Titrating the effects of mitochondrial complex I impairment in the cell physiology.

The mitochondrial oxidative phosphorylation system consists of five multimeric enzymes (complexes I-V). NADH dehydrogenase or complex I (CI) is affected in most of the mitochondrial diseases and in some neurodegenerative disorders. We have studied the physiological consequences of a partial CI inhibition at the cellular level. We used a genetic model (40% CI-inhibited human-ape xenomitochondrial cybrids) and a drug-induced model (0-100% CI-inhibited cells using different concentrations of rotenone). We observed a quantitative correlation between the level of CI impairment and cell respiration, cell growth, free radical production, lipid peroxidation, mitochondrial membrane potential, and apoptosis. We showed that cell death was quantitatively associated with free radical production rather than with a decrease in respiratory chain function. The results obtained with human xenomitochondrial cybrid cells were compatible with those observed in rotenone-induced 40% CI-inhibited cells. At high concentrations (5-6-fold higher than the concentration necessary for 100% CI inhibition), rotenone showed a second toxic effect at the level of microtubule assembly, which also led to apoptosis. The correlation found among all the parameters studied helped clarify the physiological consequences of partial CI inhibitions at the cellular level.     

Structure of the KcsA potassium channel from Streptomyces lividans: a site-directed spin labeling study of the second transmembrane segment.

KcsA is a prokaryotic potassium channel. The present study employs cysteine scanning mutagenesis and site-directed spin labeling to investigate the structure of the second transmembrane segment (residues 82-120) in functional tetrameric channels reconstituted in lipid bilayers. Spin-spin interactions are observed between nitroxide side chains at symmetry-related sites close to the 4-fold axis of symmetry. To aid in quantitative analysis of these interactions, a new diamagnetic analogue of the nitroxide side chain is used to prepare magnetically dilute samples with constant structure. Using constraints imposed by the spin-spin interactions, a packing model for this segment is deduced that is in excellent agreement with the recently reported crystal structure [Doyle, D., et al. (1998) Science 280, 69-77]. The relatively immobilized state of the nitroxide side chains suggests that the channel is rigid on the electron paramagnetic resonance time scale. Moreover, the poor sulfhydryl reactivity of the cysteine at many locations indicates that the channel is not subject to the low-frequency fluctuations that permit reaction of buried cysteines. At sites expected to be located in the pore, the accessibility of the side chains to collision with O(2) or nickel(II) ethylenediaminediacetate is low. This inaccessibility, together with the generally low mobility of the side chains throughout the sequence, makes it difficult to detect the presence of the pore based on these measurements. However, the presence of a solvated pore can be directly demonstrated using a polarity parameter deduced from the EPR spectra recorded at low temperature. These measurements also reveal the presence of a polarity gradient in the phospholipid bilayer.     

The effect of sperm preparation and co-incubation time on in vitro fertilization of Bos indicus oocytes.

The objective of the present study was to evaluate the effect of various methods of sperm selection and various sperm-oocyte co-incubation times on in vitro fertilization (IVF) of zebu (Bos indicus) oocytes. Frozen semen from one ejaculate of a single bull was used for all treatments and replicates. After thawed, sperm was subjected to one of the three treatments: 45 and 90% discontinuous Percoll gradient, swim-up and washing by centrifugation. In all treatments, the spermatozoa were incubated with in vitro matured oocytes for 3, 6, 12 and 18h. After co-incubation oocytes were transferred to the culture medium and culture for 44h, when the cleavage was evaluated. The uncleavaged oocytes were fixed and stained to determine penetration, pronucleus formation and polyspermy. The sperm selection method did not influence (P<0.05) polyspermy, pronucleus formation, penetration and cleavage rates. No interaction between method of selection and sperm-oocyte co-incubation time was observed (P>0.05). However, sperm-oocyte co-incubation time affected fertilization. The lower penetration (26.5%) and cleavage rates (13.1%) were obtained at 3-h period. The penetration and cleavage percentages increased (P<0.05) progressively at 6h (63.3 and 54.4%) and 12h (77.6 and 67.6%). No differences (P>0.05) were observed between 12 and 18h of incubation for penetration and cleavage rates. The incidence of polyspermy and pronucleus formation was similar (P>0.05) for all time points. It is concluded that the methods used in this study for sperm selection do not affect fertilization; therefore, they all can be used for bovine IVF. In addition, regardless the method used better fertilization results were obtained when sperm and oocytes were co-incubated for 12h, and the prolongation of that time for up to 18h had no detrimental effect on fertilization.     

Introduced plants in the indigenous Pharmacopoeia of Northern South America

The intellectual property rights debate focuses on the flow of germplasm from developing tropical countries to developed temperate nations. Few investigators have addressed the converse. We discuss the abundance and importance of introduced plants in pharmacopoeias of northern South America. Introduced species commonly are employed as medicines throughout the region and include at least 216 Eurasian, North American, African, and Pacific species. Among the Shuar of lowland Ecuador, four introduced plants (Citrus aurantium, Cymbopogon citratus, Saccharum officinarum, and Zingiber officinale) are included in their most commonly prescribed remedies. The widespread use of introduced plants is due, in part, to the medicinal value of plants whose primary use is for food (e.g., Musa X paradisiacal Similarly, many introduced ornamentals also have therapeutic value (e.g., Hedychium coronarium). Other species have been introduced specifically as medicines (e.g., Aloe vera). Restrictions on the flow of germplasm and plant knowledge may protect the economic interests of governments and national industry. If applied bilaterally, however, constraints on the movement of plants will limit the continued evolution of traditional medicinal systems in areas where they are most needed.     

Aeromedical transport: its hidden problems.

Air transport can move patients safely and rapidly over long distances. However, changes in altitude can have disastrous effects because diminished ambient air pressure may allow gases in closed spaces and tissues to expand rapidly. Even pressurized commercial aircraft do not maintain sea-level pressure: cabin pressures equal to those at yp to 8000 ft may be experienced, diminishing oxygen tension in proportion. Air transport is absolutely contraindicated for patients with untreated pneumothorax, gas gangrene, or air trapped in the cranium and those who have recently undergone abdominal surgery. Special considerations including a planned low-altitude flight are warrented for patients who are anemic, in respiratory or cardiac distress, or immobilized in casts, or who have been engaged in underwater diving immediately before the flight.