RESUMO
OBJECTIVE: To describe the epidemiology of pediatric type 1 diabetes over 50 years in Canterbury, New Zealand. Further, to explore variation in case presentation according to age, gender, ethnicity, urban/rural character, socio-economic deprivation and immunogenetic features. RESEARCH DESIGN AND METHODS: Prospective ascertainment of cases commenced in 1982, and incident cases presenting 1970-1982 were ascertained retrospectively from clinical records. Eligibility criteria included diagnosis of type 1 diabetes by a physician and commencement of insulin therapy at diagnosis and age less than 15 years. Data collection included name, hospital number, date of birth, date of diagnosis, and date of initiation of insulin treatment. Full address at diagnosis was assigned an urban-rural classification, and a deprivation score. HLA-DQ susceptibility alleles and diabetes associated autoantibodies were determined. RESULTS: The incidence of type 1 diabetes increased more than 5-fold (3.9% per annum) over 50 years for the entire cohort. The mean for 5-year periods, starting from 1970, increased from 5.3 to 29.0 cases per 100,000 person years. Incidence was greatest in the 10-14 year age group. The cohort is predominantly European (89.4%), but there has been an increase in cases identifying as New Zealand Maori in the last three decades. Weak evidence was found for reduced incidence of type 1 diabetes in rural regions (adjusted IRR = 0.70, 95%CI 0.52 to 0.91, p = 0.011). CONCLUSIONS: The incidence of type 1 diabetes in children aged less than 15 years continues to increase with time. Incidence was significantly affected by age, ethnicity, and urban/rural characterization of address at diagnosis.
Assuntos
Diabetes Mellitus Tipo 1 , Adolescente , Criança , Diabetes Mellitus Tipo 1/diagnóstico , Diabetes Mellitus Tipo 1/epidemiologia , Humanos , Incidência , Nova Zelândia/epidemiologia , Estudos Prospectivos , Estudos RetrospectivosRESUMO
AIMS: To examine the impact of intensive management of type 1 diabetes (T1D) from diagnosis on HbA1c 12 months from diagnosis. METHODS: HbA1c measured 12 months after diagnosis for 70 consecutively newly diagnosed children with T1D following implementation of an intensive management protocol was compared with 70 children consecutively diagnosed immediately pre-implementation. Intensive management involved carbohydrate counting and flexible insulin dosing from first meal with subcutaneous insulin, targeted blood glucose levels from 4-8mmol/L irrespective of time of day, avoidance of twice daily insulin regimen and promotion of continuous glucose monitoring (CGM). HbA1c, diabetes technology use and insulin regimen at 12 months post-diagnosis were compared. RESULTS: The post-intensive management implementation cohort had an improved mean HbA1c of 58.2±15.3mmol/mol vs 63.7±10.7mmol/mol at 12 months (p=0.014). The proportion of young people with diabetes meeting a target HbA1c of <53mmol/mol at 12 months improved from 11% to 40% (p=<0.001). There was a reduction of twice daily insulin regimen from 66% to 11% (p=<0.001), and increased CGM use from 57% to 76% (p=0.02). CONCLUSION: Intensive management when implemented with consistent messaging from the multi-disciplinary team resulted in clinic-wide improvements in HbA1c and the proportion meeting HbA1c targets.
Assuntos
Diabetes Mellitus Tipo 1 , Criança , Humanos , Adolescente , Diabetes Mellitus Tipo 1/diagnóstico , Diabetes Mellitus Tipo 1/tratamento farmacológico , Hemoglobinas Glicadas , Glicemia , Automonitorização da Glicemia/métodos , Estudos Prospectivos , Nova Zelândia , Insulina/uso terapêutico , Hipoglicemiantes/uso terapêuticoRESUMO
The hydroxyproline-rich glycoproteins (HRGPs) are the major structural proteins of the extracellular matrix of algae and land plants. They are characterized by a rigid polyproline type II (PPII) conformation and extensive O-glycosylation of 4(R)-hydroxy-l-proline (Hyp) residues, which is a unique post-translational modification of proteins. The functional consequences of HRGP glycosylation remains unclear, but they have been implicated in contributing to their structural rigidity. Here, we have investigated the effects of naturally occurring beta-O-galactosylation of Hyp residues on the conformational stability of the PPII helix. In a series of well-defined model peptides Ac-(l-proline)(9)-NH(2) (1), Ac-(Hyp)(9)-NH(2) (2), and Ac-[Hyp(beta-d-galactose)](9)-NH(2) (3) we demonstrate that contiguous O-glycosylation of Hyp residues causes a dramatic increase in the thermal stability of the PPII helix according to analysis of thermal melting curves. This represents the first quantitative data on the contributions of glycosylation to stabilizing the PPII conformation. Molecular modeling indicates the increase in conformational stability may be due to a regular network of interglycan and glycan-peptide hydrogen bonds, in which the carbohydrate residues form a hydrophilic "overcoat" of the PPII helix. Evidence of this shielding effect of the amide backbone may be provided by analysis of the circular dichroism bands, which indicates an increase in the rho value of 3 relative to 1 and 2. This study gives further insight into the effects of naturally occurring Hyp beta-O-linked glycans on the PPII conformation as found in HRGPs in plant cell walls and also indicates that polyproline sequences may be suitable for the development of molecular scaffolds for the presentation of glycan structures.
Assuntos
Galactose/química , Hidroxiprolina/química , Peptídeos/síntese química , Glicosilação , Modelos Moleculares , Peptídeos/química , Conformação Proteica , EstereoisomerismoRESUMO
The conformations of peptides and proteins are often influenced by glycans O-linked to serine (Ser) or threonine (Thr). (2S,4R)-4-Hydroxyproline (Hyp), together with L-proline (Pro), are interesting targets for O-glycosylation because they have a unique influence on peptide and protein conformation. In previous work we found that glycosylation of Hyp does not affect the N-terminal amide trans/cis ratios (K(trans/cis)) or the rates of amide isomerization in model amides. The stereoisomer of Hyp--(2S,4S)-4-hydroxyproline (hyp)--is rarely found in nature, and has a different influence both on the conformation of the pyrrolidine ring and on K(trans/cis). Glycans attached to hyp would be expected to be projected from the opposite face of the prolyl side chain relative to Hyp; the impact this would have on K(trans/cis) was unknown. Measurements of (3)J coupling constants indicate that the glycan has little impact on the C(gamma)-endo conformation produced by hyp. As a result, it was found that the D-galactose residue extending from a C(gamma)-endo pucker affects both K(trans/cis) and the rate of isomerization, which is not found to occur when it is projected from a C(gamma)-exo pucker; this reflects the different environments delineated by the proline side chain. The enthalpic contributions to the stabilization of the trans amide isomer may be due to disruption of intramolecular interactions present in hyp; the change in enthalpy is balanced by a decrease in entropy incurred upon glycosylation. Because the different stereoisomers--Hyp and hyp--project the O-linked carbohydrates in opposite spatial orientations, these glycosylated amino acids may be useful for understanding of how the projection of a glycan from the peptide or protein backbone exerts its influence.
Assuntos
Glicopeptídeos/química , Hidroxiprolina/química , Prolina/análogos & derivados , Prolina/química , Amidas/química , Glicopeptídeos/metabolismo , Glicosilação , Hidroxiprolina/metabolismo , Estrutura Molecular , Estereoisomerismo , TermodinâmicaRESUMO
A method is described for in situ hybridization of riboprobes to free-floating brain sections. Brain sections are hybridized and processed free-floating in buffer, i.e., without attachment to a support such as a slide. To withstand the extra wear compared with sections processed on-slide, the brain tissue must be well fixed (4% paraformaldehyde) and sections cut at thickness of typically 40 microm. Sections were exposed to a prehybridization treatment before a riboprobe is added to form the hybridization solution. Riboprobes were prepared from cDNA via an in vitro transcription reaction and are labeled with digoxigenin. The sections are subsequently processed to remove nonspecific binding and the digoxigenin label detected via an antibody conjugated to alkaline phosphatase. This method may be readily combined with neuronal tracing and is ideal for further processing by immunohistochemistry to detect specific proteins.
Assuntos
Encéfalo/citologia , Encéfalo/metabolismo , Hibridização In Situ/métodos , Sondas RNA , Animais , Digoxigenina/metabolismo , Técnicas Histológicas , Microtomia , Neurônios/citologia , Neurônios/metabolismo , RNA Mensageiro/análise , Ratos , Fixação de TecidosRESUMO
AIM: Landmark studies, including the Diabetes Control and Complications Trial, have demonstrated the need for intensive management and improvement of glycaemic control in children and adolescents with Type 1 Diabetes (T1DM). Our aim was to determine what clinical resources were present in New Zealand to manage diabetes in children and adolescents and compare this with international recommendations, via the Paediatric Society of New Zealand clinical network. METHOD: All 21 District Health Board (DHB) secondary care sites in New Zealand managing children and adolescents with diabetes were invited to complete a survey about the specialist services they provided in 2012. RESULTS: All of the identified 21 sites (encompassing 20 centres) replied. These centres managed 1,587 children and adolescents with diabetes up to 18 years of age (>95% with T1DM), including 251 (16%) on insulin pumps. Average clinic HbA1c was not available for many centres. Staffing for specialists (general paediatricians or paediatric endocrinologists) was low (median 0.2/100 patients, range 0.1-0.4), but was relatively higher in diabetes nurses (median 0.7/100 patients, range 0.1-1.8). Despite the psychological and social burden of diabetes, the two allied health disciplines (psychology services and social worker) were the hardest to quantify as dedicated resource in these disciplines did not exist in all but three centres. CONCLUSIONS: This survey suggests that the majority of clinical services providing care for children with diabetes in New Zealand are significantly under-resourced.
Assuntos
Serviços de Saúde da Criança/estatística & dados numéricos , Diabetes Mellitus Tipo 1/terapia , Admissão e Escalonamento de Pessoal/estatística & dados numéricos , Adolescente , Criança , Serviços de Saúde da Criança/organização & administração , Pré-Escolar , Diabetes Mellitus Tipo 1/epidemiologia , Humanos , Incidência , Lactente , Recém-Nascido , Nova Zelândia/epidemiologia , Equipe de Assistência ao Paciente/estatística & dados numéricos , Prevalência , Inquéritos e Questionários , Adulto JovemRESUMO
This study is focused on sequence analysis of peptidomimetic helical oligoureas by means of tandem mass spectrometry, to build a basis for de novo sequencing for future high-throughput combinatorial library screening of oligourea foldamers. After the evaluation of MS/MS spectra obtained for model compounds with either MALDI or ESI sources, we found that the MALDI-TOF-TOF instrument gave more satisfactory results. MS/MS spectra of oligoureas generated by decay of singly charged precursor ions show major ion series corresponding to fragmentation across both CO-NH and N'H-CO urea bonds. Oligourea backbones fragment to produce a pattern of a, x, b, and y type fragment ions. De novo decoding of spectral information is facilitated by the occurrence of low mass reporter ions, representative of constitutive monomers, in an analogous manner to the use of immonium ions for peptide sequencing.
Assuntos
Peptidomiméticos/química , Ureia/química , Sequência de Aminoácidos , Modelos Moleculares , Dobramento de Proteína , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Espectrometria de Massas em Tandem/métodosRESUMO
The structure of fused C-glucosylproline hybrid (GlcProH) has been studied in detail computationally. A systematic molecular mechanics/Monte Carlo search has been performed in order to cover the entire conformational space of GlcProH. This has been followed by density-functional (DFT B3LYP) calculations in the gas phase and in aqueous solution, using the polarizable continuum model (PCM). In the gas phase, a large excess of the cis conformation with respect to the prolyl amide bond is found. This is reversed in aqueous solution where the calculations show 80% trans conformers, which is in accordance with experimental data. Thus, the PCM model is capable of accurately predicting cis-trans ratios. The free energy of solvation is not correlated with the dipole moment. Hence, a model (such as PCM) is required that takes into account the complete charge distribution. The reversal of the cis-trans ratio between gas phase and solution also emphasizes the effects of different free energies of solvation for the distinct conformers. Nevertheless, the energy difference between the cis and trans conformers is very small in solution (0.18 kcal/mol). Intramolecular hydrogen bonding is found to stabilize the cis conformers exclusively, which is a result of the rigid geometry of the fused rings. This can be contrasted to related more flexible molecules that show hydrogen bonding for both cis and trans isomers. The hydrogen bonding is at least partially responsible for the preferential stabilization of the cis conformers in the gas phase and a very small cis-trans energy difference in solution.
Assuntos
Carboidratos/química , Prolina/química , Modelos Moleculares , Conformação MolecularRESUMO
The administration of antipsychotic drugs to human patients or experimental animals leads to significant weight gain, which is widely presumed to be driven by hyperphagia; however, the contribution from energy expenditure remains unclear. These studies aim to examine the contribution of shifts in energy expenditure, particularly those involving centrally mediated changes in thermogenesis, to the body weight gain associated with the administration of olanzapine to female Sprague Dawley rats. Olanzapine (6 mg/kg/day orally) caused a transient increase in food intake but a maintained increase in body weight. When pair-fed rats were treated with olanzapine, body weight continued to rise compared to vehicle-treated rats, consistent with a reduction in energy expenditure. Brown adipose tissue (BAT) temperature, measured using biotelemetry devices, decreased immediately after the onset of olanzapine treatment and remained depressed, as did physical activity. UCP1 expression in interscapular BAT was reduced following chronic olanzapine treatment. An acute injection of olanzapine was preceded by an injection of a retrograde tracer into the spinal cord to evaluate the nature of the olanzapine-activated neural pathway. Levels of Fos protein in a number of spinally projecting neurons within discrete hypothalamic and brainstem sites were elevated in olanzapine-treated rats. Some of these neurons in the perifornical region of the lateral hypothalamus (LHA) were also Orexin A positive. These data collectively show a significant impact of thermogenesis (and physical activity) on the weight gain associated with olanzapine treatment. The anatomical studies provide an insight into the central neuroanatomical substrate that may subserve the altered thermogenic responses brought about by olanzapine.
Assuntos
Tecido Adiposo Marrom/fisiologia , Antipsicóticos/efeitos adversos , Benzodiazepinas/farmacologia , Peso Corporal/efeitos dos fármacos , Termogênese/fisiologia , Aumento de Peso/efeitos dos fármacos , Tecido Adiposo Marrom/efeitos dos fármacos , Animais , Ingestão de Energia/efeitos dos fármacos , Feminino , Hipotálamo/efeitos dos fármacos , Hipotálamo/fisiologia , Bulbo/efeitos dos fármacos , Bulbo/fisiologia , Atividade Motora/efeitos dos fármacos , Olanzapina , Núcleo Hipotalâmico Paraventricular/efeitos dos fármacos , Núcleo Hipotalâmico Paraventricular/fisiologia , Ratos , Ratos Sprague-Dawley , Termogênese/efeitos dos fármacosRESUMO
We describe the synthesis of a fused bicyclic C-glucosylproline hybrid (GlcProH) from commercially available 2,3,4,6-tetra-O-benzyl-d-glucopyranose. The GlcProH was incorporated into the model peptides Ac-GlcProH-NHMe and Ac-Gly-GlcProH-NHMe. Postsynthetic modifications can be introduced via derivatization of the carbohydrate scaffold. Conformational analysis of the GlcProH-modified model peptides shows that while the conformation of GlcProH remains fixed, the prolyl N-terminal amide equilibrium (Kt/c) can be varied with different modifications of the carbohydrate scaffold. Simple N-acyl derivatives studied by NMR spectroscopy showed that in CD3OD there was an increase in the cis-amide content as the sugar substituents changed from benzyl (10%) to hydroxyl (22%) to acetate (36%). Similar effects were observed in DMSO-d6. The exact nature of the influence is unclear, but it most likely arises through intramolecular interactions between sugar groups and the peptidic amide backbone. Overall, our GlcProH demonstrates variation in Kt/c through tuning of the carbohydrate scaffold: a new concept in proline peptidomimetics.
Assuntos
Amidas/química , Prolina/química , Ácidos Carboxílicos/síntese química , Ácidos Carboxílicos/química , Éteres/química , Flúor/química , Glucose/química , Glicosilação , Espectroscopia de Ressonância Magnética , Metilação , Estrutura Molecular , Solventes , EstereoisomerismoRESUMO
The nature of muscle efferent fibre activation during whole body cooling was investigated in urethane-anaesthetized rats. Multiunit efferent activity to the gastrocnemius muscle was detected when the trunk skin was cooled by a water-perfused jacket to below 36.0 +/- 0.7 degrees C. That efferent activity was not blocked by hexamethonium (50 mg kg(-1), i.v.) and was not associated with movement or electromyographic activity. Cold-induced efferent activity enhanced the discharge of afferent filaments from the isotonically stretched gastrocnemius muscle, demonstrating that it was fusimotor. Fusimotor neurons were activated by falls in trunk skin temperature, but that activity ceased when the skin was rewarmed, regardless of how low core temperature had fallen. While low core temperature alone was ineffective, a high core temperature could inhibit the fusimotor response to skin cooling. Fusimotor activation by skin cooling was often accompanied by desynchronization of the frontal electroencephalogram (EEG), but was not a simple consequence of cortical arousal, in that warming the scrotum desynchronized the EEG without activating fusimotor fibres. Inhibition of neurons in the rostral medullary raphé by microinjections of glycine (0.5 m, 120-180 nl) reduced the fusimotor response to skin cooling by 95 +/- 3%, but did not prevent the EEG response. These results are interpreted as showing a novel thermoregulatory reflex that is triggered by cold exposure. It may underlie the increased muscle tone that precedes overt shivering, and could also serve to amplify shivering. Like several other cold-defence responses, this reflex depends upon neurons in the rostral medullary raphé.
Assuntos
Temperatura Baixa , Neurônios Motores gama/fisiologia , Núcleos da Rafe/fisiologia , Reflexo/fisiologia , Temperatura Cutânea , Potenciais de Ação/fisiologia , Animais , Temperatura Corporal/fisiologia , Eletroencefalografia , Eletromiografia , Bloqueadores Ganglionares/farmacologia , Glicina/farmacologia , Hexametônio/farmacologia , Hipotermia Induzida , Masculino , Músculo Esquelético/inervação , Músculo Esquelético/fisiologia , Neurônios Eferentes/fisiologia , Ratos , Ratos Sprague-Dawley , Estremecimento/fisiologiaRESUMO
The rostral ventrolateral medulla (RVLM) is essential for the generation of sympathetic nerve activity. The RVLM receives a substantial innervation from the hypothalamic paraventricular nucleus (PVN). Activation of P2X purinoceptors via ATP has been shown to mediate fast excitatory synaptic neurotransmission. There is mounting evidence to suggest the presence of P2X purinoceptors in hypothalamic nuclei, including the PVN. In this study, we determined whether P2X1-P2X6 purinoceptor subtypes were present on PVN neurones that projected to the RVLM. Injection of the retrogradely transported tracer, rhodamine-tagged microspheres, into the pressor region of the RVLM was used to identify the neurones in the PVN that innervated the RVLM. P2X1-P2X6 purinoceptors were detected by immunohistochemistry. Double-labelled neurones were quantified and expressed as a proportion of the retrogradely labelled neurones. The proportions of double-labelled neurones for each of the P2X purinoceptor subtypes varied, on average, from 14 to 29%. The P2X3 purinoceptor subtype was found to be the dominant purinoceptor subtype present on PVN neurones projecting to the RVLM. Additionally it was apparent that more than one P2X purinoceptor subtype was present on the PVN neurones projecting to the RVLM, since the sum of the average percentages of double-labelled neurones for each P2X purinoceptor subtype exceeded 100%. These findings highlight the presence of the P2X1-P2X6 purinoceptors on PVN neurones projecting to the RVLM. The results suggest a potential role for ATP in the PVN in the regulation of sympathetic nerve activity.
Assuntos
Bulbo/metabolismo , Neurônios/metabolismo , Núcleo Hipotalâmico Paraventricular/metabolismo , Receptores Purinérgicos P2/análise , Animais , Masculino , Bulbo/química , Vias Neurais/química , Vias Neurais/metabolismo , Neurônios/química , Núcleo Hipotalâmico Paraventricular/química , Ratos , Ratos Sprague-Dawley , Receptores Purinérgicos P2X , Receptores Purinérgicos P2X2 , Receptores Purinérgicos P2X3 , Receptores Purinérgicos P2X4 , Receptores Purinérgicos P2X5 , Transmissão Sináptica/fisiologiaRESUMO
The present study investigated the effect of acute thermal stimulation in conscious rats on the production of Fos, a marker of increased neuronal activity, in spinally projecting and nitrergic neurons in the hypothalamic paraventricular nucleus (PVN). The PVN contains a high concentration of nitrergic neurons, as well as neurons that project to the intermediolateral cell column (IML) of the spinal cord that can directly influence sympathetic nerve activity (SNA). During thermal stimulation, the PVN is activated, but it is unknown whether spinally projecting PVN neurons and the nitrergic neurons are involved. Compared with controls, rats exposed to an environmental temperature of 39 degrees C for 1 h had a 10-fold increase in the number of cells producing Fos in the PVN (133 +/- 23 vs. 1,336 +/- 43, respectively, P < 0.0001). Of the spinally projecting neurons in the PVN of heated rats (98 +/- 10), over 20% expressed Fos. Additionally, of the nitrergic neurons (NADPH-diaphorase positive) located in the parvocellular PVN (723 +/- 17), 40% also expressed Fos (P < 0.0001 compared with controls). Finally, there was a significant increase in the number of spinally projecting neurons in the PVN that were nitrergic and expressed Fos after heat exposure (12%) compared with controls (0.1%) (P < 0.0001). These results suggest that spinally projecting and nitrergic neurons in the PVN may contribute to the central pathways activated by thermal stimulation.