HLA Class I and Class II Polymorphism in the Population of Rijeka, Croatia

The aim of the study was to examine frequencies of HLA-A, -B, -DR antigens and haplotypes in population of Rijeka and to compare them with general Croatian and European populations. The subjects were 117 unrelated healthy blood donors. The antigens with the highest frequencies were: A2 (27.2%), A9 (16.3%), B5 (14.8%), B12 (11.8%), B18 (11.8%), DR5 (21.6%) and DR6 (13.8%). Comparison of HLA antigens frequencies has shown statistically significant difference in 1 antigen with Croatian population and in 8 antigens with European population. The HLA haplotypes with high frequencies included HLA-A2, B5 (6.84%), HLA-A2, B12 (6.84%), HLA-A2, B18 (6.84%), HLA-B12, DR2 (9.78%) and HLA-B18, DR5 (6.84%). The antigen B5 showed strongest association with DR5 (6.41%; LD = 1.30) as in general Croatian and in some European populations. The results have shown great diversity of HLA haplotypes in Rijeka population which can be the result of admixture with neighborhood immigrating populations during the history

Chemical synthesis and biological activities of 3-alkyl pyridinium polymeric analogues of marine toxins

Two new large poly-1,3-dodecylpyridinium salts, APS12 and APS12-2 of 12.5- and 14.7-kDa size, respectively, were synthesised and tested for their pore-forming and transfection capabilities in HEK 293 and undifferentiated mouse ES cells using patch-clamp recording, Ca2+ imaging and flow cytometry. Polymerisation reactions were enhanced by microwaves, and the product sizes were controlled by altering the irradiation time. This method can also be applied to obtain polymers with variable linking chains as shown by the preparation of poly-(1,3-octylpyridinium) salt of 11.9-kDa size. Molecular weights of the final products were determined using ESIMS analysis, which also indicated the products to be amongst the largest macro-cycles ever recorded, up to a 900-membered ring. Anti-bacterial, haemolytic and anti-acetylcholinesterase activities were also reported for the two dodecyl pyridinium polymers. These biological activities are characteristic to the structurally related marine toxin, poly-APS

EPR and FTIR studies reveal the importance of highly ordered sterol-enriched membrane domains for ostreolysin activity.

Ostreolysin is a cytolytic protein from the edible oyster mushroom (Pleurotus ostreatus), which recognizes specifically and binds to raft-like sterol-enriched membrane domains that exist in the liquid-ordered phase. Its binding can be abolished by micromolar concentrations of lysophospholipids and fatty acids. The membrane activity of ostreolysin, however, does not completely correlate with the ability of a certain sterol to induce the formation of a liquid-ordered phase, suggesting that the protein requires an additional structural organization of the membrane to exert its activity. The aim of this study was to further characterize the lipid membranes that facilitate ostreolysin binding by analyzing their lipid phase domain structure. Fourier-transformed infrared spectroscopy (FTIR) and electron paramagnetic resonance (EPR) were used to analyze the ordering and dynamics of membrane lipids and the membrane domain structure of a series of unilamellar liposomes prepared by systematically changing the lipid components and their ratios. Our results corroborate the earlier conclusion that the average membrane fluidity of ostreolysin-susceptible liposomes alone cannot account for the membrane activity of the protein. Combined with previous data computer-aided interpretation of EPR spectra strongly suggests that chemical properties of membrane constituents, their specific distribution, and physical characteristics of membrane nanodomains, resulting from the presence of sterol and sphingomyelin (or a highly ordered phospholipid, dipalmitoylphosphatidylcholine), are essential prerequisites for ostreolysin membrane binding and pore-formation

Synthetic analogs of stryphnusin isolated from the marine sponge : Stryphnus fortis inhibit acetylcholinesterase with no effect on muscle function or neuromuscular transmission

The marine secondary metabolite stryphnusin (1) was isolated from the boreal sponge Stryphnus fortis, collected off the Norwegian coast. Given its resemblance to other natural acetylcholinesterase antagonists, it was evaluated against electric eel acetylcholinesterase and displayed inhibitory activity. A library of twelve synthetic phenethylamine analogs, 2a-7a and 2b-7b, containing tertiary and quaternary amines respectively were synthesized to investigate the individual structural contributions to the activity. Compound 7b was the strongest competitive inhibitor of both acetylcholinesterase and butyrylcholinesterase with IC50 values of 57 and 20 μM, respectively. This inhibitory activity is one order of magnitude higher than the positive control physostigmine, and is comparable with several other marine acetylcholinesterase inhibitors. The physiological effect of compound 7b on muscle function and neuromuscular transmission was studied and revealed a selective mode of action at the investigated concentration. This data is of importance as the interference of therapeutic acetylcholinesterase inhibitors with neuromuscular transmission can be problematic and lead to unwanted side effects. The current findings also provide additional insights into the structure-activity relationship of both natural and synthetic acetylcholinesterase inhibitors

Isolation and Synthesis of Pulmonarins A and B, Acetylcholinesterase Inhibitors from the Colonial Ascidian Synoicum pulmonaria

Pulmonarins A and B are two new dibrominated marine acetylcholinesterase inhibitors that were isolated and characterized from the sub-Arctic ascidian Synoicum pulmonaria collected off the Norwegian coast. The structures of natural pulmonarins A and B were tentatively elucidated by spectroscopic methods and later verified by comparison with synthetically prepared material. Both pulmonarins A and B displayed reversible, noncompetitive acetylcholinesterase inhibition comparable to several known natural acetylcholinesterase inhibitiors. Pulmonarin B was the strongest inhibitor, with an inhibition constant (Ki) of 20 μM. In addition to reversible, noncompetitive acetylcholinesterase inhibition, the compounds displayed weak antibacterial activity but no cytotoxicity or other investigated bioactivities