Blood derived stem cells: an ameliorative therapy in veterinary ophthalmology

Stem cell technology has evoked considerable excitement among people interested in the welfare of animals, as it has suggested the potential availability of new tools for several pathologies, including eye disease, which in many cases is considered incurable. One such example is ulcerative keratitis, which is very frequent in horses. Because some of these corneal ulcers can be very severe, progress rapidly and, therefore, can be a possible cause of vision loss, it is important to diagnose them at an early stage and administer an appropriate treatment, which can be medical, surgical, or a combination of both. The therapeutic strategy should eradicate the infection in order to reduce or stop destruction of the cornea. In addition, it should support the corneal structures and control the uveal reaction, and the pain associated with it, in order to minimize scarring. In this study, we address how stem cells derived from peripheral blood can be used also in ophthalmological pathologies. Our results demonstrate that this treatment protocol improved eye disease in four horse cases, including corneal ulcers and one case of retinal detachment. In all cases, we detected a decrease in the intense inflammatory reaction as well as the restoration of the epithelial surface of the central cornea.     

Liposomes as a Model for the Biological Membrane: Studies on Daunorubicin Bilayer Interaction

In this study the interaction of the antitumoral drug daunorubicin with egg phosphatidylcholine (EPC) liposomes, used as a cell membrane model, was quantified by determination of the partition coefficient (K _p). The liposome/aqueous-phase K _p of daunorubicin was determined by derivative spectrophotometry and measurement of the zeta-potential. Mathematical models were used to fit the experimental data, enabling determination of K _p. In the partition of daunorubicin within the membrane both superficial electrostatic and inner hydrophobic interactions seem to be involved. The results are affected by the two types of interaction since spectrophotometry measures mainly hydrophobic interactions, while zeta-potential is affected by both interpenetration of amphiphilic charged molecules in the bilayer and superficial electrostatic interaction. Moreover, the degree of the partition of daunorubicin with the membrane changes with the drug concentration, due mainly to saturation factors. Derivative spectrophotometry and zeta-potential variation results, together with the broad range of concentrations studied, revealed the different types of interactions involved. The mathematical formalism applied also allowed quantification of the number of lipid molecules associated with one drug molecule.     

The complexity of cardiolipin in health and disease

Cardiolipin, the signature phospholipid of mitochondria, is a lipid dimer that is important for a diverse range of mitochondrial activities beyond the process of ATP production. Thus not surprisingly, derangements in cardiolipin metabolism are now appreciated to contribute to an assortment of pathological conditions. A comprehensive inventory of enzymes involved in cardiolipin biosynthesis and remodeling was just recently obtained. Post-biosynthesis, the acyl chain composition of cardiolipin is modified by up to three distinct remodeling enzymes that produce either a homogeneous tissue-specific mature form of cardiolipin or alternatively 'bad' cardiolipin that has been linked to mitochondrial dysfunction. In this review, we initially focus on the newly identified players in cardiolipin metabolism and then shift our attention to how changes in cardiolipin metabolism contribute to human disease.     

How cellulose-based leaf toughness and lamina density contribute to long leaf lifespans of shade-tolerant species

Cell wall fibre and lamina density may interactively affect leaf toughness and leaf lifespan. Here, we tested this with seedlings of 24 neotropical tree species differing in shade tolerance and leaf lifespan under standardized field conditions (140-867 d in gaps; longer in shade). We quantified toughness with a cutting test, explicitly seeking a mechanistic linkage to fibre. Lamina density, but not fracture toughness, exhibited a plastic response to gaps vs shade, while neither trait was affected by leaf age. Toughness corrected for lamina density, a recently recognized indicator of material strength per unit mass, was linearly correlated with cellulose content per unit dry mass. Leaf lifespan was positively correlated with cellulose and toughness in shade-tolerant species but only weakly in gap-dependent species. Leaf lifespan was uncorrelated with lamina thickness, phenolics and tannin concentrations. In path analysis including all species, leaf lifespan was directly enhanced by density and toughness, and indirectly by cellulose via its effect on toughness. Different suites of leaf traits were correlated with early seedling survival in gaps vs shade. In conclusion, cellulose and lamina density jointly enhance leaf fracture toughness, and these carbon-based physical traits, rather than phenolic-based defence, explain species differences in herbivory, leaf lifespan and shade survival.     

Evaluation of aflatoxin B1 and ochratoxin A in interacting mixed cultures of Aspergillus sections Flavi and Nigri on peanut grains

The influence of inoculum size on the colony-forming units, production of aflatoxin B₁ (AFB₁) and ochratoxin A (OTA) was determined when Aspergillus flavus and A. niger aggregate strains were cultured alone and in pairs on irradiated peanut grains at 28°C and 0.97 water activity (a_W). The results showed a marked influence of inoculum factor on fungal counts, AFB₁ and OTA production in single and paired cultures. Fungal counts of the A. niger aggregate strain in interacting cultures at 7, 14 and 21?days of incubation were significantly higher than those observed in the A. flavus strain, except in the mixed culture with 10² spores/ml of both strains. In all mixed culture assays, the AFB₁ production was significantly reduced in comparison with the accumulation of mycotoxin in single cultures. A total inhibition in AFB₁ production was observed in some interactions as 10² spores/ml of A. flavus and 10³ spores/ml of A. niger aggregate strain at 7 and 14?days, among others. With regard to OTA production, a stimulation in the interacting cultures was observed at all inoculum sizes and incubation period. The highest levels of OTA accumulation were observed at 14?days for all interacting cultures. The maximum level was reach in the culture 10³ spores/ml of A. niger aggregate and 10⁴ spores/ml of A. flavus (p?<?0.001). These results suggest that, under optimal environmental conditions in peanut grains, the interaction between A. flavus and A. niger aggregate strains could result in an inhibition of AFB₁ and in a stimulation of OTA production.     

Mitochondrial mislocalization and altered assembly of a cluster of Barth syndrome mutant tafazzins

None of the 28 identified point mutations in tafazzin (Taz1p), which is the mutant gene product associated with Barth syndrome (BTHS), has a biochemical explanation. In this study, endogenous Taz1p was localized to mitochondria in association with both the inner and outer mitochondrial membranes facing the intermembrane space (IMS). Unexpectedly, Taz1p does not contain transmembrane (TM) segments. Instead, Taz1p membrane association involves a segment that integrates into, but not through, the membrane bilayer. Residues 215-232, which were predicted to be a TM domain, were identified as the interfacial membrane anchor by modeling four distinct BTHS mutations that occur at conserved residues within this segment. Each Taz1p mutant exhibits altered membrane association and is nonfunctional. However, the basis for Taz1p dysfunction falls into the following two categories: (1) mistargeting to the mitochondrial matrix or (2) correct localization associated with aberrant complex assembly. Thus, BTHS can be caused by mutations that alter Taz1p sorting and assembly within the mitochondrion, indicating that the lipid target of Taz1p is resident to IMS-facing leaflets.     

Symmetry breaking in interspecific Drosophila hybrids is not due to developmental noise

Hybrids from crosses of different species have been reported to display decreased developmental stability when compared to their pure species, which is conventionally attributed to a breakdown of coadapted gene complexes. Drosophila subobscura and its close relative D. madeirensis were hybridized in the laboratory to test the hypothesis that genuine fluctuating asymmetry, measured as the within-individual variance between right and left wings that results from random perturbations in development, would significantly increase after interspecific hybridization. When sires of D. subobscura were mated to heterospecific females following a hybrid half-sib breeding design, F1 hybrid females showed a large bilateral asymmetry with a substantial proportion of individuals having an asymmetric index larger than 5% of total wing size. Such an anomaly, however, cannot be plainly explained by an increase of developmental instability in hybrids but is the result of some aberrant developmental processes. Our findings suggest that interspecific hybrids are as able as their parents to buffer developmental noise, notwithstanding the fact that their proper bilateral development can be harshly compromised. Together with the low correspondence between the co-variation structures of the interindividual genetic components and the within-individual ones from a Procrustes analysis, our data also suggest that the underlying processes that control (genetic) canalization and developmental stability do not share a common mechanism. We argue that the conventional account of decreased developmental stability in interspecific hybrids needs to be reappraised.     

Multiple interactions of FbsA, a surface protein from Streptococcus agalactiae, with fibrinogen: affinity, stoichiometry, and structural characterization

Streptococcus agalactiae is an etiological agent of several infective diseases in humans. We previously demonstrated that FbsA, a fibrinogen-binding protein expressed by this bacterium, elicits a fibrinogen-dependent aggregation of platelets. In the present communication, we show that the binding of FbsA to fibrinogen is specific and saturable, and that the FbsA-binding site resides in the D region of fibrinogen. In accordance with the repetitive nature of the protein, we found that FbsA contains multiple binding sites for fibrinogen. By using several biophysical methods, we provide evidence that the addition of FbsA induces extensive fibrinogen aggregation and has noticeable effects on thrombin-catalyzed fibrin clot formation. Fibrinogen aggregation was also found to depend on FbsA concentration and on the number of FbsA repeat units. Scanning electron microscopy evidentiated that, while fibrin clot is made of a fine fibrillar network, FbsA-induced Fbg aggregates consist of thicker fibers organized in a cage-like structure. The structural difference of the two structures was further indicated by the diverse immunological reactivity and capability to bind tissue-type plasminogen activator or plasminogen. The mechanisms of FbsA-induced fibrinogen aggregation and fibrin polymerization followed distinct pathways since Fbg assembly was not inhibited by GPRP, a specific inhibitor of fibrin polymerization. This finding was supported by the different sensitivity of the aggregates to the disruptive effects of urea and guanidine hydrochloride. We suggest that FbsA and fibrinogen play complementary roles in contributing to thrombogenesis associated with S. agalactiae infection.