Pigment containing lipid vesicles. I. Preparation and characterization of chlorophyll a-lecithin vesicles.

Vesicles obtained by sonication of chlorophyll a-lecithin mixtures dispersed in anaqueous medium closely resemble the well-characterized vesicles similarly prepared from pure lipids. They are bounded by one spherical lipid bilayer which contains the chlorophyll a. Appropriate conditions for sonication prevent substantial degradation of the membrane constituents. Up to one chlorophyll a molecule per 55 lecithins can be incorporated into membranes. The average Stokes' radius of the vesicles determined by analytical sieve chromatography is 102 +/- 5 A and independent of the chloropyll a content. The membrane is visible in the electron-microscope when the vesicles are treated with osmium tetroxide prior to negative staining. The osmium fixation is, however, not strong enough to allow for a preparation of the vesicles for thin sectioning (dehydration, embedding in epoxide).     

pH shifts evoked by neuronal stimulation in slices of rat hippocampus

Extracellular pH was measured with ion-selective microelectrodes in 500-microns thick slices of the CA1 region of the rat hippocampus. The center of the slice was 0.24 pH units more acidic than the surface, thus creating a decreasing pH gradient from the surface to the center, most likely owing to increased anaerobic metabolism. Stimulation at various frequencies created a transient alkaline shift of 0.03 pH unit, followed by a sustained acidification (0.05 pH unit above baseline). The same pattern was seen in both cell body and dendritic layers. The presence of the alkaline shift in slices in vitro is especially significant, implying that it is not due to alterations in blood flow. The HCO3- -Cl- transport inhibitor SITS and the carbonic anhydrase inhibitor acetazolamide increased the alkaline shift to 0.06 and 0.23, respectively. The acid shift was influenced by the Na+-H+ transport inhibitor amiloride, with a reduction of about 50%. Possible mechanisms for these stimulus-evoked changes as discussed. The most likely cause for the alkaline shift is bicarbonate accumulation in the extracellular space. Hydrogen ion and lactic acid release are seen as the major factors contributing to the sustained acid shift.     

Safety and economic aspects of continuous mammalian cell culture.

Mammalian cell cultures are the most appropriate host cells for recombinant DNA derived products if complex protein structures have to be synthesized in their native form. Due to their physiological behaviour they grow either adherent or in suspension. For the attachment of adherent cells, microcarriers or wire springs can be applied to increase the internal surface of the bioreactor. Both systems provide a simplified media exchange but, however, show some limitations in scale up. In contrast, suspension culture systems as homogeneous systems independent of any carrier have not shown any limitation in scale up. Because most cell lines which are of commercial interest grow in suspension, this technology is best advanced and used in batch and continuous mode. Although mammalian cell cultures are sensitive to hydrodynamic shear forces, technologies for deep tank production are developed which allow stirrer tip speed of up to 1.5 m s-1 sufficient for oxygen uptake, suspension of cells and homogeneous supply with nutrients. For long term bioprocesses without selection pressure it has to be considered that transformed cell lines might show genetic instability due to their variations of chromosomes. In addition, sterile technology becomes an important factor in long term bioprocesses. The decision as to which cell culture system should be chosen, whether batch or continuous processes should be applied essentially is based on the capital investment, the amount of material to be produced, genetic stability of the production cell line, reliability of sterile technology and the flexibility required in the production plant. Under the assumption that 20 kg of a protein have to be produced per year and the same product concentrations in the harvest fluid are reached in the batch process and for instance in the chemostat, it can be considered that the capital investment for one 10,000 l batch process and a 2 x 2,000 l continuous process, necessary to produce the amount of material, is comparable. Risk of microbial contamination or technical failure can be considered to be fairly low in the batch process. The economic risk for long term bioprocess in the chemostat can be expected to be medium and high in the perfusion system which is in the large scale not technically fully satisfactory. In addition, due to the longer down time period after contaminations and the start up of the continuous process, the annual yield of the batch process can be considered to be higher.(ABSTRACT TRUNCATED AT 400 WORDS)     

CD62/P-selectin recognition of myeloid and tumor cell sulfatides.

CD62, also called PADGEM protein, GMP-140, or P-selectin, is a granule membrane protein of endothelial cells and platelets that is mobilized to the plasma membrane following exposure to mediators such as thrombin, histamine, complement components, or peroxides. Data presented to date suggest that one ligand of CD62 includes CD15 (Lewis x determinant) and sialic acid. We show here that sulfatides, heterogeneous 3-sulfated galactosyl ceramides, are an apparently unrelated ligand of CD62. Sulfatides are expressed on the plasma membrane of, and are excreted by, granulocytes, and constitute the principal ligand for CD62 on the plasma membrane of some tumor cells. CD62 binds to sulfatides adsorbed to plastic as avidly as it binds to myeloid or tumor cells. We find that granulocytes excrete sulfatides at a rate predicted to allow them to be rapidly released from CD62 once they have exited the bloodstream.     

Iron and heme contents of the extracellular hemoglobins and chlorocruorins of annelids

1. A survey of the literature on the extracellular hemoglobins and chlorocruorins of over 30 species of annelids, covering the last 30 years, shows that the range of iron content is 0.211-0.265 wt.% (mean = 0.228 +/- 0.013, N = 28) and the range of the heme content is 1.83-3.64 wt.% (mean = 2.60 +/- 0.38, N = 29). 2. There is relatively little scatter in the values of the experimental iron contents and only one of the 28 values is clearly outside the standard deviation range. 3. The values of heme contents are much more scattered, with seven values, clearly outside the standard deviation limits. 4. The aberrant cases are discussed and it is noted that the mean heme content of 2.60 wt.% corresponds to an iron content of 0.236 wt.% in excellent agreement with the mean iron content of 0.228 wt.%. 5. This result suggests strongly that experimental values of iron and heme contents outside the ranges of 0.211-0.243 and 2.3-2.7 wt.%, respectively, corresponding to a minimum molecular mass outside the range 23,000-26,000, should be regarded with caution.