Behind the Link between Copper and Angiogenesis: Established Mechanisms and an Overview on the Role of Vascular Copper Transport Systems

Angiogenesis critically sustains the progression of both physiological and pathological processes. Copper behaves as an obligatory co-factor throughout the angiogenic signalling cascades, so much so that a deficiency causes neovascularization to abate. Moreover, the progress of several angiogenic pathologies (e.g. diabetes, cardiac hypertrophy and ischaemia) can be tracked by measuring serum copper levels, which are being increasingly investigated as a useful prognostic marker. Accordingly, the therapeutic modulation of body copper has been proven effective in rescuing the pathological angiogenic dysfunctions underlying several disease states. Vascular copper transport systems profoundly influence the activation and execution of angiogenesis, acting as multi-functional regulators of apparently discrete pro-angiogenic pathways. This review concerns the complex relationship among copper-dependent angiogenic factors, copper transporters and common pathological conditions, with an unusual accent on the multi-faceted involvement of the proteins handling vascular copper. Functions regulated by the major copper transport proteins (CTR1 importer, ATP7A efflux pump and metallo-chaperones) include the modulation of endothelial migration and vascular superoxide, known to activate angiogenesis within a narrow concentration range. The potential contribution of prion protein, a controversial regulator of copper homeostasis, is discussed, even though its angiogenic involvement seems to be mainly associated with the modulation of endothelial motility and permeability

Copper as a Target for Treatment of Neuroblastoma: Molecular and Cellular Mechanisms

none2Urso E; Maffia MUrso, Emanuela; Maffia, Michel

Nanostructures for SERS in living cell

Surface-enhanced Raman spectroscopy (SERS) has received renewed interest in recent years in fields such as trace analysis, biorelated diagnosis, and living cell study. However, the interference of impurities left on the surface from the preparation process of substrates limits to some extent the application of SERS. In the present paper, we propose a method to prepare clean SERS substrates by a combined method of hydrothermal green synthesis and thermal treatment to obtain a clean and impurity-free surface for SERS measurements, suitable for cells growth. The goal of such activity was the study of the membrane proteome, with special attention to prion protein (PrPC), in its physiological ambient. SERS has been used to evidence the PrPC-Cu(II) interaction in a rat neuroblastoma cell line (B104), known to overexpress the cellular prion protein PrPC

Physiological role of Prion Protein in Copper homeostasis and angiogenic mechanisms of endothelial cells

Abstract The Prion Protein (PrP) is mostly known for its role in prion diseases, where its misfolding and aggregation can cause fatal neurodegenerative conditions such as the bovine spongiform encephalopathy and human Creutzfeldt–Jakob disease. Physiologically, PrP is involved in several processes including adhesion, proliferation, differentiation and angiogenesis, but the molecular mechanisms behind its role remain unclear. PrP, due to its well-described structure, is known to be able to regulate copper homeostasis; however, copper dyshomeostasis can lead to developmental defects. We investigated PrP-dependent regulation of copper homeostasis in human endothelial cells (HUVEC) using an RNA-interference protocol. PrP knockdown did not influence cell viability in silenced HUVEC (PrPKD) compared to control cells, but significantly increased PrPKD HUVEC cells sensitivity to cytotoxic copper concentrations. A reduction of PrPKD cells reductase activity and copper ions transport capacity was observed. Furthermore, PrPKD-derived spheroids exhibited altered morphogenesis and their derived cells showed a decreased vitality 24 and 48 hours after seeding. PrPKD spheroid-derived cells also showed disrupted tubulogenesis in terms of decreased coverage area, tubule length and total nodes number on matrigel, preserving unaltered VEGF receptors expression levels. Our results highlight PrP physiological role in cellular copper homeostasis and in the angiogenesis of endothelial cells

Behind the Link between Copper and Angiogenesis: Established Mechanisms and an Overview on the Role of Vascular Copper Transport Systems

Angiogenesis critically sustains the progression of both physiological and pathological processes. Copper behaves as an obligatory co-factor throughout the angiogenic signalling cascades, so much so that a deficiency causes neovascularization to abate. Moreover, the progress of several angiogenic pathologies (e.g. diabetes, cardiac hypertrophy and ischaemia) can be tracked by measuring serum copper levels, which are being increasingly investigated as a useful prognostic marker. Accordingly, the therapeutic modulation of body copper has been proven effective in rescuing the pathological angiogenic dysfunctions underlying several disease states. Vascular copper transport systems profoundly influence the activation and execution of angiogenesis, acting as multi-functional regulators of apparently discrete pro-angiogenic pathways. This review concerns the complex relationship among copper-dependent angiogenic factors, copper transporters and common pathological conditions, with an unusual accent on the multi-faceted involvement of the proteins handling vascular copper. Functions regulated by the major copper transport proteins (CTR1 importer, ATP7A efflux pump and metallo-chaperones) include the modulation of endothelial migration and vascular superoxide, known to activate angiogenesis within a narrow concentration range. The potential contribution of prion protein, a controversial regulator of copper homeostasis, is discussed, even though its angiogenic involvement seems to be mainly associated with the modulation of endothelial motility and permeability

Potential impact of RFID-based tracing systems on the integrity of pharmaceutical products

Radio Frequency Identification (RFID) is going to play a crucial role as auto-identification technology in a wide range of applications such as healthcare, logistics, supply chain management, ticketing, et cetera. The use of electromagnetic waves to identify, trace, and track people or goods allows solving many problems related to auto-identification devices based on optical reading (i.e. bar code). Currently, high interest is concentrated on the use of Radio Frequency (RF) solutions in healthcare and pharmaceutical supply chain, in order to improve drugs flow transparency and patients’ safety. Unfortunately, there is a possibility that drug interaction with electromagnetic fields (EMFs) generated by RF devices, such as RFID readers, deteriorate the potency of bioactive compounds. This chapter proposes an experimental multidisciplinary approach to investigate potential alterations induced by EMFs on drug molecular structure and performance. To show the versatility of this approach, some experimental results obtained on two biological pharmaceuticals (peptide hormone-based) are discussed

EFFECTS OF PRION PROTEIN FRAGMENTS ON THEPROLIFERATION OF B104 NEUROBLASTOMA CELLS

Transmissible Spongiform Encephalopathies (TSEs) are a group of neurological disorders whose aetiological agent is believed to be PrPsc, an aberrant variant of the normal cellular prion protein PrPc, a GPI- anchored protein highly expressed in neuronal cells. Unlike PrPc, PrPsc is characterized by a higher β-sheet content (Pan et al., Proc. Natl. Acad. Sci. 90:10962-10966, 1993). Regarding the mechanisms of conversion of the cellular isoform in the aberrant one, a number of studies have been performed about a short peptide based on the sequence 106-126 of PrPc. It exhibits a prevalent β-sheet structure and forms amyloid fibrillar aggregates; since it has been shown to induce apoptosis in cultured cells, it might constitute the toxic core of PrPsc. In parallel, other peptides reproducing different portions of PrPc have been analysed, but their toxicity was found to be much less than with PrP[106-126] (Brown et al., J. of Inorg. Biochem., 98: 133-143, 2004). In the present work, we evaluated by MTT test the effect of several synthetic prion protein fragments on B104 neuroblastoma cells. These peptides (PrP[173-195], PrP[180-195] and several analogues) reproduce a region of PrPc corresponding to helic-2, that could be implicated in PrPc-PrPsc conversion because of its structural instability and its tendency to assume beta structure (Tizzano et al., Proteins 59(1): 72-9, 2005). We found a remarkable reduction of cell viability after 48 h of incubation of cells with PrP[173-195], so we suggest a reasonable role for this region in the anomalous misfolding of the scrapie isoform

Effect of the flame retardant tris (1,3-dichloro-2-propyl) phosphate (TDCPP) on Na+-K+-ATPase and Cl− transport in HeLa cells

Tris (1, 3-dichloro-2-propyl) phosphate (TDCPP) is one of the most diffused phosphorus flame retardants in the environment and is highly persistent and abundant in residential dust samples. To date the cellular targets and mechanisms underlying its toxic effects are not completely understood. The aim of this work was to study the effects of TDCPP on ion transport mechanisms fundamental for the cellular ionic homeostasis, such as Na+-K+-ATPase and Cl- transport. HeLa cells were used as experimental model. TDCPP showed a dose-dependent effect on cell viability in cells exposed for 24h as assessed by MTT test (IC50 = 52.5 µM). The flame retardant was able to exert a dose and time-dependent inhibition on the Na+-K+-ATPase activity. A short-term exposure (1h) was able to exert a significant inhibition at 75 and 100 µM TDCPP, suggesting that TDCPP is able to directly interfere with the Na+-K+-ATPase phosphate catalytic activity. The sensitivity of the pump to lower TDCPP concentrations increased with the increase of the time of exposure. Following 24h exposure a significant inhibition of about 40 % was evident already at 10 µM and the IC50 value observed was 12.8 ± 6.0 µM. Moreover, TDCPP was also able to impair the NKCC mediated Cl- transport in HeLa cells, as assessed in YFP-H148Q/I152L-expressing HeLa cells. Following 1h exposure TDCPP significantly inhibited the transport by about 30%. The kinetic analysis demonstrated a noncompetitive mechanism of inhibition. In conclusion, results demonstrated the impairment of ion transport mechanisms fundamental for ion homeostasis by TDCPP on HeLa cells