T cell independent Thy-1 allo-antibody response with the use of transgenic mice

We have introduced a mouse Thy-1.1 gene into the germline of Thy-1.2 mice. The introduced gene was shown to be expressed at very high levels in thymocytes when compared with the endogenous gene. Transgenic thymocytes were shown to evoke a higher than normal primary anti-Thy-1.1 antibody response in plaque-forming cell (PFC) assays. This result suggests that a direct quantitative interaction of the Thy-1 antigen activates the B cell response.     

The novel flightless-I gene brings together two gene families, actin- binding proteins related to gelsolin and leucine-rich-repeat proteins involved in Ras signal transduction

The Drosophila melanogaster gene flightless-I, involved in gastrulation and muscle degeneration, has Caenorhabditis elegans and human homologues. In these highly conserved genes, two previously known gene families have been brought together, families encoding the actin- binding proteins related to gelsolin and the leucine-rich-repeat (LRR) group of proteins involved in protein-protein interactions. Both these gene families exhibit characteristics of molecular changes involving replication slippage and exon shuffling. Phylogenetic analyses of 19 amino acid sequences of 6 related protein types indicate that actin- associated proteins related to gelsolin are monophyletic to a common ancestor and include flightless proteins. Conversely, comparison of 24 amino acid sequences of LRR proteins including the flightless proteins indicates that flightless proteins are members of a structurally related subgroup. Included in the flightless cluster are human and mouse rsp-1 proteins involved in suppressing v-Ras transformation of cells and the membrane-associated yeast (Saccharomyces cerevisae) adenylate cyclase whose analogous LRRs are required for interaction with Ras proteins. There is a strong possibility that ligands for this group could be related and that flightless may have a similar role in Ras signal transduction. It is hypothesized that an ancestral monomeric gelsolin precursor protein has undergone at least four independent gene reorganization events to account for the structural diversity of the extant family of gelsolin-related proteins and that gene duplication and exon shuffling events occurred prior to or at the beginning of multicellular life, resulting in the evolution of some members of the family soon after the appearance of actin-type proteins.      

Magnesium-induced inner membrane aggregation in heart mitochondria: prevention and reversal by carboxyatractyloside and bongkrekic acid

Mg(2+) at an optimal concentration of 2mM (ph 6.5) induces large increases (up to 30 percent) in the optical density of bovine heart mitochondria incubated under conditions of low ionic strength (< approx. 0.01). The increases are associated with aggregation (sticking together) of the inner membranes and are little affected by changes in the energy status of the mitochondria. Virtually all of a number of other polyvalent cations tested and Ag(+) induce increases in mitochondrial optical density similar to those induced by Mg(2+), their approximate order of concentration effectiveness in respect to Mg(2+) being: La(3+) > Pb(2+) = Cu(2+) > Cd(2+) > Zn(2+) > Ag(+) > Mn(2+) > Ca(2+) > Mg(2+). With the exception of Mg(2+), all of these cations appear to induce swelling of the mitochondria concomitant with inner membrane aggregation. The inhibitors of the adenine nucleotide transport reaction carboxyatratyloside and bongkrekic acid are capable of preventing and reversing Mg(2+)-induced aggregation at the same low concentration required for complete inhibition of phosphorylating respiration, suggesting that they inhibit the aggregation by binding to the adenine nucleotide carrier. The findings are interpreted to indicate (a) that the inner mitochondrial membrane is normally prevented from aggregating by virtue of its net negative outer surface change, (b) that the cations induce the membrane to aggregate by binding at its outer surface, decreasing the net negative charge, and (c) that carboxyatractyloside and bongkrekic acid inhibit the aggregation by binding to the outer surface of the membrane, increasing the net negative charge.     

Adiposity,blood pressure,and carotid intima‐media thickness in greek adolescents

Objective:

In children and adolescents with cardiovascular risk factors, the assessment of subclinical target‐organ damage is of paramount importance. This study investigated factors associated with carotid intima‐media thickness (cIMT) in adolescents.

Design and Methods:

A cross‐sectional study was performed in 448 apparently healthy adolescents recruited from schools (mean age 14 ± 2.2 years, 211 boys), which involved cIMT measurements (common carotid artery) and assessment of lipid profile, glucose, and blood pressure (BP).

Results:

The prevalence of overweight/obesity was 28.1%/12.7% and of BP ≥95th percentile 19.6%. Left cIMT was correlated with age (r = 0.10), waist circumference (WC) (0.15), and BP (0.21/0.13, systolic/diastolic) (all P < 0.05). Right cIMT was correlated with waist to hip ratio (WHR) (0.10), whereas the mean of left and right cIMT was correlated with WC (0.12), WHR (0.12), and systolic BP (0.14) (all P < 0.05). After the age of 13 years, boys tended to have higher cIMT than girls, which was significant in the 13‐15 years subgroup (P < 0.05). In stepwise multivariate analysis (independent variables: age, gender, WC, WHR, body mass index z‐score, lipid parameters, glucose, BP), left cIMT was independently associated with systolic BP; right cIMT with WHR; mean left and right cIMT with WC. Adolescents with BP ≥90th percentile had higher left cIMT than those <90th percentile (0.63 ± 0.09 vs. 0.61 ± 0.09 mm respectively, P < 0.05).

Conclusion:

Central adiposity and systolic BP appear to be independently associated with increased cIMT values in apparently healthy adolescents. Left side cIMT appears to be superior to right side measurements in terms of association with cardiovascular risk factors.     

Experimental infection of house finches with Mycoplasma gallisepticum

Mycoplasma gallisepticum (MG) has caused an endemic upper respiratory and ocular infection in the eastern house finch (Carpodacus mexicanus) after the epidemic first described in 1994. The disease has been studied by a number of investigators at a population level and reports describe experimental infection in group-housed MG-free house finches. Because detailed observation and evaluation of individual birds in group-housed passerines is problematic, we studied individually housed house finches that were experimentally inoculated with the finch strain of MG in a controlled environment. To accomplish this, a study was conducted spanning the period of November 2001-April 2002 with 20 MG-free (confirmed by the rapid plate agglutination assay and polymerase chain reaction [PCR] assay) eastern house finches captured in the Cayuga Basin area of central New York (USA) in the summer of 2001. After a period of acclimatization and observation (12 wk), 20 finches were inoculated with a 0.05-ml aliquot of MG (3.24 x 10(5) colony-forming units/ml) via bilateral conjunctival sac instillations. Two additional finches acted as controls and were inoculated in the same manner with preservative-free sterile saline solution. After inoculation, all finches except the controls exhibited clinical signs of conjunctivitis within 2-6 days. The progression of the disease was evaluated by several methods, including PCR, behavioral observations, and physical examination including eye scoring, body weight, and body condition index. Over a period of 21 wk, MG-infected finches developed signs of disease and recovered (80%), developed signs of disease and progressed to become chronically infected (15%), or died (5%). We hypothesize that the high survival rate and recovery of these finches after infection was associated with the use of controlled environmental conditions, acclimatization, a high plane of nutrition, and low stocking (housing) density, all of which are factors documented to be important in the outcome of MG infections in domestic poultry and other species.     

Rates and patterns of mitochondrial DNA sequence evolution in fringilline finches (fringilla spp.) and the greenfinch (Carduelis chloris)

Rates and patterns of evolution in partial sequences of five mitochondrial genes (cytochrome b, ATPase 6, NADH dehydrogenase subunit 5, tRNA(Glu), and the control region) were compared among taxa in the passerine bird genera Fringilla and Carduelis. Rates of divergence do not vary significantly among genes, even in comparisons with the control region. Rate variation among lineages is significant only for the control region and NADH dehydrogenase subunit 5, and patterns of variation are consistent with the expectations of neutral theory. Base composition is biased in all genes but is stationary among lineages, and there is evidence for directional mutation pressure only in the control region. Despite these similarities, patterns of substitution differ among genes, consistent with alternative regimes of selective constraint. Rates of nonsynonymous substitution are higher in NADH dehydrogenase subunit 5 than in other protein-coding genes, and transitions exist in elevated proportions relative to transversions. Transitions appear to accumulate linearly with time in tRNA(Glu), and despite exhibiting the highest overall rate of divergence among species, there are no transversional changes in this gene. Finally, for resolving phylogenetic relationships among Fringilla taxa, the combined protein-coding data are broadly similar to those of the control region in terms of phylogenetic informativeness and statistical support.      

Metabolic phenotyping of the Crohn's disease-like IBD etiopathology in the TNF(ΔARE/WT) mouse model

The underlying biochemical consequences of inflammatory bowel disease (IBD) on the systemic and gastrointestinal metabolism have not yet been fully elucidated but could help to better understand the disease pathogenesis and to identify tissue-specific markers associated with the different disease stages. Here, we applied a metabonomic approach to monitor metabolic events associated with the gradual development of Crohn's disease (CD)-like ileitis in the TNF(ΔARE/WT) mouse model. Metabolic profiles of different intestinal compartments from the age of 4 up to 24 weeks were generated by combining proton nuclear magnetic resonance ((1)H NMR) spectroscopy and liquid chromatography-mass spectrometry (LC-MS). From 8 weeks onward, mice developed CD similar to the immune and tissue-related phenotype of human CD with ileal involvement, including ileal histological abnormalities, reduced fat mass and body weight, as well as hallmarks of malabsorption with higher energy wasting. The metabonomic approach highlighted shifts in the intestinal lipid metabolism concomitant to the histological onset of inflammation. Moreover, the advanced disease status was characterized by a significantly altered metabolism of cholesterol, triglycerides, phospholipids, plasmalogens, and sphingomyelins in the inflamed tissue (ileum) and the adjacent intestinal parts (proximal colon). These results describe different biological processes associated with the disease onset, including modifications of the general cell membrane composition, alteration of energy homeostasis, and finally the generation of inflammatory lipid mediators. Taken together, this provides novel insights into IBD-related alterations of specific lipid-dependant processes during inflammatory states.     

Aberrant expression of the autoantigen heterogeneous nuclear ribonucleoprotein-A2 (RA33) and spontaneous formation of rheumatoid arthritis-associated anti-RA33 autoantibodies in TNF-alpha transgenic mice

Human TNF-alpha transgenic (hTNFtg) mice develop erosive arthritis closely resembling rheumatoid arthritis (RA). To investigate mechanisms leading to pathological autoimmune reactions in RA, we examined hTNFtg animals for the presence of RA-associated autoantibodies including Abs to citrullinated epitopes (anti-cyclic citrullinated peptide), heterogeneous nuclear ribonucleoprotein (hnRNP)-A2 (anti-RA33), and heat shock proteins (hsp) (anti-hsp). Although IgM anti-hsp Abs were detected in 40% of hTNFtg and control mice, IgG anti-hsp Abs were rarely seen, and anti-cyclic citrullinated peptide Abs were not seen at all. In contrast, >50% of hTNFtg mice showed IgG anti-RA33 autoantibodies, which became detectable shortly after the onset of arthritis. These Abs were predominantly directed to a short epitope, which was identical with an epitope previously described in MRL/lpr mice. Incidence of anti-RA33 was significantly decreased in mice treated with the osteoclast inhibitor osteoprotegerin and also in c-fos-deficient mice lacking osteoclasts. Pronounced expression of hnRNP-A2 and a smaller splice variant was seen in joints of hTNFtg mice, whereas expression was low in control animals. Although the closely related hnRNP-A1 was also overexpressed, autoantibodies to this protein were infrequently detected. Because expression of hnRNP-A2 in thymus, spleen, brain, and lung was similar in hTNFtg and control mice, aberrant expression appeared to be restricted to the inflamed joint. Finally, immunization of hTNFtg mice with recombinant hnRNP-A2 or a peptide harboring the major B cell epitope aggravated arthritis. These findings suggest that overproduction of TNF-alpha leads to aberrant expression of hnRNP-A2 in the rheumatoid joint and subsequently to autoimmune reactions, which may enhance the inflammatory and destructive process.