is present in many decapods in different isoforms, whose specific biological functions are still poorly understood. Here we report on the first chemical synthesis of three distinct isoforms of the cHH of Astacus leptodactylus carried out by solid phase peptide synthesis coupled to native chemical ligation. The synthetic 72 amino acid long peptide amides, containing L- or D-Phe3 and (Glp1, D-Phe3) were tested for their biological activity by means of homologous in vivo bioassays. The hyperglycemic activity of the D-isoforms was significantly higher than that of the L-isoform, while the presence of the N-terminal Glp residue had no influence on the peptide activity. The results show that the presence of D-Phe3 modifies the cHH functionality, contributing to the diversification of the hormone pool

Neuropeptides controlling reproduction and growth in Crustacea: a molecular approach

La review descrive l'identificazione molecolare dei neuropeptidi dei crostacei essenziali per la crescita e la riproduzion

A method for rapid and high-yield production of the tick-borne encephalitis virus E and DIII recombinant proteins in E. coli with preservation of the antigenic properties

Tick-borne encephalitis virus (TBEV) is a member of the Flavivirus genus and is the main pathogenic arbovirus circulating in Europe, Russia and China. The envelope (E) protein is exposed on the viral surface and is the main antigen that is employed in diagnostic tests based on the detection of protein-specific antibodies from serum samples of infected individuals. The high degree of similarity among the E proteins of flaviviruses can, in some cases, lead to cross-reactivity and false-positive results in serological tests. Increased specificity in the detection of positive sera for different Flavivirus infections is often obtained by using a portion of the E protein, namely, the DIII domain. Different strategies and expression systems have been described for E and DIII protein production. Here, we present the optimization of an easy and fast method for TBEV E and DIII antigen production and partial purification from E. coli inclusion bodies. The antigenic properties of the produced antigens are retained, as validated by ELISAs with anti-TBEV murine sera as well as sera from infected human patients. The potential applications of both proteins as diagnostic reagents were confirmed

BIOMARKERS FOR THE DIAGNOSIS OF MULTIPLE SCLEROSIS

The invention relates to biological markers for use in the diagnosis of multiple sclerosis and the use of said markers for distinguishing between patients with multiple sclerosis and patients with other neurological diseases. The invention further relates to a method for the diagnosis of multiple sclerosis using said biological markers

Gene expression analysis of coffea arabica seeds processed under different post-harvest processing methods

The mode of coffee processing, either the wet or dry method, determines the characteristic flavour and establishes the differences in quality of the final green coffee produced. The present study focused mainly on identifying the differential gene expression in green coffee seeds of Brazilian arabica coffee (Coffea arabica L.) among samples prepared under three different post-harvest treatments (natural, washed and semi washed method) and grown in two different locations. Expression levels of 16 genes of interest were measured. These genes are involved in various cellular, metabolic and biochemical activities influencing levels of certain compounds, such as lipids, carbohydrates, caffeine and chlorogenic acid, associated with quality characteristics of the beverage. Microarray experiments were designed with cDNA probe sequences. Microarray data was analyzed to identify the differences in gene expression between two altitudes and between two variables: location and post-harvest treatment. Cluster analysis was carried out with samples showing similar patterns, which are characteristic to the group. With this approach, it was possible to identify the important genes in C. arabica seeds that have differential (increased or decreased) expression levels. It was also seen that between the location and treatments, location profoundly impacts the levels of gene expression in samples