Synthesis of (20S)-[7,7,21,21-2H4]-3beta-(tert-butyldimethylsilanyloxy)-20-methyl-pregn-5-en-21-ol,a useful intermediate for the preparation of deuterated isotopomers of sterols

(20S)-[7,7,21,21-2H(4)]-3beta-(tert-Butyldimethylsilanyloxy)-20-methyl-pregn-5-en-21-ol, an intermediate for the preparation of deuterated isotopomers of sterols to be used as standards for biomedical studies, was prepared by reduction with dichloroaluminum deuteride of ethyl (20S)-3beta-(tert-butyldimethylsilanyloxy)-7-oxo-pregn-5-en-20-carboxylate. Using controlled experimental conditions, it has also been shown that the dichloroaluminum hydride reduction of a 7-keto steroid affords the corresponding 7beta-hydroxy derivative in a highly stereoselective manner.     

Inhibition of CD73 improves B cell-mediated anti-tumor immunity in a mouse model of melanoma

CD73 is a cell surface enzyme that suppresses T cell-mediated immune responses by producing extracellular adenosine. Growing evidence suggests that targeting CD73 in cancer may be useful for an effective therapeutic outcome. In this study, we demonstrate that administration of a specific CD73 inhibitor, adenosine 5'-(α,β-methylene)diphosphate (APCP), to melanoma-bearing mice induced a significant tumor regression by promoting the release of Th1- and Th17-associated cytokines in the tumor microenvironment. CD8(+) T cells were increased in melanoma tissue of APCP-treated mice. Accordingly, in nude mice APCP failed to reduce tumor growth. Importantly, we observed that after APCP administration, the presence of B cells in the melanoma tissue was greater than that observed in control mice. This was associated with production of IgG2b within the melanoma. Depletion of CD20(+) B cells partially blocked the anti-tumor effect of APCP and significantly reduced the production of IgG2b induced by APCP, implying a critical role for B cells in the anti-tumor activity of APCP. Our results also suggest that APCP could influence B cell activity to produce IgG through IL-17A, which significantly increased in the tumor tissue of APCP-treated mice. In support of this, we found that in melanoma-bearing mice receiving anti-IL-17A mAb, the anti-tumor effect of APCP was ablated. This correlated with a reduced capacity of APCP-treated mice to mount an effective immune response against melanoma, as neutralization of this cytokine significantly affected both the CD8(+) T cell- and B cell-mediated responses. In conclusion, we demonstrate that both T cells and B cells play a pivotal role in the APCP-induced anti-tumor immune response.     

Search for effective natural enemies of <Emphasis Type="Italic">Tetranychus evansi</Emphasis> (Acari: Tetranychidae) in northwest Argentina

The mite Tetranychus evansi Baker & Pritchard (Tetranychidae) probably originated in South America. Because of its importance as a tomato pest in Africa, an extensive project has been conducted to detect potentially effective natural enemies in South America for the classical biological control of the pest in Africa. A search for the natural enemies of T. evansi was conducted in the Province of Tucumán, northwestern Argentina, in December 2004, and this report describes the results. One hundred predatory mites of the family Phytoseiidae referring to 11 species were collected on 11 examined species of solanaceous plants. The most abundant phytoseiid species collected were Neoseiulus californicus (McGregor) and Euseius concordis (Chant). Adults and immatures of those species, as well as of Neoseiulus idaeus Denmark & Muma, Phytoseiulus fragariae Denmark & Schicha and Proprioseiopsis cannaensis (Muma) were found in association with T. evansi, suggesting that they were developing on the pest. However, because of the possible biological differences between populations of a given species, biological studies evaluating T. evansi as a prey for those predators seem desirable.     

Fecundity of the Hypostomus affinis (Siluriformes,Loricariidae) in the Lajes Reservoir,Rio de Janeiro,Brazil

The armored catfish Hypostomus affinis is a widespread Loricariidae fish in the Lajes Reservoir (22 degrees 42'-22 degrees 50' S; 43 degrees 53'-44 degrees 05' W), the largest lentic environment in the Rio de Janeiro State, but little is known about their reproductive biology. One hundred and twenty five females, captured from January 1996 to December 1997, were used to assess ovarian development (29 were used to determine fecundity). The oocyte distribution by size-diameter classes revealed the occurrence of two modes suggesting an asynchronic development of the ovaries. The total fecundity ranged from 1235 to 4304, averaging 2374. An exponential relationship between fecundity and total length was determined, and a direct linear relationship was found between fecundity and total weight and gonad weight.     

Antibiotic resistance in strains of Listeria spp. from meat products

The susceptibility or resistance to 26 antimicrobial agents was determined for 64 strains of Listeria monocytogenes and 102 strains of L. innocua isolated from Italian meat products. Some strains of L. monocytogenes were found to be resistant to tetracycline, erythromycin, co-trimoxazole and clindamycin. No plasmids were found in any L. monocytogenes strain. Five strains of L. innocua contained a 7.9 kbp plasmid, but these isolates were not resistant to any antibiotic in common and treatment with curing agents could not eliminate resistance to antibiotics. These results suggest that antibiotic resistance was not likely to be plasmid mediated in our strains.     

Vasorelaxant effect of the flavonoid galangin on isolated rat thoracic aorta

Here we investigated the effect of the flavonoid galangin in isolated rat thoracic aortic rings. Galangin (0.1-100 microM) induced relaxation in rings pre-contracted with phenylephrine (PE 1 microM) or with KCl (100 mM) or pre-treated with the nitric oxide synthase inhibitor Nomega-nitro-L-arginine methyl ester (L-NAME, 100 microM), the cyclooxygenase inhibitor indomethacin (10 microM) and the adenylate cyclase inhibitor, SQ 22,536 (100 microM). In another set of experiments, rat aortic rings were incubated with galangin (1-100 microM) and the contractile responses to PE (0.001-3 microM) or to KCl (60 mM) were evaluated. We also evaluated the effect of galangin (100 microM) on PE (10 microM)-induced contraction in a Ca2+-free medium. Galangin relaxed aortic rings with or without endothelium. Galangin effect was significantly inhibited by L-NAME. Galangin inhibited the contractile response to PE, either in presence or in absence of external calcium, and to KCl. In the end, we also found that galangin caused nitric oxide (NO) release from aortic rings and abolished the increase in [Ca2+]i triggered by PE or KCl in aortic smooth muscle cells, either in presence and in absence of external Ca2+. Our results suggest that galangin reduces the contractility of rat aortic rings through an endothelium-dependent mechanism, involving NO, and also through an endothelium-independent mechanism, inhibiting calcium movements through cell membranes.     

Interaction pattern of Arg 62 in the A-pocket of differentially disease-associated HLA-B27 subtypes suggests distinct TCR binding modes

The single amino acid replacement Asp116His distinguishes the two subtypes HLA-B*2705 and HLA-B*2709 which are, respectively, associated and non-associated with Ankylosing Spondylitis, an autoimmune chronic inflammatory disease. The reason for this differential association is so far poorly understood and might be related to subtype-specific HLA:peptide conformations as well as to subtype/peptide-dependent dynamical properties on the nanoscale. Here, we combine functional experiments with extensive molecular dynamics simulations to investigate the molecular dynamics and function of the conserved Arg62 of the α1-helix for both B27 subtypes in complex with the self-peptides pVIPR (RRKWRRWHL) and TIS (RRLPIFSRL), and the viral peptides pLMP2 (RRRWRRLTV) and NPflu (SRYWAIRTR). Simulations of HLA:peptide systems suggest that peptide-stabilizing interactions of the Arg62 residue observed in crystal structures are metastable for both B27 subtypes under physiological conditions, rendering this arginine solvent-exposed and, probably, a key residue for TCR interaction more than peptide-binding. This view is supported by functional experiments with conservative (R62K) and non-conservative (R62A) B*2705 and B*2709 mutants that showed an overall reduction in their capability to present peptides to CD8+ T cells. Moreover, major subtype-dependent differences in the peptide recognition suggest distinct TCR binding modes for the B*2705 versus the B*2709 subtype.