1,720,982 research outputs found
Melanocortins and their receptors and antagonists
No full textThe melanocortins are a group of small protein hormones derived by post-translational cleavage of the proopiomelanocortin (POMC) gene product. The known melanocortin hormones include alpha-melanocyte stimulating hormone (MSH), beta-MSH, gamma-MSH and adrenocorticotropic hormone (ACTH). Five melanocortin receptors (MCIR through to MC5R) have been identified and most of these show tissue-specific expression patterns, as well as different binding affinities for each of the melanocortin hormones. The central melanocortin system consists of alpha-MSH, agouti-related protein (AGRP), MC3R and MC4R. AGRP and alpha-MSH are believed to be the natural antagonist and agonist respectively of MC3R and MC4R. This central melanocortin system is thought to play a fundamental role in the control of feeding and body weight. Knock-out mice models and genetic studies have pointed to the importance of the melanocortins in complex human pathways such as pigmentation, lipolysis, food intake, thermogenesis, sexual behaviour, memory and inflammatory response. Recently the melanocortins and their receptors have been the target for drug-based treatment of human physiological processes. MC3R and MC4R are likely targets for controlling body weight; MCIR may be used in the treatment of inflammation and MC2R for the treatment of glucocortical deficiency. A role for MCSR still remains unclear, but the evidence suggests an exocrine gland function
Mouse models of obesity
No full textInsights into the etiology of human obesity have arisen from the study of animal models. Animal models of obesity are also important for the development of future treatments of obesity. An agouti mouse mutation resulting in obese, yellow mice was described over a century ago and in 1992 agouti was cloned, making it the first obesity gene characterized at the molecular level. The lethal yellow mouse mutation is one of five dominant agouti mutations and is an excellent model for human obesity. The molecular categorization of agouti was responsible for the elucidation of the melanocortin system's involvement in hypothalamic weight regulation. As genetic knowledge increases many transgenic mice have been created with genes either over-expressed or deleted, models which further enhance the understanding of obesity
Agouti signal protein regulation in human melanoma cells
No full textProduction of the pigment eumelanin is controlled by alpha-melanocyte stimulating hormone (alpha-MSH) stimulation of melanocortin 1 receptor (Mc1r), whereas production of pheomelanin results from agouti antagonism of alpha-MSH signalling through Mc1r. The role of agouti in mouse pigmentation has been extensively investigated but a role for agouti signalling protein (ASIP) in human pigmentation has not been determined. To determine whether ASIP regulates known melanogenic genes in humans, ASIP was over-expressed in a human melanoma cell line. Levels of mRNA and protein were measured in genes known to be up or down-regulated by agouti in the mouse, namely microphthalmia (Mitf), tyrosinase (Tyr), tyrosinase-related protein 1 (Tyrp1), dopachrome tautomerase (Dct), Mc1r, silver, initiation transcription factor 2 (Itf2) and mini chromosome maintenance protein 6 (Mcm6). These melanogenic genes were not found to be significantly up or down-regulated by ASIP at the transcriptional level in human melanoma cells. However, ASIP down-regulation of tyrp1 was observed at the translational level. To identify novel genes that may be regulated by ASIP in melanoma cells, microarrays were used to determine differences in gene expression between the control and ASIP transfected melanoma cells. The expression level of human RNAs were determined by microarray analysis using a 19,200 cDNA and a 19,200 oligonucleotide array representing 13,000 and 18,864 individual genes, respectively. Genes observed to be modulated by ASIP were confirmed by quantitative real-time polymerase chain reaction. Results identify five genes, namely PPARbeta, eIF-4B, RRM2, MINOR and EVI2B that are down-regulated by ASIP, indicating a likely role for ASIP in human melanogenesis
A polymorphism study of the human Agouti gene and its association with MC1R
No full textTo determine whether the Agouti Signalling Protein (ASP) gene is associated with skin and hair coloration in humans, the complete coding region of ASP was screened for polymorphisms. Analysis of ASP in Caucasian, African-American, Spanish Basque, Hispanic, Apache and Australian Aboriginal populations revealed no amino acid substitutions. A single polymorphism in the 3' untranslated region occurred at a frequency of 0.2 in African-Americans. Variants of the Melanocortin 1 Receptor (MC1R) gene have been found to be associated with red hair and fair skin in humans. Red hair individuals are usually compound heterozygotes or homozygous for one of a number of MC1R polymorphisms associated with red hair. Some individuals however are heterozygous for only one of these polymorphisms and dizygotic twins can be concordant for MC1R variants but discordant for hair colour. A recent study has also identified rare redheads carrying no MC1R variants indicating that polymorphisms of the human MC1R gene are required but not sufficient for the red hair phenotype. To address the question of whether ASP also contributes to the red hair phenotype, individuals previously identified as having unexpected MC1R genotypes were screened for polymorphisms at the ASP locus. No polymorphisms were found in any of these individuals. Results indicate that the human ASP gene is unlikely to function in normal human pigmentation in the same way as MC1R
Interrogation of multimeric DNA amplification products by competitive primer extension using bst DNA polymerase (large fragment)
No full textLinear dsDNA composed of tandem repeats may be exponentially amplified by the strongly strand-displacing Bst DNA polymerase (large fragment) and two primers specific for opposite strands. When the repetitive DNA is derivedfrom rolling circle replication of a circular template, the reaction is termed cascade rolling circle amplification (CRCA). We have developed a variant of CRCA in which one primer is attached to the surface of a microwell and the other is labeled, thus enabling detection of amplified material using an ELISA-like protocol. The circular template is derived by annealing and ligation of a padlock on target DNA. It was found that there was good correlation between the synthesis of amplified material and signal. The specificity of the reaction with respect to single-nucleotide polymorphisms was investigated, and it was found that Bst DNA polymerase is prone to extension from primers with mismatched 3' ends. Reliable single nucleotide specificity was only obtained when pre-synthesized amplified material was interrogated by competitive primer extension
Gene polymorphisms and their effects in the melanocortin system
No full textIn addition to its role in human pigmentation, components of the melanocortin system regulate appetite, energy homeostasis and hormone production. Recent studies have suggested possible roles of this system in immunity, transmission of pain signals, and reproductive potential. A number of polymorphisms have been identified in genes of the melanocortin system and are associated with pigmentation in humans, as well as being causative of disorders of adrenal hormone production and obesity. This review gives an outline of these polymorphisms, their functional significance and possible application to or impact on diagnosis and pharmacotherapy based on melanocortin pathways
Agouti: From mouse to man, from skin to fat
No full textThe agouti protein regulates pigmentation in the mouse hair follicle producing a black hair with a subapical yellow band. Its effect on pigmentation is achieved by antagonizing the binding of alpha-melanocyte stimulating hormone (alpha-MSH) to melanocortin 1 receptor (Mc1r), switching melanin synthesis from eumelanin (black/brown) to phaeomelanin (red/yellow). Dominant mutations in the non-coding region of mouse agouti cause yellow coat colour and ectopic expression also results in obesity, type 11 diabetes, increased somatic growth and tumourigenesis. At least some of these pleiotropic effects can be explained by antagonism of other members of the melanocortin receptor family by agouti protein. The yellow coat colour is the result of agouti chronically antagonizing the binding of alpha-MSH to Mc1r and the obese phenotype results from agouti protein antagonizing the binding of alpha-MSH to Mc3r and/or Mc4r. Despite the existence of a highly homologous agouti protein in humans, agouti signal protein (ASIP), its role has yet to be defined. However it is known that human ASIP is expressed at highest levels in adipose tissue where it may antagonize one of the melanocortin receptors. The conserved nature of the agouti protein combined with the diverse phenotypic effects of agouti mutations in mouse and the different expression patterns of human and mouse agouti, suggest ASIP may play a role in human energy homeostasis and possibly human pigmentation
Solid-phase Amplification for Detection of C282y and H63D Hemochromatosis (HFE) Gene Mutations
No full textBackground: There is a need for simple, rapid, and inexpensive methods for the detection of single-nucleotide polymorphisms. Our aim was to develop a single-tube ELISA-like PCR assay and evaluate it by detecting the common C282Y and H63D mutations found in the hemochromatosis gene (HFE) by use of clinical samples.\ud
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Methods: The method, termed solid-phase amplification (SPA), involves dual liquid- and solid-phase amplification of a target sequence by the use of two PCR primers, one of which is in two forms: the first is covalently immobilized to the wall of a microwell, and the second is free in solution. During allele-specific amplification, both the free and solid-phase amplicons are labeled by incorporation of digoxigenin (DIG)-dUTP. The amount of surface-bound amplicon is determined colorimetrically by the use of an alkaline phosphatase-anti-DIG-Fab conjugate and p-nitrophenyl phosphate.\ud
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Results: Two different amplicon-labeling methods were evaluated. Analysis of 173 clinical samples for the C282Y and H63D HFE point mutations with SPA revealed that only one sample was incorrectly diagnosed, apparently because of operator error, when compared with conventional restriction fragment length polymorphism assay results.\ud
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Conclusions: The SPA assay has potential for medium-scale mutation detection, having the advantage of being manipulatively simple and immediately adaptable for use in clinical laboratories with existing ELISA instrumentation
Going Beyond Counting First Authors in Author Co-citation Analysis
Get PDFThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
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