Natural anthraquinones probed as Type I and Type II photosensitizers: singlet oxygen and superoxide anion production.

The photosensitizing properties of six anthraquinones (AQs): soranjidiol (1), soranjidiol-1-methyl ether (2), rubiadin (3), rubiadin-1-methyl ether (4), damnacanthal (5) and damnacanthol (6), isolated from leaves and stems of Heterophyllaea pustulata Hook. f. (Rubiaceae) were studied. By means of photobiological and photophysical methods in vitro, the type of photosensitization that these metabolites are capable of producing was determined. Whereas the photosensitized generation of superoxide anion radical (O(2)(-)) (Type I) was evaluated in leukocyte suspensions, singlet molecular oxygen ((1)O(2)) production (Type II) was examined in organic solution. In addition, the quantum yield of (1)O(2) (Phi) in chloroform was measured for those AQs that generate it. It was established that 4 behaves exclusively as a Type I photosensitizer. By contrast, the others AQs act by both types of mechanisms, among which 5 showed the largest Phi of (1)O(2).

Functional expression of H4 histamine receptor in human natural killer cells, monocytes, and dendritic cells.

We describe here the protein expression of H4 histamine receptor in cells of the innate immune system, which include NK cells, monocytes, and dendritic cells (DCs). Anti-H4R specifically stained permeabilized NK cells, THP-1 clone 15 monocytes, and DCs. This binding was inhibited by incubating anti-H4R Ab with its corresponding peptide. Histamine induced NK cells, THP-1 clone 15 cells, and DCs chemotaxis with high affinity. The ED(50) chemotactic effect was 5 nM, 6.8 nM, and 2.7 nM for NK cells, THP-1 clone 15 cells, and DCs, respectively. Thioperamide, an H3R/H4R antagonist, inhibited histamine-induced chemotaxis in all these cells. However, histamine failed to induce the mobilization of [Ca(2+)](i) in NK cells and THP-1 clone 15 cells, but it induced calcium fluxes in DCs. Using a new method of detecting NK cell-mediated cytolysis, it was observed that NK cells efficiently lysed K562 target cells and that histamine did not affect this NK cell activity. In summary, this is the first demonstration of the protein expression of H4 receptor in NK cells. Also, the results of the chemotactic effects of histamine on NK cells and THP-1 cells are novel. These results may shed some light on the colocalization of cells of innate immune arm at sites of inflammation. They are also important for developing drugs that target H4R for the treatment of various disorders, such as autoimmune and immunodeficient diseases.

Emergence of phenotypic variants upon mismatch repair disruption in Pseudomonas aeruginosa.

MutS is part of the bacterial mismatch repair system that corrects point mutations and small insertions/deletions that fail to be proof-read by DNA polymerase activity. In this work it is shown that the disruption of the P. aeruginosa mutS gene generates the emergence of diverse colony morphologies in contrast with its parental wild-type strain that displayed monomorphic colonies. Interestingly, two of the mutS morphotypes emerged at a high frequency and in a reproducible way and were selected for subsequent characterization. One of them displayed a nearly wild-type morphology while the other notably showed, compared with the wild-type strain, increased production of pyocyanin and pyoverdin, lower excretion of LasB protease and novel motility characteristics, mainly related to swarming. Furthermore, it was reproducibly observed that, after prolonged incubation in liquid culture, the pigmented variant consistently emerged from the mutS wild-type-like variant displaying a reproducible event. It is also shown that these P. aeruginosa mutS morphotypes not only displayed an increase in the frequency of antibiotic-resistant mutants, as described for clinical P. aeruginosa mutator isolates, but also generated mutants whose antibiotic-resistant levels were higher than those measured from spontaneous resistant mutants derived from wild-type cells. It was also found that both morphotypes showed a decreased cytotoxic capacity compared to the wild-type strain, leading to the emergence of invasive variants. By using mutated versions of a tetracycline resistance gene, the mutS mutant showed a 70-fold increase in the reversion frequency of a +1 frameshift mutation with respect to its parental wild-type strain, allowing the suggestion that the phenotypical diversity generated in the mutS population could be produced in part by frameshift mutations. Finally, since morphotypical diversification has also been described in clinical isolates, the possibility that this mutS diversification was related to the high frequency hypermutability observed in P. aeruginosa CF isolates is discussed.