In vitro evaluation of the safe margin, antithrombotic and antiproliferative actions for the treatment of restenosis: Nitric oxide donor and polymers

Drug‐eluting stents (DES) were developed to combat the problem of in‐stent restenosis, and evaluating the biological activity from DES systems is critical for its safety and efficacy. To test the cytotoxicity of nitric oxide (NO) donor‐containing polymers for their potential use in DES applications, S‐nitrosoglutathione (GSNO) or in combination with poly(vinyl alcohol) (PVA) and poly(vinyl pyrrolidone) (PVP) in an aqueous polymeric solution (PVA/PVP/GSNO) was investigated using Balb/c 3T3 and Rabbit arterial smooth muscle (RASM) cells. The sensitivity of 3T3 cells to the cytotoxicity effects induced by GSNO was higher than that of RASM cells, while RASM cells were more susceptible to alterations in membrane permeability. Cell growth assays showed that GSNO and PVA/PVP/GSNO induced antiproliferative effects in RASM cells. Moreover, the presence of polymers can reduce the cytotoxicity and enhance the antiproliferative effects of GSNO. Dose‐dependent inhibition of platelet aggregation was similar for both PVA/PVP/GSNO (EC50 of 3.4 ± 2.3 µM) and GSNO (EC50 of 2.8 ± 1.1 µM) solutions. Platelet adhesion assays showed that the inhibition caused by GSNO (EC50 of 5.0 mM) was dependent on the presence of plasma. These results demonstrate that the methodology adopted here is suitable to establish safety margins and evaluate the antithrombotic potential and antiproliferative effects of NO‐eluting biomaterials and polymeric solutions for the new cardiovascular devices, and also to emphasize the importance of using more specific cell lines in these evaluations. Copyright © 2011 John Wiley & Sons, Ltd.     

Tenascin-C in the extracellular matrix promotes the selection of highly proliferative and tubulogenesis-defective endothelial cells

The extracellular matrix (ECM) contains important cues for tissue homeostasis and morphogenesis. The matricellular protein tenascin-C (TN-C) is overexpressed in remodeling tissues and cancer. In the present work, we studied the effect of different ECM—which exhibited a significant diversity in their TN-C content—in endothelial survival, proliferation and tubulogenic differentiation: autologous (endothelial) ECM devoid of TN-C, but bearing large amounts of FN; fibroblast ECM, bearing both high TN-C and FN contents; and finally, glioma-derived matrices, usually poor in FN, but very rich in TN-C. HUVECs initially adhered to the immobilized matrix produced by U373 MG glioma cells, but significantly detached and died by anoikis (50 to 80%) after 24 h, as compared with cells incubated with endothelial and fibroblast matrices. Surviving endothelial cells (20 to 50%) became up to 6-fold more proliferative and formed 74–97% less tube-like structures in vitro than cells grown on non-tumoral matrices. An antibody against the EGF-like repeats of tenascin-C (TN-C) partially rescued cells from the tubulogenic defect, indicating that this molecule is responsible for the selection of highly proliferative and tubulogenic defective endothelial cells. Interestingly, by using defined substrata, in conditions that mimic glioma and normal cell ECM composition, we observed that fibronectin (FN) modulates the TN-C-induced selection of endothelial cells. Our data show that TN-C is able to modulate endothelial branching morphogenesis in vitro and, since it is prevalent in matrices of injured and tumor tissues, also suggest a role for this protein in vascular morphogenesis, in these physiological contexts.     

Fifty shades of brown: Macroevolution of plumage brightness in the Furnariida,a large clade of drab Neotropical passerines

Both natural and sexual selection are thought to affect the evolution of bird color. Most studies of the topic have focused on sexually dichromatic taxa and showy plumages, which are expected to be more influenced by social selection and usually result in increased conspicuousness. However, many bird clades display dull brown or gray plumages that vary greatly in brightness (lightness), but little in hue (shade). Here, we examine the macroevolution of brightness in one such clade, the Furnariida. We make comparisons across light environments, body parts, monochromatic lineages, and each sex of dichromatic lineages. We found that support for models including light environments is greater for the dorsum than for the venter, and that brightness evolution is more constrained in the former than in the latter. Plumages in this clade have evolved to be darker in darker habitats, consistent with natural selection for increased crypsis. Finally, the features of brightness macroevolution are broadly similar across the sexes of the dichromatic clade, challenging the view that sexual dichromatism is driven by different evolutionary processes acting in each sex. We conclude that, in the Furnariida, light environments and dorsal–ventral variation are more important than sex as axes of color evolution.     

Long-term impacts of selective logging on two Amazonian tree species with contrasting ecological and reproductive characteristics: inferences from Eco-gene model simulations

The impact of logging and subsequent recovery after logging is predicted to vary depending on specific life history traits of the logged species. The Eco-gene simulation model was used to evaluate the long-term impacts of selective logging over 300 years on two contrasting Brazilian Amazon tree species, Dipteryx odorata and Jacaranda copaia. D. odorata (Leguminosae), a slow growing climax tree, occurs at very low densities, whereas J. copaia (Bignoniaceae) is a fast growing pioneer tree that occurs at high densities. Microsatellite multilocus genotypes of the pre-logging populations were used as data inputs for the Eco-gene model and post-logging genetic data was used to verify the output from the simulations. Overall, under current Brazilian forest management regulations, there were neither short nor long-term impacts on J. copaia. By contrast, D. odorata cannot be sustainably logged under current regulations, a sustainable scenario was achieved by increasing the minimum cutting diameter at breast height from 50 to 100 cm over 30-year logging cycles. Genetic parameters were only slightly affected by selective logging, with reductions in the numbers of alleles and single genotypes. In the short term, the loss of alleles seen in J. copaia simulations was the same as in real data, whereas fewer alleles were lost in D. odorata simulations than in the field. The different impacts and periods of recovery for each species support the idea that ecological and genetic information are essential at species, ecological guild or reproductive group levels to help derive sustainable management scenarios for tropical forests.     

First Baseline of Circulating Genotypic Lineages of Mycobacterium tuberculosis in Patients from the Brazilian Borders with Argentina and Paraguay

Background

At the triple border Brazil/Paraguay/Argentina there is easy mobility from one city to another for economic and tourism activities. This constant and fast population mobility is mainly to visit Iguazu Falls, in the Iguazu River, on the border of the Brazilian state of Paraná and the Argentina. As the incidence of tuberculosis is high in this setting, our study aimed to establish a first baseline of circulating genotypic lineages of Mycobacterium tuberculosis.

Methodology/Principal Findings

This study included 120 patients from 10 cities in southwestern Paraná, Brazil with pulmonary symptoms, from July 2009 to July 2011. Information about sex, age, clinical features and address was collected by reviewing the national tuberculosis notification database. Of these, 96 (80%) isolates were identified as M. tuberculosis and 22 (22.9%) were drug resistant (20, 20.8% INH mono-resistant and 2, 2.1% multidrug-resistant). All isolates were subjected to genotyping by Spoligotyping and MIRU-VNTR typing. The distribution of the isolates analyzed by spoligotyping revealed 30 distinct patterns. The four mainly detected clades were Latin American and Mediterranean (LAM), ill-defined T, Haarlem (H) and S. The MIRU-VNTR showed 85 distinct patterns. Spoligotyping combined to MIRU-VNTR allowed 90 distinct patterns.

Conclusions/Significance

Our study demonstrated that there is significant molecular diversity in circulating M. tuberculosis, with predominance of the LAM and T clades in cities of southwestern Paraná, Brazil, bordering Argentina and Paraguay.     

Development and characterization of highly polymorphic long TC repeat microsatellite markers for genetic analysis of peanut

Background

The immune system has paradoxical roles during cancer development and the prognostic significance of immune modulating factors is controversial. The aim of this study was to determine the expression of cyclooxygenase 2 (COX-2), transforming growth factor-beta (TGF- beta), interleukin-10 (IL-10) and their prognostic significance in breast cancers. Ki67 was included as a measure of growth fraction of tumor cells.

Methods

On immunohistochemical stained slides from 38 breast cancer patients, we performed digital video analysis of tumor cell areas and adjacent tumor stromal areas from the primary tumors and their corresponding lymph node metastases. COX-2 was recorded as graded staining intensity.

Results

The expression of TGF-beta, IL-10 and Ki67 were recorded in tumor cell areas and adjacent tumor stromal areas. In both primary tumors and metastases, the expression of COX-2 was higher in the tumor stromal areas than in the tumor cell areas (both P < 0.001). High stromal staining intensity in the primary tumors was associated with a 3.9 (95% CI 1.1-14.2) times higher risk of death compared to the low staining group (P = 0.036). The expression of TGF-beta was highest in the tumor cell areas of both primary tumors and metastases (both P < 0.001). High stromal expression of TGF-beta was associated with increased mortality. For IL-10, the stromal expression was highest in the primary tumors (P < 0.001), whereas in the metastases the expression was highest in tumor cell areas (P < 0.001). High IL-10 expression in tumor- and stromal cell areas of primary tumors predicted mortality. Ki67 was higher expressed in tumor stromal areas of the metastases, and in tumor cell areas of the primary tumors (P < 0.001). Ki67 expression in tumor cell areas and stromal areas of the metastases was independently associated with breast cancer mortality.

Conclusions

Stromal expression of COX-2, TGF-beta and Ki67 may facilitate tumor progression in breast cancer.     

The comparison of immobility time in experimental rat swimming models

Rat swimming models have been used in studies about stress and depression. However, there is no consensus about interpreting immobility (helplessness or adaptation) in the literature. In the present study, immobility time, glucose and glycogen mobilization, corticosterone and the effect of desipramine and diazepam were investigated in two different models: swimming stress and the forced swimming test. Immobility time was lower in swimming stress than in the forced swimming test. Both swimming models increased corticosterone levels in comparison with control animal levels. Moreover, swimming stress induced higher corticosterone levels than the forced swimming test did [F(2,14)=59.52; p<0.001]. Liver glycogen content values differed from one another (swimming stress相似文献