Evaluating Plasma Skimming with Whole Blood in Small Gap Region Imitating Clearance of Blood Pumps.

Plasma skimming is the phenomenon whereby the discharge hematocrit is lower than feed hematocrit naturally occurring in the microvessels with Poiseuille flow. It has been studied in Poiseuille flow extensively. Besides, plasma skimming has also been observed and investigated in blood pumps due to its potential to prevent hemolysis by skimming blood cells out of the small gap. However, whether plasma skimming occurs in blood pumps with whole blood has not been verified. Additionally, the independent influence of rotational speed and gap size has not been clarified. Therefore, in order to lay the foundation of applying plasma skimming to the development of blood pumps and also investigate the influence of rotational speed and gap size on plasma skimming respectively, we designed a simplified geometric device which not only imitates the flow inside clearances of blood pumps, but also provides different rotational speed and gap size conditions. We first conducted the verification tests of plasma skimming using whole blood with an initial hematocrit of 44% and the gap size was varied from 10 µm to 240 µm with 10 µm interval. The plasma skimming was verified occurring when the gap was less than 70 µm at a rotational speed of 800 rpm. Since plasma skimming was confirmed, we employed 30% hematocrit blood and performed the following tests to evaluate the influence of rotational speed of 600 rpm, 700 rpm, and 800 rpm respectively. As a result, the hematocrit of sampled blood declined as the rotational speed increased from 600 rpm to 800 rpm. And there was the lowest hematocrit of 16% when the gap was adjusted to 50 µm gap size at 800 rpm. This study further promotes the possibility of applying plasma skimming to the blood pumps with higher hemocapability.

Rejuvenation of Sequoia sempervirens by Repeated Grafting of Shoot Tips onto Juvenile Rootstocks in Vitro: Model for Phase Reversal of Trees.

Repeated grafting of 1.5-centimeter long shoot tips from an adult Sequoia sempervirens tree onto fresh, rooted juvenile stem cuttings in vitro resulted in progressive restoration of juvenile traits. After four successive grafts, stem cuttings of previously adult shoots rooted as well, branched as profusely, and grew with as much or more vigor as those of seedling shoots. Reassays disclosed retention for 3 years of rooting competence at similar levels as originally restored. Adventitious shoot formation was remanifested and callus development was depressed in stem segments from the repeatedly grafted adult. The reversion was associated with appearance and disappearance of distinctive leaf proteins. Neither gibberellic acid nor N(6)-beneyladenine as nutrient supplements duplicated the graft effects.

Repression of asexual embryogenesis in vitro by some plant growth regulators.

A factor that represses asexual embryogenesis has been observed in the Rutaceae, with particularly high concentrations in the naturally monoembryonic cultivars. This investigation was an initial step towards identifying the factor. Citrus reticulata Blanco Ponkan mandarin nucellus explants and Daucus carota L. 'Queen Anne's Lace' callus were employed to examine effects of known plant growth regulators and to determine possible identity of one or more of them with the repressive factor. The chalazal halves of ovules of C. media L. 'Citron of Commerce' were used as control repressor source. Embryo initiation and growth of both test tissues were depressed markedly by 2,4-D, abscisic acid and ethephon. Slight inhibitions were obtained with IAA, kinetin and gibberellic acid. Recovery from the repressor did not occur readily in Citrus nucellus following recultures in citron-ovule-free medium; carrot callus resumed normal embryogenesis immediately upon transfer to suppressor-free medium. The repression by natural sources apparently involved the combined action of some or all natural hormones that are generically related to the above.

Effects of ethephon, ethylene, and 2,4-dichlorophenoxyacetic Acid on asexual embryogenesis in vitro.

Asexual embryogenesis in Daucus carota L. ;Queen Anne's Lace' callus was suppressed by Ethephon, ethylene, and 2,4-dichlorophenoxyacetic acid (2,4-D). The Ethephon effect could be attributed to volatile and nonvolatile substances. The volatile component was probably entirely ethylene. Ethylene was liberated in the cultures in direct proportion to Ethephon added to the medium. Autoclaving of Ethephon caused a substantial decrease of measurable ethylene. Continuous exposure of callus to 5 mul/l ethylene depressed somatic cell embryogenesis, but not markedly. Depression of embryogenesis by 2,4-D was unrelated to ethylene evolution.

Quantitative analysis of the fate of exogenous DNA in Nicotiana protoplasts.

After a 5-hour incubation of protoplasts of Nicotiana tabacum L. ;Xanthi' with (3)H-DNA (7.26 mug/ml) from N. tabacum L. ;Xanthi nc' 3.5% of the initial radioactivity was found in acid-insoluble substances of the protoplasts. The addition of DEAE-dextran and poly-l-lysine to the incubation medium nearly doubled radioactivity adsorption. The absorption was inhibited by 2,4-dinitrophenol, KCN, and low temperature (0 C); this inhibition could not be reversed by exogenous ATP. About 500 tobacco plants established from protoplasts of a normally tobacco-mosaic virus-susceptible cultivar that had been allowed to absorb DNA prepared from a resistant cultivar did not show transfer of the virus-resistant gene.A detailed analysis was performed of the disposition of exogenous DNA in plant protoplasts, by employing Escherichia coli(3)H-DNA and Nicotiana glutinosa protoplasts. In 5 to 20 hours, about 10% of the (3)H-DNA entered the protoplasts. Competition experiments between the (3)H-DNA and unlabeled DNA or thymidine showed that the entry occurred as undegraded (3)H-DNA. Examination of intraprotoplast fractions revealed that 60 to 80% of the absorbed radioactivity resided in the "soluble" fraction of the cytoplasm and 20% in the nuclear fraction. The mitochondrion fraction also contained measurable radioactivity. Sizing on sucrose density gradients showed that the bulk of the absorbed E. coli DNA had been depolymerized. Of the incorporated radioactivity, 15% was accountable as DNA, exogenous as well as resynthesized, and 15% as RNA, protein, and other cell constituents. DNA/DNA hybridization test indicated that 17.6% of the re-extractable (3)H-DNA retained homology with the E. coli DNA; this was equivalent to 2.6% of the absorbed radioactivity. Resynthesized receptor protoplast DNA was represented by a fraction at least 1.7% of the total absorbed radioactivity. The amount of bacterial DNA remaining in protoplasts suggests that each protoplast retained 2.3 x 10(-15)g donor DNA, or approximately half of the E. coli genome.

Tissue culture propagation of tropical foliage plants.

Procedures were established for clonal multiplication in vitro of Cordyline terminalis Kunth, Dracaena godseffiana Hort., Scindapsus aureus Engler, and Syngonium podophyllum Schott. Shoot tips of actively growing terminals were selected as explants for Cordyline and Dracaena, and lateral buds were employed for Scindapsus and Syngonium. The basal nutrient medium contained Murashige and Skoog salts, 3% sucrose, 100 mg per 1 i-inositol, and 0.4 mg per 1 thiamine-HCl. The optima with respect to auxin, cytokinin, adenine sulfate-2H2O, and NaH2PO4-H2O addenda were determined. Also assessed were the influences of certain physical qualities of the nutrient medium and of the light intensity of the culture environment. The multiplication of each of the four plants was achieved by repeatedly subculturing the shoots that arose in vitro. Rates of plant increase per year per explant were calculated conservatively to be as follows: Syngonium, 5,000:Scindapsus, 100,000; Dracena, 300,000; and Cordyline, 500,000.

Influence of the nutrient medium on the recovery of dividing cells from tobacco protoplasts.

Systematic tests resulted in a nutrient solution containing the following, in milligrams per liter, for the culture of protoplasts isolated from Nicotiana tabacum L. callus cells: Murashige and Skoog salts (T. Murashige and F. Skoog, 1962. Physiol. Plant. 15: 473-497); sucrose, 15,000; mannitol, 110,000; alpha-naphthaleneacetic acid, 0.6; kinetin, 0-0.1; thiamine.HCl, 10; pyridoxine.HCl, 10; nicotinic acid, 5; myo-inositol, 100; and glycine, 2. In this medium, regeneration of cell wall has been observed in 85% and resumption of cell division among 35% of the protoplast isolates.

Evaluation of parameters in the isolation of viable protoplasts from cultured tobacco cells.

A systematic evaluation disclosed the following conditions to be optimum for the isolation of viable protoplasts from cultured cells of Nicotiana tabacum L. ;Bright Yellow' grown in liquid suspensions: (a) the cell culture in the early phase of cell number increase, (b) an enzyme mixture of 1% cellulase "Onozuka" and 0.2% Macerozyme, (c) an enzyme solution pH of 4.7 or 5.7, (d) a 2- to 3-hr incubation period, (e) 5 ml of enzyme solution per 500 mg cells and contained in a 50-ml Delong flask, (f) agitation on a gyrotory shaker at 50 rpm, and (g) 0.3 to 0.8 m mannitol as osmoticum in the cell enzyme mixture. The incubation temperature may be varied from 22 to 37 C. The procedure enabled 30% of the tobacco cells to form protoplasts, 80% of which regenerated cell walls in 4 days and 40% resumed cell division activity when returned to cell culture medium.

Starch accumulation in shoot-forming tobacco callus cultures.

Microscopic histochemical examinations of cultured tobacco callus disclosed a strong correlation between starch accumulation and shoot initiation. The accumulation started before any observable organized development and was heaviest in cells of loci which ultimately gave rise to organ primordia. Treatment of tissue cultures with gibberellin prevented starch accunmulation and organ formation.