Regulation of VEGF-induced endothelial cell migration by mitochondrial reactive oxygen species

Endothelial migration is a crucial aspect of a variety of physiologic and pathologic conditions including atherosclerosis and vascular repair. Reactive oxygen species (ROS) function as second messengers during endothelial migration. Multiple intracellular sources of ROS are regulated by cellular context, external stimulus, and the microenvironment. However, the predominant source of ROS during endothelial cell (EC) migration and the mechanisms by which ROS regulate cell migration are incompletely understood. In this study, we tested the hypothesis that mitochondria-derived ROS (mtROS) regulate EC migration. In cultured human umbilical vein endothelial cells, VEGF increased mitochondrial metabolism, promoted mtROS production, and induced cell migration. Either the targeted mitochondrial delivery of the antioxidant, vitamin E (Mito-Vit-E), or the depletion of mitochondrial DNA abrogated VEGF-mediated mtROS production. Overexpression of mitochondrial catalase also inhibited VEGF-induced mitochondrial metabolism, Rac activation, and cell migration. Furthermore, these interventions suppressed VEGF-stimulated EC migration and blocked Rac1 activation in endothelial cells. Constitutively active Rac1 reversed Mito-Vit-E-induced inhibition of EC migration. Mito-Vit-E also attenuated carotid artery reendothelialization in vivo. These results provide strong evidence that mtROS regulate EC migration through Rac-1.     

Umbilical Cord Milking Improves Transition in Premature Infants at Birth

Background

Umbilical cord milking (UCM) improves blood pressure and urine output, and decreases the need for transfusions in comparison to immediate cord clamping (ICC). The immediate effect of UCM in the first few minutes of life and the impact on neonatal resuscitation has not been described.

Methods

Women admitted to a tertiary care center and delivering before 32 weeks gestation were randomized to receive UCM or ICC. A blinded analysis of physiologic data collected on the newborns in the delivery room was performed using a data acquisition system. Heart rate (HR), SpO₂, mean airway pressure (MAP), and FiO₂ in the delivery room were compared between infants receiving UCM and infants with ICC.

Results

41 of 60 neonates who were enrolled and randomized had data from analog tracings at birth. 20 of these infants received UCM and 21 had ICC. Infants receiving UCM had higher heart rates and higher SpO₂ over the first 5 minutes of life, were exposed to less FiO₂ over the first 10 minutes of life than infants with ICC.

Conclusions

UCM when compared to ICC had decreased need for support immediately following delivery, and in situations where resuscitation interventions were needed immediately, UCM has the advantage of being completed in a very short time to improve stability following delivery.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT01434732","term_id":"NCT01434732"}}NCT01434732     

Enhanced inhibition of polymerization of sickle cell hemoglobin in the presence of recombinant mutants of human fetal hemoglobin with substitutions at position 43 in the gamma-chain

Four recombinant mutants of human fetal hemoglobin [Hb F (alpha2gamma2)] with amino acid substitutions at the position 43 of the gamma-chain, rHb (gammaD43L), rHb (gammaD43E), rHb (gammaD43W), and rHb (gammaD43R), have been expressed in our Escherichia coli expression system and used to investigate their inhibitory effect on the polymerization of deoxygenated sickle cell hemoglobin (Hb S). Oxygen-binding studies show that rHb (gammaD43E), rHb (gammaD43W), and rHb (gammaD43R) exhibit higher oxygen affinity than human normal adult hemoglobin (Hb A), Hb F, or rHb (gammaD43L), and all four rHbs are cooperative in binding O2. Proton nuclear magnetic resonance (NMR) studies of these four rHbs indicate that the quaternary and tertiary structures around the heme pockets are similar to those of Hb F in both deoxy (T) and liganded (R) states. Solution light-scattering experiments indicate that these mutants remain mostly tetrameric in the liganded (R) state. In equimolar mixtures of Hb S and each of the four rHb mutants (gammaD43L, gammaD43E, gammaD43R, and gammaD43W), the solubility (Csat) of each of the pairs of Hbs is higher than that of a similar mixture of Hb S and Hb A, as measured by dextran-Csat experiments. Furthermore, the Csat values for Hb S/rHb (gammaD43L), Hb S/rHb (gammaD43E), and Hb S/rHb (gammaD43R) mixtures are substantially higher than that for Hb S/Hb F. The results suggest that these three mutants of Hb F are more effective than Hb F in inhibiting the polymerization of deoxy-Hb S in equimolar mixtures.     

Development of a minimal defined medium for recombinant human interleukin-3 production by Streptomyces lividans 66

A systematic approach was developed to identify and optimize the essential amino acids in defined minimal medium for the production of recombinant human interleukin 3 (rHuIL-3) by Streptomyces lividans. Starvation trials were carried out initially to narrow down the number of probable essential amino acids from an initial number of 20 to 8. Then a screening mixture experiment was designed and performed with the eight identified amino acids and distance-based multivariate analysis was employed to rank the probable essential amino acids regarding both growth and product formation. Following this procedure, the search was narrowed to four amino acids (Asp, Leu, Met, and Phe). Finally, a mixture design experiment known as the simplex lattice design was carried out and the composition of the optimum minimal medium was found (Asp 53%, Met 5%, and Phe 42%).     

Cardiac high molecular weight calmodulin-binding protein is homologous to calpastatin I and calpastatin II

Calpastatin is an endogenous inhibitor of calpain, which has been implicated in various physiological and pathological processes. In the present study we determined the molecular and inhibitory properties of HMWCaMBP, calpastatin I, and calpastatin II. Western blot analysis with antibodies raised against either full length HMWCaMBP or internal peptides that are common to all isoforms showed that all three homologs have common antigenic epitopes. However, additional Western blot analysis with N-terminal specific antibodies showed that all three proteins are different at the N-terminal end. HMWCaMBP is clearly different from two other homologues, calpastatin I and II, at the N-terminal end. In addition, HMWCaMBP also showed the same affinities for m-calpain as calpastatin I and calpastatin II. Our findings suggest that HMWCaMBP is the homolog of calpastatin and may be a CaM-binding form of calpastatin.     

Kinship and Deep History: Exploring Connections between Culture Areas, Genes, and Languages

Large-scale geographic variation in kinship systems may have deep roots. A number of authors now argue for an "emerging synthesis," with genetic, linguistic, and archeological findings coming together to paint a consistent picture of large-scale population spreads in prehistory. This article explores a social structural dimension of this synthesis: Major culture areas based on variation in kinship systems correspond closely—yet not perfectly—to genetic and linguistic clusters identified by other researchers. Thus it may be possible to reconstruct: (1) a set of "primary" culture areas corresponding to major population blocs and associated with ancient demic expansions and parallel transmission of genes and culture, and (2) a smaller set of overlying "secondary" culture areas of more recent origin that do not map onto genetic subdivisions and result from changes in subsistence or political economy independent of large-scale demic expansions. I also review latitudinal variation in kinship systems. [Keywords: culture areas, demic expansions, kinship (prehistory), protolanguages]