Oral phosphatidylcholine pretreatment alleviates the signs of experimental rheumatoid arthritis

Introduction  

Phosphatidylcholine and phosphatidylcholine-derived metabolites exhibit anti-inflammatory properties in various stress conditions. We hypothesized that dietary phosphatidylcholine may potentially function as an anti-inflammatory substance and may decrease inflammatory activation in a chronic murine model of rheumatoid arthritis (collagen-induced arthritis).     

Profiling the chlorogenic acids of aster by HPLC-MS(n)

Using HPLC-MS(n), 33 chlorogenic acids were identified in an aqueous-alcoholic extract of Aster ageratoides Turcz. flower buds. These were three isomers each of p-coumaroylquinic acid, caffeoylquinic acid, feruloylquinic acid, dicaffeoylquinic acid and diferuloylquinic acid, and six isomers each of p-coumaroyl-caffeoylquinic acid, p-coumaroyl-feruloylquinic acid and caffeoyl-feruloylquinic acid. Only the caffeoylquinic acids and dicaffeoylquinic acids have been reported previously in Asteraceae. Three of the six p-coumaroyl-feruloylquinic acids (3-feruloyl-4-p-coumaroylquinic acid, '3-feruloyl-5-p-coumaroylquinic acid and 4-feruloyl-5-p-coumaroylquinic acid) have not been observed previously in nature. Cis-5-p-coumaroylquinic acid was identified at a concentration ca 25% that of the more common trans isomer. The feruloylquinic acids and diferuloylquinic acids dominated the mono- and di-acyl chlorogenic acid fractions, respectively, making this plant material a useful source of these commercially non-available substances. These 33 chlorogenic acids were not detected in the leaves or stem of A. ageratoides Turcz., or in the flower buds of A. ageratoides Turcz. var. Gerla or A. kalimeris indica (L) Sch. Bip. Only the feruloylquinic acids were detected in the root of A. ageratoides Turcz. It was not possible to detect any 1-acyl chlorogenic acids, any chlorogenic acids with a succinic acid substituent, or any chlorogenic acids based on muco-quinic acid.     

Synthesis and biological activity of conjugates between paclitaxel and the cell delivery vector penetratin

Synthesis of paclitaxel-penetratin (pAntp) constructs, in which the 2'- or 7-position of paclitaxel was used as the attachment site for linker connecting the drug and peptide moieties, is described. Paclitaxel-2'-pAntp[43-58]-NH(2)3b and paclitaxel-2'-pAntp[52-58]-NH(2)3c showed excellent antitumour activity against human lung and breast cancer cell lines. These conjugates were highly soluble and stable with a half-life of >8h under cell culture conditions. The drug-peptide conjugates may be therapeutically useful due to improved pharmaceutical properties.     

Reduced cardiac output is associated with decreased mitochondrial efficiency in the non-ischemic ventricular wall of the acute myocardial-infarcted dog

Cardiogenic shock is the leading cause of death among patients hospitalized with acute myocardial infarction (MI).Understanding the mechanisms for acute pump failure is therefore important.The aim of this study is to examine in anacute MI dog model whether mitochondrial bio-energetic function within non-ischemic wall regions are associated withpump failure.Anterior MI was produced in dogs via ligation of left anterior descending (LAD) coronary artery,thatresulted in an infract size of about 30% of the left ventricular wall.Measurements of hemodynamic status,mitochondrialfunction,free radical production and mitochondrial uncoupling protein 3 (UCP3) expression were determined over 24h period.Hemodynamic measurements revealed a>50% reduction in cardiac output at 24 h post infarction when com-pared to baseline.Biopsy samples were obtained from the posterior non-ischemic wall during acute infarction.ADP/Oratios for isolated mitochondria from non-ischemic myocardium at 6 h and 24 h were decreased when compared to theADP/O ratios within the same samples with and without palmitic acid (PA).GTP inhibition of (PA)-stimulated state 4respiration in isolated mitochondria from the non-ischemic wall increased by 7% and 33% at 6 h and 24 h post-infarctionrespectively when compared to sham and pre-infarction samples.This would suggest that the mitochondria are uncoupledand this is supported by an associated increase in UCP3 expression observed on western blots from these same biopsysamples.Blood samples from the coronary sinus measured by electron paramagnetic resonance (EPR) methods showedan increase in reactive oxygen species (ROS) over baseline at 6 h and 24 h post-infarction.In conclusion,mitochondrialbio-energetic ADP/O ratios as a result of acute infarction are abnormal within the non-ischemic wall.Mitochondria ap-pear to be energetically uncoupled and this is associated with declining pump function.Free radical production may beassociated with the induction of uncoupling proteins in the mitochondria.     

Chimeric Virus as a Source of the Potato Leafroll Virus Antigen

Large quantities of potato leafroll virus (PLRV) antigen are difficult to obtain because this virus accumulates in plants at a low titer. To overcome this problem, we constructed a binary vector containing chimeric cDNA, in which the coat protein (CP) gene of the crucifer infecting tobacco mosaic virus (crTMV) was substituted for the coat protein gene of PLRV. The PLRV movement protein (MP) gene, which overlaps completely with the CP gene, was doubly mutated to eliminate priming of the PLRV MP translation from ATG codons with no changes to the amino acid sequence of the CP. The untranslated long intergenic region located upstream of the CP gene was removed from the construct. Transcribed powerful tobamovirus polymerase of the produced vector synthesized PLRV CP gene that was, in turn, translated into the protein. CP PLRV packed RNAs from the helical crTMV in spherical virions. Morphology, size and antigenic specificities of the wild-type and chimeric virus were similar. The yield of isolated chimera was about three orders higher than the yield of native PLRV. The genetic manipulations facilitated the generation of antibodies against the chimeric virus, which recognize the wild-type PLRV.     

Fluorescent Labeling Preserving OCP Photoactivity Reveals Its Reorganization during the Photocycle

Orange carotenoid protein (OCP), responsible for the photoprotection of the cyanobacterial photosynthetic apparatus under excessive light conditions, undergoes significant rearrangements upon photoconversion and transits from the stable orange to the signaling red state. This is thought to involve a 12-Å translocation of the carotenoid cofactor and separation of the N- and C-terminal protein domains. Despite clear recent progress, the detailed mechanism of the OCP photoconversion and associated photoprotection remains elusive. Here, we labeled the OCP of Synechocystis with tetramethylrhodamine-maleimide (TMR) and obtained a photoactive OCP-TMR complex, the fluorescence of which was highly sensitive to the protein state, showing unprecedented contrast between the orange and red states and reflecting changes in protein conformation and the distances from TMR to the carotenoid throughout the photocycle. The OCP-TMR complex was sensitive to the light intensity, temperature, and viscosity of the solvent. Based on the observed Förster resonance energy transfer, we determined that upon photoconversion, the distance between TMR (donor) bound to a cysteine in the C-terminal domain and the carotenoid (acceptor) increased by 18 Å, with simultaneous translocation of the carotenoid into the N-terminal domain. Time-resolved fluorescence anisotropy revealed a significant decrease of the OCP rotation rate in the red state, indicating that the light-triggered conversion of the protein is accompanied by an increase of its hydrodynamic radius. Thus, our results support the idea of significant structural rearrangements of OCP, providing, to our knowledge, new insights into the structural rearrangements of OCP throughout the photocycle and a completely novel approach to the study of its photocycle and non-photochemical quenching. We suggest that this approach can be generally applied to other photoactive proteins.     

Isolation and characterization of BetaM protein encoded by ATP1B4--a unique member of the Na,K-ATPase β-subunit gene family

ATP1B4 genes represent a rare instance of the orthologous gene co-option that radically changed functions of encoded BetaM proteins during vertebrate evolution. In lower vertebrates, this protein is a β-subunit of Na,K-ATPase located in the cell membrane. In placental mammals, BetaM completely lost its ancestral role and through acquisition of two extended Glu-rich clusters into the N-terminal domain gained entirely new properties as a muscle-specific protein of the inner nuclear membrane possessing the ability to regulate gene expression. Strict temporal regulation of BetaM expression, which is the highest in late fetal and early postnatal myocytes, indicates that it plays an essential role in perinatal development. Here we report the first structural characterization of the native eutherian BetaM protein. It should be noted that, in contrast to structurally related Na,K-ATPase β-subunits, the polypeptide chain of BetaM is highly sensitive to endogenous proteases that greatly complicated its isolation. Nevertheless, using a complex of protease inhibitors, a sample of authentic BetaM was isolated from pig neonatal skeletal muscle by a combination of ion-exchange and lectin-affinity chromatography followed by SDS–PAGE. Results of the analysis of the BetaM tryptic digest using MALDI-TOF and ESI-MS/MS mass spectrometry have demonstrated that native BetaM in neonatal skeletal muscle is a product of alternative splice mRNA variant B and comprised of 351 amino acid residues. Isolated BetaM protein was also characterized by SELDI-TOF mass spectrometry before and after deglycosylation. This allowed us to determine that the carbohydrate moiety of BetaM has molecular mass 5.9 kDa and consists of short high-mannose type N-glycans. The results of direct analysis of the purified native eutherian BetaM protein provide first insights into structural properties underlying its entirely new evolutionarily acquired functions.     

Draft genome sequence of the anoxygenic filamentous phototrophic bacterium Oscillochloris trichoides subsp. DG-6

Oscillochloris trichoides is a mesophilic, filamentous, photoautotrophic, nonsulfur, diazotrophic bacterium which is capable of carbon dioxide fixation via the reductive pentose phosphate cycle and possesses no assimilative sulfate reduction. Here, we present the draft genome sequence of Oscillochloris trichoides subsp. DG-6, the type strain of the species, which has permitted the prediction of genes for carbon and nitrogen metabolism and for the light-harvesting apparatus.