Hypolipidemic and antiatherosclerotic potential of Pleurotus ostreatus, cultived by submerged fermentation in the high-fat diet fed rats

The ability of Pleurotus ostreatus biomass, cultived by submerged fermentation, to produce beneficial effect on lipid profile and macrophages activity during a high-fat diet (HFD) for a long-term intake was investigated. Blood samples were collected through cardiac puncture to measure the plasma cholesterol, triglycerides, low-density protein (LDL), high-density protein (HDL), aspartate aminotransferase (AST) activity, urea-blood urea nitrogen (BUN)/creatinine ratio of rats fed on an HFD for 4 months. Dosage of lipid hydroperoxides was carried out on methanolic extract of liver tissue. Peritoneal macrophages activity was evaluated in relation to the superoxide anion, hydrogen peroxide and nitric oxide production, phagocytosis and lysosomal volume. The administration of P. ostreatus significantly altered the lipid profile and oxidative stress as related to the LDL and triglycerides decrease and inhibitory effects on superoxide anion and hydrogen peroxide production. All findings of this study lead us to suggest that the P. ostreatus maybe a beneficial agent in the hyperlipidemia and atherosclerosis treatments.     

Three new species of Fufius Simon, 1888 (Araneae,Cyrtaucheniidae) from Brazil with the redescription of Fufius funebris Vellard, 1924 and description of the female of Fufius lucasae Guadanucci & Indicatti, 2004

The mygalomorph neotropical genus Fufius Simon, 1888 comprises ten species, distributed from Guatemala in Central America to southeastern Brazil, in South America. Most of the species were described from northern South America, in the Amazonian region. Only F. funebris Vellard, 1924 and F. lucasae Guadanucci & Indicatti, 2004 are known from regions more to the south of the continent. Herein we describe three new Brazilian species, Fufius minusculus sp. n. and F. jalapensis sp. n. from the state of Tocantins, and F. candango sp. n. from Distrito Federal. The female of F. lucasae is described for first time and the male and female of F. funebris are redescribed based on specimens collected at the type locality.     

Cholesterol reduction ameliorates glucose-induced calcium handling and insulin secretion in islets from low-density lipoprotein receptor knockout mice

Aims/hypothesis

Changes in cellular cholesterol level may contribute to beta cell dysfunction. Islets from low density lipoprotein receptor knockout (LDLR^−/−) mice have higher cholesterol content and secrete less insulin than wild-type (WT) mice. Here, we investigated the association between cholesterol content, insulin secretion and Ca^2 + handling in these islets.

Methods

Isolated islets from both LDLR^−/− and WT mice were used for measurements of insulin secretion (radioimmunoassay), cholesterol content (fluorimetric assay), cytosolic Ca^2 + level (fura-2AM) and SNARE protein expression (VAMP-2, SNAP-25 and syntaxin-1A). Cholesterol was depleted by incubating the islets with increasing concentrations (0–10 mmol/l) of methyl-beta-cyclodextrin (MβCD).

Results

The first and second phases of glucose-stimulated insulin secretion (GSIS) were lower in LDLR^−/− than in WT islets, paralleled by an impairment of Ca^2 + handling in the former. SNAP-25 and VAMP-2, but not syntaxin-1A, were reduced in LDLR^−/− compared with WT islets. Removal of excess cholesterol from LDLR^−/− islets normalized glucose- and tolbutamide-induced insulin release. Glucose-stimulated Ca^2 + handling was also normalized in cholesterol-depleted LDLR^−/− islets. Cholesterol removal from WT islets by 0.1 and 1.0 mmol/l MβCD impaired both GSIS and Ca^2 + handling. In addition, at 10 mmol/l MβCD WT islet showed a loss of membrane integrity and higher DNA fragmentation.

Conclusion

Abnormally high (LDLR^−/− islets) or low cholesterol content (WT islets treated with MβCD) alters both GSIS and Ca^2 + handling. Normalization of cholesterol improves Ca^2 + handling and insulin secretion in LDLR^−/− islets.     

Comparison of vitrification and slow cooling for umbilical tissues

The tissue cryopreservation maintains the cellular metabolism in a quiescence state and makes the conservation possible for an indefinite period of time. The choice of an appropriate cryopreservation protocol is essential for maintenance of cryopreserved tissue banks. This study evaluated 10 samples of umbilical cord, from which small fragments of tissue (Wharton’s jelly and cord lining membrane) were subjected to two protocols of cryopreservation: slow cooling and vitrification. The samples were frozen for a period of time ranging from 5 to 78 days. The efficiency of cryopreservation was evaluated by testing cell viability, histological analysis, cell culture, cytogenetic analysis and comparison with the results of the fresh samples. The results showed that the slow cooling protocol was more efficient than the vitrification for cryopreservation of umbilical cord tissue, because it has caused fewer changes in the structure of tissue (edema and degeneration of the epithelium) and, despite the significant decrease cell viability compared to fresh samples, the ability of cell proliferation in vitro was preserved in most samples. In conclusion, this study showed that it is possible to cryopreserve small fragments of tissue from the umbilical cord and, to obtain viable cells capable of proliferation in vitro after thawing, contributing to the creation of a frozen tissue bank.     

Demographic bottlenecks in tropical plant regeneration: A comparative analysis of causal influences

Mortality factors that act sequentially through the demographic transitions from seed to sapling may have critical effects on recruitment success. Understanding how habitat heterogeneity influences the causal factors that limit propagule establishment in natural populations is central to assess these demographic bottlenecks and their consequences. Bamboos often influence forest structure and dynamics and are a major factor in generating landscape complexity and habitat heterogeneity in tropical forests. To understand how patch heterogeneity influences plant recruitment we studied critical establishment stages during early recruitment of Euterpe edulis, Sloanea guianensis and Virola bicuhyba in bamboo and non-bamboo stands in the Brazilian Atlantic forest. We combined observational studies of seed rain and seedling emergence with seed addition experiments to evaluate the transition probabilities among regeneration stages within bamboo and non-bamboo stands. The relative importance of each mortality factor was evaluated by determining how the loss of propagules affected stage-specific recruitment success. Our results revealed that the seed addition treatment significantly increased seedling survivorship for all three species. E. edulis seedling survival probability increased in the addition treatment in the two stand types. However, for S. guianensis and V. bicuhyba this effect depended strongly on artificially protecting the seeds, as both species experienced increased seed and seedling losses due to post-dispersal seed predators and herbivores. Propagules of all three species had a greater probability of reaching subsequent recruitment stages when protected. The recruitment of large-seeded V. bicuhyba and E. edulis appears to be much more limited by post-dispersal factors than by dispersal limitation, whereas the small-seeded S. guianensis showed an even stronger effect of post-dispersal factors causing recruitment collapse in some situations. We demonstrated that E. edulis, S. guianensis and V. bicuhyba are especially susceptible to predation during early compared with later establishment stages and this early stage mortality can be more crucial than stand differences as determinants of successful regeneration. Among-species differences in the relative importance of dispersal vs. establishment limitation are mediated by variability in species responses to patch heterogeneity. Thus, bamboo effects on the early recruitment of non-bamboo species are patchy and species-specific, with successional bamboo patches exerting a far-reaching influence on the heterogeneity of plant species composition and abundance.     

Shadoo/PrP (Sprn0/0/Prnp0/0) double knockout mice

Shadoo (Sho) is a brain glycoprotein with similarities to the unstructured region of PrP^C. Frameshift alleles of the Sho gene, Sprn, are reported in variant Creutzfeldt-Jakob disease (vCJD) patients while Sprn mRNA knockdown in PrP-null (Prnp^0/0) embryos produces lethality, advancing Sho as the hypothetical PrP-like “pi” protein. Also, Sho levels are reduced as misfolded PrP accumulates during prion infections. To penetrate these issues we created Sprn null alleles (Daude et al., Proc. Natl. Acad. Sci USA 2012; 109(23): 9035–40). Results from the challenge of Sprn null and TgSprn transgenic mice with rodent-adapted prions coalesce to define downregulation of Sho as a “tracer” for the formation of misfolded PrP. However, classical BSE and rodent-adapted BSE isolates may behave differently, as they do for other facets of the pathogenic process, and this intriguing variation warrants closer scrutiny. With regards to physiological function, double knockout mice (Sprn^0/0/Prnp^0/0) mice survived to over 600 d of age. This suggests that Sho is not pi, or, given the accumulating data for many activities for PrP^C, that the pi hypothesis invoking a discrete signaling pathway to maintain neuronal viability is no longer tenable.     

P-I Snake Venom Metalloproteinase Is Able to Activate the Complement System by Direct Cleavage of Central Components of the Cascade

Background

Snake Venom Metalloproteinases (SVMPs) are amongst the key enzymes that contribute to the high toxicity of snake venom. We have recently shown that snake venoms from the Bothrops genus activate the Complement system (C) by promoting direct cleavage of C-components and generating anaphylatoxins, thereby contributing to the pathology and spread of the venom. The aim of the present study was to isolate and characterize the C-activating protease from Bothrops pirajai venom.

Results

Using two gel-filtration chromatography steps, a metalloproteinase of 23 kDa that activates Complement was isolated from Bothrops pirajai venom. The mass spectrometric identification of this protein, named here as C-SVMP, revealed peptides that matched sequences from the P-I class of SVMPs. C-SVMP activated the alternative, classical and lectin C-pathways by cleaving the α-chain of C3, C4 and C5, thereby generating anaphylatoxins C3a, C4a and C5a. In vivo, C-SVMP induced consumption of murine complement components, most likely by activation of the pathways and/or by direct cleavage of C3, leading to a reduction of serum lytic activity.

Conclusion

We show here that a P-I metalloproteinase from Bothrops pirajai snake venom activated the Complement system by direct cleavage of the central C-components, i.e., C3, C4 and C5, thereby generating biologically active fragments, such as anaphylatoxins, and by cleaving the C1-Inhibitor, which may affect Complement activation control. These results suggest that direct complement activation by SVMPs may play a role in the progression of symptoms that follow envenomation.     

Pharmacological Inhibition of polysialyltransferase ST8SiaII Modulates Tumour Cell Migration

Polysialic acid (polySia), an α-2,8-glycosidically linked polymer of sialic acid, is a developmentally regulated post-translational modification predominantly found on NCAM (neuronal cell adhesion molecule). Whilst high levels are expressed during development, peripheral adult organs do not express polySia-NCAM. However, tumours of neural crest-origin re-express polySia-NCAM: its occurrence correlates with aggressive and invasive disease and poor clinical prognosis in different cancer types, notably including small cell lung cancer (SCLC), pancreatic cancer and neuroblastoma. In neuronal development, polySia-NCAM biosynthesis is catalysed by two polysialyltransferases, ST8SiaII and ST8SiaIV, but it is ST8SiaII that is the prominent enzyme in tumours. The aim of this study was to determine the effect of ST8SiaII inhibition by a small molecule on tumour cell migration, utilising cytidine monophosphate (CMP) as a tool compound. Using immunoblotting we showed that CMP reduced ST8iaII-mediated polysialylation of NCAM. Utilizing a novel HPLC-based assay to quantify polysialylation of a fluorescent acceptor (DMB-DP3), we demonstrated that CMP is a competitive inhibitor of ST8SiaII (K _i = 10 µM). Importantly, we have shown that CMP causes a concentration-dependent reduction in tumour cell-surface polySia expression, with an absence of toxicity. When ST8SiaII-expressing tumour cells (SH-SY5Y and C6-STX) were evaluated in 2D cell migration assays, ST8SiaII inhibition led to significant reductions in migration, while CMP had no effect on cells not expressing ST8SiaII (DLD-1 and C6-WT). The study demonstrates for the first time that a polysialyltransferase inhibitor can modulate migration in ST8SiaII-expressing tumour cells. We conclude that ST8SiaII can be considered a druggable target with the potential for interfering with a critical mechanism in tumour cell dissemination in metastatic cancers.     

RKIP Inhibition in Cervical Cancer Is Associated with Higher Tumor Aggressive Behavior and Resistance to Cisplatin Therapy

Cervical cancer is one of the most common cancers in women worldwide, being high-risk group the HPV infected, the leading etiological factor. The raf kinase inhibitory protein (RKIP) has been associated with tumor progression and metastasis in several human neoplasms, however its role on cervical cancer is unclear. In the present study, 259 uterine cervix tissues, including cervicitis, cervical intraepithelial lesions and carcinomas, were analyzed for RKIP expression by immunohistochemistry. We found that RKIP expression was significantly decreased during malignant progression, being highly expressed in non-neoplastic tissues (54% of the samples; 73/135), and expressed at low levels in the cervix invasive carcinomas (∼15% (19/124). Following in vitro downregulation of RKIP, we observed a viability and proliferative advantage of RKIP-inhibited cells over time, which was associated with an altered cell cycle distribution and higher colony number in a colony formation assay. An in vitro wound healing assay showed that RKIP abrogation is associated with increased migratory capability. RKIP downregulation was also associated with an increased vascularization of the tumors in vivo using a CAM assay. Furthermore, RKIP inhibition induced cervical cancer cells apoptotic resistance to cisplatin treatment. In conclusion, we described that RKIP protein is significantly depleted during the malignant progression of cervical tumors. Despite the lack of association with patient clinical outcome, we demonstrate, in vitro and in vivo, that loss of RKIP expression can be one of the factors that are behind the aggressiveness, malignant progression and chemotherapy resistance of cervical cancer.     

Evidence of Phenotypic and Genetic Relationships between Sociality,Emotional Reactivity and Production Traits in Japanese Quail

The social behavior of animals, which is partially controlled by genetics, is one of the factors involved in their adaptation to large breeding groups. To understand better the relationships between different social behaviors, fear behaviors and production traits, we analyzed the phenotypic and genetic correlations of these traits in Japanese quail by a second generation crossing of two lines divergently selected for their social reinstatement behavior. Analyses of results for 900 individuals showed that the phenotypic correlations between behavioral traits were low with the exception of significant correlations between sexual behavior and aggressive pecks both at phenotypic (0.51) and genetic (0.90) levels. Significant positive genetic correlations were observed between emotional reactivity toward a novel object and sexual (0.89) or aggressive (0.63) behaviors. The other genetic correlations were observed mainly between behavioral and production traits. Thus, the level of emotional reactivity, estimated by the duration of tonic immobility, was positively correlated with weight at 17 and 65 days of age (0.76 and 0.79, respectively) and with delayed egg laying onset (0.74). In contrast, a higher level of social reinstatement behavior was associated with an earlier egg laying onset (-0.71). In addition, a strong sexual motivation was correlated with an earlier laying onset (-0.68) and a higher number of eggs laid (0.82). A low level of emotional reactivity toward a novel object and also a higher aggressive behavior were genetically correlated with a higher number of eggs laid (0.61 and 0.58, respectively). These results bring new insights into the complex determinism of social and emotional reactivity behaviors in birds and their relationships with production traits. Furthermore, they highlight the need to combine animal welfare and production traits in selection programs by taking into account traits of sociability and emotional reactivity.