Translational control of polyamine metabolism by CNBP is required for Drosophila locomotor function.

Microsatellite expansions of CCTG repeats in the cellular nucleic acid-binding protein (CNBP) gene leads to accumulation of toxic RNA and have been associated with myotonic dystrophy type 2 (DM2). However, it is still unclear whether the dystrophic phenotype is also linked to CNBP decrease, a conserved CCHC-type zinc finger RNA-binding protein that regulates translation and is required for mammalian development. Here, we show that depletion of Drosophila CNBP in muscles causes ageing-dependent locomotor defects that are correlated with impaired polyamine metabolism. We demonstrate that the levels of ornithine decarboxylase (ODC) and polyamines are significantly reduced upon dCNBP depletion. Of note, we show a reduction of the CNBP-polyamine axis in muscles from DM2 patients. Mechanistically, we provide evidence that dCNBP controls polyamine metabolism through binding dOdc mRNA and regulating its translation. Remarkably, the locomotor defect of dCNBP-deficient flies is rescued by either polyamine supplementation or dOdc1 overexpression. We suggest that this dCNBP function is evolutionarily conserved in vertebrates with relevant implications for CNBP-related pathophysiological conditions.

Anti-inflammatory role of curcumin in retinal disorders (Review).

Curcumin [1,7-bis-(4-hydroxy-3-methoxyphenyl)-hepta-1,6-diene-3,5-dione], the main component of turmeric (Curcuma longa, a flowering plant of the ginger family, Zingiberaceae), is known to possess different pharmacological activities, particularly anti-inflammatory and antioxidant properties. Since an underlying inflammatory process exists in several ocular conditions, such as anterior uveitis, glaucoma, age-related macular degeneration (AMD) and diabetic retinopathy (DR), the aim of the present review was to summarize the pleiotropic effects exerted by this molecule, focusing in particular on its beneficial role in retinal diseases. The anti-inflammatory activity of curcumin has also been described in numerous systemic inflammatory pathologies and tumors. Specifically, the biological, pharmaceutical and nutraceutical properties of curcumin are associated with its ability to downregulate the expression of the following genes: IκBα, cyclooxygenase 2, prostaglandin E2, interleukin (IL)-1, IL-6, IL-8 and tumor necrosis factor-α. According to this finding, curcumin may be useful in the treatment of some retinal disorders. In DR, proliferative vitreoretinopathy and AMD, beneficial effects have been observed following treatment with curcumin, including slowing down of the inflammatory process. Despite the aforementioned evidence, the main disadvantage of this substance is that it possesses a low solubility, as well as poor oral bioavailability due to its reduced absorption, rapid metabolism and rapid elimination. Therefore, several curcumin analogues have been synthesized and tested over the years, in order to improve the possible obtainable therapeutic effects. The purpose of the present review was to identify new aspects that could guide future research on this important traditional medicine, which is a well-tolerated natural product, and is widely considered safe and economical.

The antioxidant, aged garlic extract, exerts cytotoxic effects on wild-type and multidrug-resistant human cancer cells by altering mitochondrial permeability.

Aged garlic extract (AGE) has been shown to possess therapeutic properties in cancer; however its mechanisms of action are unclear. In this study, we demonstrate by MTT assay that AGE exerts an anti-proliferative effect on a panel of both sensitive and multidrug-resistant (MDR) human cancer cell lines and enhances the effects of hyperthermia (42˚C) on M14 melanoma cells. The evaluation of the mitochondrial activity in whole cancer cells treated with AGE, performed by cytofluorimetric analysis in the presence of the lipophilic cationic fluorochrome JC-1, revealed the occurrence of dose-dependent mitochondrial membrane depolarization. Membrane potential was measured by the TPP+ selective electrode. In order to shed light on its mechanisms of action, the effects of AGE on isolated rat liver mitochondria were also examined. In this regard, AGE induced a mitochondrial membrane hyperpolarization of approximately 15 mV through a mechanism that was similar to that observed with the ionophores, nigericin or salinomycin, by activating an exchange between endogenous K+ with exogenous H+. The prolonged incubation of the mitochondria with AGE induced depolarization and matrix swelling, indicative of mitochondrial permeability transition induction that, however, occurs through a different mechanism from the well-known one. In particular, the transition pore opening induced by AGE was due to the rearrangement of the mitochondrial membranes following the increased activity of the K+/H+ exchanger. On the whole, the findings of this study indicate that AGE exerts cytotoxic effects on cancer cells by altering mitochondrial permeability. In particular, AGE in the mitochondria activates K+/H+ exchanger, causes oxidative stress and induces mitochondrial permeability transition (MPT).

An eudesman derivative from Verbesina persicifolia D.C. as a natural mild uncoupler in liver mitochondria. A new potential anti-obesity agent?

4ß-cinnamoyloxy,1ß,3α-dihydroxyeudesm-7,8-ene (CDE) extracted from Verbesina persicifolia induces bioenergetic collapse in rat liver mitochondria (RLM), monitored as a fall in the respiratory control index and ADP/O values. This fall in energy is accompanied by a protonophore effect and membrane potential (Δψ) collapse, demonstrating that CDE behaves as a typical uncoupling agent. However, when examining the effect of CDE in detail, we found that it acts as a "mild" uncoupler because it drops Δψ and increases respiratory state 4. The proposed mechanism is based on the interaction of CDE with membrane protein cytochrome C oxidase, which is implicated in proton permeability, and with the respiratory chain for the generation of reactive oxygen species which mediate and regulate the activity of the above membrane protein. Considering the energy collapse, "mild" uncoupling, and the fact that CDE is largely used in folk medicines, this extract may be viewed as a potentially effective anti-obesity drug and a natural lead compound for developing new natural uncouplers against obesity.

Lysosomotropic compounds and spermine enzymatic oxidation products in cancer therapy (review).

Apoptotic pathways represent the mechanisms of programmed cell death that counteract initiation and progression of cancer. New therapeutic targets are currently being explored on the basis of our detailed knowledge of the mechanisms and factors involved in apoptosis. In recent years, numerous proteins have been identified, which act as tumour suppressors or as oncoproteins in caspase-independent programmed cell death mechanisms, in which lysosomes are implicated for their lysosomal functions in cancer, mainly attributed to lysosomal proteinases, particularly the cathepsins. If cathepsins are released from the lysosomal lumen into the cytoplasm they initiate a number of processes that may cause either apoptotic or non-apoptotic (necrotic) cell death. The release of cathepsin D into the cytoplasm by vacuolar-type ATPase (V-ATPase) inhibitors produces the characteristic signs of apoptotic cell death, including caspase-3 activation and DNA laddering. For the destabilisation of the lysosomal membrane, two methods are available having therapeutic potential: the formation of reactive oxygen species (ROS) by irradiation or by enzymatic reactions and the lysosomal membrane permeabilisation by lysosomotropic compounds. Findings also suggest that the deregulation of polyamine metabolism or cytotoxic metabolites generated from the oxidative deamination of spermine by amine oxidases in association with lysosomotropic compounds may induce apoptosis. Cross-resistance of cells to cytotoxic actions of a wide variety of natural and synthetic anticancer drugs is the well-known phenomenon called multidrug resistance (MDR), due to glycoprotein P that functions as an ATP-dependent pump. The sensitisation of tumour cells to anticancer drugs by lysosomotropic compounds, and particularly the sensitisation of MDR-resistant cells recommend scrutinizing the potential of lysosomotropic drugs in cancer therapy.

Hyperthermia enhances cytotoxicity of amine oxidase and spermine on drug-resistant LoVo colon adenocarcinoma cells.

Hyperthermia is currently receiving widespread attention when associated with other therapeutic modalities, such as irradiation or chemotherapy, in the treatment of cancer. The occurrence of resistance to cytotoxic pharmacological agents in tumor cells, associated with several phenotypic alterations, is one of the major obstacles to successful anticancer chemotherapy. We investigated a new strategy to overcome multidrug resistance (MDR) cancer cells, using bovine serum amine oxidase (BSAO), which forms toxic products from spermine (H2O2 and aldehydes). The cytotoxicity of the products was evaluated in drug-sensitive (LoVo WT) and multidrug-resistant (LoVo DX) colon adenocarcinoma cells at 37 and 42 degrees C, using a clonogenic cell survival assay. Cytotoxicity was considerably enhanced at 42 degrees C. Both toxic species contributed to the thermal enhancement of cytotoxicity induced by BSAO and spermine. Cytotoxicity was eliminated in the presence of catalase and aldehyde dehydrogenase (ALDH). An interesting finding was that BSAO and spermine at <1 microM, which were non toxic at 37 degrees C, became cytotoxic at 42 degrees C and resemble thermosensitizers. Cell survival results and electron microscopy investigations suggest that, at 42 degrees C, LoVo DX cells are not resistant to the cytotoxic enzymatic oxidation products of spermine, as was already demonstrated in these cells at 37 degrees C. Moreover, microscopy modifications caused by both toxic products were more pronounced in LoVo DX than in LoVo WT cells, where morphological cytoplasmatic alterations were shown. Our findings suggest that hyperthermia combined with the enzymatic toxic oxidation products of spermine might be a promising anticancer strategy, mainly against MDR tumor cells.

Anti-tumoral effect of native and immobilized bovine serum amine oxidase in a mouse melanoma model.

Bovine serum amine oxidase (BSAO) oxidatively deaminates polyamines containing primary amine groups, spermidine and spermine, to form the cytotoxic products hydrogen peroxide and aldehyde(s). Polyamines are present at elevated levels in many tumor tissues. The aims of the study were to evaluate the anti-tumoral activities of native and immobilized BSAO in mouse melanoma and also to determine the mechanism of tumor cell death. C57BL mice received a subcutaneous injection of B16 melanoma cells to induce formation of tumors, prior to antitumor treatments with native and immobilized BSAO. The enzyme was immobilized in a poly(ethylene glycol) (PEG) biocompatible matrix. Antitumor treatments consisted of a single injection of enzyme into the tumor. When immobilized BSAO (2.5mU) was injected into the tumor, there was a marked decrease of 70% of the tumor growth. This was compared with a decrease of only 32% of tumor size when the same amount of native BSAO was administered. The type of cell death was analysed in tumors that were treated with native or immobilized BSAO. When tumors were treated with immobilized BSAO, there was induction of a high level of apoptosis (around 70%), compared to less than 10% with the native enzyme. Apoptotic cell death was assessed by nuclear chromatin condensation using Hoechst staining and labelling of externalized phosphatidylserine using Annexin V. However, native BSAO, probably due to a burst of cytotoxic products, induced a high level of necrosis of about 40%, compared to less than 10% with immobilized BSAO. In conclusion, the advantage is that immobilized BSAO can act by allowing the slow release of cytotoxic products, which induces tumor cell death by apoptosis rather than necrosis.