Liver fatty acid-binding protein in two cases of human lipid storage

Summary FABPs in the various tissues play an important role in the intracellular fatty acid transport and metabolism. Reye's syndrome (RS) and multisystemic lipid storage (MLS) are human disorders characterized by a disturbance of lipid metabolism of unknown etiology. We investigated for the first time L-FABP in these two conditions. Affinity purified antibodies against chicken L-FABP were raised in rabbits, and found to cross-react specifically with partially purified human L-FABP. L-FABP content in liver samples of two patients with RS and MLS was investigated by immuno-histochemistry, SDS-PAGE and ELISA. L-FABP immuno-histochemistry showed increased reactivity in the liver of RS patient and normal reactivity in MLS liver. L-FABP increase in RS liver was confirmed by densitometry of SDS-PAGE and ELISA method. By these two methods the increase amounted to 180% and 199% (p < 0.02), respectively, as compared to controls. A possible role of L-FABP in the pathogenesis of RS is discussed.     

Two 2[5H]-Furanones as Possible Signaling Molecules in Lactobacillus helveticus

Two 2[5H]-furanones, in association with medium-chain fatty acids, were released in whey by Lactobacillus helveticus exposed to oxidative and heat stresses. This species plays an important role in cheese technology, particularly for Swiss-type cheeses and Grana cheese. Moreover, it significantly contributes to cheese ripening by means of an early autolysis and the release of enzymes during processing. Experimental evidence of the involvement of the two 2[5H]-furanones, detected by a gas chromatography-mass spectrometry/solid-phase microextraction technique, in the autolysis phenomenon has been obtained. Zymograms performed by using renaturing sodium dodecyl sulfate-polyacrylamide gels were used to detect the bioactivity of the supernatants containing the two furanones on fresh cells of the same strain. In addition to bands corresponding to known autolysins, new autolysins were detected concomitant with the exposure of Lactobacillus helveticus to the supernatants, which can be regarded as conditioned media (CM), and to a commercial furanone, 5-ethyl-3-hydroxy-4-methyl-2[5H]-furanone (HEMFi), having spectral data similar to those of the newly described 2[5H]-furanones. Morphological changes were observed when fresh cells were exposed to CM containing the two 2[5H]-furanones and HEMFi. The two furanones produced by Lactobacillus helveticus, which met a number of criteria to be included in cell-cell signaling molecules, have a presumptive molecular mass lower than those of already known 3[2H]-furanones having an autolytic activity and being produced by gram-negative bacteria. Moreover, they present a different chemical structure with respect to the furanones already identified as products of Lactococcus lactis subsp. cremoris or to those identified in some cheeses with Lactobacillus helveticus as a starter culture.     

Anthocyanins Double the Shelf Life of Tomatoes by Delaying Overripening and Reducing Susceptibility to Gray Mold

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Fluoro‐Sorafenib (Regorafenib) effects on hepatoma cells: Growth inhibition,quiescence, and recovery

To evaluate the growth‐inhibitory properties of the potent multi‐kinase antagonist Regorafenib (Fluoro‐Sorafenib), which was synthesized as a more potent Sorafenib, a Raf inhibitor and to determine whether similar mechanisms were involved, human hepatoma cell lines were grown in the presence or absence of Regorafanib and examined for growth inhibition. Western blots were performed for Raf targets, apoptosis, and autophagy. Regorafenib inhibited growth of human Hep3B, PLC/PRF/5, and HepG2 cells in a concentration‐ and time‐dependent manner. Multiple signaling pathways were altered, including MAP kinases phospho‐ERK and phospho‐JNK and its target phospho‐c‐Jun. There was evidence for apoptosis by FACS, cleavage of caspases and increased Bax levels; as well as induction of autophagy, as judged by increased Beclin‐1 and LC3 (II) levels. Prolonged drug exposure resulted in cell quiescence. Full growth recovery occurred after drug removal, unlike with doxorubicin chemotherapy. Regorafenib is a potent inhibitor of cell growth. Cells surviving Regorafenib treatment remain viable, but quiescent and capable of regrowth following drug removal. The reversibility of tumor cell growth suppression after drug removal may have clinical implications. J. Cell. Physiol. 228: 292–297, 2013. © 2012 Wiley Periodicals, Inc.     

Identification of Fungal Species Associated with Cladode Spot of Prickly Pear and Their Sensitivity to Chitosan

México is the most important producer of prickly pear (Opuntia ficus‐indica) in the world. There are several fungal diseases that can have a negative impact on their yields. In this study, there was a widespread fungal richness on cladodes spot of prickly pears from México. A total of 41 fungi isolates were obtained from cladodes spot; 11 of them were morphologically different. According to the pathogenicity test, seven isolates caused lesions on cladodes. The morphological and molecular identification evidenced the isolation of Colletotrichum gloeosporioides, Alternaria alternata, Fusarium lunatum, Curvularia lunata. All these species caused similar symptoms of circular cladodes spot. However, it is noticeable that some lesions showed perforation and detachment of affected tissues by Fusarium lunatum. To our knowledge, this is the first report of the Fusarium lunatum as phytopathogenic fungus of cladodes of prickly pear. The chitosan inhibited the mycelium growth in the seven isolates of phytopathogenic fungi. Chitosan applications diminished the disease incidence caused by C. gloeosporioies and F. lunatum in 40 and 100%, respectively. Likewise, the lesion severity index in cladodes decreased. There are no previous reports about the application of chitosan on cladodes of prickly pears for the control of phytopathogenic fungi. Therefore, this research could contribute to improve the strategies for the management of diseases in prickly pear.     

Microcin E492 Amyloid Formation Is Retarded by Posttranslational Modification

Microcin E492, a channel-forming bacteriocin with the ability to form amyloid fibers, is exported as a mixture of two forms: unmodified (inactive) and posttranslationally modified at the C terminus with a salmochelin-like molecule, which is an essential modification for conferring antibacterial activity. During the stationary phase, the unmodified form accumulates because expression of the maturation genes mceIJ is turned off, and microcin E492 is rapidly inactivated. The aim of this work was to demonstrate that the increase in the proportion of unmodified microcin E492 augments the ability of this bacteriocin to form amyloid fibers, which in turn decreases antibacterial activity. To this end, strains with altered proportions of the two forms were constructed. The increase in the expression of the maturation genes augmented the antibacterial activity during all growth phases and delayed the loss of activity in the stationary phase, while the ability to form amyloid fibers was markedly reduced. Conversely, a higher expression of microcin E492 protein produced concomitant decreases in the levels of the modified form and in antibacterial activity and a substantial increase in the ability to form amyloid fibers. The same morphology for these fibers, including those formed by only the unmodified version, was observed. Moreover, seeds formed using exclusively the nonmodified form were remarkably more efficient in amyloid formation with a shorter lag phase, indicating that the nucleation process is probably improved. Unmodified microcin E492 incorporation into amyloid fibers was kinetically more efficient than the modified form, probably due to the existence of a conformation that favors this process.     

Gene Expression Analysis of PTEN Positive Glioblastoma Stem Cells Identifies DUB3 and Wee1 Modulation in a Cell Differentiation Model

The term astrocytoma defines a quite heterogeneous group of neoplastic diseases that collectively represent the most frequent brain tumors in humans. Among them, glioblastoma multiforme represents the most malignant form and its associated prognosis is one of the poorest among tumors of the central nervous system. It has been demonstrated that a small population of tumor cells, isolated from the brain neoplastic tissue, can reproduce the parental tumor when transplanted in immunodeficient mouse. These tumor initiating cells are supposed to be involved in cancer development and progression and possess stem cell-like features; like their normal counterpart, these cells remain quiescent until they are committed to differentiation. Many studies have shown that the role of the tumor suppressor protein PTEN in cell cycle progression is fundamental for tumor dynamics: in low grade gliomas, PTEN contributes to maintain cells in G1 while the loss of its activity is frequently observed in high grade gliomas. The mechanisms underlying the above described PTEN activity have been studied in many tumors, but those involved in the maintenance of tumor initiating cells quiescence remain to be investigated in more detail. The aim of the present study is to shed light on the role of PTEN pathway on cell cycle regulation in Glioblastoma stem cells, through a cell differentiation model. Our results suggest the existence of a molecular mechanism, that involves DUB3 and WEE1 gene products in the regulation of Cdc25a, as functional effector of the PTEN/Akt pathway.     

Bioconversion of Agave tequilana fructans by exo-inulinases from indigenous Aspergillus niger CH-A-2010 enhances ethanol production from raw Agave tequilana juice

Agave tequilana fructans are the source of fermentable sugars for the production of tequila. Fructans are processed by acid hydrolysis or by cooking in ovens at high temperature. Enzymatic hydrolysis is considered an alternative for the bioconversion of fructans. We previously described the isolation of Aspergillus niger CH-A-2010, an indigenous strain that produces extracellular inulinases. Here we evaluated the potential application of A. niger CH-A-2010 inulinases for the bioconversion of A. tequilana fructans, and its impact on the production of ethanol. Inulinases were analyzed by Western blotting and thin layer chromatography. Optimal pH and temperature conditions for inulinase activity were determined. The efficiency of A. niger CH-A-2010 inulinases was compared with commercial enzymes and with acid hydrolysis. The hydrolysates obtained were subsequently fermented by Saccharomyces cerevisiae to determine the efficiency of ethanol production. Results indicate that A. niger CH-A-2010 predominantly produces an exo-inulinase activity. Optimal inulinase activity occurred at pH 5.0 and 50 °C. Hydrolysis of raw agave juice by CH-A-2010 inulinases yielded 33.5 g/l reducing sugars, compared with 27.3 g/l by Fructozyme^® (Novozymes Corp, Bagsværd, Denmark) and 29.4 g/l by acid hydrolysis. After fermentation of hydrolysates, we observed that the conversion efficiency of sugars into ethanol was 97.5 % of the theoretical ethanol yield for enzymatically degraded agave juice, compared to 83.8 % for acid-hydrolyzed juice. These observations indicate that fructans from raw Agave tequilana juice can be efficiently hydrolyzed by using A. niger CH-A-2010 inulinases, and that this procedure impacts positively on the production of ethanol.     

Embryology and embryogenesis of Spiranthes L.C.M. Richard (Orchidaceae): S. spiralis (L.) Cheval and S. aestivalis (Poiret) L.C.M. Richard

Abstract

The various stages of female gametophyte development and embryogenesis in S. spiralis and S. aestivalis are described. In both species the reproductive cycle is sexual. Some peculiarities are present: the female gametophyte is usually 6-7-8-nucleate; after double fertilization a single endospermatic cell is formed; the proembryo appears differentiated and is made up of different cells in the chalazal and micropylar ends; a single basal cell in the proembryo acts as suspensor.