Mechanisms regulating adrenomedullin gene expression in the left ventricle: role of mechanical load

Xenin is a 25 amino acid peptide produced by specific endocrine cells of the duodenal mucosa. Xenin has multiple biological actions in the gastrointestinal tract. It modulates intestinal motility, affects exocrine pancreatic secretion, and gastric secretion of acid. In the present investigation, we studied plasma concentration of xenin in volunteers after modified sham feeding and after meals of different composition. Plasma xenin concentrations were determined by radioimmunoassay in unextracted plasmas and after acidic extraction using C-18 Sep-Pak chromatography and after neutral extraction using affinity filtration. Both extraction methods were followed by C 18 r.p. HPLC chromatography. Xenin plasma concentrations in unextracted and in extracted plasma rose significantly after modified sham feeding when the food was brought to the volunteers from another room immediately before sham feeding started. When the volunteers had the opportunity to observe the preparation of the meal, xenin plasma concentrations during fasting were high and no further rise was observed after sham feeding. Isocaloric feeding resulted in elevated xenin concentrations in unextracted plasma and after high-pressure liquid chromatography. The methods of extraction, acidic or neutral, did not affect the results. CONCLUSION: Cephalic factors, investigated by modified sham feeding, stimulate release of xenin into the circulation. Xenin may participate in the central nervous regulation of gastrointestinal function.     

Agonist-dependent trafficking of alpha2-adrenoceptor subtypes: dependence on receptor subtype and employed agonist

Many G protein-coupled receptors (GPCRs) are internalized from the plasma membrane after agonist exposure. Previously, marked agonist-induced internalization of human alpha2A- and alpha2B-adrenergic receptors (AR) was observed in transfected neuronal rat pheochromocytoma (PC12) cells; alpha2A- and alpha2B-AR were internalized into partly distinct intracellular vesicles (Olli-L?hdesm?ki et al., J. Neurosci. 19, 9281-9288, 1999). In this paper, the extent of alpha2-AR internalization was quantitated in human embryonic kidney (HEK-293) and PC12 cells by combined application of cell surface biotinylation and ELISA methods, which allow measurement of protein trafficking in intact, differentiated and undifferentiated cells. Significant subtype-specific (but not cell type-dependent) trafficking of human alpha2-AR was observed by quantitation and immunocytochemistry. Agonist-induced sequestration of alpha2B-AR was markedly reduced after blocking the formation of clathrin-coated vesicles by hyperosmotic sucrose pretreatment. The sequestration of alpha2A-AR was partly inhibited after sucrose pretreatment but could be further reduced after inhibiting the formation of both clathrin-coated and caveolin vesicles by combined pretreatment with hyperosmotic sucrose and filipin. Differences were also observed in the recycling of alpha2A- and alpha2B-AR. The extent of maximal agonist-induced sequestration in PC12 cells was not directly dependent on relative agonist efficacy.     

Aging in Pinus sylvestris L. seeds: changes in viability and lipids

Aging of Scots pine seeds (Pinus sylvestris L.) leads to changes in seed quality, such as loss of germinability, delayed growth and abnormality in developing seedlings. The knowledge of biochemical changes responsible for these aging processes is plentiful in some seeds, which are of world-wide interest, but for pine seeds these studies are rare. The aim of the present study, was to analyse pine seeds of varying ages in order to identify biochemical changes occurring in aged pine seeds, and to see if a correlation existed between these results and traditionally used seed-quality parameters, such as germinability and electrolyte leakage.     

beta-oxidation - strategies for the metabolism of a wide variety of acyl-CoA esters

Living organisms are exposed to a number of different fatty acids and their various derivatives arising either via endogenous synthesis or from exogenous sources. These hydrophobic compounds can play specific metabolic, structural or endocrinic functions in the organisms before their elimination, which can be metabolism to CO(2) or to more polar lipid metabolites allowing their excretion. Quantitatively, one of the major pathways metabolizing fatty acids is beta-oxidation, which consists of a set of four reactions operating at the carbons 2 or 3 of acyl-CoA esters and shortening of the acyl-chain. To allow the beta-oxidation of acyl groups with various steric variants to proceed, different strategies have been developed. These strategies include evolution of beta-oxidation enzymes as paralogues showing specificity with respect to either chain-length or modified acyl-chain, metabolic compartmentalization in eukaryotic cells, controlling of substrate transport across membranes, development of auxiliary enzyme systems, acquisition of enzymes with adaptive active sites and recruiting and optimizing enzymes from non-homologous sources allowing them to catalyze a parallel set of reactions with different substrate specificities.     

Using heterozygosity–fitness correlations to study inbreeding depression in an isolated population of white‐tailed deer founded by few individuals

A heterozygosity–fitness correlations (HFCs) may reflect inbreeding depression, but the extent to which they do so is debated. HFCs are particularly likely to occur after demographic disturbances such as population bottleneck or admixture. We here study HFC in an introduced and isolated ungulate population of white‐tailed deer Odocoileus virginianus in Finland founded in 1934 by four individuals. A total of 422 ≥ 1‐year‐old white‐tailed deer were collected in the 2012 hunting season in southern Finland and genotyped for 14 microsatellite loci. We find significant identity disequilibrium as estimated by g₂. Heterozygosity was positively associated with size‐ and age‐corrected body mass, but not with jaw size or (in males) antler score. Because of the relatively high identity disequilibrium, heterozygosity of the marker panel explained 51% of variation in inbreeding. Inbreeding explained approximately 4% of the variation in body mass and is thus a minor, although significant source of variation in body mass in this population. The study of HFC is attractive for game‐ and conservation‐oriented wildlife management because it presents an affordable and readily used approach for genetic monitoring that allowing identification of fitness costs associated with genetic substructuring in what may seem like a homogeneous population.     

The Early-Onset Myocardial Infarction Associated PHACTR1 Gene Regulates Skeletal and Cardiac Alpha-Actin Gene Expression

The phosphatase and actin regulator 1 (PHACTR1) locus is a very commonly identified hit in genome-wide association studies investigating coronary artery disease and myocardial infarction (MI). However, the function of PHACTR1 in the heart is still unknown. We characterized the mechanisms regulating Phactr1 expression in the heart, used adenoviral gene delivery to investigate the effects of Phactr1 on cardiac function, and analyzed the relationship between MI associated PHACTR1 allele and cardiac function in human subjects. Phactr1 mRNA and protein levels were markedly reduced (60%, P<0.01 and 90%, P<0.001, respectively) at 1 day after MI in rats. When the direct myocardial effects of Phactr1 were studied, the skeletal α-actin to cardiac α-actin isoform ratio was significantly higher (1.5-fold, P<0.05) at 3 days but 40% lower (P<0.05) at 2 weeks after adenovirus-mediated Phactr1 gene delivery into the anterior wall of the left ventricle. Similarly, the skeletal α-actin to cardiac α-actin ratio was lower at 2 weeks in infarcted hearts overexpressing Phactr1. In cultured neonatal cardiac myocytes, adenovirus-mediated Phactr1 overexpression for 48 hours markedly increased the skeletal α-actin to cardiac α-actin ratio, this being associated with an enhanced DNA binding activity of serum response factor. Phactr1 overexpression exerted no major effects on the expression of other cardiac genes or LV structure and function in normal and infarcted hearts during 2 weeks’ follow-up period. In human subjects, MI associated PHACTR1 allele was not associated significantly with cardiac function (n = 1550). Phactr1 seems to regulate the skeletal to cardiac α-actin isoform ratio.     

Peroxisomal membrane channel Pxmp2 in the mammary fat pad is essential for stromal lipid homeostasis and for development of mammary gland epithelium in mice

To understand the functional role of the peroxisomal membrane channel Pxmp2, mice with a targeted disruption of the Pxmp2 gene were generated. These mice were viable, grew and bred normally. However, Pxmp2^−/− female mice were unable to nurse their pups. Lactating mammary gland epithelium displayed secretory lipid droplets and milk proteins, but the size of the ductal system was greatly reduced. Examination of mammary gland development revealed that retarded mammary ductal outgrowth was due to reduced proliferation of epithelial cells during puberty. Transplantation experiments established the Pxmp2^−/− mammary stroma as a tissue responsible for suppression of epithelial growth. Morphological and biochemical examination confirmed the presence of peroxisomes in the mammary fat pad adipocytes, and functional Pxmp2 was detected in the stroma of wild-type mammary glands. Deletion of Pxmp2 led to an elevation in the expression of peroxisomal proteins in the mammary fat pad but not in liver or kidney of transgenic mice. Lipidomics of Pxmp2^−/−mammary fat pad showed a decrease in the content of myristic acid (C₁₄), a principal substrate for protein myristoylation and a potential peroxisomal β-oxidation product. Analysis of complex lipids revealed a reduced concentration of a variety of diacylglycerols and phospholipids containing mostly polyunsaturated fatty acids that may be caused by activation of lipid peroxidation. However, an antioxidant-containing diet did not stimulate mammary epithelial proliferation in Pxmp2^−/− mice.     

Impacts of nutrient enrichment and sediment on phytoplankton community structure in the northern Baltic Sea

A three-week mesocosm experiment was conducted in order to study the effects of bottom sediment and nutrient enrichment on phytoplankton and zooplankton community structure in the Archipelago Sea, northern Baltic Sea. The transparent polyethylene enclosures included the whole water column and varied in volume from 30 to 40 m³. There were two types of enclosures: some with natural sediment as a bottom and others with a plastic bottom. The experiment was a 2 × 2 factorial design with presence of sediment and nutrient enrichment as treatment factors. Both the sediment presence and nutrient enrichment significantly increased water nutrient concentrations and the rate of primary production. However, external nutrient enrichment and the presence of sediment stimulated the growth of different phytoplankton groups, indicating that the effect of sediment was not related to nutrient fluxes alone, but involved more complex interactions. External nutrient enrichment was primarily channelled to picoplanktonic cyanobacteria, the biomass of which increased four- to fivefold due to enrichment. The presence of sediment increased the biomass of cryptophytes, chrysophytes and prasinophytes, but decreased the biomass of N₂-fixing cyanobacteria. Zooplankton biomass increased during the experiment, but was not affected by the treatments. The study shows that sediment plays a significant role in phytoplankton dynamics, underlining the importance of including sediment in shallow-water mesocosm experiments. Handling editor: J. Padisak     

Birth of correctly genotyped calves after multiplex marker detection from bovine embryo microblade biopsies

We report a method for multiplex genotyping of bovine embryo microblade biopsies. We have tested the reliability of the method and the viability of the embryos in vitro and in vivo. Two polymorphic gene markers (GHR F279Y and PRLR S18N) associated with milk production traits and one marker for sex diagnosis (ZFX/ZFY) were genotyped simultaneously with a method that combines nested PCR and allelic discrimination. To test the accuracy of genotyping, in the first experiment the genotypes of 134 biopsies from in vitro produced embryos were compared to genotypes determined from the corresponding embryos after biopsy. The method proved to be highly accurate as only in three cases (two for PRLR S18N and one for GHR F279Y) out of 395 genotypes the genotype was in disagreement between the two samples. The viability of similarly biopsied embryos was tested in parallel: after 24-hr culture 94.6% of embryos recovered in vitro. In the second experiment, a total of 150 in vivo-produced embryos were biopsied on Day 7 and genotyped. After the genotyping results were obtained on Day 8, female embryos were selected for transfer. From a total of 57 selected embryos 43 were transferred individually and 14 as pairs. After single embryo transfers, 19 recipients became pregnant and after embryo transfers in pairs one became pregnant. The success of genotyping was tested with the genotypes of donors and bulls and also from the hair samples of born calves. All calves were females and of the same genotypes determined from the biopsy.