Inhibition of early tumor growth requires Jα18-positive (natural killer T) cells

The role of natural killer T (NKT) cells in the immune response to tumor cells has been largely unexplored. As a model of adoptive tumor immunotherapy, cells from the draining lymph nodes of mice immunized with a tumor-specific or irrelevant antigen were transferred to naive recipients with established tumor. Inhibition of early tumor growth (day 4) required the transfer of both CD8(+) and Jalpha18(+) (NKT) cells from immunized animals without regard to immunogen. In contrast, CD8(+) cells, but not Jalpha18(+) cells, were necessary for the inhibition of late tumor growth (day 8). Thus, the developing tumor changes in sensitivity to NKT-mediated events and the role for NKT cells cannot be replaced by the presence of tumor-specific cells during early tumor growth. This suggests that recruitment/activation of Jalpha18(+) NKT cells is an important consideration during the immune therapy of early stage tumors.

Part 1. Growth, carcass and meat quality parameters of male goats: effects of genotype and liveweight at slaughter

Male kids (110) from six goat genotypes, i.e. Boer x Angora (BA), Boer x Feral (1317), Boer x Saanen (BS), Feral x Feral (FF), Saanen x Angora (SA) and Saanen x Feral (SF) and two slaughter weight groups, i.e. Capretto and Chevon (liveweight at slaughter 14-22 and 30-35 kg, respectively) were compared for growth, carcass and meat quality characteristics. Due to their better growth rate, kids from BS and SF genotypes reached the required liveweight for slaughter earlier than kids from other Genotypes used in the study. Chevon kids had a significantly (P < 0.05) lower average daily gain (119 g per day) compared to Capretto kids (171 g per day). SA, SF and FF kids deposited more internal fat in comparison to kids from other genotypes. The dressing percentage of kids ranged from 51 to 54%, with significant differences between genotypes. BS and SF kids had longer carcasses. while BF kids had larger eye muscle area compared to other genotypes. Goat carcasses had a thin subcutaneous fat cover (1.6-2.2 mm). Genotype had a significant (P < 0.05) influence on cooking loss, pigment concentration and muscle colour parameters (CIE L*, a* and b* values). As denoted by the higher V and fibre optic probe values and lower subjective muscle score, the longissimus muscle colour was lighter for BS kids than other genotypes. Cooked meat from the BF kids had lower shear force values and better sensory scores compared to other genotypes. A significant (P < 0.05) decrease in muscle tenderness was observed from Capretto to Chevon carcasses, whereas cooked meat from these two slaughter weight groups was equally accepted (P > 0.05) by the panellists. (C) 2003 Elsevier Science B.V. All rights reserved.

Genetic disruption of the growth hormone receptor does not influence motoneuron survival in the developing mouse

In the rodent central nervous system (CNS) during the five days prior to birth, both growth hormone (GH) and its receptor (GHR) undergo transient increases in expression to levels considerably higher than those found postnatally. This increase in expression coincides with the period of neuronal programmed cell death (PCD) in the developing CNS. To evaluate the involvement of growth hormone in the process of PCD, we have quantified the number of motoneurons in the spinal cord and brain stem of wild type and littermate GHR-deficient mice at the beginning and end of the neuronal PCD period. We found no change in motoneuron survival in either the brachial or lumbar lateral motor columns of the spinal cord or in the trochlear, trigeminal, facial or hypoglossal nuclei in the brain stem. We also found no significant differences in spinal cord volume, muscle fiber diameter, or body weight of GHR-deficient fetal mice when compared to their littermate controls. Therefore, despite considerable in vitro evidence for GH action on neurons and glia, genetic disruption of GHR signalling has no effect on prenatal motoneuron number in the mouse, under normal physiological conditions. This may be a result of compensation by the signalling of other neurotrophic cytokines.

Constitutively active signal transducer and activator of transcription 5 can replace the requirement for growth hormone in adipogenesis of 3T3-F442A preadipocytes

Although it is the best characterized in vitro model of GH action, the mechanisms used by GH to induce differentiation of murine 3T3-F442A preadipocytes remain unclear. Here we have examined the role of three transcriptional regulators in adipogenesis. These regulators are either rapidly induced in response to GH [Stra13, signal transducer and activator of transcription (Stat) 3] or of central importance to GH signaling (Stat5). Retroviral transfection of 3T3-F442A preadipocytes was used to increase expression of Stra13, Stat3, and Stat5a. Only Stat5a transfection increased the expression of adipogenic markers peroxisome proliferator-activated receptor gamma, CCAAT enhancer binding protein (C/EBP)alpha, and adipose protein 2/fatty acid-binding protein in response to GH, as determined by quantitative RT-PCR. Transfection with constitutively active Stat3 and Stat5a revealed that constitutively active Stat5a but not Stat3 was able to replace the GH requirement for adipogenesis. Constitutively active Stat5a but not Stat3 was able to increase the formation of lipid droplets and expression of alpha-glycerol phosphate dehydrogenase toward levels seen in mature adipocytes. Constitutively active Stat5a was also able to increase the expression of transcripts for C/EBPalpha to similar levels as GH, and of C/EBPbeta, peroxisome proliferator-activated receptor gamma, and adipose protein 2/fatty acid-binding protein transcripts to a lesser extent. An in vivo role for GH in murine adipogenesis is supported by significantly decreased epididymal fat depot size in young GH receptor-deleted mice, before manifestation of the lipolytic actions of GH. We conclude that Stat5 is a critical factor in GH-induced, and potentially prolactin-induced, murine adipogenesis.

Epitope map for a growth hormone receptor agonist monoclonal antibody, MAb 263

Monoclonal antibody (MAb) 263 is a widely used monoclonal antibody that recognizes the extracellular domain (ECD) of the GH receptor. It has been shown to act as a GH agonist both in vitro and in vivo, and we report here that it must be divalent to exert its effect on the full-length receptor. To understand the mechanism of its agonist action, we have determined the precise epitope for this antibody using a novel random PCR mutagenesis approach together with expression screening in yeast. A library of 5200 clones of rabbit GH receptor ECD mutants were screened both with MAb 263 and with an anticarboxy-tag antibody to verify complete ECD expression. Sequencing for clones that expressed complete ECD but were not MAb 263 positive identified 20 epitope residues distributed in a discontinuous manner throughout the ECD. The major part of the epitope, as revealed after mapping onto the crystal structure model of the ECD molecule, was located on the side and upper portion of domain 1, particularly within the D - E strand disulfide loop 79 - 96. Molecular dynamics docking of an antibody of the same isotype as MAb 263 was used to dock the bivalent antibody to the 1528-Angstrom(2) epitope and to visualize the likely consequences of MAb binding. The minimized model enables the antibody to grasp two receptors in a pincer-like movement from opposite sides, facilitating alignment of the receptor dimerization domains in a manner similar to, but not identical with, GH.

Human melanoblasts in culture: Expression of BRN2 and synergistic regulation by fibroblast growth factor-2, stem cell factor, and endothelin-3

The BRN2 transcription factor (POU3F2, N-Oct-3) has been implicated in development of the melanocytic lineage and in melanoma. Using a low calcium medium supplemented with stem cell factor, fibroblast growth factor-2, endothelin-3 and cholera toxin, we have established and partially characterised human melanocyte precursor cells, which are unpigmented, contain immature melanosomes and lack L-dihydroxyphenylalanine reactivity. Melanoblast cultures expressed high levels of BRN2 compared to melanocytes, which decreased to a level similar to that of melanocytes when cultured in medium that contained phorbol ester but lacked endothelin-3, stem cell factor and fibroblast growth factor-2. This decrease in BRN2 accompanied a positive L-dihydroxyphenylalanine reaction and induction of melanosome maturation consistent with melanoblast differentiation seen during development. Culture of primary melanocytes in low calcium medium supplemented with stem cell factor, fibroblast growth factor-2 and endothelin-3 caused an increase in BRN2 protein levels with a concomitant change to a melanoblast-like morphology. Synergism between any two of these growth factors was required for BRN2 protein induction, whereas all three factors were required to alter melanocyte morphology and for maximal BRN2 protein expression. These finding implicate BRN2 as an early marker of melanoblasts that may contribute to the hierarchy of melanocytic gene control.

Differences in lung function, growth parameters and frequency of hospital admissions between adolescents with cystic fibrosis-related diabetes and controls

As survival of patients with CF increases,glucose intolerance and cystic fibrosisrelated diabetes (CFRD),ar e increasingly recognised common complications. CFRD may be preceded by a pre-diabetic state. Using markers identified as being associated with CFRD may improve targeted screening. Aim: To identify features consistently predicting CFRD in paediatric patients. Patients diagnosed with CFRD between January 1997–January 2002 were compared with age and sex matched controls. Clinical,micr obiological, and hospitalisation data was collected at time of CFRD diagnosis,and at six monthly intervals for 3 yr prior to diagnosis. Eight patients with CFRD were identified,mean age 13.7 yr (S.D. 3.49) at time of diagnosis. Control patients underwent OGTT to ensure normal glucose tolerance. Patients with CFRD had a lower FEV1 up to 12 months prior to diagnosis however, this was only significant at diagnosis. There was no difference in weight and height z scores between the 2 groups; however,the decrease in weight and height z scores in the CFRD group over 3 yr prior to diagnosis was significant. Mean number of days in hospital and admissions per patient significantly increased in the CFRD group,6 months prior to diagnosis. No other significant differences were observed between the 2 groups. Conclusions: This study has shown a difference in lung function,gr owth parameters and frequency of hospital admissions between patients with CFRD and controls. These differences may be utilised as tools for targeted screening in the paediatricyadolescent population. Further larger scale studies are required to improve guidelines for targeted screening in this population.

Use of confocal scanning laser microscopy to measure the concentrations of aerial and penetrative hyphae during growth of Rhizopus oligosporus on a solid surface

In order to develop a method for use in investigations of spatial biomass distribution in solid-state fermentation systems, confocal scanning laser microscopy was used to determine the concentrations of aerial and penetrative biomass against height and depth above and below the substrate surface, during growth of Rhizopus oligosporus on potato dextrose agar. Penetrative hyphae had penetrated to a depth of 0.445 cm by 64 h and showed rhizoid morphology, in which the maximum biomass concentration, of 4.45 mg dry wt cm(-3), occurred at a depth of 0.075 cm. For aerial biomass the maximum density of 39.54 mg dry wt(-3) occurred at the substrate surface. For both aerial and penetrative biomass, there were two distinct regions in which the biomass concentration decayed exponentially with distance from the surface. For aerial biomass, the first exponential decay region was up to 0.1 cm height. The second region above the height of 0.1 cm corresponded to that in which sporangiophores dominated. This work lays the foundation for deeper studies into what controls the growth of fungal hyphae above and below the surfaces of solid substrates. (C) Wiley Periodicals, Inc.

Use of confocal microscopy to follow the development of penetrative hyphae during growth of Rhizopus oligosporus in an artificial solid-state fermentation system

Two methods were compared for determining the concentration of penetrative biomass during growth of Rhizopus oligosporus on an artificial solid substrate consisting of an inert gel and starch as the sole source of carbon and energy. The first method was based on the use of a hand microtome to make sections of approximately 0.2- to 0.4-mm thickness parallel to the substrate surface and the determination of the glucosamine content in each slice. Use of glucosamine measurements to estimate biomass concentrations was shown to be problematic due to the large variations in glucosamine content with mycelial age. The second method was a novel method based on the use of confocal scanning laser microscopy to estimate the fractional volume occupied by the biomass. Although it is not simple to translate fractional volumes into dry weights of hyphae due to the lack of experimentally determined conversion factors, measurement of the fractional volumes in themselves is useful for characterizing fungal penetration into the substrate. Growth of penetrative biomass in the artificial model substrate showed two forms of growth with an indistinct mass in the region close to the substrate surface and a few hyphae penetrating perpendicularly to the surface in regions further away from the substrate surface. The biomass profiles against depth obtained from the confocal microscopy showed two linear regions on log-linear plots, which are possibly related to different oxygen availability at different depths within the substrate. Confocal microscopy has the potential to be a powerful tool in the investigation of fungal growth mechanisms in solid-state fermentation. (C) 2003 Wiley Periodicals, Inc.

A yield architecture framework to explain adaptation of pearl millet to environmental stress

Functional knowledge of the physiological basis of crop adaptation to stress is a prerequisite for exploiting specific adaptation to stress environments in breeding programs. This paper presents an analysis of yield components for pearl millet, to explain the specific adaptation of local landraces to stress environments in Rajasthan, India. Six genotypes, ranging from high-tillering traditional landraces to low-tillering open-pollinated modern cultivars, were grown in 20 experiments, covering a range of nonstress and drought stress patterns. In each experiment, yield components (particle number, grain number, 100 grain mass) were measured separately for main shoots, basal tillers, and nodal tillers. Under optimum conditions, landraces had a significantly lower grain yield than the cultivars, but no significant differences were observed at yield levels around 1 ton ha(-1). This genotype x environment interaction for grain yield was due to a difference in yield strategy, where landraces aimed at minimising the risk of a crop failure under stress conditions, and modem cultivars aimed at maximising yield potential under optimum conditions. A key aspect of the adaptation of landraces was the small size of the main shoot panicle, as it minimised (1) the loss of productive tillers during stem elongation; (2) the delay in anthesis if mid-season drought occurs; and (3) the reduction in panicle productivity of the basal tillers under stress. In addition, a low investment in structural panicle weight, relative to vegetative crop growth rate, promoted the production of nodal tillers, providing a mechanism to compensate for reduced basal tiller productivity if stress occurred around anthesis. A low maximum 100 grain mass also ensured individual grain mass was little affected by environmental conditions. The strategy of the high-tillering landraces carries a yield penalty under optimum conditions, but is expected to minimise the risk of a crop failure, particularly if mid-season drought stress occurs. The yield architecture of low-tillering varieties, by contrast, will be suited to end-of-season drought stress, provided anthesis is early. Application of the above adaptation mechanisms into a breeding program could enable the identification of plant types that match the prevalent stress patterns in the target environments. (C) 2003 E.J. van Oosterom. Published by Elsevier Science B.V. All rights reserved.

Relationships and estimates of longitudinal growth stress in Eucalyptus dunnii at different ages

O objetivo desse trabalho foi obter estimativas da tensão de crescimento longitudinal em árvores vivas e verificar a sua relação com algumas características da madeira de Eucalyptus dunnii Maiden aos oito, treze, quinze e dezenove anos de idade. O material foi proveniente da Empresa Procopiak Compensados e Embalagens S.A., localizada no Município de Canoinhas, Santa Catarina. Os níveis de tensão longitudinal de crescimento foram mensurados indiretamente pelo método do "CIRAD-Forêt", na árvore viva, e estimados a partir do módulo de elasticidade dinâmico e do módulo de elasticidade obtido no ensaio de tração paralela à grã. A deformação residual longitudinal (DRL) e as estimativas das tensões de crescimento longitudinais apresentaram tendência de aumento, na média, com a idade do material. A DRL apresentou correlação, positiva e significativa, com todas as estimativas das tensões de crescimento longitudinais, sendo de maior magnitude aos 13, 15 e 19 anos de idade. A densidade básica apresentou maiores correlações, positivas e significativas, com o módulo de elasticidade dinâmico, estimado no sentido longitudinal, para a madeira na condição de saturação e a 12% de umidade, em todas as idades avaliadas. Todas as estimativas das tensões de crescimento longitudinais apresentam elevadas correlações, positivas e significativas, entre si.

Leaf consumption and duration of instars of the cassava defoliator Erinnyis ello (L., 1758) (Lepidoptera, Sphingidae)

The objectives of this research were to evaluate leaf consumption and the developmental time of the larvae of Erynnyis ello (L., 1758) (Lepidoptera, Sphingidae) reared on cassava, in order to obtain information for the integrated management of this pest. The larvae were reared on excised cassava leaves in Petri dishes and later in gerbox, and kept in chambers at 24 ± 2 ºC and 75 ± 10% RH. The total leaf area consumed by the larva to complete its development was 589.67 cm2; each of the five instars consumed, respectively: 1.89 cm2; 5.74 cm2; 17.48 cm2; 76.66 cm2; and 487.90 cm2. The consumption by the first three instars was insignificant, and did not reach 5% altogether; the 4th represented 13%; the 5th presented a consumption significantly higher, about 82.7%. The total time for the larval development was 22.61 days, and the duration for each of the five larval instar was, respectively: 4.35; 3.19; 3.32; 4.52; and 4.94 days. The pre-pupal period lasted 2.29 days. Since the highest consumption is by the 5th instar larva, the control should be applied before this age to avoid heavier damages to the cassava crop.

Efeito da restrição hídrica sobre o crescimento de clones de eucalipto

O estabelecimento e o crescimento inicial de espécies florestais no campo são fortemente afetados pela disponibilidade de água no solo e pela época de plantio, por isso, o presente trabalho estuda o impacto do déficit hídrico no crescimento de mudas de dois clones do híbrido Eucalyptus grandis x Eucalyptus urophylla, ambos submetidos a 4 níveis de déficit hídrico, em duas épocas de plantio. O estudo foi realizado na área experimental do Núcleo de Estudos e Difusão de Tecnologia em Florestas, Recursos Hídricos e Agricultura Sustentável (NEDTEC), do Centro de Ciências Agrárias da Universidade Federal do Espírito Santo (CCA-UFES), localizado no município de Jerônimo Monteiro. O trabalho foi realizado em duas épocas distintas, sendo a primeira no período de 09 de fevereiro a 09 de junho de 2009 e a segunda no período de 11 de julho a 07 de novembro de 2009, visando à realização das observações em diferentes condições de regime de radiação, déficit de pressão do vapor do ar, temperatura, umidade relativa do ar e velocidade do vento. O delineamento experimental utilizado foi o inteiramente ao acaso em parcelas subdivididas 2 x 4, alocando-se os 4 níveis de déficits hídricos na parcela principal e as 2 épocas nas subparcelas, com três repetições. Os manejos hídricos aplicados foram: Déficit 0 (D0) sem déficit, Déficit 1(D1) corte da irrigação aos 30 dias de experimentação, permanecendo até o final do experimento, Déficit 2 (D2) corte da irrigação aos 30 dias de experimentação, suspensão da irrigação por 60 dias e posterior retomada da irrigação por mais 30 dias; Déficit 3 (D3) corte da irrigação aos 60 dias de experimentação, prolongando até o final do experimento. Os dados experimentais foram submetidos à análise de variância, e quando significativas, as médias foram comparadas pelo teste de média Tukey a 5% de probabilidade, para cada clone estudado. Com este trabalho, foi possível avaliar o impacto de diferentes déficits hídricos, no crescimento inicial das plantas, em duas épocas do ano e avaliar o incremento no desenvolvimento das plantas durante a aplicação dos tratamentos, com retiradas de amostras médias de cada tratamento a cada 30 dias. As variáveis medidas nos dois experimentos foram altura total da planta, diâmetro ao nível do coleto, número de folhas, área foliar, matéria seca de folhas, matéria seca de haste e ramos, matéria seca de raízes e matéria seca total. Foram avaliadas as variáveis climáticas durante todo o período experimental, nas duas épocas, a fim de determinar a condição do clima em cada época. Para os dois clones estudados, em geral, os déficits hídricos promoveram a redução das variáveis morfológicas estudadas e a época experimental foi o fator que mais influenciou a redução do crescimento das plantas. Sendo que a Época 1 foi a que proporcionou resultados superiores, e a Época 2 foi a que prejudicou mais o desenvolvimento das plantas, reduzindo significativamente todas as variáveis morfológicas em todos os déficits hídricos, inclusive o D0.

Efeito da restrição hídrica sobre o crescimento de clones de eucalipto

O estabelecimento e o crescimento inicial de espécies florestais no campo são fortemente afetados pela disponibilidade de água no solo e pela época de plantio, por isso, o presente trabalho estuda o impacto do déficit hídrico no crescimento de mudas de dois clones do híbrido Eucalyptus grandis x Eucalyptus urophylla, ambos submetidos a 4 níveis de déficit hídrico, em duas épocas de plantio. O estudo foi realizado na área experimental do Núcleo de Estudos e Difusão de Tecnologia em Florestas, Recursos Hídricos e Agricultura Sustentável (NEDTEC), do Centro de Ciências Agrárias da Universidade Federal do Espírito Santo (CCA-UFES), localizado no município de Jerônimo Monteiro. O trabalho foi realizado em duas épocas distintas, sendo a primeira no período de 09 de fevereiro a 09 de junho de 2009 e a segunda no período de 11 de julho a 07 de novembro de 2009, visando à realização das observações em diferentes condições de regime de radiação, déficit de pressão do vapor do ar, temperatura, umidade relativa do ar e velocidade do vento. O delineamento experimental utilizado foi o inteiramente ao acaso em parcelas subdivididas 2 x 4, alocando-se os 4 níveis de déficits hídricos na parcela principal e as 2 épocas nas subparcelas, com três repetições. Os manejos hídricos aplicados foram: Déficit 0 (D0) sem déficit, Déficit 1(D1) corte da irrigação aos 30 dias de experimentação, permanecendo até o final do experimento, Déficit 2 (D2) corte da irrigação aos 30 dias de experimentação, suspensão da irrigação por 60 dias e posterior retomada da irrigação por mais 30 dias; Déficit 3 (D3) corte da irrigação aos 60 dias de experimentação, prolongando até o final do experimento. Os dados experimentais foram submetidos à análise de variância, e quando significativas, as médias foram comparadas pelo teste de média Tukey a 5% de probabilidade, para cada clone estudado. Com este trabalho, foi possível avaliar o impacto de diferentes déficits hídricos, no crescimento inicial das plantas, em duas épocas do ano e avaliar o incremento no desenvolvimento das plantas durante a aplicação dos tratamentos, com retiradas de amostras médias de cada tratamento a cada 30 dias. As variáveis medidas nos dois experimentos foram altura total da planta, diâmetro ao nível do coleto, número de folhas, área foliar, matéria seca de folhas, matéria seca de haste e ramos, matéria seca de raízes e matéria seca total. Foram avaliadas as variáveis climáticas durante todo o período experimental, nas duas épocas, a fim de determinar a condição do clima em cada época. Para os dois clones estudados, em geral, os déficits hídricos promoveram a redução das variáveis morfológicas estudadas e a época experimental foi o fator que mais influenciou a redução do crescimento das plantas. Sendo que a Época 1 foi a que proporcionou resultados superiores, e a Época 2 foi a que prejudicou mais o desenvolvimento das plantas, reduzindo significativamente todas as variáveis morfológicas em todos os déficits hídricos, inclusive o D0.