Monitoring and isolation of blood dendritic cells from apheresis products in healthy individuals: a platform for cancer immunotherapy

The fundamental role of dendritic cells (DC in initiating and directing the primary immune response is well established. Furthermore, it is now accepted that DC may be useful in new vaccination strategies for preventing certain malignant and infectious diseases. As blood DC (BDC physiology differs from that of the DC homologues generated in vitro from monocyte precursors, it is becoming more relevant to consider BDC for therapeutic interventions. Until recently, protocols for the isolation of BDC were laborious and inefficient; therefore, their use for investigative cancer immunotherapy is not widespread. In this study, we carefully documented BDC counts, yields and subsets during apheresis (Cobe Spectra), the initial and essential procedure in creating a BDC isolation platform for cancer immunotherapy. We established that an automated software package (Version 6,0 AutoPBPC) provides an operator-independent reliable source of motionuclear cells (MNC for BDC preparation. Further, we observed that BDC might be recovered in high yields, often greater than 100% relative to the number of circulating BDC predicted by blood volume. An average of 66 million (range, 17-179) BDC per 10-1 procedure were obtained, largely satisfying the needs for immunization. Higher yields were possible on total processed blood volumes of 151. BDC were not activated by the isolation procedure and, more importantly, both BDC subsets (CD11c(+)CD123(low) and CD11c(-)CD123(high)) were equally represented. Finally, we established that the apheresis product could be used for antibody-based BDC immunoselection and demonstrated that fully functional BDC can be obtained by this procedure. (C) 2002 Published by Elsevier Science B.V.

A subset-oriented algorithm for minimizing the number of steps required for synthesis of cyclic-peptide libraries

Libraries of cyclic peptides are being synthesized using combinatorial chemistry for high throughput screening in the drug discovery process. This paper describes the min_syn_steps.cpp program (available at http://www.imb.uq.edu.au/groups/smythe/tran), which after inputting a list of cyclic peptides to be synthesized, removes cyclic redundant sequences and calculates synthetic strategies which minimize the synthetic steps as well as the reagent requirements. The synthetic steps and reagent requirements could be minimized by finding common subsets within the sequences for block synthesis. Since a brute-force approach to search for optimum synthetic strategies is impractically large, a subset-orientated approach is utilized here to limit the size of the search. (C) 2002 Elsevier Science Ltd. All rights reserved.

delta N-15 values of tropical savanna and monsoon forest species reflect root specialisations and soil nitrogen status

A large number of herbaceous and woody plants from tropical woodland, savanna, and monsoon forest were analysed to determine the impact of environmental factors (nutrient and water availability, fire) and biological factors (microbial associations, systematics) on plant delta(15)N values. Foliar delta(15)N values of herbaceous and woody species were not related to growth form or phenology, but a strong relationship existed between mycorrhizal status and plant delta(15)N. In woodland and savanna, woody species with ectomycorrhizal (ECM) associations and putative N-2-fixing species with ECM/arbuscular (AM) associations had lowest foliar delta(15)N values (1.0-0.6parts per thousand), AM species had mostly intermediate delta(15)N values (average +0.6parts per thousand), while non-mycorrhizal Proteaceae had highest delta(15)N values (+2.9 to +4.1parts per thousand). Similar differences in foliar delta(15)N were observed between AM (average 0.1 and 0.2parts per thousand) and non-mycorrhizal (average +0.8 and +0.3parts per thousand) herbaceous species in woodland and savanna. Leguminous savanna species had significantly higher leaf N contents (1.8-2.5% N) than non-fixing species (0.9-1.2% N) indicating substantial N acquisition via N-2 fixation. Monsoon forest species had similar leaf N contents (average 2.4% N) and positive delta(15)N values (+0.9 to +2.4parts per thousand). Soil nitrification and plant NO3- use was substantially higher in monsoon forest than in woodland or savanna. In the studied communities, higher soil N content and nitrification rates were associated with more positive soil delta(15)N and plant delta(15)N. In support of this notion, Ficus, a high NO3- using taxa associated with NO3- rich sites in the savanna, had the highest delta(15)N values of all AM species in the savanna. delta(15)N of xylem sap was examined as a tool for studying plant delta(15)N relations. delta(15)N of xylem sap varied seasonally and between differently aged Acacia and other savanna species. Plants from annually burnt savanna had significantly higher delta(15)N values compared to plants from less frequently burnt savanna, suggesting that foliar N-15 natural abundance could be used as marker for assessing historic fire regimes. Australian woodland and savanna species had low leaf delta(15)N and N content compared to species from equivalent African communities indicating that Australian biota are the more N depauperate. The largest differences in leaf delta(15)N occurred between the dominant ECM Australian and African savanna (miombo) species, which were depleted and enriched in N-15, respectively. While the depleted delta(15)N of Australian ECM species are similar to those of previous reports on ECM species in natural plant communities, the N-15-enriched delta(15)N of African ECM species represent an anomaly.

Effect of changing data collection parameters on statistical motor unit number estimates

The effect of number of samples and selection of data for analysis on the calculation of surface motor unit potential (SMUP) size in the statistical method of motor unit number estimates (MUNE) was determined in 10 normal subjects and 10 with amyotrophic lateral sclerosis (ALS). We recorded 500 sequential compound muscle action potentials (CMAPs) at three different stable stimulus intensities (10–50% of maximal CMAP). Estimated mean SMUP sizes were calculated using Poisson statistical assumptions from the variance of 500 sequential CMAP obtained at each stimulus intensity. The results with the 500 data points were compared with smaller subsets from the same data set. The results using a range of 50–80% of the 500 data points were compared with the full 500. The effect of restricting analysis to data between 5–20% of the CMAP and to standard deviation limits was also assessed. No differences in mean SMUP size were found with stimulus intensity or use of different ranges of data. Consistency was improved with a greater sample number. Data within 5% of CMAP size gave both increased consistency and reduced mean SMUP size in many subjects, but excluded valid responses present at that stimulus intensity. These changes were more prominent in ALS patients in whom the presence of isolated SMUP responses was a striking difference from normal subjects. Noise, spurious data, and large SMUP limited the Poisson assumptions. When these factors are considered, consistent statistical MUNE can be calculated from a continuous sequence of data points. A 2 to 2.5 SD or 10% window are reasonable methods of limiting data for analysis. Muscle Nerve 27: 320–331, 2003

Distribution of Chlamydia pneumoniae DNA in atherosclerotic carotid arteries: significance for sampling procedures

Despite extensive efforts to confirm a direct association between Chlamydia pneumoniae and atherosclerosis, different laboratories continue to report a large variability in detection rates. In this study, we analyzed multiple sections from atherosclerotic carotid arteries from 10 endartectomy patients to determine the location of C. pneumoniae DNA and the number of sections of the plaque required for analysis to obtain a 95% confidence of detecting the bacterium. A sensitive nested PCR assay detected C. pneumoniae DNA in all patients at one or more locations within the plaque. On average, 42% (ranging from 5 to 91%) of the sections from any single patient had C. pneumoniae DNA present. A patchy distribution of C. pneumoniae in the atherosclerotic lesions was observed, with no area of the carotid having significantly more C. pneumoniae DNA present. If a single random 30-mum-thick section was tested, there was only a 35.6 to 41.6% (95% confidence interval) chance of detecting C. pneumoniae DNA in a patient with carotid artery disease. A minimum of 15 sections would therefore be required to obtain a 95% chance of detecting all true positives. The low concentration and patchy distribution of C. pneumoniae DNA in atherosclerotic plaque appear to be among the reasons for inconsistency between laboratories in the results reported.

A randomized trial of aggressive lipid reduction for improvement of myocardial ischemia, symptom status, and vascular function in patients with coronary artery disease not amenable to intervention

PURPOSE: To determine the effects of aggressive lipid lowering on markers of ischemia, resistance vessel function, atherosclerotic burden, and Symptom status in patients with symptomatic coronary artery disease. METHODS: Sixty consecutive patients with coronary artery disease that was unsuitable for revascularization were assigned randomly to either usual therapy of lipids for patients with a low-density lipoprotein (LDL) cholesterol target level <116 mg/dL, or to a, more aggressive lipid-lowering strategy involving up to 80 mg/d of atorvastatin, with a target LDL cholesterol level <77 mg/dL. The extent and severity of inducible ischemia (by dobutamine echocardiography), vascular function.(brachial artery reactivity), atheroma burden (carotid intima-media thickness), and symptom status were evaluated blindly at baseline and after 12 weeks of treatment. RESULTS: After 12 weeks of treatment, patients in the aggressive therapy group had a significantly greater decrease in mean (+/- SD) LDL cholesterol level than those in the usual care group (29 +/- 38 mg/dL vs. 7 +/- 24 mg/dL, P = 0.03). Patients in the aggressive therapy group had a reduction in the number of ischemic wall segments (mean between-group difference of 1.3; 95% confidence interval: 0.1 to 2.0; P = 0.04), flow-mediated dilatation (mean between-group difference of 5.9%; 95% confidence interval: 2.5% to 9.4%; P = 0.001), and angina score after 12 weeks. There were no significant changes in atherosclerotic burden in either group. CONCLUSION: Patients with symptomatic coronary artery disease who are treated with aggressive lipid lowering have improvement of symptom status and ischemia that appears to reflect improved vascular function but not atheroma burden. Am J Med. 2003;114:445-453. (C) 2003 by Excerpta Medica Inc.

Sport-specific practice and the development of expert decision-making in team ball sports

The role of sport-specific practice in the development of decision-making expertise in the sports of field hockey, netball, and basketball was examined. Fifteen expert decision-makers and 13 experienced non-expert athletes provided detailed information about the quantity and type of sport-specific and other related practice activities they had undertaken throughout their careers. Experts accumulated more hours of sport-specific practice from age 12 years onwards than did non-experts, spending on average some 13 years and 4,000 hours on concentrated sport-specific practice before reaching international standard. A significant negative correlation existed between the number of additional activities undertaken and the hours of sportspecific training required before attaining expertise, suggesting a functional role for activities other than sport-specific training in the development of expert decision-making.

Epizootic activity of Murray Valley encephalitis and Kunjin viruses in an aboriginal community in the Southeast Kimberley region of Western Australia: Results of mosquito fauna and virus isolation studies

We undertook annual surveys of flavivirus virus activity in the community of Billiluna of Western Australia in the southeast Kimberley region between 1989 and 2001. Culex annulirostris was the dominant mosquito species, particularly in years of above average rains and flooding. Murray Valley encephalitis (MVE) virus was isolated in 8 of the 13 years of the study from seven mosquito species, but more than 90% of the isolates were from Cx. annulirostris. The results suggest that MVE virus is epizootic in the region, with activity only apparent in years with average or above average rainfall and increased numbers of Cx. annulirostris. High levels of MVE virus activity and associated human cases were detected only once (in 1993) during the survey period. Activity of MVE virus could only be partially correlated with wet season rainfall and flooding, suggesting that a number of other factors must also be considered to accurately predict MVE virus activity at such communities.

Balancing the number and size of sites: An economic approach to the optimal design of cluster samples

The design of randomized controlled trials entails decisions that have economic as well as statistical implications. In particular, the choice of an individual or cluster randomization design may affect the cost of achieving the desired level of power, other things being equal. Furthermore, if cluster randomization is chosen, the researcher must decide how to balance the number of clusters, or sites, and the size of each site. This article investigates these interrelated statistical and economic issues. Its principal purpose is to elucidate the statistical and economic trade-offs to assist researchers to employ randomized controlled trials that have desired economic, as well as statistical, properties. (C) 2003 Elsevier Inc. All rights reserved.

Genotype and environment effects on dynamics of harvest index during grain filling in sorghum

An approach based on a linear rate of increase in harvest index (141) with time after anthesis has been used as a simple means-to predict grain growth and yield in many crop simulation models. When applied to diverse situations, however, this approach has been found to introduce significant error in grain yield predictions. Accordingly, this study was undertaken to examine the stability of the HI approach for yield prediction in sorghum [Sorghum bicolor (L.) Moench]. Four field experiments were conducted under nonlimiting water. and N conditions. The experiments were sown at times that ensured a broad range in temperature and radiation conditions. Treatments consisted of two population densities and three genotypes varying in maturity. Frequent sequential harvests were used to monitor crop growth, yield, and the dynamics of 111. Experiments varied greatly in yield and final HI. There was also a tendency for lower HI with later maturity. Harvest index dynamics also varied among experiments and, to a lesser extent, among treatments within experiments. The variation was associated mostly with the linear rate of increase in HI and timing of cessation of that increase. The average rate of HI increase was 0.0198 d(-1), but this was reduced considerably (0.0147) in one experiment that matured in cool conditions. The variations found in IN dynamics could be largely explained by differences in assimilation during grain filling and remobilization of preanthesis assimilate. We concluded that this level of variation in HI dynamics limited the general applicability of the HI approach in yield prediction and suggested a potential alternative for testing.

Low temperature induced spikelet sterility in rice. II. Effects of panicle and root temperatures

Low temperatures impose restrictions on rice (Oryza sativa L.) production at high latitudes. This study is related to low temperature damage that can arise mid-season during the panicle development phase. The objective of this study was to determine whether low temperature experienced by the root, panicle, or foliage is responsible for increased spikelet sterility. In temperature-controlled glasshouse experiments, water depth, and water and air temperatures, were changed independently to investigate the effects of low temperature in the root, panicle, and foliage during microspore development on spikelet sterility. The total number of pollen and number of engorged pollen grains per anther, and the number of intercepted and germinated pollen grains per stigma, were measured. Spikelet sterility was then analysed in relation to the total number of pollen grains per spikelet and the efficiency with which these pollen grains became engorged, were intercepted by the stigma, germinated, and were involved in fertilisation. There was a significant combined effect of average minimum panicle and root temperatures on spikelet sterility that accounted for 86% of the variation in spikelet sterility. Total number of pollen grains per anther was reduced by low panicle temperature, but not by low root temperature. Whereas engorgement efficiency ( the percentage of pollen grains that were engorged) was determined by both root and panicle temperature, germination efficiency (the percentage of germinated pollen grains relative to the number of engorged pollen grains intercepted by the stigma) was determined only by root temperature. Interception efficiency (i.e. percentage of engorged pollen grains intercepted by the stigma), however, was not affected by either root or panicle temperature. Engorgement efficiency was the dominant factor explaining the variation in spikelet sterility. It is concluded that both panicle and root temperature affect spikelet sterility in rice when the plant encounters low temperatures during the microspore development stage.

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.

Ecofisiologia, fenologia e adaptação de espécies florestais arbóreas nas condições edafoclimáticas de Viana-ES

A intensa atividade humana devasta grandes extensões de florestas nativas, seja para expansão da agricultura seja para suprir a crescente demanda do mercado por madeira, de uma forma ou de outra os pequenos fragmentos florestais remanescentes sofrem constantes pressões antrópicas, contudo, o uso de espécies arbóreas adaptadas pode contribuir para a proteção desses recursos naturais. Buscou-se neste estudo avaliar os padrões fenológicos de 109 espécies florestais arbóreas pertencentes a 37 famílias e 82 gêneros, dentre nativas e exóticas, com nove anos de idade, para diferenciar as mais adaptadas e com potencial de serem utilizadas em programas de reflorestamento. O estudo foi realizado na Fazenda Experimental do Incaper, em Jucuruaba, município de Viana-ES, (UTM E-345524, N- 7741039). Foram realizadas análises químicas do solo na área plantada e os dados climatológicos obtidos na estação meteorológica de Viana. O estudo baseou-se na observação do número de plantas sobreviventes de cada espécie e da avaliação do seu crescimento. Foram realizadas observações das fenofases de brotação, senescência de folhas, floração e frutificação. As avaliações fenológicas foram realizadas em intervalos mensais, no período de novembro de 2012 a outubro de 2013. Realizou-se a medição da altura das árvores, diâmetro à altura do peito (DAP), índice de enfolhamento, taxa de sobrevivência e cálculo do ICC (Índice Combinado de Crescimento), bem como a determinação das espécies mais adaptadas. Das 109 espécies estudadas, 64,22% apresentaram adaptação funcional e estrutural às condições de solo e clima da região experimental, 42,22 % floresceram e frutificaram e 90% apresentaram senescência e brotação acompanhando a sazonalidade climática. Vinte e nove espécies apresentaram ICC maior do que o ICC médio. A maioria das espécies destacou-se como alternativa para recuperação da cobertura vegetal local, com destaque para Inga uruguensis e Schizolobium amazonicum. As análises de fluorescência da clorofila revelaram que o aparato fotossintético da Schizolobium amazonicum foi capaz de protegê-la da fotoinibição e promover boa conversão da energia luminosa em fotoquímica.

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.