Rapid, sequential changes in surface morphology of PC12 pheochromocytoma cells in response to nerve growth factor

The effect of nerve growth factor (NGF), a substance that promotes the differentiation and maintenance of certain neurons, was studied via scanning electron microscopy utilizing the PC12 clonal NGF-responsive pheochromocytoma cell line. After 2-4 d of exposure to NGF, these cells acquire many of the properties of normal sympathic neurons. However, by phase microscopy, no changes are discernible within the first 12-18 h. Since the primary NGF receptor appears to be a membrane receptor, it seemed likely that some of the initial responses to the factor may be surface related. PC12 cells maintained without NGF are round to ovoid and have numerous microvilli and small blebs. After the addition of NGF, there is a rapidly initiated sequential change in the cell surface. Ruffles appear over the dorsal surface of the cells with 1 min, become prominent by 3 min, and almost disappear by 7 min. Microvilli, conversely, disappear as the dorsal ruffles become prominent. Ruffles are seen at the the periphery of cell at 3 min, are prominent on most of the cells by 7 min and are gone by 15 min. The surface remains smooth from 15 min until 45 min when large blebs appear. The large blebs are present on most cells at 2 h and are gone by 4 h. The surface remains relatively smooth until 6-7 h of NGF treatment, when microvilli reappear as small knobs. These microvilli increase in both number and length to cover the cell surface by 10 h. These changes were not observed with other basic proteins, with α-bungarotoxin (which binds specifically to PC12 membranes), and were not affected by an RNA synthesis inhibitor that blocks initiation of neurite outgrowth. Changes in the cell surface architecture appear to be among the earlist NGF responses yet detected and may represent or reflect primary events in the mechanism of the factor’s action.     

Transformation of LMTK− cells with purified HLA class I genes

The expression of two different HLA class I genes was observed after transformation of LMTK- cells. The corresponding class I molecules reacted differentially with monomorphic monoclonal antibodies (m.Ab). Absorption and elution studies of the human alloantibodies reacting with the transformed cells and cellular radioimmunoassay of these cells with polymorphic m.Ab resulted in the identification of HLA-A3 and CW3 molecules. These transformed cells were used to immunize C3H mice and induce the production of xenogeneic antisera, which, following absorption, showed polymorphic reactivity with human cells, suggesting that some of these sera could be used as typing reagents.     

Molecular markers and protein quantities as genetic descriptors in maize. I. Genetic diversity among 21 inbred lines

Twenty-one maize (Zea mays L.) inbred lines were analysed using isozyme electrophoresis, restriction fragment length polymorphism (RFLP), and two-dimensional electrophoresis of denatured proteins (2-D PAGE). Our goal was (1) to assess the genetic variability among these lines which are potential progenitors for the development of forage maize hybrids in Europe, and (2) to compare the relationship pattern revealed by the polymorphism at marker loci with the one derived from the amount of protein variability assessed by computer-assisted analysis of the 2-D electrophoregrams. Fourteen markers were obtained from isozyme polymorphism, 84 from the restriction fragment length polymorphism, and 70 from protein shifts revealed by 2-D PAGE. The Rogers' distance computed on the set of molecular markers was the most efficient to describe the pedigree relationships between lines. Quantitative protein data gave a picture of relationships between lines clearly different from the monogenic markers. When unrelated pairs of lines were considered, the Rogers' distance was weakly correlated to distances based on quantitative variations in the amount of protein which may be consistent with their polygenic control and the occurrence of gene interactions.     

Increase of capsular material thickness following in vivo growth of virulent Streptococcus suis serotype 2 strains

Abstract Protein profile and capsular material thickness of Streptococcus suis serotype 2 strains were compared after in vitro and in vivo growth. Three virulent and one avirulent strains were used. These strains were grown in Brain Heart Infusion (BHI) broth, cells were collected by centrifugation, resuspended in a sterile saline solution and injected in diffusion chambers. The devices were then inserted in rat abdomens for 17 h. In vitro grown strains were also inoculated into fresh BHI broth and cultivated for 17 h at 37°C. In vivo as well as in vitro grown bacteria were harvested by centrifugation, processed in a French pressure cell, treated with lysozyme and centrifuged to collect cell proteins for SDS-PAGE analysis. Transmission electron microscopy using polycationic ferritin labeling to stabilize capsular material was also carried out. No significant modification was noted in the protein profile for any strain after in vivo growth except for a 39 kDa protein of one virulent strain. On the other hand, an increase in thickness of capsular material was noted for the three in vivo grown virulent strains while no change was noted for the avirulent strain. This increase in capsular material thickness of virulent strains was accompanied by an increased resistance to killing by pig polymorphonuclear leukocytes. The capacity to produce more capsular material in vivo seems to be an attribute of some virulent S. suis serotype 2 strains.     

Two new disposable bioreactors for plant cell culture: The wave and undertow bioreactor and the slug bubble bioreactor

The present article describes two novel flexible plastic-based disposable bioreactors. The first one, the WU bioreactor, is based on the principle of a wave and undertow mechanism that provides agitation while offering convenient mixing and aeration to the plant cell culture contained within the bioreactor. The second one is a high aspect ratio bubble column bioreactor, where agitation and aeration are achieved through the intermittent generation of large diameter bubbles, "Taylor-like" or "slug bubbles" (SB bioreactor). It allows an easy volume increase from a few liters to larger volumes up to several hundred liters with the use of multiple units. The cultivation of tobacco and soya cells producing isoflavones is described up to 70 and 100 L working volume for the SB bioreactor and WU bioreactor, respectively. The bioreactors being disposable and pre-sterilized before use, cleaning, sterilization, and maintenance operations are strongly reduced or eliminated. Both bioreactors represent efficient and low cost cell culture systems, applicable to various cell cultures at small and medium scale, complementary to traditional stainless-steel bioreactors.