In Xenopus laevis eggs, cisternae shells which surround cortical granules (CG) are part of a cortical endoplasmic reticulum (ER) network. In this paper the origin of such ER shells has been studied in full-grown, progesterone-exposed Xenopus oocytes. Furthermore, the possible role of the cortical ER in the activation process has been investigated by pricking maturing oocytes. It has been shown that in full-grown ovarian oocytes ER CG shells are absent and ER cisternae are extensively and randomly distributed throughout the peripheral cytoplasm, where they appear to be continuous with annulate lamellae (AL). Following hormone treatment, the AL completely disaggregate and the ER cisternae gradually migrate to the cortex where they surround the CG constituting the typical cortical network described in uterine eggs. Furthermore, it has been found that 8 h after progesterone treatment (before the first polar body extrusion) the response to pricking (CG exocytosis) occurs only at the animal half; there is no observable response in the vegetal half. At this time ER shells surround CG only in the animal hemisphere. A complete CG exocytosis occurs following the first polar body emission, when the cortical ER is well organized in the whole oocyte cortex. The correlation between the differentiation of the cortical ER and the arousal in the oocyte of the ability to respond to a pricking stimulus is discussed in the light of an involvement of the cortical ER in the propagation of CG exocytosis.
The combination of an increase in the cAMP-phosphodiesterase activity of h-prune and its interaction with nm23-H1 have been shown to be key steps in the induction of cellular motility in breast cancer cells. Here we present the molecular mechanisms of this interaction. The region of the nm23-h-prune interaction lies between S120 and S125 of nm23, where missense mutants show impaired binding; this region has been highly conserved throughout evolution, and can undergo serine phosphorylation by casein kinase I. Thus, the casein kinase I d-e specific inhibitor IC261 impairs the formation of the nm23-hprune complex, which translates 'in vitro' into inhibition of cellular motility in a breast cancer cellular model. A competitive permeable peptide containing the region for phosphorylation by casein kinase I impairs cellular motility to the same extent as IC261. The identification of these two modes of inhibition of formation of the nm23-H1-h-prune protein complex pave the way toward new challenges, including translational studies using IC261 or this competitive peptide 'in vivo' to inhibit cellular motility induced by nm23-H1-h-prune complex formation during progression of breast cancer.
The aim of this study was to determine the prevalence of sarcosporidiosis in semi-intensively bred cattle in northwestern Italy. A diagnostic protocol was setup in which infected animals were identified by rapid histological examination of the esophagus, diaphragm, and heart and the detected Sarcocystis spp. were subsequently typed using conventional electron microscopy in combination with molecular techniques. Sarcosporidia cysts were detected in 78.1% of the animals and were seen most often in the esophagus. The cattle is intermediate host for Sarcocystis hominis (final host, humans and some primates), Sarcocystis cruzi (final host, domestic and wild canids), and Sarcocystis hirsuta (final host, wild and domestic cats).All these three species of Sarcocystis were identified, variously associated, with the following prevalence: S. cruzi (74.2%), S. hirsuta (1.8%), and S. hominis (42.7%). Furthermore, a new S. hominis-like (prevalence 18.5%), characterized by hook-like structures of villar protrusion and a different sequence of the 18S rRNA gene, was identified. The cattle sheds testing positive for zoonotic Sarcocystis were assessed for risk factors contributing to the maintenance of the parasite's life cycle. Significant associations emerged between consumption of raw meat by the farm owner, mountain pasturing, and absence of a sewerage system on the farm and cattle breed. Our study demonstrates that sarcosporidiosis may constitute a public health problem in Italy and indicates several issues to be addressed when planning surveillance and prevention actions. The applied diagnostic approach revealed that cattle can harbor a further type of Sarcocystis, of which life cycle and zoonotic potential should be investigated.
BackgroundCancer stem cells (CSCs) are considered the cell subpopulation responsible for breast cancer (BC) initiation, growth, and relapse. CSCs are identified as self-renewing and tumor-initiating cells, conferring resistance to chemo- and radio-therapy to several neoplasias. Nowadays, th (about 10mM)e pharmacological targeting of CSCs is considered an ineludible therapeutic goal. The antidiabetic drug metformin was reported to suppress in vitro and in vivo CSC survival in different tumors and, in particular, in BC preclinical models. However, few studies are available on primary CSC cultures derived from human postsurgical BC samples, likely because of the limited amount of tissue available after surgery. In this context, comparative oncology is acquiring a relevant role in cancer research, allowing the analysis of larger samples from spontaneous pet tumors that represent optimal models for human cancer.MethodsIsolation of primary canine mammary carcinoma (CMC) cells and enrichment in stem-like cell was carried out from fresh tumor specimens by culturing cells in stem-permissive conditions. Phenotypic and functional characterization of CMC-derived stem cells was performed in vitro, by assessment of self-renewal, long-lasting proliferation, marker expression, and drug sensitivity, and in vivo, by tumorigenicity experiments. Corresponding cultures of differentiated CMC cells were used as internal reference. Metformin efficacy on CMC stem cell viability was analyzed both in vitro and in vivo.ResultsWe identified a subpopulation of CMC cells showing human breast CSC features, including expression of specific markers (i.e. CD44, CXCR4), growth as mammospheres, and tumor-initiation in mice. These cells show resistance to doxorubicin but were highly sensitive to metformin in vitro. Finally, in vivo metformin administration significantly impaired CMC growth in NOD-SCID mice, associated with a significant depletion of CSCs.ConclusionsSimilarly to the human counterpart, CMCs contain stem-like subpopulations representing, in a comparative oncology context, a valuable translational model for human BC, and, in particular, to predict the efficacy of antitumor drugs. Moreover, metformin represents a potential CSC-selective drug for BC, as effective (neo-)adjuvant therapy to eradicate CSC in mammary carcinomas of humans and animals.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-015-1235-8) contains supplementary material, which is available to authorized users.
In the egg of the anuran Discoglossus pictus, the site of fertilization is restricted to the central portion of an animal hemisphere indentation (the dimple). Previous studies showed that the acrosome reaction of D. pictus sperm is triggered in the jelly, and yet sperm arrive at the dimple surface with the plasma membrane at an early stage of vesiculation. Reactivity of the dimple surface with specific lectins suggests that fucose might be utilized as a marker of glycoproteins located at the dimple surface. In this paper, proteins of the egg surface were labeled with the membrane impermeable sulfo-NHS-biotin. Four main bands of 200, 230, 260, and 270 kDa labeled only at the dimple surface, although they were detected in the cortex of the whole egg. The 270-kDa band reacted with Galanthus nivalis agglutinin only in the cortex of the dimple, suggesting that this band is differently glycosylated according to its localization. The alpha-l-fucose-specific lectin Ulex europaeus agglutinin I was utilized both in lectin blotting and in affinity chromatography and cross-reacted with the 200- and 270/260-kDa bands. Furthermore, two polypeptides were obtained by exposure of intact eggs to lysylendoproteinase C. They were also reactive to Ulex europaeus agglutinin I. The 200- and 270/260-kDa bands were eluted from the acrylamide gels and adsorbed to polystyrene beads. An assay for sperm binding to 200-kDa glycoprotein-bound beads was developed. Sperm stuck to the beads before but not after Ca-ionophore treatment. When the beads were coated with the 270/260-kDa glycoproteins, binding occurred after ionophore treatment. In these assays, the 200- and 270/260-kDa glycoproteins competitively inhibited sperm binding to the beads coated with the corresponding glycoprotein. These results indicate that the assayed glycoproteins, located either in the glycocalyx or in the plasma membrane of the fertilization site, are involved in sperm binding.
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