Cytotoxic palladium complexes of bioreductive quinoxaline N1,N4-dioxide prodrugs

Four new palladium(II) complexes with the formula Pd(L)₂, where L are quinoxaline-2-carbonitrile N¹,N⁴-dioxide derivatives, were synthesized as a contribution to the chemistry and pharmacology of metal compounds with this class of pharmacologically interesting bioreductive prodrugs. Compounds were characterized by elemental, conductometric and thermogravimetric analyses, fast atom bombardment mass spectrometry (FAB-MS) and electronic, Fourier transform infrared (FTIR) and ¹H-nuclear magnetic resonance spectroscopies. The complexes were subjected to cytotoxic evaluation on V79 cells in hypoxic and aerobic conditions. In addition, a preliminary study on interaction with plasmid DNA in normoxia was performed. Complexes showed different in vitro biological behavior depending on the nature of the substituent on the quinoxaline ring. Pd(L1)₂ and Pd(L2)₂, where L1 is 3-aminoquinoxaline-2-carbonitrile N¹,N⁴-dioxide and L2 is 3-amino-6(7)-methylquinoxaline-2-carbonitrile N¹,N⁴-dioxide, showed non selective cytotoxicity, being cytotoxic either in hypoxic or in aerobic conditions. On the other hand, Pd(L3)₂, where L3 is 3-amino-6(7)-chloroquinoxaline-2-carbonitrile N¹,N⁴-dioxide, resulted in vitro more potent cytotoxin in hypoxia (P = 5.0 μM) than the corresponding free ligand (P = 9.0 μM) and tirapazamine (P = 30.0 μM), the first bioreductive cytotoxic drug introduced into clinical trials. In addition, it showed a very good selective cytotoxicity in hypoxic conditions, being non-cytotoxic in normoxia. Its hypoxic cytotoxicity relationship value, HCR, was of the same order than those of other hypoxia selective cytotoxins (i.e., Mitomycine C, Misonidazole and the N-oxide RB90740). Interaction of the complexes with plasmid DNA in normoxia showed dose dependent ability to relax the negative supercoiled forms via different mechanisms. Pd(L2)₂ introduced a scission event in supercoiled DNA yielding the circular relaxed form. Meanwhile, both Pd(L1)₂ and Pd(L3)₂ produced the loss of negative supercoils rendering a family of topoisomers with reduced electrophoretic mobility. Pd(L3)₂ showed a more marked effect than Pd(L1)₂. Indeed, for the highest doses assayed, Pd(L3)₂ was even able to introduce positive supercoils on the plasmid DNA.     

Comparative genomic analysis of dinucleotide repeats in Tritryps

The protozoans Trypanosoma cruzi, Trypanosoma brucei and Leishmania major (Tritryps), are evolutionarily ancient eukaryotes which cause worldwide human parasitosis. They present unique biological features. Indeed, canonical DNA/RNA cis-acting elements remain mostly elusive. Repetitive sequences, originally considered as selfish DNA, have been lately recognized as potentially important functional sequence elements in cell biology. In particular, the dinucleotide patterns have been related to genome compartmentalization, gene evolution and gene expression regulation. Thus, we perform a comparative analysis of the occurrence, length and location of dinucleotide repeats (DRs) in the Tritryp genomes and their putative associations with known biological processes. We observe that most types of DRs are more abundant than would be expected by chance. Complementary DRs usually display asymmetrical strand distribution, favoring TT and GT repeats in the coding strands. In addition, we find that GT repeats are among the longest DRs in the three genomes. We also show that specific DRs are non-uniformly distributed along the polycistronic unit, decreasing toward its boundaries. Distinctive non-uniform density patterns were also found in the intergenic regions, with predominance at the vicinity of the ORFs. These findings further support that DRs may control genome structure and gene expression.     

New in situ mouse model to quantify alveolar epithelial fluid clearance

Because the availability of transgenic micemakes it possible to examine the contribution of single genes to invivo function, we developed a simple in situ mouse model that can beused to quantify isosmolar alveolar epithelial fluid clearance (AFC). Mice were killed, a tracheostomy was done, and then a test solution ofa 5% isosmolar albumin solution with 0.1 µCi of¹²⁵I-labeled albumin was instilledvia the trachea into the distal air spaces of both lungs. Afterinstillation, the lungs were inflated to 7 cmH₂O with 100%O₂ and maintained at 37°C byplacing the animals under an infrared lamp. AFC was measured by theprogressive increase in concentration of labeled and unlabeled proteinover 1 h. The results indicated the following.1) Basal, unstimulated AFC in mouselungs was significantly faster than in ex vivo rat lungs (27 ± 5%in in situ mice vs. 11 ± 3% in ex vivo rat lungs; P < 0.05).2) Comparison of equivalent doses(10⁴ M) of -adrenergicagonist (isoproterenol) and₂-adrenergic agonists(terbutaline and salmeterol) indicated that stimulated clearanceoccurred only in presence of isoproterenol.3) Because atenolol, a specific₁-antagonist, abolished theeffect of isoproterenol, the -adrenergic stimulation appears to bemediated by ₁-receptors. Therate of AFC in nonperfused mouse lungs was significantly faster than inprior studies of nonperfused lungs in rats and sheep. Interestingly,the stimulated clearance rate in mice was similar to the fast rates ofAFC that we recently reported in patients recovering from hydrostaticpulmonary edema. This in situ model is a unique experimentalpreparation that can be readily used to quantify isosmolar epithelialfluid clearance in mice.

     

Abnormal osteogenesis in osteoporotic patients is reflected by altered mesenchymal stem cells dynamics.

Bone marrow contains a population of mesenchymal stem cells with the ability to differentiate into cells that form bone, cartilage, adipose, and other connective tissues. Stem cells can be isolated from bone marrow aspirates and expanded in vitro. Presently, most stem cells studies have been performed in cells obtained from "healthy" control subjects. The goal of this study was to compare the functional characteristics of mesenchymal stem cells derived from "healthy" control and osteoporotic postmenopausal women to better understand the mechanisms involved in the pathogenesis of this disease. Osteoporotic and control stem cells have similar morphology and size and express similar cell surface antigens as evidenced by their reactivity with cell specific monoclonal antibodies. Mesenchymal stem cells from osteoporotic women differ from controls in having a lower growth rate than control cells, being refractory to the mitogenic effect of IGF-1, and exhibiting a deficient ability to differentiate into the osteogenic linage as evidenced by the alkaline phosphatase activity and calcium phosphate deposition. We conclude that in osteoporosis stem cell growth, proliferative response and osteogenic differentiation are significantly affected. Also, the study of mesenchymal stem cells from osteoporotic postmenopausal women may provide a better understanding of the mechanisms involved in the pathogenesis of the osteoporosis. It may also serve to test in vitro in rapid manner novel new therapeutic strategies.     

Induction of smooth muscle alpha-actin in vascular smooth muscle cells by arginine vasopressin is mediated by c-Jun amino-terminal kinases and p38 mitogen-activated protein kinase

Exposure of vascular smooth muscle cells to arginine vasopressin (AVP) increases smooth muscle alpha-actin (SM-alpha-actin) expression through activation of the SM- alpha-actin promoter. The goal of this study was to determine the role of the mitogen-activated protein kinase (MAP kinase) family in regulation of SM-alpha-actin expression. AVP activated all three MAP kinase family members: ERKs, JNKs, and p38 MAP kinase. Inhibition of JNKs or p38 decreased AVP-stimulated SM-alpha-actin promoter activity, whereas inhibition of ERKs had no effect. A 150-base pair region of the promoter containing two CArG boxes was sufficient to mediate regulation by vasoconstrictors. Mutations in either CArG box decreased AVP-stimulated promoter activity. Electrophoretic mobility shift assays using oligonucleotides corresponding to either CArG box resulted in a complex of similar mobility whose intensity was increased by AVP. Antibodies against serum response factor (SRF) completely super-shifted this complex, indicating that SRF binds to both CArG boxes. Overexpression of SRF increased basal promoter activity, but activity was still stimulated by AVP. AVP stimulation rapidly increased SRF phosphorylation. These data indicate that both JNKs and p38 participate in regulation of SM- alpha-actin expression. SRF, which binds to two critical CArG boxes in the promoter, represents a potential target of these kinases.     

Ecoepidemiology,short history and control of Chagas disease in the endemic countries and the new challenge for non-endemic countries

Chagas disease is maintained in nature through the interchange of three cycles: the wild, peridomestic and domestic cycles. The wild cycle, which is enzootic, has existed for millions of years maintained between triatomines and wild mammals. Human infection was only detected in mummies from 4,000-9,000 years ago, before the discovery of the disease by Carlos Chagas in 1909. With the beginning of deforestation in the Americas, two-three centuries ago for the expansion of agriculture and livestock rearing, wild mammals, which had been the food source for triatomines, were removed and new food sources started to appear in peridomestic areas: chicken coops, corrals and pigsties. Some accidental human cases could also have occurred prior to the triatomines in peridomestic areas. Thus, triatomines progressively penetrated households and formed the domestic cycle of Chagas disease. A new epidemiological, economic and social problem has been created through the globalisation of Chagas disease, due to legal and illegal migration of individuals infected by Trypanosoma cruzi or presenting Chagas disease in its varied clinical forms, from endemic countries in Latin America to non-endemic countries in North America, Europe, Asia and Oceania, particularly to the United States of America and Spain. The main objective of the present paper was to present a general view of the interchanges between the wild, peridomestic and domestic cycles of the disease, the development of T. cruzi among triatomine, their domiciliation and control initiatives, the characteristics of the disease in countries in the Americas and the problem of migration to non-endemic countries.     

Effect of slice thickness on brain magnetic resonance image texture analysis

Background  

The accuracy of texture analysis in clinical evaluation of magnetic resonance images depends considerably on imaging arrangements and various image quality parameters. In this paper, we study the effect of slice thickness on brain tissue texture analysis using a statistical approach and classification of T1-weighted images of clinically confirmed multiple sclerosis patients.     

Legacies in Switchgrass Resistance to and Recovery from Drought Suggest That Good Years Can Sustain Plants Through Bad Years

The warm-season perennial switchgrass (Panicum virgatum) is a candidate bioenergy crop. To be successful, switchgrass production must be maintained on low-quality landscapes with minimal inputs while facing future climates that are expected to be more extreme and more variable. We propose that antecedent rainfall constrains how plants respond to drought, as well as subsequently recover from drought. To test this idea, we examined how six switchgrass genotypes responded to a 1-year severe drought and then recovered under normal rainfall in the following year. These plants had previously grown for 3 years under a range of dry to wet rainfall levels in a shallow-soil common garden with no fertilizer. Plants previously exposed to drought produced less biomass, and basal area after the severe drought was relieved compared to previously well-watered plants. In addition, there were legacy effects caused by plant size: plants that were larger pre-drought were more likely to survive the severe drought, and plants that were larger during the severe drought recovered more biomass, basal area, and tillers post-drought. Although genotypes differed somewhat in their responses, the size constraint was consistent across genotypes. These findings suggest that we can establish more drought-resilient switchgrass stands by, for example, planning for initial irrigation or planting during a wet year to allow plants to grow larger prior to experiencing drought. Additional studies are needed to understand whether these rainfall and size legacies persist or are transient.