BIO-NAIL: a bacterial cellulose dressing as a new alternative to preserve the nail bed after avulsion.

Avulsion of the nail plate is the most accomplished surgical procedure among the nail apparatus surgeries. Since it is not possible to use the removed nail to cover the nail bed, some materials have been suggested, however, they are generally not available for use and often at a high cost. To evaluate the use of Bacterial Cellulose (BC) dressing as a biological nail (Bio-Nail) after partial or total avulsion of the nail plate. Twenty-six candidates for nail avulsion, were randomized into two groups: Control, using Vaseline with gauze (11 patients) and Experimental group, BC group, using the Bio-Nail (15 patients). The patients were followed up during the 180-day period. The distribution of the patients in the groups was homogeneous for both sociodemographic and clinical data. The occurrence of infection (1 case in the control group) was not statistically relevant. The BC group had lower pain intensity (p = 0.011) with earlier temporal resolution when compared to the control group (p = 0.003). The BC group presented earlier reepithelization (p = 0.022) and better quantitative (p = 0.021) and qualitative conditions (p = 0.011) for the exudate. Regarding satisfaction, all the patients were satisfied. Good preservation of the nail plate area was observed in the BC group at the end of the 180-day period (p = 0.024). Average time of BC dressing permanence was 16.4 ± 7.1 days. BC showed to be appropriate as a dressing after partial or total avulsion of the nail plate. BC is a Bio-Nail promising for nail bed healing.

Training and practice of cytotechnologists: a discussion forum focused on Europe.

OBJECTIVES: To discuss the role and training of cytotechnologists (CTs) in Europe, to identify areas of good practice and to provide an informed opinion to those providing guidelines for training and practice in Europe. METHODS: All members of the Editorial Advisory Board of Cytopathology were invited to take part in a 'discussion forum' for which six topics were circulated in advance concerning the roles of CTs with regard to: (1) pre-screening slides; (2) 'signing out' reports; (3) carrying out ancillary techniques; (4) supervising laboratory staff; (5) taking part in rapid on-site evaluation (ROSE) of fine needle aspirates (FNAs); and (6) whether CTs were trained specifically in cytopathology or in general histopathology. Notes of the meeting were circulated by email and a final report was agreed by 22 participants from 17 predominantly European countries. RESULTS: Training for CTs throughout Europe was variable, especially for non-gynaecological cytology, which was inconsistent with the range of activities required. The participants recommended graduate entry, preliminary training in general laboratory technology, and subsequent training to take account of the probability and, in some centres, the reality of primary cervical cancer screening changing from cytology to human papillomavirus (HPV) testing. They further recommended that CTs should perform HPV tests and take part in ROSE for FNAs, and they supported the European Federation of Cytology Societies developing guidelines for training and practice. CONCLUSION: With CT training added to a university-based education in laboratory or biomedical science, a career in cytotechnology should be an attractive option involving a diverse range of laboratory and clinically based activities.

Fish oil supplementation in obese rats ameliorates metabolic syndrome response.

Accumulation of visceral adipose tissue is associated with metabolic syndrome (MS), insulin resistance, and dyslipidemia. Here we examined several morphometric and biochemical parameters linked to MS in a rodent litter size reduction model, and how a 30-day fish oil (FO) supplementation affected these parameters. On day 3 post-birth, pups were divided into groups of ten or three. On day 22, rats were split into control (C) and small litter (SL) until 60 days old. Then, after metabolic disturbance and obesity were confirmed, FO supplementation started for 30 days and the new groups were named control (C), FO supplemented (FO), obese (Ob), and obese FO supplemented (ObFO). Comparison was performed by Student t-test or 2-way ANOVA followed by Tukey post hoc test. At the end of the 60-day period, SL rats were hyperphagic, obese, hypoinsulinemic, normoglycemic, and had high visceral fat depot and high interleukin (IL)-6 plasma concentration. Obese rats at 90 days of age were fatter, hyperphagic, hyperglycemic, hypertriacylgliceromic, hipoinsulinemic, with low innate immune response. IL-6 production ex vivo was higher, but in plasma it was not different from the control group. FO supplementation brought all biochemical changes to normal values, normalized food intake, and reduced body weight and fat mass in obese rats. The innate immune response was improved but still not as efficient as in lean animals. Our results suggested that as soon MS appears, FO supplementation must be used to ameliorate the morpho- and biochemical effects caused by MS and improve the innate immune response.

Effects of depot growth hormone replacement on thyroid function and volume in adults with congenital isolated growth hormone deficiency.

BACKGROUND: Conflicting data exist on the effects of GH replacement therapy (GHRT) on thyroid function and thyroid volume (TV) in GH-deficient (GHD) patients. AIM: The aim of this study was to assess the effects of GHRT on thyroid function and TV in adults with congenital lifetime isolated GHD (IGHD). SUBJECTS AND METHODS: We studied 20 GH-naïve adults with IGHD due to a homozygous mutation of the GHRH-receptor gene at baseline, after 6-month depot- GH replacement therapy (pGH), and 6-month washout (6mo). Total T(3), free T(4) (FT(4)), reverse T(3) (rT(3)), TSH, IGF-I, SHBG, and TV were measured; body surface area-corrected TV (CTV) was calculated. RESULTS: IGF-I and T(3) increased pGH. T(3) levels remained elevated at 6mo. GHRT did not significantly change FT(4), rT(3), TSH, and SHBG. TV and CTV increased pGH and remained elevated at 6mo. CONCLUSIONS: GHRT in IGHD adults caused an increase in serum T(3) levels and TV, suggesting an important role of the GH-IGF-I axis in thyroid function.

Fish oil supplementation in obese rats ameliorates metabolic syndrome response

Accumulation of visceral adipose tissue is associated with metabolic syndrome (MS), insulin resistance, and dyslipidemia. Here we examined several morphometric and biochemical parameters linked to MS in a rodent litter size reduction model, and how a 30-day fish oil (FO) supplementation affected these parameters. On day 3 post-birth, pups were divided into groups of ten or three. On day 22, rats were split into control (C) and small litter (SL) until 60 days old. Then, after metabolic disturbance and obesity were confirmed, FO supplementation started for 30 days and the new groups were named control (C), FO supplemented (FO), obese (Ob), and obese FO supplemented (ObFO). Comparison was performed by Student t-test or 2-way ANOVA followed by Tukey post hoc test. At the end of the 60-day period, SL rats were hyperphagic, obese, hypoinsulinemic, normoglycemic, and had high visceral fat depot and high interleukin (IL)-6 plasma concentration. Obese rats at 90 days of age were fatter, hyperphagic, hyperglycemic, hypertriacylgliceromic, hipoinsulinemic, with low innate immune response. IL-6 production ex vivo was higher, but in plasma it was not different from the control group. FO supplementation brought all biochemical changes to normal values, normalized food intake, and reduced body weight and fat mass in obese rats. The innate immune response was improved but still not as efficient as in lean animals. Our results suggested that as soon MS appears, FO supplementation must be used to ameliorate the morpho- and biochemical effects caused by MS and improve the innate immune response.

Natural killer cell subpopulations in putative resistant individuals and patients with active Mycobacterium tuberculosis infection.

Herein, we intended to perform flow-cytometric analyses of peripheral blood NK-cell subsets in patients with active tuberculosis (TB) and those putative resistant subjects displaying positive tuberculin skin test (TST+) and compared with TST- healthy controls. Our findings demonstrated distinct phenotypic features in TST+ as compared with TB. While lower values of NK-cells with increased frequency of CD3-CD16+ CD56- and CD3-CD16-CD56+ subsets besides lower frequency of CD3-CD16+ CD56+ NK-cells was observed in TST+, unaltered levels of NK-cells with increased levels of CD3-CD16+ CD56- NK-cells with lower frequency of CD3-CD16+ CD56+ NK-cells was found in TB. Additional analysis highlighted a shift towards increased levels of CD3-CD16-/+CD56bright NK-cells as the hallmark of TST+, whereas unaltered frequency was observed in TB. Increased levels of CD3+CD56+ cells were observed in both TST+ and TB. Further focusing on the monocyte/NK-cell network, we have reported that enhanced frequency of CD14+ CD16+ monocytes particularly observed in TST+. Outstanding were the distinct correlation profiles observed between CD3-CD16-CD56+ NK-cells and CD3+ CD56+ cells CD14+ CD16+ monocytes for TST+ and TB. These data suggested that high levels of CD3-CD16-CD56+ NK-cells aside CD14+ CD16+ monocytes as well as non-concurrent increment of CD3+ CD56+ cells, may be involved in protective mechanisms in putative tuberculosis-resistant individuals. On the other hand, the basal levels of macrophage-like monocytes despite its positive correlation with increased levels of CD3+ CD56+ cells may count for the lack of the protective immunity in patients with active tuberculosis. Further studies focusing on the cytokine profiling of peripheral blood innate immunity cells before and after chemotherapeutic treatment are currently under evaluation.

Annual assessment of the sea urchin (Paracentrotus lividus) humoral innate immune status: Tales from the north Portuguese coast.

Innate immune status of the sea urchin Paracentrotus lividus population from two different rocky shore beaches in the northern Portuguese coast was evaluated for a period of one year. Although some ecological studies regarding the effect of toxics on the immune parameters of the sea urchin were made in Portuguese waters, there is a current lack of knowledge concerning their immune status all over the year. In perspective of a changing ecosystem in these waters due to global warming and colonization of new species, it is important to assess the status of the major species living in the area. In this way, immune parameters such as total protein content, nitric oxide concentration, haemolytic activity, protease activity, lysozyme concentration and bactericidal activity were evaluated in the perivisceral coelomic fluid, and were correlated with the gonadal index of the population and water parameters. Also, the spawning period can upset some immune status parameters, and others such as haemolytic activity and bactericidal activity against Vibrio anguillarum, showed a clear correlation with the gonad maturation status. The knowledge of the basal immune status of the species could serve as ecological indicator of some stress agent or contaminant into the field; also, coelomic fluid is suggested as good quality marker to assess the immune status of sea urchins.

Immunological and microbiological periodontal profiles in isolated growth hormone deficiency.

BACKGROUND: Growth hormone (GH) has been identified as an important regulator of the immune response. We have previously shown that adults with isolated GH deficiency (IGHD) due to a mutation in the GH releasing hormone receptor (GHRHR) gene, have a greater chance of having periodontitis. However, the interaction of GH with periodontal tissues is still unknown, and this population has emerged as a unique model to investigate this issue. Therefore, we evaluated the microbiological and immunological periodontal profiles of such individuals. METHODS: Nineteen IGHD and 19 controls matched by age, sex, diabetes, and smoking status, were enrolled in this case-control study. Periodontal clinical parameters (probing depth [PD] and clinical attachment loss [AL]) were measured at six sites per tooth. Immune mediators (C-reactive protein, matrix metalloproteinase [MMP]-8, MMP-9, interleukin [IL]-1α, IL-6, IL-8, tumor necrosis factor [TNF]-α, adiponectin, and leptin) were analyzed by enzyme-linked immunosorbent assay (ELISA) in the gingival crevicular fluid (GCF) in four non-adjacent sites for each participant (two with PD ≤3 mm [shallow sites] and two with PD ≥7 mm or the worst PD found in the mouth [deep sites]). Bacterial quantification (Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia) of subgingival biofilm samples collected from these same sites was performed by quantitative real-time polymerase chain reaction (qPCR). RESULTS: IGHD individuals presented higher values of PD and AL, and increased levels of CRP, IL-8, MMP-8, and adiponectin in the GCF. Bacterial quantification did not identify differences between the two groups. CONCLUSION: IGHD alters the local immune response in periodontal pockets leading to greater attachment loss, and GH stands out as an important hormone to be evaluated in the pathogenesis of periodontitis.

Influence of the asymmetric excited state decay on coherent population trapping.

Electromagnetically induced transparency (EIT) is an optical phenomenon which allows a drastic modification of the optical properties of an atomic system by applying a control field. It has been largely studied in the last decades and nowadays we can find a huge number of experimental and theoretical related studies. Recently a similar phenomenon was also shown in quantum dot molecules (QDM), where the control field is replaced by the tunneling rate between quantum dots. Our results show that in the EIT regime, the optical properties of QDM and the atomic system are identical. However, here we show that in the strong probe field regime, i.e., "coherent population trapping" (CPT) regime, it appears a strong discrepancy on the optical properties of both systems. We show that the origin of such difference relies on the different decay rates of the excited state of the two systems, implying in a strong difference on their higher order nonlinear susceptibilities. Finally, we investigate the optical response of atom/QDM strongly coupled to a cavity mode. In particular, the QDM-cavity system has the advantage of allowing a better narrowing of the width of the dark state resonance in the CPT regime when compared with atom-cavity system.

Reproductive performance of red-winged tinamous (Rhynchotus rufescens) using different mating strategies in captivity - case report / [Desempenho reprodutivo de perdizes (Rhynchotus rufescens) utilizando diferentes estratégias de acasalamento em cativeiro - relato de caso]

The aim of this study was to evaluate the reproductive performance of red-winged tinamous in captivity using different mating systems during three breeding seasons. The data were collected from August to March during three breeding seasons of the species. Ten couples and four groups of red-winged tinamous were randomly selected for formation of the monogamous and polygynous mating system, respectively, with egg collection from both systems. The traits evaluated were number of eggs per female, egg breaking and egg fertilization. Data were statistically analyzed by the least square method and logistic regression. Animals in couples mating strategy were superior (P<0.05) in terms of number of eggs per female compared to the group mating strategy. High egg breaking rate (P<0.05) in the red-winged tinamous groups was observed. Eggs of animals in couples mating strategy were 5.24 and 11.5 more likely to be fertilized than those of group strategy in two breeding seasons. In general, the reproductive efficiency in terms of low egg breaking, high egg production and fertility was observed for red-winged tinamous from the monogamous mating system.

O objetivo deste estudo foi avaliar o desempenho reprodutivo dos perdizes de asas vermelhas em cativeiro utilizando diferentes sistemas de acasalamento durante três épocas de reprodução. Os dados foram coletados de agosto a março, durante três épocas de reprodução da espécie. Dez casais e quatro grupos de perdizes de asas vermelhas foram selecionados aleatoriamente para a formação do sistema de acasalamento monogâmico e poliginoso, respectivamente, com coleta de ovos de ambos os sistemas. Os traços avaliados foram o número de ovos por fêmea, a quebra dos ovos e a fertilização dos ovos. Os dados foram analisados estatisticamente pelo método dos mínimos quadrados e pela regressão logística. Os animais na estratégia de acasalamento de casais foram superiores (P<0,05) em termos de número de óvulos por fêmea, em comparação com a estratégia de acasalamento do grupo. Foi observada uma alta taxa de quebra de ovos (P<0,05) nos grupos perdizes de asas vermelhas. Ovos de animais em estratégia de acasalamento de casais foram 5,24 e 11,5 mais propensos a serem fertilizados do que os de estratégia de grupo em duas épocas de reprodução. Em geral, a eficiência reprodutiva em termos de baixa quebra de ovos, alta produção de ovos e fertilidade foi observada para os tinâmicos de asas vermelhas do sistema de acasalamento monogâmico.

GH response to hypoglycemia and clonidine in the GH-releasing hormone resistance syndrome.

GH secretion by the pituitary is the result of the balance between the stimulatory effect of GHRH and the inhibitory effect of SS. Patients with mutations in GHRH receptor (GHRH-R) gene (GHRH-R) offer a unique model to study the mechanism of action of different GH secretion stimuli. In the past, we have demonstrated a small but significant GH response to a GH secretagogue (GHRP-2) in a homogenous cohort of patients with severe GH deficiency (GHD) due to a homozygous null mutation in GHRH-R (IVS1+1G-->A). Now, we sought to determine if we could detect a GH response to hypoglycemia (ITT: insulin tolerance test) or clonidine (CL) in these patients. Nine young GHD subjects underwent both ITT and CL tests, and 2 additional subjects underwent only CL test. There was a small but significant GH increase during ITT, but not during CL test. These results indicate that a minimal albeit significant GH response to ITT can occur despite complete lack of GHRH-R function.

Postnatal cerebellar granule cells of the white rat (Rattus norvegicus): a quantitative study, using design-based stereology.

A stereological study was carried out on postnatal cerebellar granule cells of rats aged 6 and 10 days, for detecting whether and how much they would differ from those of young adult rats. The following parameters were estimated: number-weighted mean volume of the nucleus and of the soma; mean total surface area of the soma; mean absolute volumes per cell of total cytoplasm, mitochondria, Golgi apparatus, and cytosol; mean surface density of the rough endoplasmic reticulum (RER); mean total surface area of the RER. These values were compared between the two postnatal ages. In addition, those values were also analysed in comparison to the ones depicted in young adult rats (60 days), already published by our team, in order to detect similarities between them. It was noticed that, between 6 and 10 days, the mean surface density of the RER was the only parameter that did not change significantly. The comparison of each of the postnatal ages with 60 days revealed that, with the exception of the absolute volume of Golgi apparatus, significant differences were displayed concerning other organelles and cellular compartments. It was concluded that, although fine structural differences have been disclosed, from the stereological point of view postnatal granule cells at 10 days were practically similar to the young adult ones at 60 days. Some potential physiological implications have been considered.

Genetic parameters for weight, morphometry and oviposition of Africanized honeybee queens using simulated data / [Parâmetros genéticos para peso, morfometria e oviposição de abelhas rainhas africanizadas sob simulação]

This study aimed to estimate genetic parameters for simulated data of body weight (BW), abdominal width (AW), abdominal length (AL), and oviposition. Simulation was performed based on real data collected at apiaries in the region of Campo das Vertentes, Minas Gerais, Brazil. Genetic evaluations were performed using single- and two-trait models and (co)variance components were estimated by the restricted maximum likelihood method. The heritability for BW, AW, AL and oviposition were 0.54, 0.47, 0.31 and 0.66, respectively. Positive genetic correlations of high magnitude were obtained between BW and AW (0.80), BW and oviposition (0.69), AW and oviposition (0.82), and AL and oviposition (0.96). The genetic correlations between BW and AL (0.11) and between AW and AL (0.26) were considered moderate and low. In contrast, the phenotypic correlations were positive and high between BW and AW (0.97), BW and AL (0.96), and AW and AL (0.98). Phenotypic correlations of low magnitude and close to zero were obtained for oviposition with AL (0.02), AW (-0.02), and BW (-0.03). New studies involving these characteristics should be conducted on populations with biological data in order to evaluate the impact of selection on traits of economic interest.(AU)

Objetivou-se estimar parâmetros genéticos para dados simulados de peso corporal (PC), largura abdominal (LA), comprimento abdominal (CA) e oviposição (OV). A simulação foi conduzida com base em dados reais, coletados em apiários da região do Campo das Vertentes, Minas Gerais. As estimativas das análises genéticas foram realizadas por modelos uni e bicaracterísticos, sendo os componentes de (co) variância estimados pelo método da máxima verossimilhança restrita. As herdabilidades para PC, LA, CA e OV foram de 0,54, 0,47, 0,31 e 0,66 respectivamente. As correlações genéticas foram positivas e de alta magnitude para PC e LA (0,80), PC e OV (0,69), LA e OV (0,82) e CA e OV (0,96). Para PC e CA (0,11) e LA e CA (0,26), as correlações genéticas foram moderadas e de baixa magnitude. As correlações fenotípicas foram positivas e de alta magnitude para PC e LA (0,97), PC e CA (0,96) e LA e CA (0,98). Para OV e CA (0,02), OV e LA (-0,02) e OV e PC (-0,03), foram encontradas correlações fenotípicas de magnitude baixa e próximas de zero. Novos estudos devem ser realizados em populações com dados biológicos, a fim de se observar o impacto da seleção em características de interesse econômico.(AU)

Behavioral activities of white-lipped peccary (Tayassu pecari, Link, 1795) in a semi-intensive farming system in Brazil / [Atividades comportamentais de queixadas (Tayassu pecari, Link, 1795) em sistema semi-intensivo de criação no Brasil]

This study aimed to describe the main behaviors of white-lipped peccaries (Tayassu pecari) expressed in a semi-intensive production system in Brazil. The behavioral activities of two groups of peccaries allocated in two paddocks were assessed, totaling 17 adult animals and eventual pups in different developmental stages. The animals were visually observed using the ad libitum sampling and scan sample methods during the adaptation and experimental phases, respectively. The animals were evaluated in the morning, afternoon, and at night. The behavioral activities observed by ad libitum sampling comprised feeding, pool contact, moving, staying inside the handling area, resting, mud bathing, mutual rubbing, rubbing, aggression, sexual behavior, copulation, foraging, breastfeeding, sleeping, territorial marking, interactions, birthing, gnawing, and other behaviors. All behaviors observed by the ad libitum sampling method were confirmed in the experimental period by scan sample, except for territorial marking and birthing. The effects of time of day were statistically significant (p <0.05) for eating, pool contact, movement, staying inside the handling area, resting, mud bathing and sleeping. These results contribute to the understanding of instinctive animal habits and to the development of nutritional, environmental and health management protocols that meet the requirements of peccaries.(AU)

Objetivou-se descrever os principais comportamentos que os queixadas (Tayassu pecari) expressam em sistema de produção semi-intensivo no Brasil, bem como suas diferenças de frequências comportamentais em função do período do dia. Foram avaliadas as atividades comportamentais de dois grupos de queixadas alocados em dois piquetes, totalizando 17 animais adultos e eventuais filhotes em diferentes estágios comportamentais. Na fase adaptativa e experimental os animais foram observados visualmente pelos métodos ad libitum e scan sample, respectivamente. Os animais foram avaliados nos períodos da manhã, tarde e noite. As atividades comportamentais observadas pelo método ad libitum foram alimentação, contato com a piscina, movimentação, interior do brete de manejo, descanso, lameando, esfregação mútua, esfregar-se, agressão, comportamento sexual, cópula, fuçando, amamentação, dormindo, marcando território, interações, manilha, parição, roendo e outros comportamentos. Todos os comportamentos observados pelo método ad libitum foram confirmados no período experimental scan sample, exceto para as variáveis marcando território e parição. O efeito do período do dia foi estatisticamente significativo (P<0,05) sobre as frequências dos comportamentos alimentação, contato com a piscina, movimentação, interior do brete de manejo, descanso, lameando e dormindo. Os resultados contribuem para a compreensão dos hábitos instintivos dos animais e, com isso, colaborar com a elaboração de protocolos de manejo nutricional, ambiental e sanitário, buscando-se suprir as necessidades dos queixadas. Futuras pesquisas envolvendo queixadas em cativeiro fomentarão os estudos voltados aos aspectos biológicos, conservacionistas e produtivos.(AU)

Growth hormone-releasing peptide-2 stimulates GH secretion in GH-deficient patients with mutated GH-releasing hormone receptor.

GH-releasing peptides (GHRPs) are synthetic peptides that bind to specific receptors and thereby stimulate the secretion of pituitary GH. In vivo it is uncertain whether these peptides act directly on somatotroph cells or indirectly via release of GHRH from the hypothalamus. In this study we compared the pituitary hormone response to GHRP-2 in 11 individuals with isolated GH deficiency (GHD) due to a homozygous mutation of the GHRH receptor (GHRH-R) gene and in 8 normal unrelated controls. Basal serum GH levels were lower in the GHD group compared with controls [0.11 +/- 0.11 (range, <0.04 to 0.38) vs. 0.59 +/- 0.76 microg/L (range, 0.04-2.12 microg/L); P = 0.052]. After GHRP-2 administration there was a 4.5-fold increase in serum GH relative to baseline values in the GHD group (0.49 +/- 0.41 vs. 0.11 +/- 0.11 microg/L; P = 0.002), which was significantly less than the 79-fold increase in the control group (46.8 +/- 17.6 vs. 0.59 +/- 0.76 microg/L; P = 0.008). Basal and post-GHRP-2 serum levels of ACTH, cortisol, and PRL were similar in both groups. Basal levels of serum TSH were significantly higher in the GHD group than in the control group (3.23 +/- 2.21 vs. 1.37 +/- 0.34 microIU/mL; P = 0.003). TSH levels in both groups did not change after GHRP-2 administration. These results suggest that an intact GHRH signaling system is not an absolute requirement for GHRP-2 action on GH secretion and that GHRP-2 has a GHRH-independent effect on pituitary somatotroph cells.

Familial dwarfism due to a novel mutation of the growth hormone-releasing hormone receptor gene.

Isolated growth hormone (GH) deficiency (IGHD) is a rare cause of short stature. The same mutation of the gene encoding the growth hormone-releasing hormone receptor (GHRHR) has been identified as the basis for IGHD in three families from the Indian subcontinent. The prevalence and heterogeneity of defects in the GHRHR gene are not known. Twenty-two dwarf members of a large, extended kindred containing at least 105 affected members with autosomal recessive short stature underwent extensive endocrine evaluation, which confirmed markedly reduced or undetectable serum concentrations of GH that did not increase in response to different stimuli. DNA sequences of the 13 exons and intron-exon boundaries of the GHRHR gene were determined in an index patient. A novel homozygous 5' splice site mutation (G-->A at position +1) in IVS1 was found. Thirty of the affected subjects tested were homozygous for this mutation, and 64 clinically unaffected patients were either heterozygous for the mutation (n = 41, including 9 obligate carriers) or homozygous for the wild-type sequence (n = 23). We describe a novel mutation in the GHRHR gene as cause of dwarfism in the largest kindred with familial IGHD described to date.

Effect of severe growth hormone (GH) deficiency due to a mutation in the GH-releasing hormone receptor on insulin-like growth factors (IGFs), IGF-binding proteins, and ternary complex formation throughout life.

Measurement of the insulin-like growth factors (IGFs) and their binding proteins has become commonplace in the indirect assessment of the integrity of the GH axis. However, the relative effect of GH deficiency (GHD) on each component of the IGF axis and the merit of any one parameter as a diagnostic test have not been defined in a homogeneous population across all ages. We therefore measured IGF-I, IGF-II, IGF-binding protein-1 (IGFBP-1), IGFBP-2, IGFBP-3, and acid labile subunit (ALS) in 27 GHD subjects (aged 5-82 yr) from an extended kindred in Northeast Brazil with an identical GHRH receptor mutation and in 55 indigenous controls (aged 5-80 yr). The effect of GHD on the theoretical distribution of IGFs between the IGFBPs and the ternary complex was also examined. All components of the IGF axis, measured and theoretical, showed complete separation between GHD and control subjects, except IGFBP-1 and IGFBP-2 concentrations, which did not differ. The most profound effects of GHD were on total IGF-I, IGF-I in the ternary complex, and ALS. The proportion of IGF-I associated with IGFBP-3 remained constant throughout life, but was significantly lower in GHD due to an increase in IGF-I/IGFBP-2 complexes. IGF-I in the ternary complex was determined principally by concentrations of ALS in GHD and IGFBP-3 in controls, implying that ALS has greater GH dependency. In the controls, IGF-II was associated primarily with IGFBP-3 and to a lesser extent with IGFBP-2, whereas in GHD the reverse was found. There was also a dramatic decline in the proportion of free ALS in GHD adults that was not evident in controls. As diagnostic tests, IGF-I in the ternary complex and total IGF-I provided the greatest separation between GHD and controls in childhood. Similarly, in older adults the best separation was achieved with IGF-I in the ternary complex, with free ALS being optimal in younger adults. Severe GHD not only reduces the amounts of IGFs, IGFBP-3, and ALS, but also modifies the distribution of the IGFs bound to each IGFBP. Diagnostic tests used in the investigation of GHD should be tailored to the age of the individual. In particular, measurement of IGF-I in the ternary complex may prove useful in the diagnosis of GHD in children and older adults, whereas free ALS may be more relevant to younger adults.

Quantitative age-changes in endoplasmic reticulum and nucleus of cerebellar granule cells.

A stereological study was performed on cerebellar granule cells from rats 2 to 24 months of age (eight different ages, five animals per age group) to quantify age-related alterations in the rough endoplasmic reticulum (RER). The mean surface density and the mean total surface area of the nucleus, as well as the mean absolute volume of euchromatin per cell, were also estimated to examine whether or not these had quantitative relationships with the RER. The mean surface density and the mean total surface area of RER per cell changed significantly, attaining maximum values at 24 months of 1733 microm(2)/1000 microm(3) (0.06) and 64 microm(2) (0.03), respectively, (coefficients of variation in parentheses). The corresponding values at 2 months were 706 microm(2)/1000 microm(3) (0.20) and 26 microm(2) (0.24). The mean absolute volume of the euchromatin changed significantly, with a minimum value of 57 microm(3) (0.05) occurring at 21 months. We postulate that the increase in RER may be part of a mechanism that compensates for an age-related decrease in euchromatin. An increase in the RER network may improve intracellular transport of proteins, production of which is apparently diminished with aging. The increase may also compensate for the reported decrease in calcium buffer capacity of smooth endoplasmic reticulum.

Age-related changes in the volume of somata and organelles of cerebellar granule cells.

Because cerebellar granule cells are fixed post-mitotic cells, it is expected that they undergo age-related changes like other neurons. To examine this possibility, a stereological study on granule cells of rat neocerebellar cortex was performed for an age spectrum of 2 to 24 months using eight different age groups. The nucleator method, together with point and intersection counting, was used to obtain primary data; arithmetical calculations determined the secondary data. In the soma, the absolute surface area did not change significantly; the volume did, however, exhibit a significant negative linear trend with age. Excluding dense bodies, the absolute volumes of the cytoplasmic components did not vary significantly. The absolute volume of dense bodies displayed a significant positive linear trend with age. Significant positive correlations were detected between the somatic volume and the absolute volume of either mitochondria or ground substance. It was concluded that granule cells showed a fair degree of morphological stability through 18 months. However, the observed changes warn that accompanying physiological alterations may occur, with putative effects on motor coordination.

The hepatocytes of the brown trout (Salmo trutta f. fario): a quantitative study using design-based stereology.

A stereological study was performed on brown trout hepatocytes aiming to disclose whether there are basic gender differences when minimal levels of sex hormones exist, and also to establish a platform for both interspecific comparisons and physiological correlations. We used the so-called "design-based stereology" (with no shape, size or orientation assumptions) and also some new related statistics. Two-year-old brown trout were collected in April, and the livers were fixed by perfusion. From liver slicing to microscopical field selection, systematic sampling was used. Stereology was applied at light and electron microscopy. Target parameters were the relative and total hepatocyte number, the mean individual hepatocyte volume and surface, and also both relative and total volumes, and surfaces, either of organelles or of cell compartments. Observed variability was usually high, but the precision of estimates was proved to be globally adequate facing the true biological variation amongst specimens. Females had more hepatocytes per liver (1.79x10(9) vs. 1.12x10(9)). Considering the individual hepatocytes, whereas no gender differences were detected in the cell volume, males had higher values of nuclear volume (199 vs. 151 microm3) and surface (170 vs. 131 microm2), endoplasmic reticulum volume (1,300 vs. 824 microm3), and microvilli volume (82 vs. 54 microm3) and surface (1,445 vs. 975 microm2). However, when dealing with quantities per liver, gender differences were found only in the volumes of dense bodies (56 vs. 97 mm3) and of residual cytoplasm (169 vs. 341 mm3)--both volumes were higher in females. Functional implications of data are discussed, namely that females seem to have basic structural traits for coping with the later demands of breeding. Data also support that structural remodelling of hepatocytes occurs after breeding, urging to pursue seasonal studies (namely on lysosomes). We advanced the hypothesis that genders differ in microvilli surface just to maintain an optimal physiological surface-to-volume ratio. Interspecific similarities and differences were disclosed. For example, the number of hepatocytes/cm3 of parenchyma of brown trout was much lower than those reported in rainbow trout, but in both trouts females seem to have an higher cell number. In addition, when comparing the size of hepatocytes of brown trout with that from other fish and mammals it was suggested that major interspecific differences exist.