Carbonic Anhydrase Protects Fatty Liver Grafts against Ischemic Reperfusion Damage

Carbonic anhydrases (CAs) are ubiquitous metalloenzymes that catalyze the reversible hydration of carbon dioxide to bicarbonate and a proton. CAs are involved in numerous physiological and pathological processes, including acid-base homeostasis, electrolyte balance, oxygen delivery to tissues and nitric oxide generation. Given that these processes are found to be dysregulated during ischemia reperfusion injury (IRI), and taking into account the high vulnerability of steatotic livers to preservation injury, we hypothesized a new role for CA as a pharmacological agent able to protect against ischemic damage. Two different aspects of the role of CA II in fatty liver grafts preservation were evaluated: 1) the effect of its addition to Institut Georges Lopez (IGL-1) storage solution after cold ischemia; 2) and after 24h of cold storage followed by two hours of normothermic ex-vivo perfusion. In all cases, liver injury, CA II protein concentration, CA II mRNA levels and CA II activity were determined. In case of the ex-vivo perfusion, we further assessed liver function (bile production, bromosulfophthalein clearance) and Western blot analysis of phosphorylated adenosine monophosphate activated protein kinase (AMPK), mitogen activated protein kinases family (MAPKs) and endoplasmic reticulum stress (ERS) parameters (GRP78, PERK, IRE, eIF2α and ATF6). We found that CA II was downregulated after cold ischemia. The addition of bovine CA II to IGL-1 preservation solution efficiently protected steatotic liver against cold IRI. In the case of reperfusion, CA II protection was associated with better function, AMPK activation and the prevention of ERS and MAPKs activation. Interestingly, CA II supplementation was not associated with enhanced CO₂ hydration. The results suggest that CA II modulation may be a promising target for fatty liver graft preservation.     

Vibrio communities in scleractinian corals differ according to health status and geographic location in the Mediterranean Sea

The increase in seawater temperature associated with global warming is a significant threat to coral health and is linked to increasing mass mortality events and Vibrio-related coral diseases. In the Mediterranean Sea, the endemic Cladocora caespitosa and the invasive species Oculina patagonica are the main scleractinian corals affected by mass mortalities. In this study, culturable Vibrio spp. assemblages associated with healthy and unhealthy colonies of these two shallow coral species were characterized to assess the presence of Vibrio pathogens in tissue necrosis. Vibrio communities associated with O. patagonica and C. caespitosa showed geographical differences, although these became more homogeneous in unhealthy specimens of both species. Furthermore, the number of recovered Vibrio specimens was more than five times higher in unhealthy than in healthy corals. Within these culturable vibrios, the known pathogens Vibrio mediterranei and Vibrio coralliilyticus were present in unhealthy colonies of both coral species in the two localities, suggesting that they could play a role in the health status of C. caespitosa and thus act as generalist pathogens in Mediterranean corals. Nonetheless, a clonal type of V. coralliilyticus detected in C. caespitosa was not associated with disease signs, suggesting that this species could encompass assemblages with different levels of virulence.     

The intestinal stem cell signature identifies colorectal cancer stem cells and predicts disease relapse

A frequent complication in colorectal cancer (CRC) is regeneration of the tumor after therapy. Here, we report that a gene signature specific for adult intestinal stem cells (ISCs) predicts disease relapse in CRC patients. ISCs are marked by high expression of the EphB2 receptor, which becomes gradually silenced as cells differentiate. Using EphB2 and the ISC marker Lgr5, we have FACS-purified and profiled mouse ISCs, crypt proliferative progenitors, and late transient amplifying cells to define a gene program specific for normal ISCs. Furthermore, we discovered that ISC-specific genes identify a stem-like cell population positioned at the bottom of tumor structures reminiscent of crypts. EphB2 sorted ISC-like tumor cells display robust tumor-initiating capacity in immunodeficient mice as well as long-term self-renewal potential. Taken together, our data suggest that the ISC program defines a cancer stem cell niche within colorectal tumors and plays a central role in CRC relapse.     

Localisation of the Vacuolar Proton Pump (V-H+-ATPase) and Carbonic Anhydrase II in the Human Eccrine Sweat Gland

The localisation of the vacuolar proton pump (V-H+ -ATPase) and the enzyme carbonic anhydrase II (CAII) was investigated in the human eccrine sweat gland employing standard immunohistochemical techniques after antigen retrieval using microwave heat treatment and high pressure. The high-pressure antigen retrieval unmasked the presence of V-H+ -ATPase in the clear cells of the secretory coil, with a distribution similar to that previously observed for CAII. However, the dark cells were unreactive to both antibodies. In addition, heat and high-pressure antigen retrieval demonstrated the presence of CAII in the apical zone of luminal cells of the reabsorptive duct, a location not previously reported. The localisation of V-H+ -ATPase and CAII in the secretory coil clear cells suggests that the formation of HCO3- and H+ by carbonic anhydrase II and the transport of H+ by V-H+ -ATPase may play an role in sweat fluid secretion. Their presence at the apex of the duct cells indicates involvement in ductal ion reabsorption.     

Empirical Evidence for Species-Specific Export of Fish Na?veté from a No-Take Marine Protected Area in a Coastal Recreational Hook and Line Fishery

No-take marine protected areas (MPAs) are assumed to enhance fisheries catch via the “spillover” effect, where biomass is exported to adjacent exploited areas. Recent studies in spearfishing fisheries suggest that the spillover of gear-naïve individuals from protected to unprotected sites increases catch rates outside the boundaries of MPAs. Whether this is a widespread phenomenon that also holds for other gear types and species is unknown. In this study, we tested if the distance to a Mediterranean MPA predicted the degree of vulnerability to hook and line in four small-bodied coastal fish species. With the assistance of underwater video recording, we investigated the interaction effect of the distance to the boundary of an MPA and species type relative to the latency time to ingest a natural bait, which was considered as a surrogate of fish naïveté or vulnerability to fishing. Vulnerability to angling increased (i.e., latency time decreased) within and near the boundary of an MPA for an intrinsically highly catchable species (Serranus scriba), while it remained constant for an intrinsically uncatchable control species (Chromis chromis). While all of the individuals of S. scriba observed within the MPA and surrounding areas were in essence captured by angling gear, only one fifth of individuals in the far locations were captured. This supports the potential for the spillover of gear-naïve and consequently more vulnerable fish from no-take MPAs. Two other species initially characterized as intermediately catchable (Coris julis and Diplodus annularis) also had a shorter latency time in the vicinity of an MPA, but for these two cases the trend was not statistically significant. Overall, our results suggest that an MPA-induced naïveté effect may not be universal and may be confined to only intrinsically highly catchable fish species. This fact emphasizes the importance of considering the behavioural dimension when predicting the outcomes of MPAs, otherwise the effective contribution may be smaller than predicted for certain highly catchable species such as S. scriba.     

Analysis of genotypic diversity and relationships among Pseudomonas stutzeri strains by PCR-based genomic fingerprinting and multilocus enzyme electrophoresis.

Molecular fingerprinting procedures including random amplified polymorphic DNA-PCR (RAPD), repetitive extragenic palindromic PCR (rep-PCR) with REP, ERIC, and BOX primers and multilocus enzyme electrophoresis (MLEE) were used for genotypic characterization of 16 P. stutzeri strains originally isolated from marine, waste water, clinical and soil samples. A distinct genotype of each strain and overall great genotypic diversity were found within P. stutzeri. Cluster analysis (UPGMA) of the electrophoretic patterns of all PCR-based methods used resulted in concordant grouping of 8 strains. With the other strains conflicting clustering was noticed. The variability of clustering in PCR-based analyses suggested the occurrence of chromosomal rearrangements. When RAPD-, rep-PCR and MLEE fingerprints were used in a cluster analysis of combined electrophoretic patterns, the P. stutzeri strains could be differentiated into seven distinct genotypic groups. These results supported the subdivision of the species in several genomovars and reproduced, with higher resolution, the strain grouping after 16S rRNA phylogenetic reconstruction. The combined use of several fingerprint-based genotypic analyses results in higher resolutive strain clustering by UPGMA than each of the single ones analyzed separately. Additionally, this combination of individual typings proved to be reliable of the determination of the great genotypic diversity and relationships among the P. stutzeri strains.     

Amplification, contraction and genomic spread of a satellite DNA family (E180) in Medicago (Fabaceae) and allied genera

Background and Aims

Satellite DNA is a genomic component present in virtually all eukaryotic organisms. The turnover of highly repetitive satellite DNA is an important element in genome organization and evolution in plants. Here we assess the presence and physical distribution of the repetitive DNA E180 family in Medicago and allied genera. Our goals were to gain insight into the karyotype evolution of Medicago using satellite DNA markers, and to evaluate the taxonomic and phylogenetic signal of a satellite DNA family in a genus hypothesized to have a complex evolutionary history.

Methods

Seventy accessions from Medicago, Trigonella, Melilotus and Trifolium were analysed by PCR to assess the presence of the repetitive E180 family, and fluorescence in situ hybridization (FISH) was used for physical mapping in somatic chromosomes.

Key Results

The E180 repeat unit was PCR-amplified in 37 of 40 taxa in Medicago, eight of 12 species of Trigonella, six of seven species of Melilotus and in two of 11 Trifolium species. Examination of the mitotic chromosomes revealed that only 13 Medicago and two Trigonella species showed FISH signals using the E180 probe. Stronger hybridization signals were observed in subtelomeric and interstitial loci than in the pericentromeric loci, suggesting this satellite family has a preferential genomic location. Not all 13 Medicago species that showed FISH localization of the E180 repeat were phylogenetically related. However, nine of these species belong to the phylogenetically derived clade including the M. sativa and M. arborea complexes.

Conclusions

The use of the E180 family as a phylogenetic marker in Medicago should be viewed with caution. Its amplification appears to have been produced through recurrent and independent evolutionary episodes in both annual and perennial Medicago species as well as in basal and derived clades.     

Tripartitions do not always discriminate phylogenetic networks

Phylogenetic networks are a generalization of phylogenetic trees that allow for the representation of non-treelike evolutionary events, like recombination, hybridization, or lateral gene transfer. In a recent series of papers devoted to the study of reconstructibility of phylogenetic networks, Moret, Nakhleh, Warnow and collaborators introduced the so-called tripartition metric for phylogenetic networks. In this paper we show that, in fact, this tripartition metric does not satisfy the separation axiom of distances (zero distance means isomorphism, or, in a more relaxed version, zero distance means indistinguishability in some specific sense) in any of the subclasses of phylogenetic networks where it is claimed to do so. We also present a subclass of phylogenetic networks whose members can be singled out by means of their sets of tripartitions (or even clusters), and hence where the latter can be used to define a meaningful metric.