Heterogeneic nature of adult cardiac side population cells

Side population cells have been found in various types of adult tissue including heart and are presumed to be tissue-specific stem/progenitor cells. In the present study, we confirmed the presence of cardiac side population (cSP) cells, which showed both the Hoechst 33342 efflux ability and ABCG2 expression, in adult murine heart. Flow cytometric analysis showed that more than half of cSP cells expressed the endothelial marker VE-cadherin or the smooth muscle markers, α-smooth muscle actin and desmin. In addition, immunohistochemical analysis demonstrated that ABCG2⁺ cells were mainly localized within vascular walls. Quantitative RT-PCR analysis demonstrated that VE-cadherin⁻ cSP cells progressively expressed Nkx2.5 and cardiac troponin T with time in culture. VE-cadherin⁻ cSP cells also expressed mesodermal-mesenchymal-associated markers and differentiated into osteocytes and adipocytes. These results highlight the heterogeneic nature of cSP cells, consisting of vascular endothelial cells, smooth muscle cells, and mesenchymal stem/progenitor cells including potential cardiomyogenic cells.     

Differential association of HLA-B*2705 and B*2709 to ankylosing spondylitis correlates with limited peptide subsets but not with altered cell surface stability

In contrast to HLA-B*2705, B*2709 is weakly or not associated to ankylosing spondylitis. Both allotypes differ by a single D116H change. We compared the B*2705- and B*2709-bound peptide repertoires by mass spectrometry to quantify the effect of B*2709 polymorphism on peptide specificity. In addition, shared and differentially bound ligands were sequenced to define the structural features of the various peptide subsets. B*2705 shared 79% of its peptide repertoire with B*2709. Shared ligands accounted for 88% of the B*2709-bound repertoire. All B*2705 ligands not bound to B*2709 had C-terminal basic or Tyr residues. Most B*2709-bound peptides had C-terminal aliphatic and Phe residues, but two showed C-terminal Arg or Tyr. The B*2709-bound repertoire included 12% of peptides not found in B*2705. These had aliphatic C-terminal residues, which are also favored in B*2705. However, these peptides bound weakly B*2705 in vitro, indicating distinct contribution of secondary anchor residues in both subtypes. Differences in peptide binding did not affect the ratio of native to beta2-microglobulin-free HLA-B27 heavy chain at the cell surface. Our results suggest that weaker association of B*2709 with ankylosing spondylitis is based on differential binding of a limited subset of natural ligands by this allotype.     

Evolutionary history of the genus Rhamdia (Teleostei: Pimelodidae) in Central America

We constructed phylogenetic hypotheses for Mesoamerican Rhamdia, the only genus of primary freshwater fish represented by sympatric species across Central America. Phylogenetic relationships were inferred from analysis of 1990 base pairs (bp) of mitochondrial DNA (mtDNA), represented by the complete nucleotide sequences of the cytochrome b (cyt b) and the ATP synthase 8 and 6 (ATPase 8/6) genes. We sequenced 120 individuals from 53 drainages to provide a comprehensive geographic picture of Central American Rhamdia systematics and phylogeography. Phylogeographic analysis distinguished multiple Rhamdia mtDNA lineages, and the geographic congruence across evolutionarily independent Rhamdia clades indicated that vicariance has played a strong role in the Mesoamerican diversification of this genus. Phylogenetic analyses of species-level relationships provide strong support for the monophyly of a trans-Andean clade of three evolutionarily equivalent Rhamdia taxa: R. guatemalensis, R. laticauda, and R. cinerascens. Application of fish-based mitochondrial DNA clocks ticking at 1.3-1.5% sequence divergence per million years (Ma), suggests that the split between cis- and trans-Andean Rhamdia extends back about 8 Ma, and the three distinct trans-Andean Rhamdia clades split about 6 Ma ago. Thus the mtDNA divergence observed between cis- and trans-Andean Rhamdia species is too low to support an ancient colonization of Central America in the Late Cretaceous or Paleocene as had been hypothesized in one colonization model for Mesoamerican fishes. Rather the mtDNA data indicate that Rhamdia most likely colonized Central America in the late Miocene or Pliocene, promoting a strong role for the Isthmus of Panamá in the Mesoamerican expansion of this genus. Basal polytomies suggest that both the R. laticauda and R. guatemalensis clades spread rapidly across the Central American landscape, but differences in the average mtDNA genetic distances among clades comprising the two species, indicate that the R. laticauda spread and diversified across Mesoamerica about 1 million years before R. guatemalensis.     

Applying proteomics to tick vaccine development: where are we?

Introduction: Ticks are second to mosquitoes as a vector of human diseases and are the first vector of animal diseases with a great impact on livestock farming. Tick vaccines represent a sustainable and effective alternative to chemical acaricides for the control of tick infestations and transmitted pathogens. The application of proteomics to tick vaccine development is a fairly recent area, which has resulted in the characterization of some tick-host-pathogen interactions and the identification of candidate protective antigens.

Areas covered: In this article, we review the application and possibilities of various proteomic approaches for the discovery of tick and pathogen derived protective antigens, and the design of effective vaccines for the control of tick infestations and pathogen infection and transmission.

Expert commentary: In the near future, the application of reverse proteomics, immunoproteomics, structural proteomics, and interactomics among other proteomics approaches will likely contribute to improve vaccine design to control multiple tick species with the ultimate goal of controlling tick-borne diseases.     

Transfer of R388 derivatives by a pathogenesis-associated type IV secretion system into both bacteria and human cells

Bacterial type IV secretion systems (T4SSs) are involved in processes such as bacterial conjugation and protein translocation to animal cells. In this work, we have switched the substrates of T4SSs involved in pathogenicity for DNA transfer. Plasmids containing part of the conjugative machinery of plasmid R388 were transferred by the T4SS of human facultative intracellular pathogen Bartonella henselae to both recipient bacteria and human vascular endothelial cells. About 2% of the human cells expressed a green fluorescent protein (GFP) gene from the plasmid. Plasmids of different sizes were transferred with similar efficiencies. B. henselae codes for two T4SSs: VirB/VirD4 and Trw. A ΔvirB mutant strain was transfer deficient, while a ΔtrwE mutant was only slightly impaired in DNA transfer. DNA transfer was in all cases dependent on protein TrwC of R388, the conjugative relaxase, implying that it occurs by a conjugation-like mechanism. A DNA helicase-deficient mutant of TrwC could not promote DNA transfer. In the absence of TrwB, the coupling protein of R388, DNA transfer efficiency dropped 1 log. The same low efficiency was obtained with a TrwB point mutation in the region involved in interaction with the T4SS. TrwB interacted with VirB10 in a bacterial two-hybrid assay, suggesting that it may act as the recruiter of the R388 substrate for the VirB/VirD4 T4SS. A TrwB ATPase mutant behaved as dominant negative, dropping DNA transfer efficiency to almost null levels. B. henselae bacteria recovered from infected human cells could transfer the mobilizable plasmid into recipient Escherichia coli under certain conditions, underscoring the versatility of T4SSs.     

Synthesis of (di)nucleoside polyphosphates by the ubiquitin activating enzyme E1

Previous work from this laboratory had shown that ligases may catalyze the synthesis of (di)nucleoside polyphosphates. Here, we show that one of the enzymes of the proteasome system (E1 or the ubiquitin (Ub) activating enzyme, EC 6.3.2.19) catalyzes very effectively (k(cat) = 0.29+/-0.05 s(-1)) the transfer of AMP from the E-AMP-ubiquitin complex to tripolyphosphate or tetrapolyphosphate with formation of adenosine tetra- or pentaphosphate (p4A or p5A), respectively. Whereas the concomitant formation of AMP is stimulated by the presence of dithiothreitol in a concentration dependent manner, the synthesis of p4A is only slightly inhibited by this compound. Previous treatment of the enzyme (E1) with iodoacetamide inhibited only partially the synthesis of p4A. p4A can substitute for ATP as substrate of the reaction to generate the ubiquityl adenylate complex. A small amount of diadenosine pentaphosphate (Ap5A) was also synthesized in the presence of p4A.     

Synthetic chalcones, flavanones, and flavones as antitumoral agents: biological evaluation and structure-activity relationships

A series of synthetic chalcones, flavanones, and flavones has been synthesized and evaluated for antitumor activity against the human kidney carcinoma cells TK-10, human mammary adenocarcinoma cells MCF-7 (estrogen receptor-positive), and human colon adenocarcinoma cells HT-29. The most active series is the chalcone ones with the best results against TK-10 and HT-29 cells. Fourteen out of 53 analyzed compounds resulted very active against at least two of the studied tumoral cells. Alkaline single cell gel electrophoresis, comet assay, was performed as a study of the chromosomal aberrations promoted by the compounds on normal cells. Four active and two inactive chalcones were studied in the comet assay against normal human kidney cells (HK-2). A structure-activity relationship analysis of these compounds was performed and for 4- and 3,4-disubstituted derivatives a quantitative correlation was obtained in the case of anti-HT-29 activity.