Food-induced expression of orexin receptors in rat duodenal mucosa regulates the bicarbonate secretory response to orexin-A

Presence of appetite-regulating peptides orexin-A and orexin-B in mucosal endocrine cells suggests a role in physiological control of the intestine. Our aim was to characterize orexin-induced stimulation of duodenal bicarbonate secretion and modulation of secretory responses and mucosal orexin receptors by overnight food deprivation. Lewis x Dark Agouti rats were anesthetized and proximal duodenum cannulated in situ. Mucosal bicarbonate secretion (pH stat) and mean arterial blood pressure were continuously recorded. Orexin-A was administered intra-arterially close to the duodenum, intraluminally, or into the brain ventricles. Total RNA was extracted from mucosal specimens, reverse transcribed to cDNA and expression of orexin receptors 1 and 2 (OX1 and OX2) measured by quantitative real-time PCR. OX1 protein was measured by Western blot. Intra-arterial orexin-A (60-600 nmol.h(-1).kg(-1)) increased (P < 0.01) the duodenal secretion in fed but not in fasted animals. The OX1 receptor antagonist SB-334867, which was also found to have a partial agonist action, abolished the orexin-induced secretory response but did not affect secretion induced by the muscarinic agonist bethanechol. Atropine, in contrast, inhibited bethanechol but not orexin-induced secretion. Orexin-A infused into the brain ventricles (2-20 nmol.kg(-1).h(-1)) or added to luminal perfusate (1.0-100 nM) did not affect secretion, indicating that orexin-A acts peripherally and at basolateral receptors. Overnight fasting decreased mucosal OX1 and OX2 mRNA expression (P < 0.01) as well as OX1 protein expression (P < 0.05). We conclude that stimulation of secretion by orexin-A may involve both receptor types and is independent of cholinergic pathways. Intestinal OX receptors and secretory responses are markedly related to food intake.     

The tissue distribution of lipoprotein lipase determines where chylomicrons bind

To determine the role of LPL for binding of lipoproteins to the vascular endothelium, and for the distribution of lipids from lipoproteins, four lines of induced mutant mice were used. Rat chylomicrons labeled in vivo with [¹⁴C]oleic acid (primarily in TGs, providing a tracer for lipolysis) and [³H]retinol (primarily in ester form, providing a tracer for the core lipids) were injected. TG label was cleared more rapidly than core label. There were no differences between the mouse lines in the rate at which core label was cleared. Two minutes after injection, about 5% of the core label, and hence chylomicron particles, were in the heart of WT mice. In mice that expressed LPL only in skeletal muscle, and had much reduced levels of LPL in the heart, binding of chylomicrons was reduced to 1%, whereas in mice that expressed LPL only in the heart, the binding was increased to over 10%. The same patterns of distribution were evident at 20 min when most of the label had been cleared. Thus, the amount of LPL expressed in muscle and heart governed both the binding of chylomicron particles and the assimilation of chylomicron lipids in the tissue.     

The E3 ligase synoviolin controls body weight and mitochondrial biogenesis through negative regulation of PGC-1β

Obesity is a major global public health problem, and understanding its pathogenesis is critical for identifying a cure. In this study, a gene knockout strategy was used in post-neonatal mice to delete synoviolin (Syvn)1/Hrd1/Der3, an ER-resident E3 ubiquitin ligase with known roles in homeostasis maintenance. Syvn1 deficiency resulted in weight loss and lower accumulation of white adipose tissue in otherwise wild-type animals as well as in genetically obese (ob/ob and db/db) and adipose tissue-specific knockout mice as compared to control animals. SYVN1 interacted with and ubiquitinated the thermogenic coactivator peroxisome proliferator-activated receptor coactivator (PGC)-1β, and Syvn1 mutants showed upregulation of PGC-1β target genes and increase in mitochondrion number, respiration, and basal energy expenditure in adipose tissue relative to control animals. Moreover, the selective SYVN1 inhibitor LS-102 abolished the negative regulation of PGC-1β by SYVN1 and prevented weight gain in mice. Thus, SYVN1 is a novel post-translational regulator of PGC-1β and a potential therapeutic target in obesity treatment.     

The Spontaneous Mutation Rate in the Fission Yeast Schizosaccharomyces pombe

The rate at which new mutations arise in the genome is a key factor in the evolution and adaptation of species. Here we describe the rate and spectrum of spontaneous mutations for the fission yeast Schizosaccharomyces pombe, a key model organism with many similarities to higher eukaryotes. We undertook an ∼1700-generation mutation accumulation (MA) experiment with a haploid S. pombe, generating 422 single-base substitutions and 119 insertion-deletion mutations (indels) across the 96 replicates. This equates to a base-substitution mutation rate of 2.00 × 10⁻¹⁰ mutations per site per generation, similar to that reported for the distantly related budding yeast Saccharomyces cerevisiae. However, these two yeast species differ dramatically in their spectrum of base substitutions, the types of indels (S. pombe is more prone to insertions), and the pattern of selection required to counteract a strong AT-biased mutation rate. Overall, our results indicate that GC-biased gene conversion does not play a major role in shaping the nucleotide composition of the S. pombe genome and suggest that the mechanisms of DNA maintenance may have diverged significantly between fission and budding yeasts. Unexpectedly, CpG sites appear to be excessively liable to mutation in both species despite the likely absence of DNA methylation.     

Functional Implications of Photosystem II Crystal Formation in Photosynthetic Membranes

The structural organization of proteins in biological membranes can affect their function. Photosynthetic thylakoid membranes in chloroplasts have the remarkable ability to change their supramolecular organization between disordered and semicrystalline states. Although the change to the semicrystalline state is known to be triggered by abiotic factors, the functional significance of this protein organization has not yet been understood. Taking advantage of an Arabidopsis thaliana fatty acid desaturase mutant (fad5) that constitutively forms semicrystalline arrays, we systematically test the functional implications of protein crystals in photosynthetic membranes. Here, we show that the change into an ordered state facilitates molecular diffusion of photosynthetic components in crowded thylakoid membranes. The increased mobility of small lipophilic molecules like plastoquinone and xanthophylls has implications for diffusion-dependent electron transport and photoprotective energy-dependent quenching. The mobility of the large photosystem II supercomplexes, however, is impaired, leading to retarded repair of damaged proteins. Our results demonstrate that supramolecular changes into more ordered states have differing impacts on photosynthesis that favor either diffusion-dependent electron transport and photoprotection or protein repair processes, thus fine-tuning the photosynthetic energy conversion.     

Cost of Preventing,Managing, and Treating Human Papillomavirus (HPV)-Related Diseases in Sweden before the Introduction of Quadrivalent HPV Vaccination

Objective

Costs associated with HPV-related diseases such as cervical dysplasia, cervical cancer, and genital warts have not been evaluated in Sweden. These costs must be estimated in order to determine the potential savings if these diseases were eradicated and to assess the combined cost-effectiveness of HPV vaccination and cervical cancer screening. The present study aimed to estimate prevention, management, and treatment costs associated with cervical dysplasia, cervical cancer, and genital warts from a societal perspective in Sweden in 2009, 1 year before the quadrivalent HPV vaccination program was implemented.

Methods and Materials

Data from the Swedish cervical cancer screening program was used to calculate the costs associated with prevention (cytological cervical cancer screening), management (colposcopy and biopsy following inadequate/abnormal cytological results), and treatment of CIN. Swedish official statistics were used to estimate treatment costs associated with cervical cancer. Published epidemiological data were used to estimate the number of incident, recurrent, and persistent cases of genital warts; a clinical expert panel assessed management and treatment procedures. Estimated visits, procedures, and use of medications were used to calculate the annual cost associated with genital warts.

Results

From a societal perspective, total estimated costs associated with cervical cancer and genital warts in 2009 were €106.6 million, of which €81.4 million (76%) were direct medical costs. Costs associated with prevention, management, and treatment of CIN were €74 million; screening and management costs for women with normal and inadequate cytology alone accounted for 76% of this sum. The treatment costs associated with incident and prevalent cervical cancer and palliative care were €23 million. Estimated costs for incident, recurrent and persistent cases of genital warts were €9.8 million.

Conclusion

Prevention, management, and treatment costs associated with cervical dysplasia, cervical cancer, and genital warts are substantial. Defining these costs is important for future cost-effectiveness analyses of the quadrivalent HPV vaccination program in Sweden.     

Genetic homogeneity/heterogeneity of Propionibacterium acnes isolated from patients during cardiothoracic reoperation

Following cardiothoracic surgery, deep sternal wound infection (SWI) remains one of the most severe complications. Recently, Propionibacterium acnes has been suspected as an etiological agent of deep SWI. However, this bacterium constitutes part of the resident micro-flora of the human skin. Consequently, findings of P. acnes in invasive samples are difficult to value. The aims of this study were to develop and optimize a pulsed-field gel electrophoresis (PFGE) protocol for P. acnes, in order to investigate the genetic homogeneity/heterogeneity of P. acnes isolates from multiple tissue samples (predominantly biopsies), collected at different locations, from 12 patients during cardiothoracic reoperation. There were 24 distinguishable PFGE fingerprints identified among the P. acnes isolates (n=54). Five (42%) of the patients carried only isolates that were interpreted as presumably clonally related. From the remaining seven patients, two or three different P. acnes clones were cultured, however, from six of them, the clones were identified in multiple samples. P. acnes may be a relatively frequent etiological agent of postoperative cardiothoracic infections. Existence of several clonally related P. acnes isolates derived from multiple samples from patients suffering from deep SWI after cardiothoracic surgery has not previously been shown.     

Proteomics in prostate cancer research

The incidence of early prostate cancer (PCa) among middle-aged men has increased rapidly. For many of these men, curatively intended treatment does more harm than good. Established prognostic factors are tumor stage and grade. As a result of earlier detection a majority of patients now have nonpalpable tumors (T1c) of intermediate grade (Gleason score 6). Prostate specific antigen in serum in such cases is generally at a low level and not a reliable predictor of prognosis. Altogether there is an urgent need for adjunctive prognostic indicators. In the search for relevant tumor markers for improved patient selection an exploration of the proteome (the human proteins) could be fruitful. This paper critically reviews the use of 2-dimensional gel electrophoresis (2-DE) for proteome research. Additional steps such as image analysis and mass spectrometry are described. Techniques based on non-2-DE platforms: surface-enhanced laser desorption/ionization (SELDI), isotope coded affinity tags (ICAT) and array-based technologies are also summarized. Although labor-intensive and time-consuming, 2-DE is presently the most powerful method for analysis of cellular protein phenotype and may potentially reveal gene regulations that cannot be detected on a genetic level.     

Malleability of protein folding pathways: a simple reason for complex behaviour

Although the structures of native proteins are generally unique, the pathways by which they form are often free to vary. Some proteins fold by a multitude of different pathways, whereas others seem restricted to only one choice. An explanation for this variation in folding behaviour has recently emerged from studies of transition state changes: the number of accessible pathways is linked to the number of nucleation motifs contained within the native topology. We refer to these nucleation motifs as 'foldons', as they approach the size of an independent cooperative unit. Thus, with respect to pathway malleability and the composition of the folding funnel, proteins can be seen as modular assemblies of competing foldons. For the split beta-alpha-beta fold, these foldons are two-strand-helix motifs coupled by spatial overlap.