Sugarcane micropropagation and phenolic excretion

Sugarcane shoot formation was followed using a temporary immersion system. Plant fresh weight, plant dry weight, shoot number and phenolic excretion to the culture medium were recorded during shoot formation. Shoot number increased for 30 days of culture but formation of new shoots was greatly reduced from 31 to 40 days. Phenolic excretion also increased during the first 20 days of culture (gallic acid represented 82% total phenolics) and decreased during the last 10 days (31–40 days of culture). The most intensive period of phenolic excretion (11–20 days) preceded the most intensive period of shoot formation (21–30 days). The same relationship does not seem to exist between the accumulation of fresh and dry weights. Subculture onto fresh medium at the beginning of proliferation (10 days after culture initiation) was detrimental to shoot formation in the subsequent period (11–20 days). However, such a detrimental effect could be avoided if gallic acid was added to the medium. Addition of cysteine to the culture medium reduced both excretion of phenolics and shoot formation but not fresh weight. The use of temporary immersion systems, the increase of culture medium volume per initial explant and the addition of paclobutrazol promoted both phenolic excretion and sugarcane shoot formation. Results presented here indicate a relationship between phenolic excretion and shoot formation but not with accumulation of plant weight. This revised version was published online in June 2006 with corrections to the Cover Date.     

New contributions to propagation of pineapple (<Emphasis Type="Italic">Ananas comosus</Emphasis> L. Merr) in temporary immersion bioreactors

Summary Plantlets propagated in temporary immersion bioreactors (TIB) have better performance than those propagated by conventional methods such as micropropagation. This is as a result of a better handling of the in vitro atmosphere and the nutrition. The object of this study to further improve the cultivation conditions by introducting photomixotrophism as an intermediate link of photoautotrophic growth during ex vitro acclimatization. For this purpose the effects of light were evaluated by different parameters such as photosynthetic photon flux density (PPF), sucrose concentration, and CO₂ enrichment levels on CO₂ evolution inside the culture vessels. It was observed that CO₂ diminished upon light exposure and increased in the dark according to the photoperiod during each cycle of immersion. With this approach it was possible to increase the photomixotrophism in the pineapple plantlets propagated in TIB. It was demonstrated that light is the factor with more influence on plant quality, although under these conditions they seem to use more of the nutrients of the medium than their photoassimilates. The propagation of pineapple in TIB involves three phases: proliferation, pre-elongation, and final growth of the buds. In each phase the cultivation conditions were determined to substitute for sterilization by autoclaving, to improve the quality of the plants, to elevate the efficiency of the process, and to reduce production costs. The buds that grew in the temporary immersion bioreactor with the presence of Vitrofural (G-1) achieved the best indicators of growth. Significant increases were observed in the leaf area, dry mass of the buds, and chlorophyll contents.     

Hypoxia and acidosis impair cGMP synthesis in microvascular coronary endothelial cells

To characterize the effects of ischemia on cGMP synthesis in microvascular endothelium, cultured endothelial cells from adult rat hearts were exposed to hypoxia or normoxia at pH 6.4 or 7.4. Cellular cGMP and soluble (sGC) and membrane guanylyl cyclase (mGC) activities were measured after stimulation of sGC (S-nitroso-N-acetyl-penicillamine) or mGC (urodilatin) or after no stimulation. Cell death (lactate dehydrogenase release) was negligible in all experiments. Hypoxia at pH 6.4 induced a rapid approximately 90% decrease in cellular cGMP after sGC and mGC stimulation. This effect was reproduced by acidosis. Hypoxia at pH 7.4 elicited a less pronounced (approximately 50%) and slower reduction in cGMP synthesis. Reoxygenation after 2 h of hypoxia at either pH 6.4 or 7.4 normalized the response to mGC stimulation but further deteriorated the sGC response; normalization of pH rapidly reversed the effects of acidosis. At pH 7.4, the response to GC stimulation correlated well with cellular ATP. We conclude that simulated ischemia severely depresses cGMP synthesis in microvascular coronary endothelial cells through ATP depletion and acidosis without intrinsic protein alteration.     

Effects of epidermal growth factor on epinephrine-stimulated heart function in rodents

Epidermal growth factor (EGF) interferes with beta-adrenergic receptor (beta-AR) signaling in adipocytes and hepatocytes, which leads to decreased lipolytic and glycogenolytic responses, respectively. We studied the effect of EGF on the heart. EGF interfered with the cAMP signal generated by beta-AR agonists in cardiac myocytes. In perfused hearts, EGF decreased inotropic and chronotropic responses to epinephrine but not to 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate. Sustained epinephrine infusion induced heart contracture, which resulted in altered heart function as demonstrated by decreased inotropy and increased heart rate variability. EGF prevented all these alterations. In the whole animal (anesthetized mice), EGF administration reduced the rise in heart rate induced by a single epinephrine dose and the occurrence of Bezold-Jarisch reflex episodes induced by repeated doses. Sialoadenectomy enhanced the response to epinephrine, and EGF administration restored normal response. All these results suggest that, by interfering with beta-AR signaling, EGF protects the heart against the harmful effects of epinephrine.     

Steady-state chlorophyll fluorescence (Fs) measurements as a tool to follow variations of net CO2 assimilation and stomatal conductance during water-stress in C3 plants

Water stress experiments were performed with grapevines ( Vitis vinifera L.) and other C₃ plants in the field, in potted plants in the laboratory, and with detached leaves. It was found that, in all cases, the ratio of steady state chlorophyll fluorescence (Fs) normalized to dark-adapted intrinsic fluorescence (Fo) inversely correlated with non-photochemical quenching (NPQ). Also, at high irradiance, the ratio Fs/Fo was positively correlated with CO₂ assimilation in air, with electron transport rate calculated from fluorescence, and with stomatal conductance, but no clear correlation was observed with qP. The significance of these relationships is discussed. The ratio Fs/Fo, measured with a portable instrument (PAM-2000) or with a remote sensing FIPAM system, provides a good method for the early detection of water stress, and may become a useful guide to irrigation requirements.     

Comparison of plantain plantlets propagated in temporary immersion bioreactors and gelled medium during in vitro growth and acclimatization

The current work compared the physiological characteristics of plantain (Musa AAB) plantlets micropropagated in temporary immersion bioreactors (TIB) and on a gelled medium (GM). The plantlets were evaluated during in vitro growth (in the shoot elongation phase) and at the end of ex vitro acclimatization. TIB improved rooting and gave rise to longer shoots and higher dry mass. Respiration rate was the highest at the beginning of shoot elongation in both the TIB and GM plantlets. Photosynthetic rate in TIB was significantly higher than in GM from the midpoint of acclimatization, whereas a pyruvate kinase (PK) activity was lower. Starch accumulation was ca. two fold higher in corms than in leaves and always higher in the TIB than GM plantlets. The higher expression of genes coding for carbon metabolism enzymes PK and phosphoenolpyruvate carboxylase (PEPC) in TIB than in PM indicates a more important role of an autotrophic metabolism in the TIB plantlets when compared to the GM ones. The accumulated reserves were used during the first days of acclimatization leading to the higher survival rates and to the better plant quality of the TIB plantlets.     

Characterization of a Novel Water Pocket Inside the Human Cx26 Hemichannel Structure

Connexins (Cxs) are a family of vertebrate proteins constituents of gap junction channels (GJCs) that connect the cytoplasm of adjacent cells by the end-to-end docking of two Cx hemichannels. The intercellular transfer through GJCs occurs by passive diffusion allowing the exchange of water, ions, and small molecules. Despite the broad interest to understand, at the molecular level, the functional state of Cx-based channels, there are still many unanswered questions regarding structure-function relationships, perm-selectivity, and gating mechanisms. In particular, the ordering, structure, and dynamics of water inside Cx GJCs and hemichannels remains largely unexplored. In this work, we describe the identification and characterization of a believed novel water pocket—termed the IC pocket—located in-between the four transmembrane helices of each human Cx26 (hCx26) monomer at the intracellular (IC) side. Using molecular dynamics (MD) simulations to characterize hCx26 internal water structure and dynamics, six IC pockets were identified per hemichannel. A detailed characterization of the dynamics and ordering of water including conformational variability of residues forming the IC pockets, together with multiple sequence alignments, allowed us to propose a functional role for this cavity. An in vitro assessment of tracer uptake suggests that the IC pocket residue Arg-143 plays an essential role on the modulation of the hCx26 hemichannel permeability.     

Effect of sucrose,light, and carbon dioxide on plantain micropropagation in temporary immersion bioreactors

In vitro physiology and carbon metabolism can be affected by the sink–source relationship. The effect of different sucrose concentrations (10, 30, and 50 g L⁻¹), light intensities (80 and 150 μmol m⁻² s⁻¹), and CO₂ levels (375 and 1,200 μmol mol⁻¹) were tested during plantain micropropagation in temporary immersion bioreactors. Activities of pyruvate kinase, phosphoenol pyruvate carboxylase, and the photosynthesis rate were recorded. From the morphological and practical point of view, the best results were obtained when plants were cultured with 30 g L⁻¹ sucrose, 80 μmol m⁻² s⁻¹ light intensity, and 1,200 μmol mol⁻¹ CO₂ concentration. This treatment improved leaf and root development, reduced respiration during in vitro culture, and increased starch level at the end of the hardening phase. In addition to that, the number of competent plants was increased from 80.0% to 91.0% at the end of the in vitro phase and the survival percentage from 95.71% to 99.80% during ex vitro hardening.     

Plant water status and genotype affect fruit respiration in grapevines

An understanding of fruit gas exchange is necessary to determine the carbon balance in grapevines, but little attention has been paid to the relationships among fruit respiration, plant water status and genetic variability. The effect of plant water status and genotype on cluster respiration was studied over two seasons (2013 and 2014) under field conditions using a whole cluster respiration chamber. Whole cluster CO₂ fluxes were measured in growing grapevines at hard-green, veraison and ripening stages under irrigated and non-irrigated conditions, and under light and dark conditions in two grapevine varieties, Tempranillo and Grenache. A direct relationship between cluster CO₂ efflux and plant water status was found at hard-green stage. Genotype influenced the fruit CO₂ efflux that resulted in higher carbon losses in Tempranillo than in Grenache. Fruit respiration rates decreased from the first berry developmental stages to ripening stage. The integration of fruit respiration rates under light and dark conditions showed the magnitude of fruit carbon losses and gains as well as interesting variety and environmental conditions effects on those processes.