Antioxidant activity of new ruthenium compounds

Many biological properties have been attributed to ruthenium complexes including anti-tumor activity and the attenuation of reperfusion damage and infarct size. In this work, we characterize the antioxidant activity of trans-[RuCl2(nic)4] where nic is 3-pyridinecarboxylic acid and trans-[RuCl2(i-nic)4] where i-nic is 4-pyridinecarboxylic acid by (i) evaluation of total antioxidant potential (TRAP); (ii) prevention of DNA damage induced by hydrogen peroxide using the alkaline comet assay; and (iii) the prevention of lipid peroxidation and cell death induced by iron in liver slices. Our results suggest that nic has stronger antioxidant potential when compared to the i-nic. Higher doses (above 200 microM) of these compounds gave genotoxic effects, but the antioxidant potential could be obtained with the use lower doses (0.1-10 microM).     

Cytologic features of clear cell carcinoma of the female genital tract. Diagnostic value of the "raspberry body" in nonexfoliative cytologic specimens

OBJECTIVE: To evaluate the presence of basement membrane stromal material in fine needle aspiration (FNA) and scrape cytologic specimens from patients with clear cell carcinoma (CCC) of the female genital tract. STUDY DESIGN: The study group consisted of 6 patients with CCC (5 ovarian and 1 cervical). Four samples corresponded to FNA specimens and 3 to scrape material obtained during intraoperative consultation for ovarian tumors. FNA was performed on a pelvic recurrence and on liver, pulmonary and lymph node metastases. The 6 cases had a complete histopathologic study. RESULTS: In addition to large, clear cells, all cases showed basement membrane stromal material that assumed several forms. The most common was globular, hyaline structures, either naked or surrounded by neoplastic epithelial cells ("raspberry bodies"). Other fragments were larger, with several spherules and elongated prolongations. Scrape material showed stromal material resembling reduplicated basement membrane material. In Diff-Quik-stained smears (QCA, Tarragana, Spain) it showed metachromatic staining with a pink to purple color. Its recognition on Papanicolaou-stained smears was more difficult since it did not stain or was gray. CONCLUSION: Basement membrane stromal material and, more precisely, "raspberry bodies," are a characteristic cytologic feature of CCC of the female genital tract. The combination of clear, atypical cells and basement membrane stroma is highly specific to this neoplasm and can be observed not only in exfoliative specimens but also in FNA and scrape samples.     

Analysis of the spatial expression pattern of seven Kip related proteins (KRPs) in the shoot apex of Arabidopsis thaliana

BACKGROUND AND AIMS: Kip-related-proteins (KRPs), negative regulators of cell division, have recently been discovered in plants but their in planta function is as yet unclear. In this study the spatial expression of all seven KRP genes in shoot apices of Arabidopsis thaliana were compared. METHODS: In situ hybridization analyses were performed on longitudinal sections of shoot apices from 2-month-old Arabidopsis plants. KEY RESULTS: The study provides evidence for different expression pattern groups. KRP1 and KRP2 expression is restricted to the endoreduplicating tissues. In contrast, KRP4 and KRP5 expression is mainly restricted to mitotically dividing cells. KRP3, KRP6 and KRP7 can be found in both mitotically dividing and endoreduplicating cells. CONCLUSION: The results suggest differential roles for the distinct KRPs. KRP1 and KRP2 might specifically be involved in the establishment of polyploidy. In contrast, KRP4 and KRP5 might be involved in regulating the progression through the mitotic cell cycle. KRP3, KRP6 and KRP7 might have a function in both types of cell cycle.     

In vitro rhizogenesis: histoanatomy of Cedrela odorata (Meliaceae) microcuttings

Cedrela odorata (Meliaceae) is considered as one of the most valuable forest tree in the tropics. Clonal propagation of this species provide an alternative method to propagate superior genotypes, being the production of good quality adventitious roots one of the most important steps in micropropagation techniques. The sequence of anatomical changes that takes place during the formation of adventitious roots in shoots of Cedrela odorata cultured in vitro is described in this study. Eigth-week-old shoots, from multiplication cultures, were rooted in Murashige and Skoog's medium (1962) with half-strength macronutrients and with 0 or 1 mg/l indole-3-butyric acid (IBA). Between 12 and 24h after the start of rooting, some cambium, phloem and interfascicular parenchyma cells became dense cytoplasm, nuclei with prominent nucleoli and the first cell divisions were observed, especially in shoots treated with auxin (dedifferentiation phase). After 3-4 days, the number of dedifferentiated cells and mitotic divisions increased considerably, and the formation of groups of some 30-40 meristematic cells (meristemoids) was observed (induction phase). The first primordial roots developed from the 4th-5th day. The vascular tissues of these primordia connected to those of the explant, and roots began to emerge from the base by day 6. Development of the primordial roots was similar in the control shoots and shoots treated with 1 mg/l IBA, although there were more roots per explant in the latter.     

Structuring detergents for extracting and stabilizing functional membrane proteins

Background

Membrane proteins are privileged pharmaceutical targets for which the development of structure-based drug design is challenging. One underlying reason is the fact that detergents do not stabilize membrane domains as efficiently as natural lipids in membranes, often leading to a partial to complete loss of activity/stability during protein extraction and purification and preventing crystallization in an active conformation.

Methodology/Principal Findings

Anionic calix[4]arene based detergents (C4Cn, n = 1–12) were designed to structure the membrane domains through hydrophobic interactions and a network of salt bridges with the basic residues found at the cytosol-membrane interface of membrane proteins. These compounds behave as surfactants, forming micelles of 5–24 nm, with the critical micellar concentration (CMC) being as expected sensitive to pH ranging from 0.05 to 1.5 mM. Both by ¹H NMR titration and Surface Tension titration experiments, the interaction of these molecules with the basic amino acids was confirmed. They extract membrane proteins from different origins behaving as mild detergents, leading to partial extraction in some cases. They also retain protein functionality, as shown for BmrA (Bacillus multidrug resistance ATP protein), a membrane multidrug-transporting ATPase, which is particularly sensitive to detergent extraction. These new detergents allow BmrA to bind daunorubicin with a Kd of 12 µM, a value similar to that observed after purification using dodecyl maltoside (DDM). They preserve the ATPase activity of BmrA (which resets the protein to its initial state after drug efflux) much more efficiently than SDS (sodium dodecyl sulphate), FC12 (Foscholine 12) or DDM. They also maintain in a functional state the C4Cn-extracted protein upon detergent exchange with FC12. Finally, they promote 3D-crystallization of the membrane protein.

Conclusion/Significance

These compounds seem promising to extract in a functional state membrane proteins obeying the positive inside rule. In that context, they may contribute to the membrane protein crystallization field.     

Extensive multiple test centre evaluation of the VecTest malaria antigen panel assay

To determine which species and populations of Anopheles transmit malaria in any given situation, immunological assays for malaria sporozoite antigen can replace traditional microscopical examination of freshly dissected Anopheles. We developed a wicking assay for use with mosquitoes that identifies the presence or absence of specific peptide epitopes of circumsporozoite (CS) protein of Plasmodium falciparum and two strains of Plasmodium vivax (variants 210 and 247). The resulting assay (VecTest Malaria) is a rapid, one-step procedure using a 'dipstick' test strip capable of detecting and distinguishing between P. falciparum and P. vivax infections in mosquitoes. The objective of the present study was to test the efficacy, sensitivity, stability and field-user acceptability of this wicking dipstick assay. In collaboration with 16 test centres world-wide, we evaluated more than 40 000 units of this assay, comparing it to the standard CS ELISA. The 'VecTest Malaria' was found to show 92% sensitivity and 98.1% specificity, with 97.8% accuracy overall. In accelerated storage tests, the dipsticks remained stable for > 15 weeks in dry conditions up to 45 degrees C and in humid conditions up to 37 degrees C. Evidently, this quick and easy dipstick test performs at an acceptable level of reliability and offers practical advantages for field workers needing to make rapid surveys of malaria vectors.     

Decline of neural tube defects cases after a folic acid campaign in Nuevo León,México

BACKGROUND: Nuevo León is a state in northeastern Mexico, near the border of Texas. Mean mortality rate from 1996-98 due to anencephaly cases was 0.6/1,000. In 1999 a surveillance program for the registry and prevention of neural tube defects (NTD) cases was initiated. METHODS: Cases were obtained from hospitals and OB-GYN clinics by immediate notification, death certificates, or fetal death registries. Only isolated cases of NTD were included. In August 1999 a folic acid campaign was initiated with the free distribution of the vitamin to low-income women with a recommendation to take a 5.0-mg pill once a week. Number of cases and rates from 1999 to 2001 were compared (chi(2) test). RESULTS: After 2 years there has been a significant reduction in the number of cases and rates. In 1999 there were 95 NTD cases and in the years 2000 and 2001 there were only 59 and 55 respectively (P < 0.001). NTD rate decreased from 1.04/1,000 in 1999 to 0.58/1,000 in 2001. Anencephaly and spina bifida rates decreased from 0.55/1,000 to 0.29/1,000 and from 0.47/1,000 to 0.22/1,000 respectively, from 1999-2001. Decrease of female cases was higher than male cases for both phenotypes. CONCLUSION: After 2 years there was a 50% decrease in the incidence of anencephaly and spina bifida cases with a significant reduction of infant mortality and disability. These results encourage us to propose the use of a single tablet of 5.0-mg of folic acid per week as an alternative to supplementation on a daily basis.     

High DNA methylation pattern intratumoral diversity implies weak selection in many human colorectal cancers

Background

It is possible to infer the past of populations by comparing genomes between individuals. In general, older populations have more genomic diversity than younger populations. The force of selection can also be inferred from population diversity. If selection is strong and frequently eliminates less fit variants, diversity will be limited because new, initially homogeneous populations constantly emerge.

Methodology and Results

Here we translate a population genetics approach to human somatic cancer cell populations by measuring genomic diversity within and between small colorectal cancer (CRC) glands. Control tissue culture and xenograft experiments demonstrate that the population diversity of certain passenger DNA methylation patterns is reduced after cloning but subsequently increases with time. When measured in CRC gland populations, passenger methylation diversity from different parts of nine CRCs was relatively high and uniform, consistent with older, stable lineages rather than mixtures of younger homogeneous populations arising from frequent cycles of selection. The diversity of six metastases was also high, suggesting dissemination early after transformation. Diversity was lower in DNA mismatch repair deficient CRC glands, possibly suggesting more selection and the elimination of less fit variants when mutation rates are elevated.

Conclusion/Significance

The many hitchhiking passenger variants observed in primary and metastatic CRC cell populations are consistent with relatively old populations, suggesting that clonal evolution leading to selective sweeps may be rare after transformation. Selection in human cancers appears to be a weaker than presumed force after transformation, consistent with the observed rarity of driver mutations in cancer genomes. Phenotypic plasticity rather than the stepwise acquisition of new driver mutations may better account for the many different phenotypes within human tumors.     

Arabidopsis thaliana cdd1 mutant uncouples the constitutive activation of salicylic acid signalling from growth defects

Arabidopsis genotypes with a hyperactive salicylic acid-mediated signalling pathway exhibit enhanced disease resistance, which is often coupled with growth and developmental defects, such as dwarfing and spontaneous necrotic lesions on the leaves, resulting in reduced biomass yield. In this article, we report a novel recessive mutant of Arabidopsis, cdd1 (constitutive defence without defect in growth and development1), that exhibits enhanced disease resistance associated with constitutive salicylic acid signalling, but without any observable pleiotropic phenotype. Both NPR1 (NON-EXPRESSOR OF PATHOGENESIS-RELATED GENES1)-dependent and NPR1-independent salicylic acid-regulated defence pathways are hyperactivated in cdd1 mutant plants, conferring enhanced resistance against bacterial pathogens. However, a functional NPR1 allele is required for the cdd1-conferred heightened resistance against the oomycete pathogen Hyaloperonospora arabidopsidis. Salicylic acid accumulates at elevated levels in cdd1 and cdd1 npr1 mutant plants and is necessary for cdd1-mediated PR1 expression and disease resistance phenotypes. In addition, we provide data which indicate that the cdd1 mutation negatively regulates the npr1 mutation-induced hyperactivation of ethylene/jasmonic acid signalling.