Cellularity and structure of fresh human coronary thrombectomy specimens; presence of cells with markers of progenitor cells

Acute coronary syndromes and acute myocardial infarctions are often related to plaque rupture and the formation of thrombi at the site of the rupture. We examined fresh coronary thrombectomy specimens from patients with acute coronary syndromes and assessed their structure and cellularity. The thrombectomy specimens consisted of platelets, erythrocytes and inflammatory cells. Several specimens contained multiple cholesterol crystals. Culture of thrombectomy specimens yielded cells growing in various patterns depending on the culture medium used. Culture in serum‐free stem cell enrichment medium yielded cells with features of endothelial progenitor cells which survived in culture for a year. Immunohistochemical analysis of the thrombi revealed cells positive for CD34, cells positive for CD15 and cells positive for desmin in situ, whereas cultured cell from thrombi was desmin positive but pancytokeratin negative. Cells cultured in endothelial cell medium were von Willebrand factor positive. The content of coronary thrombectomy specimens is heterogeneous and consists of blood cells but also possibly cells from the vascular wall and cholesterol crystals. The culture of cells contained in the specimens yielded multiplying cells, some of which demonstrated features of haematopoietic progenitor cells and which differentiated into various cell‐types.     

Effects of a 50 Hz magnetic field on Dictyostelium discoideum (Protista)

Some studies have demonstrated that a few biological systems are affected by weak, extremely low frequency (ELF) electromagnetic fields (EMFs), lower than 10 mT. However, to date there is scanty evidence of this effect on Protists in the literature. Due to their peculiarity as single-cell eukaryotic organisms, Protists respond directly to environmental stimuli, thus appearing as very suitable experimental systems. Recently, we showed the presence of propionylcholinesterase (PrChE) activity in single-cell amoebae of Dictyostelium discoideum. This enzyme activity was assumed to be involved in cell-cell and cell-environment interactions, as its inhibition affects cell aggregation and differentiation. In this work, we have exposed single-cell amoebae of D. discoideum to an ELF-EMF of about 200 microT, 50 Hz, for 3 h or 24 h at 21 degrees C. A delay in the early phase of the differentiation was observed in 3 h exposed cells, and a significant decrease in the fission rate appeared in 24 h exposed cells. The PrChE activity was significantly lower in 3 h exposed cells than in the controls, whereas 24 h exposed cells exhibited an increase in this enzyme activity. However, such effects appeared to be transient, as the fission rate and PrChE activity values returned to the respective control values after a 24 h stay under standard conditions.     

Trophic ecology of parabiotic ants: Do the partners have similar food niches?

Organisms associated with another species may experience both costs and benefits from their partner. One of these costs is competition, which is the more likely if the two species are ecologically similar. Parabioses are associations between two ant species that share a nest and often attend the same food sources. Albeit parabioses are probably mutualistic, parabiotic partners may compete for food. We therefore investigated feeding niches and dietary overlap of two parabiotically associated ants in Borneo using cafeteria experiments and stable isotope analyses. The two species strongly differed in their food choices. While Crematogaster modiglianii mostly foraged at carbohydrate‐rich baits, Camponotus rufifemur preferred urea‐rich sources. Both species also consumed animal protein. The ¹⁵N concentration in Ca. rufifemur workers was consistently lower than in Cr. modiglianii. Camponotus rufifemur but not Cr. modiglianii possesses microbial endosymbionts, which can metabolize urea and synthesize essential amino acids. Its lower ¹⁵N signature may result from a relatively higher intake of plant‐based or otherwise ¹⁵N‐depleted nitrogen. Isotopic signatures of the two partners in the same parabiosis showed strongly parallel variation across nests. As we did not find evidence for spatial autocorrelation, this correlation suggests an overlap of food sources between the two ant species. Based on model simulations, we estimated a diet overlap of 22–66% for nitrogen sources and 45–74% for carbon sources. The overlap may arise from either joint exploitation of the same food sources or trophallactic exchange of food. This suggests an intense trophic interaction and potential for competition between the parabiotic partners.     

Identification of the catalytic nucleophile of the family 29 alpha-L-fucosidase from Sulfolobus solfataricus via chemical rescue of an inactive mutant

We have recently reported that a functional alpha-L-fucosidase could be expressed by a single insertional mutation in the region of overlap between the ORFs SSO11867 and SSO3060 of the hyperthermophilic Archaeon Sulfolobus solfataricus [Cobucci-Ponzano et al. J. Biol. Chem. (2003) 278, 14622-14631]. This enzyme, belonging to glycoside hydrolase family 29 (GH29), showed micromolar specificity for p-nitrophenyl-alpha-L-fucoside (pNp-Fuc) and promoted transfucosylation reactions by following a reaction mechanism in which the products retained the anomeric configuration of the substrate. The active site residues in GH29 enzymes are still unknown. We describe here the identification of the catalytic nucleophile of the reaction in the alpha-L-fucosidase from S. solfataricus by reactivation with sodium azide of the mutant Asp242Gly that shows a 10(3)-fold activity reduction on pNp-Fuc. The detailed stereochemical analysis of the fucosyl-azide produced by the mutant reactivated on pNp-Fuc revealed its inverted (beta-fucosyl azide) configuration compared with the substrate. This allows for the first time the unambiguous assignment of Asp242, and its homologous residues, as the nucleophilic catalytic residues of GH29 alpha-L-fucosidases. This is the first time that this approach is used for alpha-L-glycosidases, widening the applicability of this method.     

Socioeconomic and nutritional factors account for the association of gastric cancer with amerindian ancestry in a latin american admixed population

Gastric cancer is one of the most lethal types of cancer and its incidence varies worldwide, with the Andean region of South America showing high incidence rates. We evaluated the genetic structure of the population from Lima (Peru) and performed a case-control genetic association study to test the contribution of African, European, or Native American ancestry to risk for gastric cancer, controlling for the effect of non-genetic factors. A wide set of socioeconomic, dietary, and clinic information was collected for each participant in the study and ancestry was estimated based on 103 ancestry informative markers. Although the urban population from Lima is usually considered as mestizo (i.e., admixed from Africans, Europeans, and Native Americans), we observed a high fraction of Native American ancestry (78.4% for the cases and 74.6% for the controls) and a very low African ancestry (<5%). We determined that higher Native American individual ancestry is associated with gastric cancer, but socioeconomic factors associated both with gastric cancer and Native American ethnicity account for this association. Therefore, the high incidence of gastric cancer in Peru does not seem to be related to susceptibility alleles common in this population. Instead, our result suggests a predominant role for ethnic-associated socioeconomic factors and disparities in access to health services. Since Native Americans are a neglected group in genomic studies, we suggest that the population from Lima and other large cities from Western South America with high Native American ancestry background may be convenient targets for epidemiological studies focused on this ethnic group.     

Computational and Experimental Studies on β-Sheet Breakers Targeting Aβ1–40 Fibrils

In this work we present and compare the results of extensive molecular dynamics simulations of model systems comprising an Aβ_1–40 peptide in water in interaction with short peptides (β-sheet breakers) mimicking the 17–21 region of the Aβ_1–40 sequence. Various systems differing in the customized β-sheet breaker structure have been studied. Specifically we have considered three kinds of β-sheet breakers, namely Ac-LPFFD-NH₂ and two variants thereof, one obtained by substituting the acetyl group with the sulfonic amino acid taurine (Tau-LPFFD-NH₂) and a second novel one in which the aspartic acid is substituted by an asparagine (Ac-LPFFN-NH₂). Thioflavin T fluorescence, circular dichroism, and mass spectrometry experiments have been performed indicating that β-sheet breakers are able to inhibit in vitro fibril formation and prevent the β sheet folding of portions of the Aβ_1–40 peptide. We show that molecular dynamics simulations and far UV circular dichroism provide consistent evidence that the new Ac-LPFFN-NH₂ β-sheet breaker is more effective than the other two in stabilizing the native α-helix structure of Aβ_1–40. In agreement with these results thioflavin T fluorescence experiments confirm the higher efficiency in inhibiting Aβ_1–40 aggregation. Furthermore, mass spectrometry data and molecular dynamics simulations consistently identified the 17–21 Aβ_1–40 portion as the location of the interaction region between peptide and the Ac-LPFFN-NH₂ β-sheet breaker.     

The electron impact, chemical ionization and fast atom bombardment positive ion mass spectra of 1,2-bis(sulfonyl)methylhydrazines.

A series of bis(sulfonyl)-1-methylhydrazines were analyzed by positive ion electron impact (EI), chemical ionization (CI) and fast atom bombardment (FAB) mass spectrometry. Since these compounds showed activity against the L1210 leukemia, an understanding of their mass spectral behavior is important should the structural characterization of metabolites be required. FAB proved to be the most useful technique, generally providing abundant protonated molecule ion peaks, in contrast to the weak peaks observed with CI (ammonia or isobutane) and the total absence of molecular ion peaks in the EI mass spectra. In addition, utilizing FAB eliminated the problem of thermal decomposition, which was very difficult to control under EI and CI experimental conditions. Fragments observed in FAB and CI mass spectra were consistent with protonation at the methyl-bearing nitrogen. One can locate the R1 and R2 moieties relative to the methyl-bearing nitrogen in FAB and CI by assigning that nitrogen as the site of protonation, with subsequent elimination of R2SO2H.     

Modeling human osteosarcoma in mice through 3AB-OS cancer stem cell xenografts

Osteosarcoma is the second leading cause of cancer‐related death for children and young adults. In this study, we have subcutaneously injected—with and without matrigel—athymic mice (Fox1nu/nu) with human osteosarcoma 3AB‐OS pluripotent cancer stem cells (CSCs), which we previously isolated from human osteosarcoma MG63 cells. Engrafted 3AB‐OS cells were highly tumorigenic and matrigel greatly accelerated both tumor engraftment and growth rate. 3AB‐OS CSC xenografts lacked crucial regulators of beta‐catenin levels (E‐cadherin, APC, and GSK‐3beta), and crucial factors to restrain proliferation, resulting therefore in a strong proliferation potential. During the first weeks of engraftment 3AB‐OS‐derived tumors expressed high levels of pAKT, beta1‐integrin and pFAK, nuclear beta‐catenin, c‐Myc, cyclin D2, along with high levels of hyperphosphorylated‐inactive pRb and anti‐apoptotic proteins such as Bcl‐2 and XIAP, and matrigel increased the expression of proliferative markers. Thereafter 3AB‐OS tumor xenografts obtained with matrigel co‐injection showed decreased proliferative potential and AKT levels, and undetectable hyperphosphorylated pRb, whereas beta1‐integrin and pFAK levels still increased. Engrafted tumor cells also showed multilineage commitment with matrigel particularly favoring the mesenchymal lineage. Concomitantly, many blood vessels and muscle fibers appeared in the tumor mass. Our findings suggest that matrigel might regulate 3AB‐OS cell behavior providing adequate cues for transducing proliferation and differentiation signals triggered by pAKT, beta1‐integrin, and pFAK and addressed by pRb protein. Our results provide for the first time a mouse model that recapitulates in vivo crucial features of human osteosarcoma CSCs that could be used to test and predict the efficacy in vivo of novel therapeutic treatments. J. Cell. Biochem. 113: 3380–3392, 2012. © 2012 Wiley Periodicals, Inc.