Regulation of adiponectin by adipose tissue-derived cytokines: in vivo and in vitro investigations in humans

Adiponectin is an adipose tissue-specific protein that is abundantly present in the circulation and suggested to be involved in insulin sensitivity and development of atherosclerosis. Because cytokines are suggested to regulate adiponectin, the aim of the present study was to investigate the interaction between adiponectin and three adipose tissue-derived cytokines (IL-6, IL-8, and TNF-alpha). The study was divided into three substudies as follows: 1) plasma adiponectin and mRNA levels in adipose tissue biopsies from obese subjects [mean body mass index (BMI): 39.7 kg/m2, n = 6] before and after weight loss; 2) plasma adiponectin in obese men (mean BMI: 38.7 kg/m2, n = 19) compared with lean men (mean BMI: 23.4 kg/m2, n = 10) before and after weight loss; and 3) in vitro direct effects of IL-6, IL-8, and TNF-alpha on adiponectin mRNA levels in adipose tissue cultures. The results were that 1) weight loss resulted in a 51% (P < 0.05) increase in plasma adiponectin and a 45% (P < 0.05) increase in adipose tissue mRNA levels; 2) plasma adiponectin was 53% (P < 0.01) higher in lean compared with obese men, and plasma adiponectin was inversely correlated with adiposity, insulin sensitivity, and IL-6; and 3) TNF-alpha (P < 0.01) and IL-6 plus its soluble receptor (P < 0.05) decreased adiponectin mRNA levels in vitro. The inverse relationship between plasma adiponectin and cytokines in vivo and the cytokine-induced reduction in adiponectin mRNA in vitro suggests that endogenous cytokines may inhibit adiponectin. This could be of importance for the association between cytokines (e.g., IL-6) and insulin resistance and atherosclerosis.     

Sexual selection and speciation in mammals,butterflies and spiders

Recently refined evolutionary theories propose that sexual selection and reproductive conflict could be drivers of speciation. Male and female reproductive optima invariably differ because the potential reproductive rate of males almost always exceeds that of females: females are selected to maximize mate 'quality', while males can increase fitness through mate 'quantity'. A dynamic, sexually selected conflict therefore exists in which 'competitive' males are selected to override the preference tactics evolved by 'choosy' females. The wide variation across taxa in mating systems therefore generates variance in the outcome of intrasexual conflict and the strength of sexual selection: monandry constrains reproductive heterozygosity and allows female choice to select and maintain particular (preferred) genes; polyandry promotes reproductive heterozygosity and will more likely override female choice. Two different theories predict how sexual selection might influence speciation. Traditional ideas indicate that increased sexual selection (and hence conflict) generates a greater diversity of male reproductive strategies to be counteracted by female mate preferences, thus providing elevated potentials for speciation as more evolutionary avenues of male-female interaction are created. A less intuitively obvious theory proposes that increased sexual selection and conflict constrains speciation by reducing the opportunities for female mate choice under polyandry. We use a comparative approach to test these theories by investigating whether two general measures of sexual selection and the potential for sexual conflict have influenced speciation. Sexual size dimorphism (across 480 mammalian genera, 105 butterfly genera and 148 spider genera) and degree of polyandry (measured as relative testes size in mammals (72 genera) and mating frequency in female butterflies (54 genera)) showed no associations with the variance in speciosity. Our results therefore show that speciation occurs independently of sexual selection.     

CL-46, a novel collectin highly expressed in bovine thymus and liver

Collectins are oligomeric molecules with C-type lectin domains attached to collagen-like regions via alpha-helical neck regions. They bind nonself glycoconjugates on the surface of microorganisms and inhibit infection by direct neutralization, agglutination, or opsonization. During the characterization of the gene encoding bovine CL-43 (43-kDa collectin), we identified a novel collectin-gene. We report the cloning and partial characterization of the novel collectin CL-46. The mRNA comprises 1188 nucleotides encoding a protein of 371 aa with an included leader peptide of 20 residues. CL-46 has two cysteine residues in the N-terminal segment, a potential N-glycosylation site in the collagen region, and an extended hydrophilic loop close to the binding site of the carbohydrate recognition domain. It is expressed in the thymus, liver, mammary gland, and tissues of the digestive system. Recombinant CL-46 corresponding to the alpha-helical neck region and the C-type lectin domain binds preferential N-acetyl-D-glucoseamine and N-acetyl-D-mannoseamine. The gene encoding CL-46 spans approximately 10 kb and consists of eight exons, with high structural resemblance to the gene encoding human surfactant protein D. It is located on the bovine chromosome 28 at position q1.8 together with the gene encoding conglutinin and CL-43. Several potential thymus-related cis-regulatory elements were identified in the 5'-upstream sequence, indicating that the expression in thymus may be modulated by signals involved in T cell development.     

Adoptive transfer of BALb/c mouse splenocytes reduces lesion severity and induces intestinal pathophysiologic changes in the Mycobacterium avium Subspecies paratuberculosis beige/scid mouse model

Successful immune reconstitution would enhance resistance of beige/scid mice to chronic infection with Mycobacterium avium subspecies paratuberculosis, but may cause damage to intestinal tissue. Therefore, we investigated the effect of adoptive transfer of BALB/c mouse splenocytes on lesion severity and intestinal physiology in beige/scid mice infected with M. paratuberculosis. Mice were inoculated intraperitoneally (i.p.) with M. paratuberculosis, and two weeks later were inoculated i.p. with viable spleen cells from immune-competent BALB/c mice. Mice were necropsied 12 weeks after infection when engraftment of lymphocytes, clinical disease, pathologic lesions, and intestinal electrophysiologic parameters were evaluated. Lymphocytes were rare in control beige/scid mice not inoculated with spleen cells. In contrast, high numbers of CD4+, CD8+, and B220+ lymphocytes were detected in the spleen of all beige/scid mice (n = 24) inoculated with spleen cells, indicating that adoptive transfer resulted in successful engraftment of donor lymphocytes (immune reconstitution). Immune reconstitution of M. paratuberculosis-infected beige/ scid mice significantly reduced the severity of clinical disease and pathologic lesions, and numbers of bacteria in the liver. However, intestinal electrophysiologic parameters studied in vitro indicated that intestinal tissues from reconstituted beige/scid mice had reduced short-circuit current responses (due to reduced ion secretion) following electrical, glucose, and forskolin stimulation. These abnormal responses suggested that neural or epithelial cells in the intestine were damaged. We conclude that successful immune reconstitution of beige/scid mice enhance their resistance to M. paratuberculosis infection, but may cause pathophysiologic changes associated with intestinal inflammation.     

RIBOSOMAL DNA AND RAPD VARIATION IN THE RARE PLANT FAMILY LACTORIDACEAE

Lactoris fernandeziana, endemic to the island of Masatierra in the Juan Fernandez Archipelago, is the only living member of the primitive angiosperm family, Lactoridaceae. The species was surveyed for ribosomal DNA (rDNA) and RAPD (Random Amplified Polymorphic DNA) variation. Previous analyses of allozymes had revealed no variation within the species. Variation was found for length in the intergenic spacer and for restriction sites in the 18S–25S genes of rDNA, and for the presence of amplified bands using 16 primers. Different rDNA repeat lengths and restriction site variants were detected within individuals as well as within and among populations. The level of variation in RAPDs is low relative to other Juan Fernandez endemic species surveyed, and nearly all variants were restricted to single populations. The rDNA length variants were distributed throughout the island, whereas the rDNA restriction site variants and RAPD markers indicated minor genetic differences among the populations.     

A new Pax gene,Pax-9, maps to mouse Chromosome 12

Members of the Pax gene family have recently been shown to play important roles in mouse embryogenesis. Of eight so far characterized Pax genes, three have been associated with mouse developmental mutants. Here we report the cloning of a new Pax gene, Pax-9. Most of the DNA sequence encoding the highly conserved paired domain has been determined and compared with previously known paired domains. This comparison classifies Pax-9 as a member of the same subgroup as Pax-1/undulated. By analysis of the segregation of a Pax-9 restriction fragment length polymorphism and a large number of simple sequence length polymorphisms in an interspecific C57BL/6 x Mus musculus mollosinus backcross, Pax-9 was mapped close to the D12Nds1 locus on the proximal part of Chromosome (Chr) 12.     

Analysis of limb patterning in BMP-7-deficient mice

Bone morphogenetic proteins `BMPs' are polypeptide signaling molecules, belonging to the TGF-β superfamily. They were originally identified by their ability to induce ectopic bone formation, but their expression patterns in embryos suggest multiple functions. BMP-7-deficient mice show among other mesodermal and skeletal patterning defects, polydactyly in the hindlimbs `Luo G, Hofmann C, Bronckers ALJJ, Sohocki M, Bradley A, Karsenty G `1995': Genes Dev 9:2808-2820; Dudley AT, Lyons KM, Robertson EJ `1995': Genes Dev 9:2795-2807'. Here we report a more detailed analysis of the limb phenotype in BMP-7-deficient mice using in situ hybridization to monitor expression of molecules implicated in patterning processes of the developing vertebrate limb. In previous studies we showed that Sonic hedgehog (Shh) was expressed normally, but Hoxd-13 expression in limb mesenchyme was lower in BMP-7 mutant limbs. Here we show that Hoxd-11 expression domains are also contracted and decreased in intensity in mutant limbs, suggesting that 5′ genes of the Hoxd cluster are coordinately downregulated, while another Bmp, Bmp-2, which can be activated by Shh, is similarly expressed. The mutant limb buds are broader than normal buds, and fibroblast growth factor Fgf-8 is expressed throughout the extended ridge. However, expression of the homeobox gene Msx-1, which has been shown to be involved in epithelial-mesenchymal interactions during limb development, was decreased in the mesenchyme of BMP-7 mutant limbs. Taken together, our data suggest that BMP-7 is involved in regulating proliferation and/or epithelial-mesenchymal interactions in the developing limb. © 1996 Wiley-Liss Inc.     

Human topoisomerase IIalpha: targeting to subchromosomal sites of activity during interphase and mitosis

Mammalian topoisomerase IIalpha (topo IIalpha) plays a vital role in the removal of topological complexities left on DNA during S phase. Here, we developed a new assay to selectively identify sites of catalytic activity of topo IIalpha with subcellular resolution. We show that topo IIalpha activity concentrates at replicating heterochromatin in late S in a replication-dependent manner and at centric heterochromatin during G2 and M phases. Inhibitor studies indicate that this cell cycle-dependent concentration over heterochromatin is sensitive to chromatin structure. We further show that catalytically active topo IIalpha concentrates along the longitudinal axis of mitotic chromosomes. Finally, we found that catalytically inert forms of the enzyme localize predominantly to splicing speckles in a dynamic manner and that this pool is differentially sensitive to changes in the activities of topo IIalpha itself and RNA polymerase II. Together, our data implicate several previously unsuspected activities in the partitioning of the enzyme between sites of activity and putative depots.