Properties of erythrocyte light refraction in diabetic patients

Since hyperglycaemia changes the erythrocyte cell membrane fluidity and impairs cell deformity, our goal was to characterize hemoglobin and red blood cell (RBC) light refractive property changes in diabetic patients. Microscopic investigation was carried out on intact and fixed RBCs. To determine the refractive index (RI): smears of peripheral blood were air dried and fixed for 3 min in methanol. Mixtures of polyvinylpyrolidine and buffer of different pH (1:1) were used as embedding media. Intact RBCs were mixed with a buffered embedding medium, placed on a slide and overlaid with a coverslip. Interference microscopy was used for RI measurements at 18 different pH (pH=2-13). The results showed that curves of the RI of diabetic patients and of a control group were of similar configuration, with one branch in the acidic portion of the pH scale, a maximum and two minima in the neutral (middle) portion, and one branch in the alkaline portion. The curves of the individuals from the control group overlapped each other. To the contrary, the curves of the diabetic patients were not uniform in the neutral portion and the alkaline portion. The curves of the diabetic patients in the neutral zone were shifted towards the alkaline end of the pH scale, and the RBC RI curves were lower in comparison to the control curves. The center maximum of the curves of diabetic patients corresponded to pH=6.6 whereas the central maximum of the control group curves was at pH=6.2-6.8. Contrary to in the diabetic group, intact RBC RI curves in the control group revealed only one significantly different minimum at pH of 7.2 in the neutral zone. Using this method it is possible to show phenotypic differences between uniform type intact and fixed cells, erythrocytes of diabetic patients and of healthy donors.     

The search for molecular determinants of LPS inhibition by proteins and peptides

Lipo-poly-saccharide (LPS) induced Gram-negative sepsis and septic shock remain lethal in up to 60 % of cases, and LPS antagonists that neutralize its endotoxic action are the subject of intensive research. The molecular motifs of specific binding of LPS by antiendotoxin proteins and peptides may lead to an understanding of LPS action at the atomic level and provide clues for the development of new immunomodulatory compounds for use as therapy in the treatment of Gram-negative bacterial sepsis. The interaction of LPS with its cognate binding proteins has been structurally elucidated in the single case of the X-ray crystallographic structure of LPS in complex with the integral outer membrane protein FhuA from E. coli K-12 (Ferguson et al., Science 1999, 282, 2215). This structure and other known structures of LPS binding proteins have been used to propose a common binding motif of LPS to proteins. Another independent source of structural information are solution structures of peptides in complex with LPS that can be determined using the transferred NOE effect. The molecular mechanisms of biological activity of bacterial endotoxins can additionally be probed by theoretical means. The growing structural knowledge is opening pathways to the design of peptides or peptidomimetics with improved antiendotoxin properties.     

High Dose Therapy With Alpha-Lipoic Acid Improves Cutaneous Vasomotor Response In Patients With Diabetic Neuropathy

The saposin C-derived peptide TX14(A) prevents onset of functional and structural disorders in the peripheral nerve of diabetic rats. We have now investigated the ability of TX14(A) to alleviate behavioral indices of abnormal pain perception in adult female rats 4-6 weeks after onset of STZ-induced diabetes. Untreated diabetic rats exhibited tactile allodynia (response threshold = 3 ± 1 g) compared to age-matched controls (10 ± 1g). A single ip injection of TX14(A) transiently alleviated tactile allodynia, with an effect that was maximal 6 hours (11 ± 1g) after injection and diminished within 48 hours. Maximal efficacy was seen with a 1 mg/kg dose while no effects were noted in control rats. Control rats exhibited a transient thermal hyperalgesia (77 ± 5% of baseline paw withdrawal latency) 15 minutes after intrathecal delivery of substance P (30 nmol) that resolved within 30 minutes. Untreated diabetic rats exhibited substance P evoked thermal hyperalgesia of similar magnitude (82 ± 5% at 15 minutes) but of greater duration (83 ± 4% at 1 hour). Intrathecal delivery of TX14(A) 30 minutes before intrathecal substance P was without effect on the transient thermal hyperalgesia in control rats (74 ± 9% at 15 minutes). In diabetic rats, the prolonged thermal hyperalgesia was abolished by prior intrathecal delivery of TX14(A), although the transient thermal hyperalgesia (72 ± 8% at 15 minutes) remained. These studies show that TX14(A) can rapidly allevate diabetes-induced allodynia and hyperalgesia for up to 48 hours.     

Enhancement of antibacterial and lipopolysaccharide binding activities of a human lactoferrin peptide fragment by the addition of acyl chain

Cationic antibacterial peptides are potentially therapeutic in the treatment of sepsis, because of their amalgamated antibacterial and lipopolysaccharide-binding activities. We prepared acyl analogues of the peptide fragment of human lactoferrin, which originally had weak antibacterial activity. It was found that 12 carbon units constitute the optimal acyl chain length, enhancing the antibacterial activity and binding of lipopolysaccharide by up to two orders of magnitude. Lactoferrin-based lipopeptides approached the activity of polymyxin B, a lipopeptide of natural origin, but were also active against Gram-positive bacteria.     

Socio-economic factors in the differential upsurge of tick-borne encephalitis in Central and Eastern Europe

Tick-borne encephalitis (TBE), the most serious widespread vector-borne disease of humans in Europe, increased from 2- to 30-fold in many Central and Eastern European countries from 1992 to 1993, coinciding with independence from Soviet rule. Unemployment and low income have been shown in Latvia to be statistically associated with high-risk behaviour involving harvest of wild foods from tick-infested forests, and also with not being vaccinated against TBE. Archival data for 1970--2005 record major changes in the agricultural and industrial sectors, and consequent changes in the abiotic and biotic environment and socio-economic conditions, which could have increased the abundance of infected ticks and the contact of humans with those ticks. For example, abandoned agricultural fields became suitable for rodent transmission hosts; use of pesticides and emissions of atmospheric industrial pollutants plummeted; wildlife hosts for ticks increased; tick populations appear to have responded; unemployment and inequality increased in all countries. These factors, by acting synergistically but differentially between and within each country, can explain the marked spatio-temporal heterogeneities in TBE epidemiology better than can climate change alone, which is too uniform across wide areas. Different degrees of socio-economic upheaval caused by political transition in Estonia, Latvia, Lithuania, Slovenia and the Czech Republic can apparently explain the marked variation in TBE upsurge. Causal linkage between national socio-economic conditions and epidemiology is strongly indicated by striking correlations across eight countries between the degree of upsurge of TBE and both poverty and household expenditure on food (R2 = 0.533 and 0.716, respectively).     

Molecular paleontology of transposable elements in the Drosophila melanogaster genome

We report here a superfamily of "cut and paste" DNA transposons called Transib. These transposons populate the Drosophila melanogaster and Anopheles gambiae genomes, use a transposase that is not similar to any known proteins, and are characterized by 5-bp target site duplications. We found that the fly genome, which was thought to be colonized by the P element <100 years ago, harbors approximately 5 million year (Myr)-old fossils of ProtoP, an ancient ancestor of the P element. We also show that Hoppel, a previously reported transposable element (TE), is a nonautonomous derivate of ProtoP. We found that the "rolling-circle" Helitron transposons identified previously in plants and worms populate also insect genomes. Our results indicate that Helitrons were horizontally transferred into the fly or/and mosquito genomes. We have also identified a most abundant TE in the fly genome, DNAREP1_DM, which is an approximately 10-Myr-old footprint of a Penelope-like retrotransposon. We estimated that TEs are three times more abundant than reported previously, making up approximately 22% of the whole genome. The chromosomal and age distributions of TEs in D. melanogaster are very similar to those in Arabidopsis thaliana. Both genomes contain only relatively young TEs (<20 Myr old), constituting a main component of paracentromeric regions.