Cystic fibrosis gene variability in two southern Brazilian Amerindian populations: analysis of the deltaF508 mutation and the KM19 and XV2C haplotypes

The frequencies of the deltaF508 deletion, the most common cystic fibrosis mutation in Europeans and European-derived populations, and the XV2C and KM19 restriction fragment length polymorphisms that are tightly linked to the CFTR locus vary among populations. To determine the distribution of these extragenic markers and of the deltaF508 mutation, we analyzed 326 chromosomes of individuals from two South American Indian populations, the Guarani and the Kaingang. The allele and haplotype frequencies differed greatly between the two populations as well as among Amerindians and normal European Brazilians and European Brazilian cystic fibrosis patients. The absence of the deltaF508 mutation and the B haplotype are in agreement with the hypothesis that the deltaF508 mutation occurred after the divergence of these two populations. This finding is useful for populations containing a large Amerindian component and helps us to understand the origins of the deltaF508 deletion, the most common cystic fibrosis mutation in Europeans and European-derived populations, as well as the different incidences of cystic fibrosis in continental groups.     

Polyphenolic compounds from Artemisia dracunculus L. inhibit PEPCK gene expression and gluconeogenesis in an H4IIE hepatoma cell line

An ethanolic extract of Russian tarragon, Artemisia dracunculus L., with antihyperglycemic activity in animal models was reported to decrease phosphoenolpyruvate carboxykinase (PEPCK) mRNA expression in STZ-induced diabetic rats. A quantitative polymerase chain reaction (qPCR) assay was developed for the bioactivity-guided purification of the compounds within the extract that decrease PEPCK expression. The assay was based on the inhibition of dexamethasone-stimulated PEPCK upregulation in an H4IIE hepatoma cell line. Two polyphenolic compounds that inhibited PEPCK mRNA levels were isolated and identified as 6-demethoxycapillarisin and 2',4'-dihydroxy-4-methoxydihydrochalcone with IC(50) values of 43 and 61 muM, respectively. The phosphoinositide-3 kinase (PI3K) inhibitor LY-294002 showed that 6-demethoxycapillarisin exerts its effect through the activation of the PI3K pathway, similarly to insulin. The effect of 2',4'-dihydroxy-4-methoxydihydrochalcone is not regulated by PI3K and dependent on activation of AMPK pathway. These results indicate that the isolated compounds may be responsible for much of the glucose-lowering activity of the Artemisia dracunculus extract.     

Assessment of vegetation and soil conditions in restored mangroves interrupted by severe tropical typhoon ‘Chan-hom’ in the Philippines

Using a space-for-time substitution approach, we investigated the effects of a typhoon on the vegetation and soil development trajectories of monospecific stands of Rhizophora mucronata mangroves of different ages (6-, 8- 10-, 11-, 12-, 17-, 18- and 50-year stands). The vegetation and soil parameters were compared to a reference system comprised of mature, natural mangrove stands. Pre-typhoon measures of vegetation and soil parameters were compared with 1-mo, 7-mo and 9-mo post-typhoon. Prior to the occurrence of the typhoon, there were clear patterns of vegetation and soil development with age of the stands. The development trajectory was however interrupted by the occurrence of the typhoon. Severe damage was more apparent in older mangrove stands (11- and 18- year stands) with very low to no damage in the younger stands. The typhoon-impacted sites experienced a number of changes, including: complete defoliation; reduced living tree densities of 61–69 %; decreased above-ground biomass of 70–79 %; increased soil nutrient levels of 40–60 %; more waterlogged soils by at least 113 % and increased soil temperature of 8–10 °C. Cumulative tree mortality, compounded by the lack of seedling recruits and unfavourable soil conditions may limit long-term recovery of the typhoon-impacted stands.     

Hypertrophic Gene Expression Induced by Chronic Stretch of Excised Mouse Heart Muscle

Altered mechanical stress and strain in cardiac myocytes induce modifications in gene expression that affects cardiac remodeling and myocyte contractile function. To study the mechanisms of mechanotransduction in cardiomyocytes, probing alterations in mechanics and gene expression has been an effective strategy. However, previous studies are self-limited due to the general use of isolated neonatal rodent myocytes or intact animals. The main goal of this study was to develop a novel tissue culture chamber system for mouse myocardium that facilitates loading of cardiac tissue, while measuring tissue stress and deformation within a physiological environment. Intact mouse right ventricular papillary muscles were cultured in controlled conditions with superfusate at 95% O₂/ 5% CO₂, and 34°C, such that cell to extracellular matrix adhesions as well as cell to cell adhesions were undisturbed and both passive and active mechanical properties were maintained without significant changes. The system was able to measure the induction of hypertrophic markers (BNP, ANP) in tissue after 2 hrs and 5 hrs of stretch. ANP induction was highly correlated with the diastolic load of the muscle but not with developed systolic load. Load induced ANP expression was blunted in muscles from muscle-LIM protein knockout mice, in which defective mechanotransduction pathways have been predicted.     

Plants and human health in the twenty-first century

The concept of growing crops for health rather than for food or fiber is slowly changing plant biotechnology and medicine. Rediscovery of the connection between plants and health is responsible for launching a new generation of botanical therapeutics that include plant-derived pharmaceuticals, multicomponent botanical drugs, dietary supplements, functional foods and plant-produced recombinant proteins. Many of these products will soon complement conventional pharmaceuticals in the treatment, prevention and diagnosis of diseases, while at the same time adding value to agriculture. Such complementation can be accelerated by developing better tools for the efficient exploration of diverse and mutually interacting arrays of phytochemicals and for the manipulation of the plant's ability to synthesize natural products and complex proteins. This review discusses the history, future, scientific background and regulatory issues related to botanical therapeutics.     

rRNA and poly-beta-hydroxybutyrate dynamics in bioreactors subjected to feast and famine cycles

Feast and famine cycles are common in activated sludge wastewater treatment systems, and they select for bacteria that accumulate storage compounds, such as poly-beta-hydroxybutyrate (PHB). Previous studies have shown that variations in influent substrate concentrations force bacteria to accumulate high levels of rRNA compared to the levels in bacteria grown in chemostats. Therefore, it can be hypothesized that bacteria accumulate more rRNA when they are subjected to feast and famine cycles. However, PHB-accumulating bacteria can form biomass (grow) throughout a feast and famine cycle and thus have a lower peak biomass formation rate during the cycle. Consequently, PHB-accumulating bacteria may accumulate less rRNA when they are subjected to feast and famine cycles than bacteria that are not capable of PHB accumulation. These hypotheses were tested with Wautersia eutropha H16 (wild type) and W. eutropha PHB-4 (a mutant not capable of accumulating PHB) grown in chemostat and semibatch reactors. For both strains, the cellular RNA level was higher when the organism was grown in semibatch reactors than when it was grown in chemostats, and the specific biomass formation rates during the feast phase were linearly related to the cellular RNA levels for cultures. Although the two strains exhibited maximum uptake rates when they were grown in semibatch reactors, the wild-type strain responded much more rapidly to the addition of fresh medium than the mutant responded. Furthermore, the chemostat-grown mutant culture was unable to exhibit maximum substrate uptake rates when it was subjected to pulse-wise addition of fresh medium. These data show that the ability to accumulate PHB does not prevent bacteria from accumulating high levels of rRNA when they are subjected to feast and famine cycles. Our results also demonstrate that the ability to accumulate PHB makes the bacteria more responsive to sudden increases in substrate concentrations, which explains their ecological advantage.     

Electrical detection of DNA hybridization: three extraction techniques based on interdigitated Al/Al2O3 capacitors

Based on interdigitated aluminum electrodes covered with Al(2)O(3) and silver precipitation via biotin-antibody coupled gold nano-labels as signal enhancement, three complementary electrical methods were used and compared to detect the hybridization of target DNA for concentrations down to the 50 pM of a PCR product from cytochrome P450 2b2 gene. Human hepatic cytochrome P450 (CYP) enzymes participate in detoxification metabolism of xenobiotics. Therefore, determination of mutational status of P450 gene in a patient could have a significant impact on the choice of a medical treatment. Our three electrical extraction procedures are performed on the same interdigitated capacitive sensor lying on a passivated silicon substrate and consist in the measurement of respectively the low-frequency inter-electrodes capacitance, the high-frequency self-resonance frequency, and the equivalent MOS capacitance between the short-circuited electrodes and the backside metallization of the silicon substrate. This study is the first of its kind as it opens the way for correlation studies and noise reduction techniques based on multiple electrical measurements of the same DNA hybridization event with a single sensor.     

Purification, cloning, and expression of a pathogen inducible UDP-glucose:Salicylic acid glucosyltransferase from tobacco

Salicylic acid (SA) plays an important role in plant disease resistance. Inoculation of tobacco leaves with incompatible pathogens triggers the biosynthesis of SA which accumulates primarily as the SA 2-O-beta-D-glucoside (SAG) and glucosyl salicylate (GS). The tobacco UDP-glucose:salicylic acid glucosyltransferase (SA GTase) capable of forming both SAG and GS was purified, characterized, and partially sequenced. It has an apparent molecular mass of 48 kDa, a pH optimum of 7.0, and an isoelectric point at pH 4.4. UDP-glucose was the sole sugar donor for the enzyme. However, SA and several phenolics served as glucose acceptors. The apparent K(m) values for UDP-glucose and SA were 0.27 and 1-2 mM, respectively. Zn(2+) and UDP inhibited its activity. The corresponding cDNA clone which encoded a protein of 459 amino acids was isolated from an SA-induced tobacco cDNA library and overexpressed in Escherichia coli. The recombinant protein catalyzed the formation of SAG and GS, and exhibited a broad specificity to simple phenolics, similar to that of the purified enzyme. Northern blot analysis showed that the SA GTase mRNA was induced both by SA and incompatible pathogens. The rapid induction timing of the mRNA by SA indicates that it belongs to the early SA response genes.     

Genetic variation at twentythree microsatellite loci in sixteen human populations

We have analysed genetic variation at 23 microsatellite loci in a global sample of 16 ethnically and geographically diverse human populations. On the basis of their ancestral heritage and geographic locations, the studied populations can be divided into five major groups, viz. African, Caucasian, Asian Mongoloid, American Indian and Pacific Islander. With respect to the distribution of alleles at the 23 loci, large variability exists among the examined populations. However, with the exception of the American Indians and the Pacific Islanders, populations within a continental group show a greater degree of similarity. Phylogenetic analyses based on allele frequencies at the examined loci show that the first split of the present-day human populations had occurred between the Africans and all of the non-African populations, lending support to an African origin of modern human populations. Gene diversity analyses show that the coefficient of gene diversity estimated from the 23 loci is, in general, larger for populations that have remained isolated and probably of smaller effective sizes, such as the American Indians and the Pacific Islanders. These analyses also demonstrate that the component of total gene diversity, which is attributed to variation between groups of populations, is significantly larger than that among populations within each group. The empirical data presented in this work and their analyses reaffirm that evolutionary histories and the extent of genetic variation among human populations can be studied using microsatellite loci.