RESUMO
The black soldier fly, Hermetia illucens, is an emerging biotechnological agent with its larvae being effective converters of organic waste into usable bio-products including protein and lipids. To date, most operations use unimproved commercial populations produced by mass rearing, without cognisance of specific breeding strategies. The genetic and phenotypic consequences of these commercial practices remain unknown and could have a significant impact on long-term population viability and productivity. The aim of this study was thus to assess the genetic and phenotypic changes during the early phases of colony establishment and domestication in the black soldier fly. An experimental colony was established from wild founder flies and a new microsatellite marker panel was developed to assess population genetic parameters along with the phenotypic characteristics of each generational cohort under captive breeding. The experimental colony was characterised by a small effective population size, subsequent loss of genetic diversity and rapid genetic and phenotypic differentiation between the generational cohorts. Ultimately, the population collapsed by the fifth generation, most likely owing to the adverse effect of inbreeding depression following the fixation of deleterious alleles. Species with r-selected life history characteristics (e.g. short life-span, high fecundity and low larval survival) are known to pose particular challenges for genetic management. The current study suggests that sufficient genetic and phenotypic variations exist in the wild population and that domestication and strain development could be achieved with careful population augmentation and selection during the early stages of colony establishment.
Assuntos
Dípteros/genética , Domesticação , Variação Genética , Animais , Dípteros/crescimento & desenvolvimento , Larva/genética , Larva/crescimento & desenvolvimento , FenótipoRESUMO
Data from the Australasian Bone Marrow Transplant Recipient Registry show a steady increase in the number of allogeneic haemopoietic stem cell transplantations (HSCT) performed annually in Australia and New Zealand. In 2012, 629 allogeneic HSCT were performed. Allogeneic HSCT is associated with numerous potential complications, including chronic graft-versus-host disease (cGVHD). The oral cavity is one of the most frequent sites affected by cGvHD, often leading to significant disability and reduced quality of life. Management strategies are often complex, of variable efficacy and influenced by the availability of various therapeutic agents, access to compounding pharmacies and associated costs. This paper summarises the current status of allogeneic HSCT in Australia and New Zealand with a focus on oral cGvHD and the associated challenges in its management.
Assuntos
Doença Enxerto-Hospedeiro/diagnóstico , Doença Enxerto-Hospedeiro/terapia , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Doenças da Boca/diagnóstico , Doenças da Boca/terapia , Austrália , Doença Crônica , Doença Enxerto-Hospedeiro/etiologia , Humanos , Doenças da Boca/etiologia , Transplante HomólogoRESUMO
Presented here is a magnetic hydrogel particle enabled workflow for capturing and concentrating SARS-CoV-2 from diagnostic remnant swab samples that significantly improves sequencing results using the Oxford Nanopore Technologies MinION sequencing platform. Our approach utilizes a novel affinity-based magnetic hydrogel particle, circumventing low input sample volumes and allowing for both rapid manual and automated high throughput workflows that are compatible with Nanopore sequencing. This approach enhances standard RNA extraction protocols, providing up to 40 × improvements in viral mapped reads, and improves sequencing coverage by 20-80% from lower titer diagnostic remnant samples. Furthermore, we demonstrate that this approach works for contrived influenza virus and respiratory syncytial virus samples, suggesting that it can be used to identify and improve sequencing results of multiple viruses in VTM samples. These methods can be performed manually or on a KingFisher automation platform.
Assuntos
COVID-19 , Sequenciamento por Nanoporos , Humanos , SARS-CoV-2 , Sequenciamento por Nanoporos/métodos , Hidrogéis , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Fenômenos MagnéticosRESUMO
There are few topical formulations used for oral medicine applications most of which have been developed for the management of dermatological conditions. As such, numerous obstacles are faced when utilizing these preparations in the oral cavity, namely enzymatic degradation, taste, limited surface area, poor tissue penetration and accidental swallowing. In this review, we discuss common mucosal diseases such as oral cancer, mucositis, vesiculo-erosive conditions, infections, neuropathic pain and salivary dysfunction, which could benefit from topical delivery systems designed specifically for the oral mucosa, which are capable of sustained release. Each condition requires distinct penetration and drug retention profiles in order to optimize treatment and minimize side effects. Local drug delivery may provide a more targeted and efficient drug-delivery option than systemic delivery for diseases of the oral mucosa. We identify those mucosal diseases currently being treated, the challenges that must be overcome and the potential of novel therapies. Novel biological therapies such as macromolecular biological drugs, peptides and gene therapy may be of value in the treatment of many chronic oral conditions and thus in oral medicine if their delivery can be optimized.
Assuntos
Sistemas de Liberação de Medicamentos , Doenças da Boca/tratamento farmacológico , Fatores Biológicos/uso terapêutico , Preparações de Ação Retardada , Terapia Genética , Humanos , Substâncias Macromoleculares/uso terapêutico , Terapia de Alvo Molecular , Mucosa Bucal/efeitos dos fármacos , Neoplasias Bucais/tratamento farmacológico , Doenças das Glândulas Salivares/tratamento farmacológicoRESUMO
Aspalathus linearis (Burm. F.) R. Dahlgren (Fabaceae) or rooibos, is a strict endemic species, limited to areas of the Cederberg (Western Cape) and the southern Bokkeveld plateau (Northern Cape) in the greater Cape Floristic Region (CFR) of South Africa. Wild rooibos, unlike the cultivated type, is variable in morphology, biochemistry, ecology and genetics, and these ecotypes are broadly distinguished into two main groups, namely, reseeders and resprouters, based on their fire-survival strategy. No previous assessment of genetic diversity or population structure using microsatellite markers has been conducted in A. linearis. This study aimed to test the hypothesis that wild rooibos ecotypes are distinct in genetic variability and that the ecotypes found in the Northern Cape are differentiated from those in the Cederberg that may be linked to a fire-survival strategy as well as distinct morphological and phytochemical differences. A phylogeographical and population genetic analyses of both chloroplast (trnLF intergenic region) and newly developed species-specific nuclear markers (microsatellites) was performed on six geographically representative wild rooibos populations. From the diversity indices, it was evident that the wild rooibos populations have low-to-moderate genetic diversity (He: 0.618-0.723; Ho: 0.528-0.704). The Jamaka population (Cederberg, Western Cape) had the lowest haplotype diversity (H = 0.286), and the lowest nucleotide diversity (π = 0.006) even though the data revealed large variations in haplotype diversity (h = 0.286-0.900) and nucleotide diversity (π = 0.006-0.025) between populations and amongst regions where wild rooibos populations are found. Our data suggests that populations of rooibos become less diverse from the Melkkraal population (Suid Bokkeveld, Northern Cape) down towards the Cederberg (Western Cape) populations, possibly indicative of clinal variation. The largest genetic differentiation was between Heuningvlei (Cederberg, Western Cape) and Jamaka (FST = 0.101) localities within the Cederberg mountainous region, and, Blomfontein (Northern Cape) and Jamaka (Cederberg) (FST = 0.101). There was also a significant isolation by distance (R2 = 0.296, p = 0.044). The presence of three main clusters is also clearly reflected in the discriminant analysis of principal components (DAPC) based on the microsatellite marker analyses. The correct and appropriate management of wild genetic resources of the species is urgently needed, considering that the wild Cederberg populations are genetically distinct from the wild Northern Cape plants and are delineated in accordance with ecological functional traits of reseeding or resprouting, respectively. The haplotype divergence of the ecotypes has also provided insights into the genetic history of these populations and highlighted the need for the establishment of appropriate conservation strategies for the protection of wild ecotypes.
RESUMO
Although active learning improves student outcomes in science, technology, engineering, and mathematics (STEM) programs, it may provoke anxiety in some students. We examined whether two psychological variables, social anxiety (psychological distress relating to the fear of negative evaluation by others) and academic self-efficacy (confidence in one's ability to overcome academic challenges), interact with student perceptions of evidence-based instructional practices (EBIPs) and associate with their final grades in a STEM-related course. Human anatomy and physiology students in community college courses rated various EBIPs for their perceived educational value and their capacity to elicit anxiety (N = 227). In general, practices causing students the most anxiety (e.g., cold calling) were reported by students as having the least educational value. When controlling for students' self-reported grade point averages, socially anxious students rated several EBIPs as more anxiety inducing, whereas high-efficacy students reported less anxiety surrounding other EBIPs. Furthermore, mediation analysis revealed that individual differences in academic self-efficacy at the beginning of the term explained some of the negative association between students' social anxiety levels and final grades in the course. Our results, obtained in a community college context, support a growing body of evidence that social anxiety and academic self-efficacy are linked with how students perceive and perform in an active-learning environment.
Assuntos
Aprendizagem Baseada em Problemas , Autoeficácia , Ansiedade , Medo , Humanos , Percepção , EstudantesRESUMO
Retinal degenerative diseases can have many possible causes and are currently difficult to treat. As an alternative to therapies that require genetic manipulation or the implantation of electronic devices, photopharmacology has emerged as a viable approach to restore visual responses. Here, we present a new photopharmacological strategy that relies on a photoswitchable excitatory amino acid, ATA. This freely diffusible molecule selectively activates AMPA receptors in a light-dependent fashion. It primarily acts on amacrine and retinal ganglion cells, although a minor effect on bipolar cells has been observed. As such, it complements previous pharmacological approaches based on photochromic channel blockers and increases the potential of photopharmacology in vision restoration.
Assuntos
Cegueira/tratamento farmacológico , Luz , Receptores de AMPA/metabolismo , Receptores de Ácido Caínico/metabolismo , Células Ganglionares da Retina/fisiologia , Potenciais de Ação/efeitos dos fármacos , Potenciais de Ação/genética , Animais , Animais Recém-Nascidos , Cegueira/genética , Cegueira/patologia , Canais de Cátion Regulados por Nucleotídeos Cíclicos/deficiência , Canais de Cátion Regulados por Nucleotídeos Cíclicos/genética , Modelos Animais de Doenças , GABAérgicos/farmacologia , Células HEK293 , Hipocampo/citologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Ácidos Fosfínicos/farmacologia , Picrotoxina/análogos & derivados , Picrotoxina/farmacologia , Piridinas/farmacologia , Receptores de Ácido Caínico/genética , Células Ganglionares da Retina/efeitos dos fármacos , Opsinas de Bastonetes/deficiência , Opsinas de Bastonetes/genética , Sesterterpenos , Proteínas rho de Ligação ao GTP/deficiência , Proteínas rho de Ligação ao GTP/genética , Receptor de GluK2 CainatoRESUMO
It is now well established that exogenous GH promotes sexual maturation and reproductive function. The possibility that this may reflect physiological actions of endogenous GH has, however, rarely been considered. Correlative changes in GH secretion and reproductive state (puberty, pregnancy, lactation, menopause and ovarian cycles) are thus the primary focus of this review. GH secretion is, for instance, elevated during major transitions in reproductive status such as puberty and pregnancy. In some cases, augmented circulating GH levels are paired with hepatic GH resistance. This interaction may permit selective activation of gonadal responses to GH without activating IGF-I-mediated systemic responses. This selective activation may also be mediated by autocrine, paracrine or intracrine GH actions, since GH is also synthesized in reproductive tissues. Correlative changes in GH secretion and reproductive state may be mediated by events at the hypothalamic, pituitary and gonadal level. In addition to direct effects on gonadal function, GH may influence reproductive activity by increasing gonadotropin secretion at the hypothalamic and pituitary level and by enhancing gonadotropin responsiveness at the gonadal level. The close association between reproductive status and the somatotrophic axis supports the physiological importance of GH in reproductive function.
Assuntos
Envelhecimento/fisiologia , Hormônio do Crescimento/fisiologia , Reprodução/fisiologia , Adolescente , Adulto , Animais , Criança , Ciclo Estral/fisiologia , Feminino , Hormônios Esteroides Gonadais/fisiologia , Gonadotropinas Hipofisárias/fisiologia , Humanos , Hipotálamo/metabolismo , Lactação/fisiologia , Fígado/metabolismo , Masculino , Menopausa/fisiologia , Ciclo Menstrual/fisiologia , Pessoa de Meia-Idade , Ovário/metabolismo , Hipófise/metabolismo , Gravidez , Ratos , Estações do Ano , Maturidade Sexual/fisiologia , Testículo/metabolismoRESUMO
The actions of growth hormone (GH) are not restricted to growth: GH modulates metabolic pathways as well as neural, reproductive, immune, cardiovascular, and pulmonary physiology. The importance of GH in most physiological systems suggests that GH deficiency at any age would be associated with significant morbidity. However, prior to the advent of recombinant GH, cadaver-derived GH was only used therapeutically to correct the height deficit, and thereby hypothetically improve quality of life (QoL), in GH-deficient children. Physicians now have access to unlimited, albeit expensive, supplies of recombinant GH, and are considering the advisability of GH replacement or supplementation in other patient populations. This paper analyses studies investigating the relationship between GH and QoL in GH-deficient children or adults, in GH-replete short children suffering from idiopathic short stature, Turner syndrome, or intrauterine growth retardation and in GH-deficient or replete elderly adults. Possible mechanisms by which GH might improve QoL at neural and somatic sites are also proposed.
Assuntos
Transtornos do Crescimento/tratamento farmacológico , Hormônio do Crescimento Humano/uso terapêutico , Qualidade de Vida , Adulto , Idoso , Envelhecimento/efeitos dos fármacos , Criança , Transtornos do Crescimento/psicologia , Hormônio do Crescimento Humano/deficiência , Humanos , Proteínas Recombinantes/uso terapêutico , Síndrome de Turner/tratamento farmacológicoRESUMO
GH, as its name suggests, is obligatory for growth and development. It is, however, also involved in the processes of sexual differentiation and pubertal maturation and it participates in gonadal steroidogenesis, gametogenesis and ovulation. It also has additional roles in pregnancy and lactation. These actions may reflect direct endocrine actions of pituitary GH or be mediated by its induction of hepatic or local IGF-I production. However, as GH is also produced in gonadal, placental and mammary tissues, it may act in paracrine or autocrine ways to regulate local processes that are strategically regulated by pituitary GH. The concept that GH is an important modulator of female reproduction is the focus of this review.
Assuntos
Hormônio do Crescimento/fisiologia , Mamíferos/fisiologia , Ovário/fisiologia , Reprodução/fisiologia , Animais , Comunicação Autócrina , Manutenção do Corpo Lúteo/fisiologia , Tubas Uterinas/fisiologia , Feminino , Hormônios Esteroides Gonadais/biossíntese , Humanos , Fator de Crescimento Insulin-Like I/fisiologia , Lactação/fisiologia , Fígado/fisiologia , Glândulas Mamárias Animais/fisiologia , Oogênese/fisiologia , Ovulação/fisiologia , Placenta/fisiologia , Gravidez , Puberdade/fisiologia , Receptores da Somatotropina/fisiologia , Útero/fisiologia , Vertebrados/fisiologia , Vitelogênese/fisiologiaRESUMO
Growth hormone (GH) regulates numerous cellular functions in many different tissues. A common receptor is believed to mediate these tissue-specific effects, suggesting that post-receptor signalling molecules or tissue sensitivity to GH may differ between tissues. Tissue sensitivity depends upon the abundance of GH receptors (GHRs), thus tissue-specific GHR regulation could enable tissue-specific GH actions. The comparative autoregulation of GHR gene transcription in central (whole brain or hypothalami) and peripheral (liver, bursa, spleen and thymus) tissues was therefore examined in domestic fowl. In all tissues, a 4.4 kb GHR gene transcript that encodes the full-length GHR was identified. The abundance of this transcript was inversely related to endogenous GH status; it was lower in males with high circulating concentrations of GH and higher in females with lower basal concentrations of plasma GH. The abundance of this transcript was also rapidly downregulated in response to a bolus systemic injection of recombinant chicken GH, designed to mimic an episodic burst of endogenous GH release. This autoregulatory response was observed within 2 h of GH administration and was of greater magnitude in the brain than in peripheral tissues. Intracerebroventricular injections of GH also downregulated GHR gene expression in the brain, although hepatic GHR transcripts were unaffected 24 h after central administration of GH. In contrast, the induction of hyposomatotropism by passive GH immunoneutralization increased the abundance of the GHR transcript in the thymus, but not in other central (brain) or peripheral (bursa, liver) tissues. GH is not the sole regulator of GHR abundance, however; hypersomatropism induced by hypothyroidism was associated with an increase in GHR mRNA. The expression of the GHR gene in the domestic fowl would thus appear to be autoregulated by GH in a tissue-specific way.
Assuntos
Encéfalo/metabolismo , Galinhas/metabolismo , Homeostase/fisiologia , Fígado/metabolismo , RNA Mensageiro/metabolismo , Receptores da Somatotropina/genética , Animais , Encéfalo/efeitos dos fármacos , Bolsa de Fabricius/efeitos dos fármacos , Bolsa de Fabricius/metabolismo , Feminino , Hormônio do Crescimento/imunologia , Hormônio do Crescimento/metabolismo , Hormônio do Crescimento/farmacologia , Homeostase/efeitos dos fármacos , Soros Imunes/administração & dosagem , Injeções Intraventriculares , Fígado/efeitos dos fármacos , Masculino , Fatores Sexuais , Baço/efeitos dos fármacos , Baço/metabolismo , Timo/efeitos dos fármacos , Timo/metabolismo , Hormônios Tireóideos/metabolismoRESUMO
GH exerts pleiotropic effects on growth and metabolism through the GH receptor. A deficiency in the GH receptor gene is thus associated with GH resistance and dwarfism. Complete GH resistance in humans, or Laron syndrome, has been associated with numerous inherited defects in the GH receptor, including point mutations, complete or partial gene deletions, and splice site alterations. Analysis of the GH receptor genes of these patients has provided considerable insight into structure-function relationships of the GH receptor. However, the relative rarity of this disease and the obvious difficulties involved in human research have prompted a search for an animal model of GH resistance. Numerous models have been proposed, including the sex-linked dwarf chicken, the guinea pig, and the Laron mouse. In this review, the characteristics and etiology of Laron syndrome and these animal models will be discussed. The insight provided by these disorders into the roles and mechanism of action of GH will also be reviewed.
Assuntos
Modelos Animais de Doenças , Nanismo Hipofisário/genética , Hormônio do Crescimento/fisiologia , Animais , Galinhas , Cobaias , Humanos , Camundongos , Receptores da Somatotropina/genéticaRESUMO
Sex-linked dwarfism (SLD) in chickens is characterized by impaired growth despite normal or supranormal plasma growth hormone (GH) levels. This resistance to GH action is thought to be due to mutations of the GH receptor (GHR) gene that reduce or prevent GH binding to target sites. The genetic lesion causing GH resistance in Cornell SLD chickens is, however, not known. Previous studies have shown that hepatic GH-binding activity is abnormally low in these birds, yet the GHR gene is transcribed into a transcript of appropriate size and abundance. Point mutations or defects in translation could therefore account for the impaired GHR activity in this strain. These possibilities were addressed in the present study. A missense mutation resulting in the substitution of serine for the conserved phenylalanine was identified in the region of the GHR cDNA encoding the extracellular domain. Translation of this mutant transcript was indicated by the presence of GHR/GH-binding protein (GHBP)-immunoreactive proteins in liver (55, 70 and 100 kDa) and serum (70 kDa) of normal (K) and SLD birds. Radiolabelled GH did not, however, bind to the hepatic membranes of most SLD chickens. Serum GH-binding activity, in contrast, was readily detectable, although at significantly lower levels than in normal birds. The missense mutation in the SLD GHR gene may thus affect targeting of GHRs to hepatic plasma membranes.
Assuntos
Galinhas/genética , Nanismo/genética , Hormônio do Crescimento/metabolismo , Mutação de Sentido Incorreto , Receptores da Somatotropina/genética , Cromossomos Sexuais , Animais , Western Blotting , Membrana Celular/metabolismo , Fígado/metabolismo , Ligação Proteica , Ensaio RadioliganteRESUMO
It is well established that GH-like proteins and mRNA are present in extrapituitary tissues, but it is not known whether this reflects ectopic transcription of the pituitary GH gene or the expression of a closely related gene. This possibility was, therefore, further investigated. Immunoreactive (IR) GH-like proteins were readily measured by RIA and immunoblotting in hypothalamic and extrahypothalamic brain tissues of the domestic fowl, in which GH-IR was similar in size and antigenicity to pituitary GH. RT-PCR of mRNA from these brain tissues, with oligonucleotide primers spanning the coding region of pituitary GH cDNA, also generated cDNA fragments identical in size (689 bp) to pituitary GH cDNA. The amplified brain cDNA sequences contained BamH1 and Rsa1 cleavage sites similar to those located in pituitary GH cDNA. These cDNA sequences also hybridized with a cDNA probe for chicken GH cDNA, producing moieties of expected size that were identical to the hybridizing moieties in pituitary tissue. The nucleotide sequences of the PCR products generated from hypothalamic and extrahypothalamic brain tissues, determined by a modified cycle dideoxy chain termination method, were also identical to pituitary GH cDNA. This homology extended over 594 bp of the hypothalamic cDNA fragment (spanning nucleotides 65 to 659 of the pituitary GH cDNA and its coding region for amino acids 4 to 201) and 550 bp of the extrahypothalamic cDNA fragment (spanning nucleotides 76 to 626 of pituitary GH cDNA and its coding region for amino acids 8 to 190). These results clearly establish that pituitary GH mRNA sequences are transcribed in hypothalamic and extrahypothalamic tissues of the chicken brain, in which GH-IR proteins are abundantly located. However, as GH mRNA could not be detected in the chicken brain by Northern blotting, its turnover may be more rapid than in pituitary tissue. The local production of GH within the brain nevertheless suggests that it has paracrine roles in modulating neural or neuroendocrine function.
Assuntos
Encéfalo/metabolismo , Hormônio do Crescimento/genética , Hipófise/metabolismo , Animais , Sequência de Bases , Northern Blotting , Galinhas , Primers do DNA/genética , Expressão Gênica , Hormônio do Crescimento/fisiologia , Hipotálamo/metabolismo , Immunoblotting , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , RNA Mensageiro/análise , Radioimunoensaio , Homologia de Sequência do Ácido NucleicoRESUMO
Although GH has no direct effect on GH release from chicken pituitary glands, GH receptor mRNA similar to that in the rabbit liver was identified by Northern blot analysis in extracts of adult chicken pituitaries. Complementary (c) DNA, reverse transcribed from chicken pituitary RNA, was amplified by the polymerase chain reaction (PCR) in the presence of 3'- and 5'-oligonucleotide primers coding for the extracellular domain of the chicken liver GH receptor and was found to contain an electrophoretically separable fragment of 500 bp, identical in size to that in chicken liver. Digestion of this pituitary cDNA with NcoI produced expected moities of 350 and 150 bp. Amplification of chicken pituitary cDNA in the presence of oligonucleotide primers for the intracellular sequence of the chicken liver GH receptor produced an electrophoretically separable fragment of approximately 800 bp, similar to that in chicken liver. This fragment was cut into expected moieties of 530 and 275 bp after digestion with EcoRI. These PCR fragments were identified in extracts of the pituitary caudal lobe, in which somatotrophs are confined and account for the majority of endocrine cell types, and in the cephalic lobe, in which somatotrophs are lacking. Translation of the GH receptor mRNA in the pituitary gland was indicated by the qualitative demonstration of radiolabelled GH-binding sites in plasma membrane preparations, in pituitary cytosol and in nuclear membranes. These results provide evidence for the expression and translation of the GH receptor gene in pituitary tissue, in which GH receptors appear to be widely distributed within cells and in different cell types. GH may therefore have paracrine, autocrine or intracrine effects on pituitary function.
Assuntos
Expressão Gênica/genética , Hipófise/embriologia , Receptores da Somatotropina/genética , Animais , Sítios de Ligação , Northern Blotting , Embrião de Galinha , Hipófise/metabolismo , Hipófise/fisiologia , Reação em Cadeia da Polimerase , Biossíntese de Proteínas/fisiologiaRESUMO
GH receptor (GHR) mRNA has been identified in peripheral (liver and muscle) and central (brain and hypothalamus) tissues of sex-linked dwarf (SLD) Leghorn chickens. Total RNA was extracted from the tissues of immature (1 week, 4 week), pubertal (16 week) and adult (> 24 weeks) SLD and K (the normally growing strain) Leghorn chickens. In both groups and all tissues, an mRNA moiety of 4.4 kb hybridized with cRNA probes derived from the rabbit hepatic GHR sequence. An additional low-abundance transcript of 2.8 kb was also identified in some tissues. An age-related increase in expression was observed in K and SLD hepatic GHR mRNA, suggesting normal regulation of SLD GHR gene transcription. Amplification of cDNA from K and SLD tissues in the presence of oligonucleotide primers coding for the intracellular or extracellular domains of the chicken GHR generated electrophoretically separable fragments of expected size. Restriction enzyme digestion of the products with EcoRI, BstNI, HaeIII, NcoI or BamHI produced smaller moieties of expected sizes in both strains. These results demonstrate that, in contrast to broiler SLDs, a GHR gene deletion is not responsible for the GHR dysfunction in Leghorn SLDs. Although the actual defect in GHR gene expression in SLD Leghorns remains to be identified, this study demonstrates that sex-linked dwarfism, like Laron dwarfism, is due to a heterogeneity of lesions.
Assuntos
Galinhas/genética , Deleção de Genes , Expressão Gênica/fisiologia , Transtornos do Crescimento/genética , Receptores da Somatotropina/genética , Cromossomos Sexuais , Animais , Sequência de Bases , Northern Blotting , Dados de Sequência Molecular , Reação em Cadeia da PolimeraseRESUMO
Thyroid hormones inhibit the synthesis and release of GH in avian species. This may represent a feedback mechanism, since GH enhances the peripheral production of tri-iodothyronine (T3). The possibility that GH may also have direct effects on thyroidal function was therefore investigated. The basal and thyrotrophin-induced release of thyroxine (T4) from incubated chicken thyroid glands was not enhanced, however, in the presence of chicken GH. Contrarily, GH impaired T4 release in a dose-related way. These actions were probably mediated by specific receptors, since binding sites for radiolabelled GH were demonstrated on the plasma membranes of chicken thyroid glands. Expression of the GH receptor gene in these tissues was also demonstrated using a cRNA probe for the rabbit liver GH receptor, which specifically hybridized with RNA moieties of 4.4 kb, 2.7 kb and 1.0 kb. Moreover, reverse transcription of thyroidal RNA and its amplification in the presence of 3'- and 5'-oligonucleotide primers coding for the extracellular or intracellular domains of the GH receptor generated electrophoretically separable fragments of 500 bp and 800 bp respectively, as would be expected from analysis of the hepatic GH receptor cDNA sequence. Digestion of the 500 bp fragment with NcoI or EcoRI also produced moieties of expected size (350 bp and 150 bp or 325 bp and 175 bp respectively), as did BamHI or HaeIII digestion of the 800 bp fragment (yielding fragments of 550 bp and 275 bp or 469 bp and 337 bp respectively). Translation of the GH receptor mRNA was also indicated by the immunocytochemical demonstration of GH receptors in thyroid follicular and parafollicular cells, using a specific polyclonal antibody raised against the chicken GH-binding protein. These results therefore provide evidence, for the first time, of GH receptor gene expression in thyroid tissue and the translation of functional GH receptors in thyroid glands. These results also demonstrate differential effects of GH on the extracellular concentrations of T3 and T4, which may permit subtle regulation within the somatotroph-thyroid axis.
Assuntos
Hormônio do Crescimento/fisiologia , Glândula Tireoide/fisiologia , Tiroxina/metabolismo , Animais , Sequência de Bases , Northern Blotting , Membrana Celular/metabolismo , Galinhas , Depressão Química , Relação Dose-Resposta a Droga , Expressão Gênica , Dados de Sequência Molecular , Sondas de Oligonucleotídeos/genética , Reação em Cadeia da Polimerase , Radioimunoensaio , Receptores da Somatotropina/genética , Receptores da Somatotropina/metabolismo , Glândula Tireoide/citologia , Glândula Tireoide/efeitos dos fármacos , Tireotropina/farmacologiaRESUMO
Growth hormone (GH) differs from other pituitary hormones in that it can affect a wide spectrum of cellular activities in many different tissues. These disparate actions are, however, mediated by a common receptor, suggesting tissue-specific differences in the post-receptor mechanisms and/or tissue sensitivities to GH stimulation may confer specificity. Tissue sensitivity depends upon the abundance of GH receptors (GHRs) and may be modulated by the amplitude and pulsatility of GH secretion. It may also be dependent upon the presence of non-signal transducing GH-binding proteins (GHBPs), which result from the alternate splicing of GHR gene transcripts. Tissue-specific autoregulation of GHRs and GHBPs could, therefore, contribute to differential tissue responsiveness to GH action. The autoregulation of GHR and GHBP gene transcription in novel central (hypothalamus, brainstem, and cortex/neocortex) and peripheral (spleen) tissues was therefore examined in adult, male Sprague-Dawley rats. For comparative purposes, GHR/GHBP gene expression was also examined in the liver, which has traditionally been considered the major GH-target site. Chronic hyposomatotropism, induced by hypophysectomy, exerted tissue-specific effects on the abundance of GHR gene products 10 days post-hypophysectomy. Both GHR and GHBP transcripts were reduced in the hypothalamus of hypophysectomized rats by 20% (P < 0.001), although neither transcript was affected in the liver, spleen, cortex/neocortex or brainstem. In contrast, 2 h after a single bolus GH injection that was designed to simulate a pulsatile increase in circulating GH concentrations, GHR and GHBP mRNA content was significantly increased by 25-30% (P < 0.001) in all brain regions and in the spleen of hypophysectomized or sham-hypophysectomized rats. Production of the two transcripts was differentially regulated, however, as GHBP, but not GHR, transcripts were increased in the liver (P < 0.001), whereas the GHR:GHBP ratio was decreased in the hypothalamus of GH-treated rats (P < 0.001). These results suggest that GHR gene transcription and splicing are acutely autoregulated in a tissue-specific way.
Assuntos
Encéfalo/metabolismo , Proteínas de Transporte/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/genética , Hormônio do Crescimento/farmacologia , Receptores da Somatotropina/genética , Animais , Hormônio do Crescimento/deficiência , Hipofisectomia , Fígado/metabolismo , Masculino , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Baço/metabolismoRESUMO
BACKGROUND: Tumor necrosis factor receptor-associated periodic syndrome (TRAPS) is an inflammatory disorder characterized by prolonged episodes of periodic fever and localized inflammation and dominantly inherited mutations in TNFRSF1A, the gene encoding the 55-kDa tumor necrosis factor receptor. To our knowledge, the cutaneous pathologic characteristics of TRAPS have not been described previously. OBJECTIVES: To characterize the dermatologic manifestations of TRAPS by clinical, microscopic, and molecular methods, and to investigate its immunophenotype. DESIGN, SETTING, AND PATIENTS: At the National Institutes of Health Clinical Center, Bethesda, Md, a tertiary care referral center, 25 patients with a clinical and molecular diagnosis of TRAPS were evaluated clinically and 10 biopsy specimens of lesional skin were examined by light microscopy and immunohistochemistry. Patients were screened for mutations in TNFRSF1A, the gene coding for the p55 tumor necrosis factor receptor. MAIN OUTCOME MEASURES: Clinical, light microscopic, and immunohistochemical features. RESULTS: The skin eruption usually lasted 4 to 21 days (mean, 13 days). Of 25 patients, 21 (84%) presented with migratory erythematous macules and patches and 10 (40%) had edematous dermal plaques. Conjunctivitis, characterized by pain and redness and/or periorbital edema, was present in 11 patients (44%). Most patients had their first skin eruption during the first 2 years of life. All patients had fever associated with the skin eruption. Most patients had associated abdominal pain (22 [88%]) and myalgia (20 [80%]). Other symptoms included arthralgia (13 [52%]), pleuritic chest pain (10 [40%]), and headache (17 [68%]). Microscopic examination of 10 biopsy specimens of lesional skin showed a superficial and deep perivascular and interstitial infiltrate of lymphocytes and monocytes. None of the biopsy specimens showed multinucleated macrophages or granulomatous or leukocytoclastic vasculitis. The results of immunohistochemistry showed a perivascular infiltrate of CD3+, CD4+, CD8+, CD68+, CD79a-, and CD20- cells. All the mutations were missense mutations in exons 2 through 4 of TNFRSF1A, directly affecting the extracellular domain of the protein. CONCLUSIONS: TRAPS is characterized by a spectrum of dermatologic findings, including migratory patches, edematous plaques, periorbital edema, and/or conjunctivitis. TRAPS is characterized by a perivascular dermal infiltrate of lymphocytes and monocytes.
Assuntos
Antígenos CD/genética , Febre Familiar do Mediterrâneo/patologia , Receptores do Fator de Necrose Tumoral/genética , Dermatopatias/patologia , Adolescente , Adulto , Criança , Pré-Escolar , Diagnóstico Diferencial , Febre Familiar do Mediterrâneo/diagnóstico , Febre Familiar do Mediterrâneo/genética , Feminino , Humanos , Imunofenotipagem , Masculino , Pessoa de Meia-Idade , Receptores Tipo I de Fatores de Necrose Tumoral , Dermatopatias/diagnóstico , Dermatopatias/genética , SíndromeRESUMO
While oral administration of therapeutic doses of phenylpropanolamine (PPA) does little to cardiovascular function in humans, intravenous doses administered to experimental animals are known to alter heart rate and blood pressure. Previous in vivo and in vitro studies have documented a beta-adrenoceptor agonist action for PPA and thus it was of interest to investigate whether these effects could be partially mediated by a direct interaction with beta-adrenoceptors. Phenylpropanolamine, [1R, 2R]-(-)-norephedrine, [1S, 2S]-(+)-norephedrine, [1S, 2R]-(+)-norpseudoephedrine, [S]-(+)-amphetamine, and [1R, 2S]-(-)-ephedrine, were compared with the known beta-adrenoceptor agonists [R]-(-)-epinephrine (EPI), [R]-(-)-norepinephrine (NE), and [R*, S*]-(+/-)-isoproterenol (ISO) for their ability to increase the intracellular concentration of cyclic-3',5'-adenosine monophosphate (cAMP) in minces of rat heart. Of the compounds investigated only NE, EPI, and ISO, as well as, forskolin, which directly stimulates adenylyl cyclase, significantly (p < 0.05) increased intracellular levels of cAMP. The other phenethylamines were without effect. The results of this study demonstrate that PPA and its diastereomers do not act directly at beta-adrenoceptors to alter cardiac function and supports the hypothesis that significant agonist activity of beta-phenethylamines at the beta-adrenoceptor requires phenolic/aryl ether substitution on the phenyl-ring (typically positions 3, 4 and/or 5).