Effects of treatment with a carbon monoxide donor and an activator of heme oxygenase 1 on the nociceptive, apoptotic and/or oxidative alterations induced by persistent inflammatory pain in the central nervous system of mice.

The activation of heme oxygenase 1 (HO-1)/carbon monoxide (CO) inhibits chronic inflammatory pain, but its role in the central nervous system (CNS) is not entirely known. We evaluated whether the treatment with an HO-1 inducer, cobalt protoporphyrin IX (CoPP), or a CO-releasing molecule, tricarbonyldichlororuthenium(II)dimer (CORM-2), modulates the nociceptive, apoptotic and/or oxidative responses provoked by persistent inflammatory pain in the CNS. In C57BL/6 male mice with peripheral inflammation caused by complete Freund's adjuvant (CFA), we assessed the effects of CORM-2 and CoPP on the expression of protein kinase B (Akt), the apoptotic protein BAX, and the antioxidant enzymes HO-1 and NADPH quinone oxidoreductase 1 (NQO1) in the periaqueductal gray matter (PAG), amygdala (AMG), ventral hippocampus (VHPC) and medial septal area (MSA). Our results showed that the increased expression of p-Akt caused by peripheral inflammation in the four analyzed brain areas was reversed by CORM-2 and CoPP therapies. Both treatments also normalized the upregulation of BAX induced by CFA on the VHPC and MSA. Oxidative stress, demonstrated with the decreased expression of HO-1 on the PAG and AMG, was normalized in CORM-2 and CoPP treated animals. CoPP also increased the expression of HO-1 on VHPC, and both treatments up-regulated the NQO1 levels on the PAG of CFA-injected animals. In conclusion, both CORM-2 and CoPP treatments inhibited the nociceptive and apoptotic responses generated by peripheral inflammation and/or potentiated the antioxidant responses in several brain areas revealing the new modulatory effects of these treatments in the CNS of animals with chronic inflammatory pain.

Evaluation of 68Ga-DOTATATE uptake at the pituitary region and the biochemical response to somatostatin analogs in acromegaly.

PURPOSE: Acromegaly is associated with many comorbidities and increased mortality. The first-line treatment is transsphenoidal surgery. However, many patients also need adjuvant drug treatment after surgery. Somatostatin analog (SSA), which suppresses GH secretion by somatotrophs by binding to the SSTR2 receptor, is the first choice. Nevertheless, 50% of patients are partially or totally resistant to SSA, so predictive factors of response are helpful to individualize drug treatment. 68GaDOTATATE PET/CT has emerged as the gold-standard method in the diagnosis and follow-up of gastroenteropancreatic neuroendocrine tumors, which also express SSTR. Our objective was to evaluate whether 68Ga-DOTATATE uptake (SUV max) at the pituitary region of patients on SSA therapy would be useful as a drug response predictor without the need of tumoral tissue. METHODS: Fifteen acromegalics patients on SSA treatment for at least 6 months were underwent to 68Ga-DOTATATE PET/CT at the nuclear medicine service. There was an SSA complete response group (n = 5), defined as GH < 1 µg/L and IFG-1 in the normal range for gender and age, and a group that did not meet these criteria (n = 10). RESULTS: As a result, we did not find out a significantly higher SUV max in the complete response group (p = 0.0576) to SSA. However, we found a significant inverse relationship between postoperative GH values and the SUVmax at the sella turcica (p = 0.0188), probably reflecting tumor SSTR2 expression. CONCLUSION: Thus, after this initial evaluation, 68GaDOTATATE PET/CT should be better studied to assess its usefulness in the follow-up of acromegalic patients.

The role of Cryptococcus neoformans histone deacetylase genes in the response to antifungal drugs, epigenetic modulators and to photodynamic therapy mediated by an aluminium phthalocyanine chloride nanoemulsion in vitro.

Cryptococcus is a globally distributed fungal pathogen that primarily afflicts immunocompromised individuals. The therapeutic options are limited and include mostly amphotericin B or fluconazole, alone or in combination. The extensive usage of antifungals allowed the selection of resistant pathogens posing threats to global public health. Histone deacetylase genes are involved in Cryptococcus virulence, and in pathogenicity and resistance to azoles in Candida albicans. Aiming to assess whether histone deacetylase genes are involved in antifungal response and in synergistic drug interactions, we evaluated the activity of amphotericin B, fluconazole, sulfamethoxazole, sodium butyrate or trichostatin A (histone deacetylase inhibitors), and hydralazine or 5- aza-2'-deoxycytidine (DNA methyl-transferase inhibitors) against different Cryptococcus neoformans strains, C. neoformans histone deacetylase null mutants and Cryptococcus gattii NIH198. The drugs were employed alone or in different combinations. Fungal growth after photodynamic therapy mediated by an aluminium phthalocyanine chloride nanoemulsion, alone or in combination with the aforementioned drugs, was assessed for the C. neoformans HDAC null mutant strains. Our results showed that fluconazole was synergistic with sodium butyrate or with trichostatin A for the hda1Δ/hos2Δ double mutant strain. Sulfamethoxazole was synergistic with sodium butyrate or with hydralazine also for hda1Δ/hos2Δ. These results clearly indicate a link between HDAC impairment and drug sensitivity. Photodynamic therapy efficacy on controlling the growth of the HDAC mutant strains was increased by amphotericin B, fluconazole, sodium butyrate or hydralazine. This is the first study in Cryptococcus highlighting the combined effects of antifungal drugs, histone deacetylase or DNA methyltransferase inhibitors and photodynamic therapy in vitro.

Comparison of real time and malachite-green based loop-mediated isothermal amplification assays for the detection of Plasmodium vivax and P. falciparum.

The current context of malaria elimination requires urgent development and implementation of highly sensitive and specific methods for prompt detection and treatment of malaria parasites. Such methods should overcome current delays in diagnosis, allow the detection of low-density infections and address the difficulties in accessing remote endemic communities. In this study, we assessed the performance of the RealAmp and malachite-green loop mediated isothermal amplification (MG-LAMP) methodologies, using microscopy and conventional nested-PCR as reference techniques. Both LAMP techniques were performed for Plasmodium genus, P. falciparum, and P. vivax identification using 136 whole blood samples collected from three communities located in the Peruvian Amazon basin. Turnaround time and costs of performing the LAMP assays were estimated and compared to that of microscopy and nested-PCR. Using nested-PCR as reference standard, we calculated the sensitivity, specificity and 95% confidence interval (CI) for all methods. RealAmp had a sensitivity of 92% (95% CI: 85-96.5%) and specificity of 100% (95% CI: 89.1-100%) for species detection; sensitivity and specificity of MG-LAMP were 94% (95% CI: 87.5-97.8%) and 100% (89.1-100%), respectively. Whereas microscopy showed 88.1% sensitivity (95% CI: 80.2-93.7%) and 100% specificity (95%: 89.1-100%). The turnaround time and costs of performing the LAMP assays were lower compared to those associated with nested-PCR but higher than those associated with microscopy. The two LAMP assays were shown to be more sensitive and simple to implement than microscopy. Both LAMP methodologies could be used as large-scale screening tests, but the MG-LAMP assay uses a simple, portable heat-block while the RealAmp requires a RealAmp machine or a real-time PCR machine. This makes the MG-LAMP an appropriate choice for malaria surveillance studies in endemic sites. Use of LAMP tests in active case detection of Plasmodium parasites could help to detect positive malaria cases early.

Preparation, characterization and activity of CuZn and Cu2 superoxide dismutase mimics encapsulated in mesoporous silica.

Encapsulation of three superoxide dismutase (SOD) functional mimics, [CuZn(dien)2(µ-Im)(ClO4)2]ClO4 (1), [Cu2(dien)2(µ-Im)(ClO4)2]ClO4 (2) (Im = imidazolate, dien = diethylenetriamine), and [CuZn(salpn)Cl2] (3) (H2salpn = 1,3-bis(salicylideneamino)propane) in mesoporous MCM-41 silica afforded three hybrid catalysts 1@MCM-41, 2@MCM-41 and 3@MCM-41. Spectroscopic and magnetic analyses of these materials confirmed the metal centers of the complexes keep the coordination sphere after insertion into the MCM-41 silica matrix. For the imidazolate-bridged complexes the silica channels restraint the relative orientation of the two metal ions. While 3@MCM-41 shows SOD activity significantly lower than the host-free complex, insertion of the imidazolate-bridged CuZn or Cu2 complexes by ion exchange onto mesoporous MCM-41 silica affords durable and recoverable supported catalysts with much better SOD activity than the free complexes. For confined imidazolate-bridged complexes, 1@MCM-41 and 2@MCM-41, the small pore size of the silica matrix improves the SOD activity more than a host with larger pores. This high SOD activity is attributed to the close-fitting of the complexes into the nanochannels of MCM-41 silica that favors the Cu active site and HImZn(or Cu) group stay in close proximity during catalysis.

68Ga-DOTATATE PET: temporal variation of maximum standardized uptake value in normal tissues and neuroendocrine tumours.

OBJECTIVES: Higher affinity of Ga compounds to somatostatin receptors (SSTRs) and PET better image resolution increased interest in Ga-labelled somatostatin analogs in the management of neuroendocrine tumours (NETs). This study aimed to evaluate the maximum standardized uptake value (SUVmax) variation in sequential somatostatin analogs-PET in NET patients and identify optimal tumour detection and characterization imaging time. METHODS: Patients with histological or biochemical NET diagnosis performed two to three PET/computed tomography (CT) scans after intravenous injection of Ga-DOTATATE: Early PET [EarlyPET: <15 minutes postinjection (p.i.)], diagnostic PET (DiagPET: 45-90 minutes p.i.) and delayed PET (DelayPE: 90-240 minutes p.i.). Up to five tumour sites and normal tissues had SUVmax determined. Time-SUVmax curves were created for the target lesions and normal organs. Ratios between tumour and liver SUVmax (SUVTU/Liver) and tumour/blood pool (SUVTU/BP) were also calculated. RESULTS: Twenty-nine patients were included, 16 female, mean age of 46.5 ± 14.3 years. Average administered activity was 129.5 ± 29.6 MBq. Kidneys SUVmax was higher in EarlyPET compared with DiagPET (P = 0.04) and DelayPET showed higher SUVmax compared with DiagPET for normal liver, pancreas and kidneys (P = 0.02). No differences were noted between EarlyPET, DiagPET and DelayPET in tumour SUVmax (P > 0.05). SUVTU/Liver and SUVTU/BP did not change between EarlyPET and DiagPET, with a slight decrease in DelayPET. CONCLUSION: Stability in tumour SUVmax values measured at different intervals independently of tumour location, as also in normal tissues as kidneys and liver suggest that a more flexible imaging protocol may be adopted.

Carbonyl-heterobimetallic Ru(II)/Fe(II)-complexes containing polypyridyl ligands: Synthesis, characterization, cellular viability assays and interactions with biomolecules.

This paper describes on the interaction studies of carbonyl heterobimetallic compounds of Ru(II)/Fe(II) containing polypyridyl ligands, with general formula ct-[RuCl(CO)(N-N)(dppf)]PF6, N-N = 1,10-phenanthroline (phen) 5; dipyrido[3,2-f:2',3'-h]quinoxaline (dpq) 6; dipyrido[3,2-a:2',3'-c]phenazine (dppz) 7; dipyrido[3,2-f:2',3'-h]quinoxalino[2,3-b]quinoxaline (dpqQX) 8 and dppf = 1,1'-bis(diphenylphosphino) ferrocene], with calf thymus DNA (ct-DNA) and bovine serum albumin (BSA). Also, it describes the cellular viability assays of these complexes in tumorigenic and non-tumorigenic cell lines. The carbonyl complexes 5-8 and their respective precursors with formula cis-[RuCl2(N-N)(dppf)], N-N = phen (1), dpq (2), dppz (3) and dpqQX (4), were characterized by elemental analysis and spectroscopic techniques (FTIR, UV-vis, 1H and 31P{1H} NMR). Also, a cyclic voltammetry study was performed for all complexes. The crystal structure of the complex 3 is presented and discussed. Spectrofluorimetric titrations shows spontaneous and strong interaction of 5-8 with BSA, through a static quenching mechanism, resulting in binding constants in the order of 104-106 L mol-1, at 310 K. Viscosity measurements and circular dichroism spectra prompts interactions of 5-8 with ct-DNA via non-classical intercalations or by an electrostatic pathway. MTT assays in breast tumor cells MDA-MB-231 and in non-tumorigenic cells MCF-10A and V79-4 cell lines revealed IC50 values ranging from 0.19 to 1.11 µmol L-1, 1.07-3.18 µmol L-1 and 1.29-3.85 µmol L-1 respectively, for complexes 5-8.

Enhanced expression of heme oxygenase-1 in the locus coeruleus can be associated with anxiolytic-like effects.

Some researchers have shown that carbon monoxide (CO) plays a role in emotional behavior modulation through intracellular 3'-5'-guanosine monophosphate mechanisms in the locus coeruleus (LC). In fact, the LC region has a high expression of the heme-oxygenase (HO) enzymes, which are responsible for the production of CO. However, the physiological mechanism by which the HO-CO pathway participates in the modulation of emotional responses in the LC still needs clarification. This study evaluates whether a systemic intraperitoneal treatment is able to alter behavioral responses (in the elevated plus-maze and the light-dark box test) and the expression of the HO-1 and HO-2 enzymes in the LC. The tested treatments are acute (3h before) or chronic (twice daily for 10days) and with a carbon monoxide releaser (tricarbonyldichlororuthenium [II] dimer, or CORM-2) or with a HO-1 inducer compound (cobalt protoporphyrin IX, CoPP). The results for the elevated plus-maze show that CO-for both acute or chronic administration of either drug-ncreased the number of entries into the open arms and the percentage of time spent in the open arms. Regarding the light-dark box test, chronic treatment with either drug increased the time spent in the light compartment. Additionally, treatment with CORM-2 or CoPP, either acutely or chronically, increased HO-1 enzyme expression in the LC cells. This study shows that systemic CO treatment can promote an anxiolytic-like effect and the expression of HO-1 enzymes in LC cells.

Association studies to transporting proteins of fac-ReI(CO)3(pterin)(H2O) complex.

A new synthetic route to acquire the water soluble complex fac-ReI(CO)3(pterin)(H2O) was carried out in aqueous solution. The complex has been obtained with success via the fac-[ReI(CO)3(H2O)3]Cl precursor complex. ReI(CO)3(pterin)(H2O) has been found to bind strongly with bovine and human serum albumins (BSA and HSA) with intrinsic-binding constants, Kb, of 6.5 × 105 M-1 and 5.6 × 105 M-1 at 310 K, respectively. The interactions of serum albumins with ReI(CO)3(pterin)(H2O) were evaluated employing UV-vis fluorescence and absorption spectroscopy and circular dichroism. The results suggest that the serum albumins-ReI(CO)3(pterin)(H2O) interactions occurred in the domain IIA-binding pocket without loss of helical stability of the proteins. The comparison of the fluorescence quenching of BSA and HSA due to the binding to the Re(I) complex suggested that local interaction around the Trp 214 residue had taken place. The analysis of the thermodynamic parameters ΔG0, ΔH0, and ΔS0 indicated that the hydrophobic interactions played a major role in both HSA-Re(I) and BSA-Re(I) association processes. All these experimental results suggest that these proteins can be considered as good carriers for transportation of ReI(CO)3(pterin)(H2O) complex. This is of significant importance in relation to the use of this Re(I) complex in several biomedical fields, such as photodynamic therapy and radiopharmacy.

Photodynamic performance of zinc phthalocyanine in HeLa cells: A comparison between DPCC liposomes and BSA as delivery systems.

Comparable intracellular concentrations (≈30pmol/10(6) cells) of bovine serum albumin-ZnPc (BSA) adduct outperformed dipalmitoyl-phosphatidyl-choline (DPPC) liposomes containing ZnPc at photodynamic-killing of human cervical cancer cells (HeLa) after only 15min of irradiation using red light (λ>620nm, 30W/cm(2)). This result could not be simply explained in terms of dye aggregation within the carrier, since in the liposomes the dye was considerably less aggregated than in bovine serum albumin, formulation that was capable to induce cell apoptosis upon red light exposure. Thus, using specific organelle staining, our cumulative data points towards intrinsic differences in intra-cellular localization depending on the cargo vehicle used, being ZnPc:BSA preferentially located in the near vicinity of the nucleus and in the Golgi structures, while the liposomal formulation ZnPc:DPPC was preferentially located in cellular membrane and cytoplasm. In addition to those differences, using real-time advanced fluorescence lifetime imaging of HeLa cells loaded with the photosensitizer contained in the different vehicles, we have found that only for the ZnPc:BSA formulation, there was no significant changes in the fluorescence lifetime of the photosensitizer inside the cells. This contrasts with the in situ≈two-fold reduction of the fluorescence lifetime measured for the liposomal ZnPc formulation. Those observations point towards the superiority of the protein to preserve dye aggregation, and its photochemical activity, post-cell uptake, demonstrating the pivotal role of the delivery vehicle at determining the ultimate fate of a photosensitizer.

Iontophoresis Improved Growth Reduction of Invasive Squamous Cell Carcinoma in Topical Photodynamic Therapy.

This study examined the potential of iontophoresis in topical photodynamic therapy (PDT) of human invasive squamous cells carcinomas (SCC). SCC was induced in nude BALB/c mice by subcutaneous injection of A431 cells. Tumor penetration and distribution of the photosensitizer tetrasulfonated zinc phthalocyanine (ZnPcS4) was investigated after 10 and 30 min of in vivo iontophoresis of a gel containing ZnPcS4. PDT was performed immediately after iontophoresis using laser at 660 nm with a dose of irradiation of 100 J/cm(2) and irradiance of 48 mW/cm(2) while tumor growth was measured for 30 days. Iontophoresis increased ZnPcS4 penetration into tumors by 6-fold after 30 min when compared with passive delivery. Confocal microscopy analysis showed that ZnPcS4 was homogeneous distributed within deep regions of the tumor after iontophoresis. Irradiation of the tumors immediately after iontophoresis showed reduction in tumor size by more than 2-fold when compared to non-treated tumors. Iontophoretic-PDT treated tumors presented large areas of necrosis. The study concluded that iontophoretic delivery of photosensitizers could be a valuable strategy for topical PDT of invasive SCC.

Side-on cupric-superoxo triplet complexes as competent agents for H-abstraction relevant to the active site of PHM.

Copper complexes with N3S donors mimic the CuM site of copper monooxygenases and react with O2 affording side-on cupric-superoxo complexes capable of H-abstraction from dihydroanthracene and THF. Spectroscopic and DFT data of the Cu-superoxos support a spin triplet ground state for the side-on complexes, as well as a hemilabile thioether.

Interaction between proteins and Ir based CO releasing molecules: mechanism of adduct formation and CO release.

Carbon monoxide releasing molecules (CORMs) have important bactericidal, anti-inflammatory, neuroprotective, and antiapoptotic effects and can be used as tools for CO physiology experiments, including studies on vasodilation. In this context, a new class of CO releasing molecules, based on pentachlorocarbonyliridate(III) derivative have been recently reported. Although there is a growing interest in the characterization of protein-CORMs interactions, only limited structural information on CORM binding to protein and CO release has been available to date. Here, we report six different crystal structures describing events ranging from CORM entrance into the protein crystal up to the CO release and a biophysical characterization by isothermal titration calorimetry, Raman microspectroscopy, and molecular dynamics simulations of the complex between a pentachlorocarbonyliridate(III) derivative and hen egg white lysozyme, a model protein. Altogether, the data indicate the formation of a complex in which the ligand can bind to different sites of the protein surface and provide clues on the mechanism of adduct formation and CO release.

Water soluble molybdenocene complexes: synthesis, cytotoxic activity and binding studies to ubiquitin by fluorescence spectroscopy, circular dichroism and molecular modeling.

Four new molybdenocene complexes, Cp2Mo(l-ascorbato), Cp2Mo(6-O-palmitoyl-l-ascorbato), [Cp2Mo(ethyl maltolato)]Cl and Cp2Mo((2S)-2-amino-3-methyl-3-thiolato-butanoato), were synthesized and structurally characterized by standard analytical methods. The cytotoxicity of these complexes was assessed on colon HT-29 and breast MCF-7 cancer cell lines using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. A higher cytotoxic activity was shown by all the new complexes on the MCF-7 cells over the Cp2MoCl2 complex. The complexes Cp2Mo(l-ascorbato), Cp2Mo(6-O-palmitoyl-l-ascorbato) and [Cp2Mo(ethyl maltolato)]Cl displayed a stronger cytotoxic activity on colon cancer HT-29 cell line, over the molybdenocene dichloride (Cp2MoCl2). In contrast, Cp2Mo((2S)-2-amino-3-methyl-3-thiolato-butanoato) exhibited proliferative properties on this cell line. Ubiquitin (Ub)-molybdenocene interactions were investigated using cyclic voltammetry, fluorescence quenching spectroscopy, circular dichroism (CD) and molecular modeling. The thermodynamic parameters (ΔH and ΔS) obtained using fluorescence quenching spectra and van't Hoff plot indicate the Ub-molybdenocene interactions are mainly hydrophobic. The CD data also support hydrophobic interactions with conformational changes in the Ub protein. Docking studies using molecular modeling revealed the amino acids involved in the Ub-molybdenocene interactions and corroborated the hydrophobic nature of the binding combined with hydrogen bonding.

Organopalladium compound 7b targets mitochondrial thiols and induces caspase-dependent apoptosis in human myeloid leukemia cells.

The advances in the treatment of chronic myeloid leukemia (CML) during the last years were also accompanied by the development of evading strategies by tumor cells, resulting in chemotherapy resistance in some patients. Patented organopalladium compounds derived from the reaction of N,N-dimethyl-1-phenethylamine (dmpa) with [1,2-ethanebis(diphenylphosphine)] (dppe) exhibited a potent antitumor activity in vivo and in vitro in melanoma cells. We showed here that the cyclopalladated derivative [Pd2(R(+))C(2), N-dmpa)2(µ-dppe)Cl2], named compound 7b, was highly effective to promote cell death in the K562 human leukemia cells and its mechanisms of action were investigated. It was shown that compound 7b was able to promote exclusively apoptotic cell death in K562 cells associated to cytochrome c release and caspase 3 activation. This cytotoxic effect was not observed in normal peripheral mononuclear blood cells. The compound 7b-induced intrinsic apoptotic pathway was triggered by the protein thiol oxidation that resulted in the dissipation of the mitochondrial transmembrane potential. The preventive effect of the dithiothreitol on the compound 7b-induced cell death and all downstream events associated to apoptosis confirmed that death signal was elicited by the thiol oxidation. These findings contribute to the elucidation of the palladacycle 7b-induced cell death mechanism and present this compound as a promising drug in the CML antitumor chemotherapy.

Peculiar reactivity of a di-imine copper(II) complex regarding its binding to albumin protein.

A set of four di-imine copper(II) complexes containing pyridine, pyrazine and/or imidazole moieties, [Cu(apyhist)H2O](2+) 1 (apyhist = 2-(1H-imidazol-4-yl)-N-(1-(pyridin-2-yl)ethylidene)ethanamine), [Cu(apzhist)OH](+) 2 (apzhist = 2-(1H-imidazol-4-yl)-N-(1-(pyrazin-2-yl)ethylidene)ethanamine), [Cu(apyepy)OH](+) 3 (apyepy = 2-(pyridin-2-yl)-N-(1-(pyridin-2-yl)ethylidene)ethanamine), and [Cu(apzepy)H2O](2+) 4 (apzepy = N-(1-(pyrazin-2-yl)ethylidene)-2-(pyridin-2-yl)ethanamine), were investigated regarding their capability of interacting with serum albumin (human, HSA and bovine, BSA), by using spectroscopic techniques, CD, UV/Vis and EPR. Like other similar di-imine copper(II) complexes, most of them showed an expected preferential insertion of the metal ion at the primary N-terminal site of the protein, very selective for copper and characterized by a CD band at 560 nm. Further insertion of the copper ion at a secondary site is expected when using an excess of the metal. However, one of these studied complexes, [Cu(apyhist)H2O](2+) 1, exhibited anomalous behaviour interacting only at this secondary metal binding site of albumin, characterized by a CD band at 370 nm, and attributed to the coordination of copper at the Cys34 pocket. Analogous experiments with HSA previously treated with N-ethyl-maleimide (NEM), that oxidizes the protein Cys34 residue and obstructs the metal coordination, verified these results. Additional data obtained by EPR spectroscopy complemented those results. DFT calculations, considering some structural and electronic characteristics of such series of di-imine ligands and of the corresponding copper complexes, suggested molecular recognition of the apyhist ligand at the protein cavity as a feasible explanation for this unexpected and peculiar behaviour of complex 1.

A metal mixture induces transformation upon antioxidant depletion in a hepatic cell line.

INTRODUCTION: Metals are ubiquitous soil, air, and water pollutants. A mixture of arsenic cadmium and lead, in particular, has commonly been found in the vicinity of smelter areas. The mixture of As-Cd-Pb has been shown to be carcinogenic, and transforming potential and oxidative stress have been proposed as principal mechanisms involved in this process. The aim of this work was to explore the role of the antioxidant barrier in the establishment of cell transformation upon chronic exposure to a metal mixture containing 2 µM NaAsO(2), 2 µM. CdCl(2), and 5 µM Pb(C(2)H(3)O(2))(2)∙3H(2)O in WRL-68 cells-a non-transformed human hepatic cell line. MATERIAL AND METHODS: In this study, we used a WRL-68 cell model of human embryonic hepatic origin treated with antioxidant inhibitors (L-Buthionine-sulfoxamine and aminotriazole) to test the role of the antioxidant barrier in the establishment of cell transformation upon chronic exposure to a metal mixture of As-Cd-Pb (2 µM NaAsO(2), 2 µM CdCl(2) and 5 µM Pb(C(2)H(3)O(2))(2)∙3H(2)O). We evaluated oxidative damage markers, including reactive oxygen species, lipid peroxidation, and genotoxicity, as well as antioxidant response markers, including glutathione concentration, catalase activity, and superoxide dismutase activity, which promote morphological transformation, which can be quantified by foci formation. RESULTS: As expected, we found an increase in the intracellular concentration of the metals after treatment with the metal mixture. In addition, treatment with the metal mixture in addition to inhibitors resulted in a large increase in the intracellular concentration of cadmium and lead. Our results describe the generation of reactive oxygen species, cytotoxicity, genotoxicity, and oxidative damage to macromolecules that occurred exclusively in cells that were morphologically transformed upon exposure to a metal mixture and antioxidant barrier inhibition. CONCLUSION: Our results show the importance of the antioxidant barrier role in the protection of cellular integrity and the transformation potential of this metal mixture via free radicals.

Biotransformation of LASSBio-579 and pharmacological evaluation of p-hydroxylated metabolite a N-phenylpiperazine antipsychotic lead compound.

Using a combination of docking and molecular dynamics simulations, we predicted that p-hydroxylation by CYP1A2 would be the main metabolic pathway for the 1-[1-(4-chlorophenyl)-1H-4pyrazolylmethyl] phenylhexahydropiperazine, LASSBio-579 (3). As the result of a screening process with strains of filamentous fungi, Cunninghamella echinulata ATCC 9244 was chosen to scale up the preparation of the p-hydroxylated metabolite (4). About 30 min after i.p. administration of (3) to rats was identified as the p-hydroxylated metabolite, confirming our in silico previsions. Chemical synthesis of the metabolite was performed and allowed its pharmacological evaluation in binding assays revealing its high affinity for D2 and D4 receptors, indicating that this metabolite should participate to the antipsychotic effect of (3) in vivo. Furthermore, we report here that both (3) and its p-hydroxylated metabolite (4) have a much lower affinity than clozapine for two receptors involved in adverse reactions. Voltammetric assays were useful to understand the redox profile of (3).

Molecular structure of tetraaqua adenosine 5'-triphosphate aluminium(III) complex: a study involving Raman spectroscopy, theoretical DFT and potentiometry.

The Alzheimer's disease is one of the most common neurodegenerative diseases that affect elderly population, due to the formation of ß-amyloid protein aggregate and several symptoms, especially progressive cognitive decline. The result is a decrease in capture of glucose by cells leading to obliteration, meddling in the Krebs cycle, the principal biochemical route to the energy production leading to a decline in the levels of adenosine 5'-triphosphate. Aluminium(III) is connected to Alzheimer's and its ion provides raise fluidity of the plasma membrane, decrease cell viability and aggregation of amyloid plaques. Studies reveal that AlATP complex promotes the formation of reactive fibrils of ß-amyloid protein and independent amyloidogenic peptides, suggesting the action of the complex as a chaperone in the role pathogenic process. In this research, one of complexes formed by Al(III) and adenosine 5'-triphosphate in aqueous solution is analyzed by potentiometry, Raman spectroscopy and ab initio calculations. The value of the logK(AlATP) found was 9.21±0.01 and adenosine 5'-triphosphate should act as a bidentate ligand in the complex. Raman spectroscopy and potentiometry indicate that donor atoms are the oxygen of the phosphate ß and the oxygen of the phosphate γ, the terminal phosphates. Computational calculations using Density Functional Theory, with hybrid functions B3LYP and 6-311++G(d,p) basis set regarding water solvent effects, have confirmed the results. Frontier molecular orbitals, electrostatic potential contour surface, electrostatic potential mapped and Mulliken charges of the title molecule were also investigated.

In vitro phototoxicity of ultradeformable liposomes containing chloroaluminum phthalocyanine against New World Leishmania species.

The use of photodynamic therapy (PDT) against cutaneous leishmaniasis (CL) based on chloroaluminum phthalocyanine (ClAlPc) is a promissory alternative therapy. The main purpose of this article was to assess the internalization and in vitro phototoxic activities of ClAlPc encapsulated in ultradeformable liposomes (UDL-ClAlPc) in Leishmania parasites and mammalian cells. Cell internalization was determined by fluorescence microscopy, cell and parasite damage by standard MTT or direct microscopic analysis and a phototoxic index (PI) was calculated as the compound activity (IC(50)) at 0 J/cm(2)/IC(50) at 17 J/cm(2). Liposomal and free ClAlPc were internalized by infected and non-infected THP-1 cells and co-localized in the mitochondria. Treatment of UDL-ClAlPc was almost 10 times more photoactive than free ClAlPc on THP-1 cells and promastigotes and intracellular amastigotes of Leishmania chagasi and Leishmania panamensis. Liposomal compounds were active on non-irradiated and irradiated cells however PI higher than 50 were calculated. PI for amphotericin B referential drug were lower than 1.2. Empty liposomes tested at the same lipid concentration of active ClPcAl-liposomes were non-toxic. Upon photodynamic treatment a nonselective-parasite activity against intracellular amastigotes were observed and loss of membrane integrity resulting in a release of parasites was detected. Further studies oriented to evaluate both the state of infection after PDT and the effectiveness of UDL as delivery vehicles of ClAlPc in CL experimental models are required.