The CCR5Delta32 allele is associated with reduced liver inflammation in hepatitis C virus infection.

CCR5Delta32 is a deletion mutation in the chemokine receptor CCR5. Liver inflammatory activity was found to be significantly reduced (P = 0.005) in Jewish Israeli patients infected with the hepatitis C virus (HCV) carrying the CCR5Delta32 allele. The CCR5Delta32 allele does not alter susceptibility to HCV infection; however, it may play a role in the progression and outcome of the disease.     

Peptidomimetic thrombin inhibitors

The central position of thrombin in the coagulation cascade has made it a popular target for discovery of novel antithrombotic agents. Starting with hirudin, a natural peptide isolated from the medicinal leech, its shorter synthetic analogue hirulog, and argatroban,the first therapeutically used synthetic small-molecule thrombin active site inhibitor, hundreds of direct thrombin inhibitors have been discovered over the last 20 years. Most of them are peptidomimetic compounds,based on the amino acid sequence of fibrinogen which binds into the thrombin active site. Since elucidation of the crystal structure of human thrombin in 1989, the structure-based design of low-molecular-weight peptidomimetic thrombin inhibitors has been greatly aided by the use of x-ray crystallographic analysis of thrombin-inhibitor complexes. The ultimate goal of most research programmes and drug optimization strategies is to develop an orally bioavailable, small-molecule,direct thrombin inhibitor that would be suitable for once or twice daily dosing. An overview of the most advanced peptidomimetic direct thrombin inhibitors bivalirudin, argatroban, ximelagatran and dabigatran is presented.     

The CCR5Δ32 allele is associated with reduced liver inflammation in hepatitis C virus infection

CCR5Δ32 is a deletion mutation in the chemokine receptor CCR5. Liver inflammatory activity was found to be significantly reduced (P = 0.005) in Jewish Israeli patients infected with the hepatitis C virus (HCV) carrying the CCR5Δ32 allele. The CCR5Δ32 allele does not alter susceptibility to HCV infection; however, it may play a role in the progression and outcome of the disease.     

Human monoclonal antibodies specific to hepatitis B virus generated in a human/mouse radiation chimera: the Trimera system.

An approach to develop fully human monoclonal antibodies in a human/mouse radiation chimera, the Trimera system, is described. In this system, functional human lymphocytes are engrafted in normal strains of mice which are rendered immuno-incompetent by lethal total body irradiation followed by radioprotection with severe combined immunodeficient (SCID) mouse bone marrow. Following transplantation, human lymphocytes colonize murine lymphatic organs and secrete human immunoglobulins. We have established this system as a tool to develop fully human monoclonal antibodies, and applied it for the generation of monoclonal antibodies specific for hepatitis B virus surface antigen. A strong memory response to hepatitis B surface antigen was elicited in Trimera engrafted with lymphocytes from human donors positive for antibodies to hepatitis B surface antigen. The human specific antibody fraction in the Trimera was 10(2)-10(3)-fold higher as compared with that found in the donors. Spleens were harvested from Trimera mice showing high specific-antibody titres and cells were fused to a human-mouse heteromyeloma fusion partner. Several stable hybridoma clones were isolated and characterized. These hybridomas produce high-affinity, IgG, anti-hepatitis B surface antigen antibodies demonstrating the potential of the Trimera system for generating fully human monoclonal antibodies. The biological function and the neutralizing activity of these antibodies are currently being tested.     

Systemic activity of ivermectin on the human body louse (Anoplura: Pediculidae)

Eighty-one to 100% of nymphs and females of the human body louse (Pediculus humanus humanus) that fed artificially on blood containing 2.5-10 ng ivermectin/ml died. The mortality of nymphs and female lice fed on rabbits treated with 200 micrograms/kg ivermectin was very high during the first two to three days, then declined sharply, reaching the level of the controls on day six. Nymphs were more sensitive than females. The average number of eggs laid by surviving females and the percentage that hatched from those eggs were lower than in controls.     

Efficient induction and selection of chloroplast-encoded antibiotic-resistant mutants in Nicotiana

A high rate of plastome-encoded mutations was induced in Nicotiana by exposing seeds to N-nitroso-N-methylurea. Seeds then were subjected to nutritional and in vitro selection procedures for systematic isolation of plastome-dependent antibiotic-resistant plants. Multiple flowering lines resistant to streptomycin, spectinomycin, lincomycin, or chloramphenicol were obtained. A detailed analysis of the streptomycin-resistant lines is presented. Sexual hybridization, cybrid formation following protoplast fusion, and in organello protein synthesis were used to rigorously assign the mutations to the chloroplast genome. The efficient rates of mutagenesis combined with the in vitro mass-screening procedures described here should facilitate investigation of fundamental aspects of chloroplast genetics in higher plants.     

Biotransformation by plant cells immobilized in cross-linked polyacrylamide-hydrazide

Plant cells were entrapped by mixing suspended MENTHA cells with linear, water soluble polyacrylamide-hydrazide chains followed by the stoichiometric addition of glyoxal as the cross linking agent (PAAH-G entrapment). In parallel, some cells were entrapped in calcium-alginate beads, as previously described. The capability of both immobilized cell systems to reduce monoterpenes was compared with freely suspended MENTHA cells. Entrapment by either alginate or PAAH-G did not impair cell vitality, as observed by fluorescein diacetate staining. Biotransformation of (-) menthone to (+) neomenthol by M-cells and of (+) pulegone to (+) isomenthone by P-cells indicated that the transformation efficiency of the cells entrapped in PAAH-G is as high as that of freely suspended cells. Moreover, the distribution of both precursor and product in the medium versus their content in the cells (or cells contained in gel-beads) showed that less monoterpenes were retained in cells entrapped in PAAH-G, as compared to the freely suspended cells. Thus prolonged incubation (e.g. 24 hr), which usually results in appreciable loss of monoterpenes from the chloroform extract of freely-suspended-cells, caused considerably less loss from the PAAH-G entrapped-cells. In a preliminary test it was shown that PAAH-G entrapped cells were capable to perform three, consecutive, batch-type monoterpene biotransformations, without significant decrease of transformation capability. The capability to immobilize living plant cells within this synthetic chemically crosslinked gel system, combined with the favourable beads/ free-medium ratio of monoterpene distribution, point towards a potential development of a continuous biotransformation process carried out by plant-cells entrapped in this system.     

Biotransformation of monoterpenes by Mentha cell lines

Previous results indicated that all tested cell suspension lines derived from various MENTHA chemotypes were capable to biotransform (-) menthone to (+)-neomentol but only some of these lines converted (+)-pulegone to (+)-isomenthone. In order to quest whether only the natural secondary metabolites or also other compounds with similarities to pulegone are biohydrogenated by MENTHA cell suspensions, we incubated such suspensions with 5 unsaturated alpha-beta; ketones. No conversion was detected when mesityl oxide, trans-6-tert. butyl pulegone or 3-isopropylidine-9-methyl-decalone-2 were incubated with MENTHA cells, while saturation of the alpha-beta double bond of 2-isopropylidine cyclohexanone and of trans-6-methyl pulegone was observed in suspensions of those cell lines which are capable of pulegone transformation. Suspensions of MENTHA cell lines which were incapable to hydrogenate pulegone did not biotransform the two latter pulegone analogues.     

Functional magnetic resonance imaging monitoring of pathological changes in rodent livers during hyperoxia and hypercapnia

Liver diseases and regeneration are associated with hemodynamic changes denoting pathological alterations. Determining and monitoring physiological and pathological liver changes is essential for diagnostic and therapeutic objectives. Our aim was to determine the feasibility of functional magnetic resonance imaging (fMRI) during hypercapnia and hyperoxia for monitoring liver pathology. Liver fMRI images were acquired in rodents following acute bleeding, partial hepatectomy, and fibrosis. Results were quantitated and confirmed by histology. Changes induced by hyperoxia and hypercapnia following hemorrhage significantly correlated with the percentage of blood loss, reflecting lower liver perfusion and diminished vessel responsiveness to gas saturation. Hepatectomy resulted in an early decline in signal intensity changes due to hyperoxia, suggesting a decrease in liver perfusion and blood content. Following hepatectomy, signal intensity changes due to hypercapnia increased, signifying a change in liver perfusion from a mainly portal to a more arterial source. Two weeks after induction of fibrosis, signal intensity changes due to hypercapnia became much lower and those due to hyperoxia were much higher than those in normal livers, reflecting the increased perfusion due to the inflammatory process as confirmed by histologic analysis. With fibrosis progression, signal intensity changes induced by hypercapnia and hyperoxia were gradually attenuated, indicating structural and functional alterations of the liver vasculature during fibrosis. CONCLUSION: In various liver pathologies, fMRI response to hypercapnia and hyperoxia is sensitive to changes in liver hemodynamic status involved in hepatic damage or recovery; thus, this technique may offer an additional noninvasive diagnostic tool for evaluation and follow-up of liver diseases by means of examining perfusion-related alterations.