Continuous-flow NMR culture system for mammalian cells

A continuous-flow NMR culture system for mammalian cells has been developed on which ³¹P-NMR experiments under complete and strictly physiologic conditions have been performed. Observations on the response of the cellular metabolism to stresses such as starvation, low temperature and changes in environmental pH monitored by ³¹P-NMR are reported. The response of the intracellular pH relative to the external pH of the growth medium is studied. We find that under the experimental conditions used there exists a ΔpH varying between less than 0.2 and more than 0.6 pH units. These results are compatible with those obtained using other techniques.     

Intracellular magnetophoresis of amyloplasts and induction of root curvature

High-gradient magnetic fields (HGMFs) were used to induce intracellular magnetophoresis of amyloplasts. The HGMFs were generated by placing a small ferromagnetic wedge into a uniform magnetic field or at the gap edge between two permanent magnets. In the vicinity of the tip of the wedge the dynamic factor of the magnetic field, (H²/2), was about 10⁹ Oe² · cm^–1, which subjected the amyloplasts to a force comparable to that of gravity. When roots of 2-d-old seedlings of flax (Linum usitatissimum L.) were positioned vertically and exposed to an HGMF, curvature away from the wedge was transient and lasted approximately 1 h. Average curvature obtained after placing magnets, wedge and seedlings on a 1-rpm clinostat for 2 h was 33 ± 5 degrees. Roots of horizontally placed control seedlings without rotation curved about 47 ± 4 degrees. The time course of curvature and changes in growth rate were similar for gravicurvature and for root curvature induced by HGMFs. Microscopy showed displacement of amyloplasts in vitro and in vivo. Studies with Arabidopsis thaliana (L.) Heynh. showed that the wild type responded to HGMFs but the starchless mutant TC7 did not. The data indicate that a magnetic force can be used to study the gravisensing and response system of roots.Abbreviations HGMF high-gradient magnetic field - emu electromagnetic units - Oe Oersted We thank Dr. John Kiss, Miami University, Ohio for providing the Arabidopsis seeds. This work was supported by NASA grant NAGW-3656     

THE LASER TOMOGRAPHICAL METHOD USINGMINIMUM OF PROJECTION FOR BIOLOGICAL OBJECT STRUCTURE STUDY

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Embryonic stem cells derived from morulae,inner cell mass,and blastocysts of mink: Comparisons of their pluripotencies

A characterization of cell lines that we derived from morulae (three lines), blastocysts (two lines), and the inner cell mass (ICM) is given. The karyotype of all the lines was normal; the genotype of four lines was XX, and four lines were genotypically XY. The pluripotencies and commitment status of the derived lines were estimated. First, there were not less than two-thirds of cells in the populations of the lines derived from morulae and the lCM with both Xs active; 70–100% of cells of the blastocyst-derived lines had one of the Xs in an inactive state. The activity of glucose-6-phosphate dehydrogenase (G6PD) in the lines (genotype XX) derived from morulae and ICM was found to be twofold higher than in lines with genotype XY, and G6PD activity was the same in the blastocyst-derived XX lines and XY lines. Second, when injected intraperitoneally into athymic mice, morulae- and ICM-derived cells gave rise to simple and complex embryoid bodies (EB) resembling to typical “cystic” mouse EBs. Third, when injected subcutaneously to athymic mice, the ICM- or morula-derived cells gave rise to typical teratomas containing derivatives of the three germ layers and components of organogenesis. Comparisons of cell lines of different derivations demonstrated that the pluripotencies of the ES cells derived from morulae or the ICM are higher than those of blastocyst derivation. © 1993 Wiley-Liss, Inc.     

Regeneration in Gastrotricha—I. Light Microscopical Observations on the Regeneration in Turbanella sp.

For the first time, the phenomenon of regeneration is reported for the phylum Gastrotricha. Adult specimens of Turbanella sp. were transected transversely and events of regeneration were observed on the light microscopical level. In both anterior and posterior fragments the wound closure was completed I day after the operation, and in 3 days the epidermis was restored entirely. The dorsal commissure of the Y-organ was rebuilt soon after wound closure. In anterior fragments cut at any level of the intestine, caudal adhesive tubes were formed anew, following the species-specific pattern. In posterior fragments, and in both fragments if transected at a pharynx level, regeneration resulted in a complete wound healing, with no new adhesive tubes being observed.     

A 13c-n.m.r. study of 1,6:2,3- and 1,6:3,4-dianhydro-β-D-hexo-pyranoses and their O-acetyl and deoxy derivatives

¹³C-N.m.r. spectra of all possible 1,6:2,3- and 1,6:3,4-dianhydro-β-D-hexo-pyranoses and their O-acetyl and deoxy derivatives are presented. Relations between chemical shifts of certain carbon atoms and the structure of the dianhydrides are outlined, and their application in structural analysis is discussed. Inversion of configuration of the oxirane ring from the endo to the exo position is associated with typical upfield-shifts for oxirane-ring carbon atoms C-2 or C-4, respectively. Possible inter-relationships between ¹³C-chemical shifts and steric and polar interactions in the dianhydro derivatives are discussed.     

Optimal Velocity and Safety of Discontinuous Conduction through the Heterogeneous Purkinje-Ventricular Junction

Slow and discontinuous wave conduction through nonuniform junctions in cardiac tissues is generally considered unsafe and proarrythmogenic. However, the relationships between tissue structure, wave conduction velocity, and safety at such junctions are unknown. We have developed a structurally and electrophysiologically detailed model of the canine Purkinje-ventricular junction (PVJ) and varied its heterogeneity parameters to determine such relationships. We show that neither very fast nor very slow conduction is safe, and there exists an optimal velocity that provides the maximum safety factor for conduction through the junction. The resultant conduction time delay across the PVJ is a natural consequence of the electrophysiological and morphological differences between the Purkinje fiber and ventricular tissue. The delay allows the PVJ to accumulate and pass sufficient charge to excite the adjacent ventricular tissue, but is not long enough for the source-to-load mismatch at the junction to be enhanced over time. The observed relationships between the conduction velocity and safety factor can provide new insights into optimal conditions for wave propagation through nonuniform junctions between various cardiac tissues.     

Uniparental Genetic Heritage of Belarusians: Encounter of Rare Middle Eastern Matrilineages with a Central European Mitochondrial DNA Pool

Ethnic Belarusians make up more than 80% of the nine and half million people inhabiting the Republic of Belarus. Belarusians together with Ukrainians and Russians represent the East Slavic linguistic group, largest both in numbers and territory, inhabiting East Europe alongside Baltic-, Finno-Permic- and Turkic-speaking people. Till date, only a limited number of low resolution genetic studies have been performed on this population. Therefore, with the phylogeographic analysis of 565 Y-chromosomes and 267 mitochondrial DNAs from six well covered geographic sub-regions of Belarus we strove to complement the existing genetic profile of eastern Europeans. Our results reveal that around 80% of the paternal Belarusian gene pool is composed of R1a, I2a and N1c Y-chromosome haplogroups – a profile which is very similar to the two other eastern European populations – Ukrainians and Russians. The maternal Belarusian gene pool encompasses a full range of West Eurasian haplogroups and agrees well with the genetic structure of central-east European populations. Our data attest that latitudinal gradients characterize the variation of the uniparentally transmitted gene pools of modern Belarusians. In particular, the Y-chromosome reflects movements of people in central-east Europe, starting probably as early as the beginning of the Holocene. Furthermore, the matrilineal legacy of Belarusians retains two rare mitochondrial DNA haplogroups, N1a3 and N3, whose phylogeographies were explored in detail after de novo sequencing of 20 and 13 complete mitogenomes, respectively, from all over Eurasia. Our phylogeographic analyses reveal that two mitochondrial DNA lineages, N3 and N1a3, both of Middle Eastern origin, might mark distinct events of matrilineal gene flow to Europe: during the mid-Holocene period and around the Pleistocene-Holocene transition, respectively.