Biophoton emission

The phenomenon of ultraweak photon emission from living systems was further investigated in order to elucidate the physical properties of this radiation and its possible source. We obtained evidence that the light has a high degree of coherence because of (1) its photon count statistics, (2) its spectral distribution, (3) its decay behavior after exposure to light illumination, and (4) its transparency through optically thick materials. Moroever, DNA is apparently at least an important source, since conformational changes induced with ethidium bromide in vivo are clearly reflected by changes of the photon emission of cells. The physical properties of the radiation are described, taking DNA as an exciplex laser system, where a stable state can be reached far from thermal equilibrium at threshold.     

Direct peptide profiling of lateral cell groups of the antennal lobes of Manduca sexta reveals specific composition and changes in neuropeptide expression during development

The paired antennal lobes are the first integration centers for odor information in the insect brain. In the sphinx moth Manduca sexta, like in other holometabolous insects, they are formed during metamorphosis. To further understand mechanisms involved in the formation of this particularly well investigated brain area, we performed a direct peptide profiling of a well defined cell group (the lateral cell group) of the antennal lobe throughout development by MALDI-TOF mass spectrometry. Although the majority of the about 100 obtained ion signals represent still unknown substances, this first peptidomic characterization of this cell group indicated the occurrence of 12 structurally known neuropeptides. Among these peptides are helicostatin 1, cydiastatins 2, 3, and 4, M. sexta-allatotropin (Mas-AT), M. sexta-FLRFamide (Mas-FLRFamide) I, II, and III, nonblocked Mas-FLRFamide I, and M. sexta-myoinhibitory peptides (Mas-MIPs) III, V, and VI. The identity of two of the allatostatins (cydiastatins 3 and 4) and Mas-AT were confirmed by tandem mass spectrometry (MALDI-TOF/TOF). During development of the antennal lobe, number and frequency of ion signals including those representing known peptides generally increased at the onset of glomeruli formation at pupal Stage P7/8, with cydiastatin 2, helicostatin 1, and Mas-MIP V being the exceptions. Cydiastatin 2 showed transient occurrence mainly during the period of glomerulus formation, helicostatin 1 was restricted to late pupae and adults, while Mas-MIP V occurred exclusively in adult antennal lobes. The power of the applied direct mass spectrometric profiling lies in the possibility of chemically identifying neuropeptides of a given cell population in a fast and reliable manner, at any developmental stage in single specimens. The identification of neuropeptides in the antennal lobes now allows to specifically address the function of these signaling molecules during the formation of the antennal lobe network.     

Mecardonia tenella (Plantaginaceae) Attracts Oil-, Perfume-, and Pollen-Gathering Bees in Southern Brazil

Floral characteristics often indicate the pollinators' functional group visiting the plant and the pollination syndromes associated with them. This idea has been challenged in the past decades due to increasing evidence that most plants, including those exhibiting floral syndromes, are visited by large arrays of species that differ in their effectiveness as pollinators. Our study focuses on Mecardonia tenella (Plantaginaceae) from the Araucaria forest of southern Brazil, which exhibits characteristics of the oil flower pollination syndrome. However, it is visited by three types of functional groups of bees: male orchid bees, oil-collecting bees, and pollen-seeking bees. The relative contribution of each functional group to the plant's reproductive success was estimated based on their pollen load, visitation frequency, and morphology. We assessed resources, phenology, and breeding system of M. tenella . Our results indicate that flowers lack nectar, but volatiles, lipids, and pollen are resources that can be gathered by visitors. This combination of floral traits and visitors' assemblage makes M. tenella a challenge to the concept of pollination syndromes. Our findings indicate that the current interactions may not reflect the circumstances under which some floral traits of this plant were selected.     

Effects of AY 9944 on low density lipoprotein metabolism in cultured human fibroblasts

Treatment of cultured human fibroblasts with the hypocholesterolemic drug AY 9944 resulted in a marked increase in low density lipoprotein internalization and degradation for concentrations up to 5 X 10(-6)M. Low density lipoprotein binding was less affected. Concentrations above 5 X 10(-6)M resulted in a relative decrease in low density lipoprotein degradation, whereas binding and internalization plateaued. The stimulation of low density lipoprotein internalization took place within the first hours of incubation of cells with the drug, which suggests a direct effect on the cell membrane. Such phenomenon could account at least partially for the hypocholesterolemic effect of the drug, besides its inhibitory effect on 7-dehydrocholesterol reductase.     

Uptake of [(3)H]glycine by synaptosomes of channel

It is known that channel catfish erythrocytes can take up glycine by several distinct transport systems. Further, glycine is an inhibitory neurotransmitter in mammalian brain and spinal cord. Consequently, the uptake of [(3)H]glycine by catfish brain was investigated and found to be a saturable process, dependent on the presence of Na(++) and Cl(--) and sensitive to temperature. A kinetic analysis of transport was performed at 22C. This showed that a high-affinity system existed which exhibited a K(m) of 5.1 (+/- 2. 1) microM. Several structural analogues of glycine were capable of inhibiting uptake in a competitive manner. The most effective inhibitor was sarcosine (IC(50) 5 36 microM). Uptake was also able to be inhibited by harmaline, a drug known to interfere with Na(+)-dependent transport processes. It is concluded that glycine transport by channel catfish brain has much in common with transport by mammalian nervous tissue which is carried out by the membrane carriers GLYT1 and GLYT2. On the other hand, synaptosomal transport differs somewhat from glycine transport by channel catfish erythrocytes.     

Quantitative polymerase chain reaction as a reliable method to determine functional lentiviral titer after ex vivo gene transfer in human mesenchymal stem cells

BACKGROUND: Human mesenchymal stem cells (hMSCs) are a promising target for ex vivo gene therapy and lentiviruses are excellent gene transfer vehicles in hMSCs since they achieve high transduction rates with long-term gene expression. Nevertheless, senescence of hMSCs may limit therapeutic applications due to time-consuming cell selection and viral titration. Here, we describe a fast and reliable method to determine functional lentiviral titer by quantitative polymerase chain reaction (qPCR) after highly efficient ex vivo gene transfer in hMSCs. METHODS: Lentivirus production was tested with different types of packaging systems. Using p24 ELISA remaining viral particles were detected in the cell culture supernatant. The lentiviral gene transfer efficiency was quantified by FACS analysis. Lentiviral titers were determined by qPCR of expressed transgenes. RESULTS: Third-generation self-inactivating vectors showed highly efficient gene transfer in hMSCs. No viral antigen was detected in the cell culture supernatant after four media changes, suggesting the absence of infectious particles after 4 days. We observed a linear correlation between virus dilution and level of transgene expression by qPCR analysis, therefore allowing viral titering by quantification of transgene expression. Finally, we demonstrated that transduced hMSCs retained their stem cell character by differentiation towards adipogenic, osteogenic and chondrogenic lineages. CONCLUSIONS: Quantification of transgene copy numbers by qPCR is a fast and reliable method to determine functional lentiviral titer after ex vivo gene transfer in hMSCs.     

Studies on well-coupled Photosystem I-enriched subchloroplast vesicles — energy-dependent switching between two different active states of the proton-translocation adenosine triphosphatase

Single-turnover flash-induced ATP synthesis coupled to natural cyclic electron flow in Photosystem I-enriched subchloroplast vesicles (from spinach) was continuously followed by the luciferin-luciferase luminescence. The ATP yield per flash was maximal (1 ATP per s per 1000 Chl) around a flash frequency of 0.5–2 Hz. It decreased both at lower and higher flash frequencies. The decrease at high flash frequency was due to limitation by the electron-transfer rate, while the decrease at low flash frequency was directly due to intrinsic properties of the ATPase itself. Carbonylcyanide-p-trifluoromethoxyphenylhydrazone (FCCP) decreased the yield at low frequency more than at high frequency. The same behaviour was observed if electron transfer was artificially mediated by pyocyanin. If the ADP concentration was increased from 40 to at least 80 μM, or if the vesicles were preincubated with 5 mM dithiothreitol (DTT), the decrease of the yield at flash frequencies below 0.5 Hz was no longer observed. Incubation with DTT increased the rates of ATP hydrolysis and synthesis at any flash frequency. The decrease of the yield could be elicited again by addition of 50 nM FCCP. It is concluded that at low levels of the protonmotive force (Δ gm_H⁺), the ATPase is converted into an active ATP-hydrolyzing state in which ATP synthesis activity is decreased due to a decreased affinity towards ADP and/or to a decreased release of newly synthesized ATP, that can be cancelled by increasing the ADP concentration or by addition of DTT in the absence of uncoupler.