Cholesteryl ester transfer protein in metabolic syndrome

Objective: Low high‐density lipoprotein cholesterol (HDL‐C), hypertriglyceridemia, and small dense‐low density lipoprotein (LDL) are key components of metabolic syndrome (MS). Cholesteryl ester transfer protein (CETP) mediates the transfer of triglycerides (TGs) from TG‐rich lipoproteins to HDL and LDL particles in exchange for cholesteryl esters, leading to low HDL‐C and small dense‐LDL. The aim of this study was to investigate the role of CETP in subjects with MS. Research Methods and Procedures: In a cross‐sectional cohort of 234 middle‐aged men and 252 women randomly selected from the Salzburg Atherosclerosis Prevention Program in Subjects at High Individual Risk (SAPHIR) study, MS was diagnosed according to the National Cholesterol Education Program guidelines. CETP mass was determined by enzyme‐linked immunosorbent assay and LDL size‐by‐gradient polyacrylamide gel electrophoresis. Results: Men and women with MS had lower HDL‐C (45 ± 7 vs. 58 ± 13 and 48 ± 10 vs. 71 ± 14 mg/dL for men and women, respectively; p < 0.001 for all) and higher TG levels (222 ± 71 vs. 98 ± 54 and 167 ± 67 vs. 90 ± 35 mg/dL for men and women, respectively; p < 0.001 for all) than healthy subjects. LDL size was lower in subjects with MS (256 ± 11 Å vs. 267 ± 11 Å and 262 ± 10 Å vs. 273 ± 8 Å for men and women, respectively; p < 0.001 for all). CETP mass was higher in men with MS (1.87 ± 0.78 vs. 1.40 ± 0.65 μg/mL; p < 0.001) but not in women (1.74 ± 0.79 vs. 1.62 ± 0.62 μg/mL). CETP mass correlated inversely with LDL size in both men and women (r = ?0.19, p < 0.01 and r = ?0.13, p < 0.05 in men and women, respectively). Discussion: MS is associated with increased CETP mass in men. Increased CETP mass may be responsible for reduced HDL‐C and reduced LDL particle diameter in MS.     

Role of protein kinase C-dependent A-kinase anchoring proteins in lysophosphatidic acid-induced cAMP signaling in human diploid fibroblasts

Previously, we reported that lysophosphatidic acid (LPA)-induced adenosine 3',5'-cyclic monophosphate (cAMP) production by human diploid fibroblasts depends on the age of the fibroblasts. In this study, we examined the role of A-kinase anchoring proteins (AKAP) in the regulation of LPA-stimulated cAMP production in senescent fibroblasts. We found that levels of protein kinase C (PKC)-dependent AKAPs, such as Gravin and AKAP79, were elevated in senescent cells. Co-immunoprecipitation experiments revealed that Gravin and AKAP79 do not associate with adenylyl cyclase type 2 (AC2) but bind to AC4/6, which interacts with calcium-dependent PKCs alpha/beta both in young and senescent fibroblasts. When the expression of Gravin and AKAP79 was blocked by small interference RNA transfection, the basal level of cAMP was greatly reduced and the cAMP status after LPA treatment was also reversed. Protein kinase A showed a similar pattern in terms of its basal activity and LPA-dependent modulation. These data suggest that Gravin and to a lesser extent, AKAP79, may play important roles in maintaining the basal AC activity and in coupling the AC systems to inhibitory signals such as Gialpha in young cells, and to stimulatory signals such as PKCs in senescent cells. This study also demonstrates that Gravin is especially important for the long-term activation of PKC by LPA in senescent cells. We conclude that LPA-dependent increased level of cAMP in senescent human diploid fibroblasts is associated with increases in Gravin levels resulting in its increased binding with and activation of calcium-dependent PKC alpha/beta and AC4/6.     

Vector systems of RNA interference

RNA interference is a mechanism of posttranslational (at the level of mRNA) gene silencing. Sequence-specific mRNA degradation is realized with the help of small interfering RNAs produced by processing of a precursor using Dicer, an enzyme from the RNAse III family. This mechanism is now widely used in vitro on cultures of mammalian cells in order to elucidate functions of individual genes by gene specific knockdown. Analogs of small interference RNAs are intensely expressed during embryogenesis. The mechanism of RNA interference plays an especially important role in embryogenesis of invertebrates. Identification of the functions of small noncoding RNAs is essential for understanding the genetic mechanisms underlying individual developmental stages. In order to integrate small interference RNAs in mammalian cells, various systems have been developed that allow both transient (for 48 h) and stable expression in vitro. These systems are considered in the present review.     

Phylogeography of the cosmopolitan marine parasite Kudoa thyrsites (Myxozoa: Myxosporea)

Kudoa thyrsites (Myxozoa: Multivalvulida) is a cosmopolitan marine parasite of fishes associated with post-mortem tissue degradation. Financial losses incurred as a result of these infections are of concern to commercial fisheries. There is conflicting evidence whether K. thyrsites represents a cryptic species complex. Myxospore morphology is very similar for K. thyrsites across its range, but preliminary genetic analyses show some differences. Kudoa thyrsites and the morphologically similar Kudoa histolytica were examined from hosts in British Columbia, Canada, Oregon, USA, Chile, England, South Africa, Australia, and Japan. We compared myxospore morphology and DNA sequences of heat shock protein 70 and the small subunit, large subunit, and internal transcribed spacer 1 of the ribosomal DNA. There was some morphological variation between regional representatives, inconsistent with genetic analyses. Phylogenetically, major separations correlated to four broad geographic regions: Japan, Australia, eastern Pacific, and eastern Atlantic. Within these regions there was little additional genetic structure. These data are evidence for regional subdivision of K. thyrsites suggesting global transplantation of fishes has yet to homogenize these distinctions. Within regions, parasite gene flow appears to be high between host species, suggesting little host specificity and minimal cryptic speciation. Our data also indicate that K. histolytica is not a valid species, as it was morphologically and genetically indistinguishable from K. thyrsites.     

Nano-hydroxyapatite-thermally denatured small intestine sub-mucosa composites for entheses applications

The objective of the present in vitro study was to estimate the adhesion strength of nanometer crystalline hydroxyapatite (HA)-small intestine sub-mucosa (SIS) composites on model implant surfaces. Techniques of thermal denaturation (60 degrees C, 20 min) of SIS were used to enhance the adhesion strength of entheses materials to underlying implants. Specifically, results indicated that the adhesion strength of thermally denatured SIS was 2-3 times higher than that for normal unheated SIS. In addition, aqua-sonicated, hydrothermally treated nano-HA dispersions enhanced the adhesion strength of SIS on implant surfaces. Importantly, results of the present study demonstrated that human skeletal muscle cell (hSkMC) numbers were not affected by thermally denaturing SIS in nano-HA composite coatings; however, they increased on aqua-sonicated nano-HA/SIS composites compared with SIS alone. Interestingly, thermally denatured SIS that contained aqua-sonicated, hydrothermally treated nano-HA decreased human osteoblasts (hOBs) numbers compared with respective unheated composites; all other composites when thermally denatured did not influence hOB numbers. Results also showed that the number of hOBs increased on nano-HA/SIS composites compared with SIS composites alone. Human mesenchymal stem cell (hMSC) numbers were not affected by the presence of nano-HA in SIS composites. For these reasons, the collective results of this in vitro study demonstrated a technique to increase the coating strength of entheses coatings on implant surfaces (using thermally denatured SIS and aqua-sonicated, hydrothermally prepared nano-HA) while, at the same time, supporting cell functions important for entheses regeneration.     

A Daphnia Parasite (Caullerya mesnili) Constitutes a New Member of the Ichthyosporea,a Group of Protists near the Animal–Fungi Divergence

ABSTRACT. Caullerya mesnili is a protozoan endoparasite in the gut epithelium of Daphnia, which causes regular epidemics in lakes throughout Europe. Its classification has remained unchanged for over a century, leaving it placed with the Haplosporidia, despite speculation that this position is incorrect. The difficulty in classifying C. mesnili stems from its few known morphological and ecological characteristics, as well as a lack of genetic markers. Here we sequenced the nuclear small subunit (SSU) and internal transcribed spacer rDNA regions of C. mesnili samples from 10 locations. Based on sequence similarities, we suggest the re‐classification of C. mesnili to the Ichthyosporea, a class of protists near the animal–fungi divergence. We report average intragenomic variation of 0.75% and 2.27% in the SSU and internal transcribed spacer regions, respectively. From electron micrographs and light microscopy of histological sections we determined that C. mesnili spores grow within the intestinal epithelium where they establish themselves intercellularly. In addition, we confirmed previous accounts regarding the high virulence of this parasite. Caullerya mesnili reduces host lifespan, the number of clutches, and the total number of offspring. This high selection pressure placed on hosts supports the importance of C. mesnili as a model parasite for the study of host–parasite biology in permanent lakes.     

The spinal muscular atrophy disease gene product, SMN: A link between snRNP biogenesis and the Cajal (coiled) body

The spliceosomal snRNAs U1, U2, U4, and U5 are synthesized in the nucleus, exported to the cytoplasm to assemble with Sm proteins, and reimported to the nucleus as ribonucleoprotein particles. Recently, two novel proteins involved in biogenesis of small nuclear ribonucleoproteins (snRNPs) were identified, the Spinal muscular atrophy disease gene product (SMN) and its associated protein SIP1. It was previously reported that in HeLa cells, SMN and SIP1 form discrete foci located next to Cajal (coiled) bodies, the so-called "gemini of coiled bodies" or "gems." An intriguing feature of gems is that they do not appear to contain snRNPs. Here we show that gems are present in a variable but small proportion of rapidly proliferating cells in culture. In the vast majority of cultured cells and in all primary neurons analyzed, SMN and SIP1 colocalize precisely with snRNPs in the Cajal body. The presence of SMN and SIP1 in Cajal bodies is confirmed by immunoelectron microscopy and by microinjection of antibodies that interfere with the integrity of the structure. The association of SMN with snRNPs and coilin persists during cell division, but at the end of mitosis there is a lag period between assembly of new Cajal bodies in the nucleus and detection of SMN in these structures, suggesting that SMN is targeted to preformed Cajal bodies. Finally, treatment of cells with leptomycin B (a drug that blocks export of U snRNAs to the cytoplasm and consequently import of new snRNPs into the nucleus) is shown to deplete snRNPs (but not SMN or SIP1) from the Cajal body. This suggests that snRNPs flow through the Cajal body during their biogenesis pathway.     

Study of small air ions concentrations versus asthma attacks in the region of Athens-Greece after the Chernobyl disaster

A consequence of the Chernobyl disaster was the isolated increase of small air ions for a limited time period over Athens-Greece. Taking advantage of this unique condition we studied the relation of log daily mean concentrations of small positive air ions, small negative air ions, NO2 and SO2 with the total daily number of asthma attacks attendances at the region's largest chest diseases hospital for 35 days following the disaster and during a same time period in the preceding four years. No correlation was found between air ionization and asthma attacks, though the study was of a rather short duration and several possible influencing factors such as aeroallergen concentrations were not taken into consideration.     

CHARACTER EVOLUTION IN POLYKRIKOID DINOFLAGELLATES1

A synthesis of available data on the morphological diversity of polykrikoid dinoflagellates allowed us to formulate a hypothesis of relationships that help explain character evolution within the group. Phylogenetic analyses of new SSU rDNA sequences from Pheopolykrikos beauchampii Chatton, Polykrikos kofoidii Chatton, and Polykrikos lebourae Herdman helped refine this hypothetical framework. Our results demonstrated that “pseudocolonies” in dinoflagellates evolved convergently at least three times independently from different Gymnodinium‐like ancestors: once in haplozoans; once in Ph. beauchampii; and at least once within a lineage containing Ph. hartmannii, P. kofoidii, and P. lebourae. The Gymnodiniales sensu stricto was strongly supported by the data, and the type species for the genus, namely Gymnodinium fuscum (Ehrenb.) F. Stein, formed the nearest sister lineage to a well‐supported Polykrikos clade. The best synapomorphy for the Polykrikos clade was the presence of two nuclei irrespective of zooid number. Two unidentified Gymnodinium species formed the nearest sister clade to Ph. beauchampii, which has four nuclei and four zooids per pseudocolony. The chain‐forming dinoflagellate G. catenatum L. W. Graham branched closely to the clade containing all members of Polykrikos and Pheopolykrikos, suggesting that an ancestral capacity toward chain formation existed before the evolution of pseudocolonies in this group. Our results also clarified the phylogenetic significance of nematocysts, ocelloids, and photosynthesis in reconstructing the evolution of polykrikoids and warnowiids. The molecular phylogenies exposed taxonomic problems associated with Polykrikos, Pheopolykrikos, and Gymnodinium, and suggested that a revision for some of these genera is warranted.