Development of an MIP based electrochemical sensor for TGF-ß1 detection and its application in liquid biopsy.

Oral cancer is one of the most common types of cancer in Europe and its large diffusion requires, together with prevention, the development of low-cost and reliable portable platforms for its diagnosis, with features of high selectivity and sensitivity. In this study, the development and characterization of a molecularly imprinted polymer (MIP)-based electrochemical sensor for TGF-ß1 detection are reported. The optimized biosensor is a potential tool for the early screening of oral cancer. A biomimetic surface has been obtained by electropolymerization of o-phenylenediamine (o-PD) on platinum electrodes, in the presence of TGF-ß1 as a template molecule. MIP synthesis, template removal and TGF-ß1 rebinding have been monitored by Differential Pulse Voltammetry (DPV). Atomic Force Microscopy (AFM) has been performed to investigate and characterize the surface morphology and the influence of the washing step on MIP and NIP (non-imprinted polymer as the control) while the thickness of the polymer layer has been measured by Scanning Transmission Electron Microscopy (STEM) analysis. The MIP sensor performance has been tested in both buffer solution and saliva samples with TGF-ß1, showing a linear response in the considered range (from 20 ng ml-1 down to 0.5 ng ml-1), an outstanding LOD of 0.09 ng mL-1 and affinity and selectivity to TGF-ß1 also in the presence of interfering molecules. The sensor was used also for the detection of target molecules in spiked saliva samples with good recovery results suggesting the possibility of the use of the proposed system for large scale fast screening in oral cancer diagnosis.

An in vivo humanized model to study homing and sequestration of Plasmodium falciparum transmission stages in the bone marrow.

Introduction: Recent evidence suggests that the bone marrow (BM) plays a key role in the diffusion of P. falciparum malaria by providing a "niche" for the maturation of the parasite gametocytes, responsible for human-to-mosquito transmission. Suitable humanized in vivo models to study the mechanisms of the interplay between the parasite and the human BM components are still missing. Methods: We report a novel experimental system based on the infusion of immature P. falciparum gametocytes into immunocompromised mice carrying chimeric ectopic ossicles whose stromal and bone compartments derive from human osteoprogenitor cells. Results: We demonstrate that immature gametocytes home within minutes to the ossicles and reach the extravascular regions, where they are retained in contact with different human BM stromal cell types. Discussion: Our model represents a powerful tool to study BM function and the interplay essential for parasite transmission in P. falciparum malaria and can be extended to study other infections in which the human BM plays a role.

Polydopamine-Coated Magnetic Iron Oxide Nanoparticles: From Design to Applications.

Magnetic iron oxide nanoparticles have been extensively investigated due to their applications in various fields such as biomedicine, sensing, and environmental remediation. However, they need to be coated with a suitable material in order to make them biocompatible and to add new functionalities on their surface. This review is intended to give a comprehensive overview of recent advantages and applications of iron oxide nanoparticles coated by polydopamine film. The synthesis method of magnetic nanoparticles, their functionalization with bioinspired materials and (in particular) with polydopamine are discussed. Finally, some interesting applications of polydopamine-coated magnetic iron oxide nanoparticles will be pointed out.

The Nitrobenzoxadiazole Derivative NBDHEX Behaves as Plasmodium falciparum Gametocyte Selective Inhibitor with Malaria Parasite Transmission Blocking Activity.

This work describes the activity of 6-((7-nitrobenzo[c][1,2,5]oxadiazol-4-yl)thio)hexan-1-ol (NBDHEX) and of its newly identified carboxylic acid metabolite on the human malaria parasite Plasmodium falciparum. NBDHEX has been previously identified as a potent cytotoxic agent against murine and human cancer cells as well as towards the protozoan parasite Giardia duodenalis. We show here that NBDHEX is active in vitro against all blood stages of P. falciparum, with the rare feature of killing the parasite stages transmissible to mosquitoes, the gametocytes, with a 4-fold higher potency than that on the pathogenic asexual stages. This activity importantly translates into blocking parasite transmission through the Anopheles vector in mosquito experimental infections. A mass spectrometry analysis identified covalent NBDHEX modifications in specific cysteine residues of five gametocyte proteins, possibly associated with its antiparasitic effect. The carboxylic acid metabolite of NBDHEX retains the gametocyte preferential inhibitory activity of the parent compound, making this novel P. falciparum transmission-blocking chemotype at least as a new tool to uncover biological processes targetable by gametocyte selective drugs. Both NBDHEX and its carboxylic acid metabolite show very limited in vitro cytotoxicity on VERO cells. This result and previous evidence that NBDHEX shows an excellent in vivo safety profile in mice and is orally active against human cancer xenografts make these molecules potential starting points to develop new P. falciparum transmission-blocking agents, enriching the repertoire of drugs needed to eliminate malaria.

Plasmodium falciparum sexual parasites regulate infected erythrocyte permeability.

To ensure the transport of nutrients necessary for their survival, Plasmodium falciparum parasites increase erythrocyte permeability to diverse solutes. These new permeation pathways (NPPs) have been extensively characterized in the pathogenic asexual parasite stages, however the existence of NPPs has never been investigated in gametocytes, the sexual stages responsible for transmission to mosquitoes. Here, we show that NPPs are still active in erythrocytes infected with immature gametocytes and that this activity declines along gametocyte maturation. Our results indicate that NPPs are regulated by cyclic AMP (cAMP) signaling cascade, and that the decrease in cAMP levels in mature stages results in a slowdown of NPP activity. We also show that NPPs facilitate the uptake of artemisinin derivatives and that phosphodiesterase (PDE) inhibitors can reactivate NPPs and increase drug uptake in mature gametocytes. These processes are predicted to play a key role in P. falciparum gametocyte biology and susceptibility to antimalarials.

Gold-Speckled SPION@SiO2 Nanoparticles Decorated with Thiocarbohydrates for ASGPR1 Targeting: Towards HCC Dual Mode Imaging Potential Applications.

Efforts are made to perform an early and accurate detection of hepatocellular carcinoma (HCC) by simultaneous exploiting multiple clinically non-invasive imaging modalities. Original nanostructures derived from the combination of different inorganic domains can be used as efficient contrast agents in multimodal imaging. Superparamagnetic iron oxide nanoparticles (SPIONs) and Au nanoparticles (NPs) possess well-established contrasting features in magnetic resonance imaging (MRI) and X-ray computed tomography (CT), respectively. HCC can be targeted by using specific carbohydrates able to recognize asialoglycoprotein receptor 1 (ASGPR1) overexpressed in hepatocytes. Here, two different thiocarbohydrate ligands were purposely designed and alternatively conjugated to the surface of Au-speckled silica-coated SPIONs NPs, to achieve two original nanostructures that could be potentially used for dual mode targeted imaging of HCC. The results indicated that the two thiocarbohydrate decorated nanostructures possess convenient plasmonic/superparamagnetic properties, well-controlled size and morphology and good selectivity for targeting ASGPR1 receptor.

Persistence of a large population of exhausted monoclonal B cells in mixed cryoglobuliemia after the eradication of hepatitis C virus infection.

PURPOSE: Functionally exhausted and mostly autoreactive B-cells with a peculiar CD21(low)CD11c(+) phenotype accumulate in several human immunological disorders including common variable immunodeficiency, HIV infection and rheumatoid arthritis. In HCV-associated mixed cryoglobulinemia (MC) there is accumulation of exhausted clonal B cells expressing a V(H)1-69-encoded cross-reactive idiotype; these cells are phenotypically heterogeneous, displaying either a CD21(low)CD11c(+) or a marginal zone (MZ)-like (IgM(+)CD27(+)CD21(+)CD11c(-)) phenotype. Irrespective of their phenotype, V(H)1-69(+) B-cells are unresponsive to the stimulation of Toll-like receptor 9 (TLR9). We investigated the fate of these cells after the eradication of HCV. METHODS: Fourteen MC patients were studied before and after antiviral therapy. V(H)1-69(+) B-cells were identified using the G6 monoclonal antibody and their phenotype and responsiveness to the stimulation of TLR9 were investigated. RESULTS: In seven virological non-responders, cryoglobulin levels and the number and phenotype of V(H)1-69(+) B cells remained substantially unchanged. By contrast, in sustained viral responders cryoglobulinemia subsided and the number of V(H)1-69(+) B cells declined. However, high proportions of MZ-like V(H)1-69(+) B cells retaining unresponsiveness to TLR9 stimulation persisted for several months in these patients. CONCLUSIONS: Clonal expansion of CD21(low) V(H)1-69(+) B cells may depend on continual stimulation by HCV, whereas their MZ-like counterparts may persist for years after the eradication of infection. Prolonged survival of exhausted MZ-like B cells after withdrawal of the initial inciting stimulus may contribute to the accumulation of autoreactive B cells in immunological disorders.

Telomere-dependent replicative senescence of B and T cells from patients with type 1a common variable immunodeficiency.

A subset of patients with common variable immunodeficiency (CVID), group 1a of the Freiburg classification, is characterized by increased B cells expressing low levels of CD21 (CD21(low) ), lymphoproliferation and autoimmunity. The CD21(low) B cells have been shown to be profoundly anergic, and defects of BCR-mediated calcium signaling and of T cells have been described in CVID 1a. We found that also the classical naïve B cells from CVID 1a patients, but not from CVID non-1a patients, proliferated poorly. The B cells of CVID 1a patients had a reduced capacity to divide reminiscent of the proliferative arrest associated with replicative senescence. Thus, we investigated whether lymphocyte dysfunction in CVID 1a was related to telomere-dependent replicative senescence, and found that both the B and the T cells from CVID 1a patients had significantly shorter telomeres compared with B and T cells from CVID non-1a patients. Telomere lengths in B and T cells were significantly correlated, indicating that the rate of telomere attrition in lymphocytes is an individual characteristic of CVID patients. Our findings suggest that telomere-dependent replicative senescence contributes to the immune dysfunction of CVID 1a patients, and may provide an important clue for a better understanding of the pathogenesis of CVID.