scholarly journals Schwann cell proliferation in vitro is under negative autocrine control.

1990 ◽  
Vol 111 (6) ◽  
pp. 2663-2671 ◽  
Author(s):  
D Muir ◽  
S Varon ◽  
M Manthorpe
Keyword(s):  
Molecular Weight ◽  
Schwann Cell ◽  
Molecular Mass ◽  
Conditioned Medium ◽  
Schwann Cells ◽  
Cell Cultures ◽  
Antiproliferative Activity ◽  
Gel Filtration ◽  
Molecular Weight Form

In healthy adult peripheral nerve, Schwann cells are believed to be generally quiescent. Similarly, cultures of isolated rat sciatic nerve Schwann cells hardly proliferate in serum-supplemented medium. The possibility that Schwann cells negatively regulate their own proliferation was supported by the demonstration that conditioned media from Schwann cell cultures inhibited the proliferation of mitogen-stimulated test cultures. The inhibition could be complete, was dose dependent, and was exhibited when the test Schwann cells were under the influence of different types of mitogens such as cholera toxin, laminin, and living neurons. The inhibition of proliferation was completely reversible and a rapid doubling of cell number resulted when treatment with conditioned medium was withdrawn from mitogen-stimulated Schwann cells. Conditioned medium from cholera toxin-stimulated and immortalized Schwann cell cultures contained less antiproliferative activity than that found in medium from quiescent Schwann cell cultures. However, media conditioned by two actively proliferating rat Schwannoma cell lines were rich sources of antiproliferative activity for Schwann cells. Unlike the mitogen-stimulated Schwann cells, whose proliferation could be inhibited completely, the immortalized and transformed Schwann cell types were nearly unresponsive to the antiproliferative activity. The antiproliferative activity in Schwann and Schwannoma cell conditioned media was submitted to gel filtration and SDS-PAGE. The activity exists in at least two distinct forms: (a) a high molecular weight complex with an apparent molecular mass greater than 1,000 kD, and (b) a lower molecular weight form having a molecular mass of 55 kD. The active 55-kD form could be derived from the high molecular weight form by gel filtration performed under dissociating conditions. The 55-kD form was further purified to electrophoretic homogeneity. These results suggest that Schwann cells produce an autocrine factor, which we designate as a "neural antiproliferative protein," which completely inhibits the in vitro proliferation of Schwann cells but not that of immortalized Schwann cells or Schwannoma lines.

scholarly journals Studies on cultured Schwann cells: the induction of myelin synthesis, and the control of their proliferation by a new growth factor

1981 ◽  
Vol 95 (1) ◽  
pp. 215-230
Author(s):  
J. P. Brockes ◽  
K. J. Fryxell ◽  
G. E. Lemke
Keyword(s):  
Molecular Weight ◽  
Growth Factor ◽  
Schwann Cell ◽  
Schwann Cells ◽  
Solid Phase ◽  
Immunological Methods ◽  
Subunit Molecular Weight ◽  
Cultured Schwann Cells

We have recently described the use of immunological methods to identify and purify rat Schwann cells. In dissociated cultures of neonatal sciatic nerve, all of the cells can be identified by antigenic criteria as either Schwann cells or fibroblasts. The fibroblasts may be removed by treatment with antiserum to the Thy-1 antigen and complement. The purified Schwann cells have been used to study the regulation of the expression of myelin components, and the stimulation of Schwann cell division by a soluble growth factor. Among the components of myelin, we have concentrated on the peripheral myelin glycoprotein P0, which constitutes 50–60% of the protein in peripheral myelin. We have studied the distribution of P0 in vitro and in vivo by immunofluorescence, immuno-autoradiography on SDS gels, and solid-phase radioimmunoassay. Our results support the hypothesis that P0 is induced specifically as a consequence of the interaction between the Schwann cell and the myelinated type of axon. The level of P0 in the myelin membrane is at least 1000-fold higher than in the Schwann cell membrane. Purified Schwann cells divide very slowly in a conventional tissue culture medium. This has allowed us to purify a new growth factor from extracts of brain and pituitary, tentatively named Glial Growth Factor (GGF). The activity resides in a basic protein with a native molecular weight of 6 × 10(4) daltons and a subunit molecular weight of 3 × 10(4) daltons, which is active at levels comparable to those of epidermal growth factor. GGF is mitogenic for Schwann cells, astrocytes and muscle fibroblasts.

scholarly journals Interleukin-4 Regulates the Expression of CD209 and Subsequent Uptake of Mycobacterium leprae by Schwann Cells in Human Leprosy

2010 ◽  
Vol 78 (11) ◽  
pp. 4634-4643 ◽  
Author(s):  
Rosane M. B. Teles ◽  
Stephan R. Krutzik ◽  
Maria T. Ochoa ◽  
Rosane B. Oliveira ◽  
Euzenir N. Sarno ◽  
...  
Keyword(s):  
Schwann Cell ◽  
Peripheral Nerve ◽  
Schwann Cells ◽  
Primary Cultures ◽  
Mycobacterium Leprae ◽  
Microbial Pathogens ◽  
Interleukin 4 ◽  
Common Mechanism ◽  
Specific Cell

ABSTRACT The ability of microbial pathogens to target specific cell types is a key aspect of the pathogenesis of infectious disease. Mycobacterium leprae, by infecting Schwann cells, contributes to nerve injury in patients with leprosy. Here, we investigated mechanisms of host-pathogen interaction in the peripheral nerve lesions of leprosy. We found that the expression of the C-type lectin, CD209, known to be expressed on tissue macrophages and to mediate the uptake of M. leprae, was present on Schwann cells, colocalizing with the Schwann cell marker, CNPase (2′,3′-cyclic nucleotide 3′-phosphodiesterase), along with the M. leprae antigen PGL-1 in the peripheral nerve biopsy specimens. In vitro, human CD209-positive Schwann cells, both from primary cultures and a long-term line, have a higher binding of M. leprae compared to CD209-negative Schwann cells. Interleukin-4, known to be expressed in skin lesions from multibacillary patients, increased CD209 expression on human Schwann cells and subsequent Schwann cell binding to M. leprae, whereas Th1 cytokines did not induce CD209 expression on these cells. Therefore, the regulated expression of CD209 represents a common mechanism by which Schwann cells and macrophages bind and take up M. leprae, contributing to the pathogenesis of leprosy.

Remyelination of Demyelinated CNS Axons by Transplanted Human Schwann Cells: The Deleterious Effect of Contaminating Fibroblasts

2001 ◽  
Vol 10 (3) ◽  
pp. 305-315 ◽  
Author(s):  
C. M. H. Brierley ◽  
A. J. Crang ◽  
Y. Iwashita ◽  
J. M. Gilson ◽  
N. J. Scolding ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
Schwann Cell ◽  
Schwann Cells ◽  
Axonal Degeneration ◽  
Autologous Transplantation ◽  
Growth Factor Receptor ◽  
Adult Rats ◽  
Demyelinating Lesions ◽  
Cell Implantation

Areas of demyelination can be remyelinated by transplanting myelin-forming cells. Schwann cells are the naturally remyelinating cells of the peripheral nervous system and have a number of features that may make them attractive for cell implantation therapies in multiple sclerosis, in which spontaneous but limited Schwann cell remyelination has been well documented. Schwann cells can be expanded in vitro, potentially affording the opportunity of autologous transplantation; and they might also be spared the demyelinating process in multiple sclerosis. Although rat, cat, and monkey Schwann cells have been transplanted into rodent demyelinating lesions, the behavior of transplanted human Schwann cells has not been evaluated. In this study we examined the consequences of injecting human Schwann cells into areas of acute demyelination in the spinal cords of adult rats. We found that transplants containing significant fibroblast contamination resulted in deposition of large amounts of collagen and extensive axonal degeneration. However, Schwann cell preparations that had been purified by positive immunoselection using antibodies to human low-affinity nerve growth factor receptor containing less than 10% fibroblasts were associated with remyelination. This result indicates that fibroblast contamination of human Schwann cells represents a greater problem than would have been appreciated from previous studies.

HEMODIALYSIS OF THE RAT

1966 ◽  
Vol 44 (5) ◽  
pp. 849-859 ◽  
Author(s):  
Sumner M. Robinson ◽  
David A. Hurwitz ◽  
Robert Louis-Ferdinand ◽  
William F. Blatt
Keyword(s):  
Molecular Weight ◽  
Gel Filtration ◽  
Low Molecular Weight

A technique is described for hemodialysis of either anesthetized or non-restrained rats. In the apparatus the dialysis plates of an autoanalyzer system are used with only minor modification. The efficiency of this method has been evaluated with regard to the clearance of saccharides, both in vitro and in vivo, as well as the extraction of nitrogenous low molecular weight moieties from circulating blood. Approximately 50% of the dialyzable material was obtained in a 1-hour dialysis. Further fractionation of the dialyzate was accomplished by gel filtration (Sephadex G-25).

Purification and characterization of prostate-specific antigen (PSA) complexed to alpha 1-antichymotrypsin: potential reference material for international standardization of PSA immunoassays

1995 ◽  
Vol 41 (9) ◽  
pp. 1273-1282 ◽  
Author(s):  
Z Chen ◽  
A Prestigiacomo ◽  
T A Stamey
Keyword(s):  
Molecular Mass ◽  
Prostate Specific Antigen ◽  
Specific Antigen ◽  
Gel Filtration ◽  
Exchange Chromatography ◽  
Molar Ratio ◽  
Free Psa ◽  
Apparent Homogeneity ◽  
International Standardization

Abstract We describe for the first time a protocol to purify to apparent homogeneity an in vitro-prepared complex of prostate-specific antigen (PSA) and alpha 1-antichymotrypsin (ACT) by using a combination of gel filtration and ion-exchange chromatography. The purity of the PSA-ACT complex was confirmed by gel electrophoresis and Western blot. The PSA-ACT complex was stable in the pH range 6.0 to 7.8; it was also stable in various matrices, temperatures, and high concentrations of salt. Purification of the PSA-ACT complex was highly reproducible. An absorptivity of 0.99 L x g-1 x cm-1 at 280 nm was assigned to the PSA-ACT complex, based on amino acid analysis. Because PSA and ACT bind in a 1:1 molar ratio, we determined the molecular mass of the PSA-ACT complex as the mass encoded by the cDNA of ACT (plus 26% carbohydrate) plus the molecular mass of PSA (28,430 Da), which totals 89,280 Da. Using this material, we made two common calibrators, one of 100% PSA-ACT complex and one of 90% PSA-ACT complex plus 10% free PSA by volume (90:10 calibrator). Substitution of these calibrators for the manufacturers' calibrators in nine commercial immunoassays substantially reduced differences between immunoassays, especially for serum PSA values between 4 and 10 micrograms/L. The 90:10 calibrator is recommended as a universal calibrator for international standardization of PSA immunoassays.

scholarly journals Isolation and Characterization of Two Proteins fromMoraxella catarrhalis That Bear a Common Epitope

1998 ◽  
Vol 66 (9) ◽  
pp. 4374-4381 ◽  
Author(s):  
John C. McMichael ◽  
Michael J. Fiske ◽  
Ross A. Fredenburg ◽  
Deb N. Chakravarti ◽  
Karl R. VanDerMeid ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Bacterial Attachment ◽  
Vaccine Candidates ◽  
Isolation And Characterization ◽  
Sds Page

ABSTRACT The UspA1 and UspA2 proteins of Moraxella catarrhalisare potential vaccine candidates for preventing disease caused by this organism. We have characterized both proteins and evaluated their vaccine potential using both in vitro and in vivo assays. Both proteins were purified from the O35E isolate by Triton X-100 extraction, followed by ion-exchange and hydroxyapatite chromatography. Analysis of the sequences of internal peptides, prepared by enzymatic and chemical cleavage of the proteins, revealed that UspA1 and UspA2 exhibited distinct structural differences but shared a common sequence including an epitope recognized by the monoclonal antibody 17C7. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), purified UspA1 exhibited a molecular weight of approximately 350,000 when unheated and a molecular weight of 100,000 after being heated for 10 min at 100°C. In contrast, purified UspA2 exhibited an apparent molecular weight of 240,000 by SDS-PAGE that did not change with the length of time of heating. Their sizes as determined by gel filtration were 1,150,000 and 830,000 for UspA1 and UspA2, respectively. Preliminary results indicate the proteins have separate functions in bacterial pathogenesis. Purified UspA1 was found to bind HEp-2 cells, and sera against UspA1, but not against UspA2, blocked binding of the O35E isolate to the HEp-2 cells. UspA1 also bound fibronectin and appears to have a role in bacterial attachment. Purified UspA2, however, did not bind fibronectin but had an affinity for vitronectin. Both proteins elicited bactericidal antibodies in mice to homologous and heterologous disease isolates. Finally, mice immunized with each of the proteins, followed by pulmonary challenge with either the homologous or a heterologous isolate, cleared the bacteria more rapidly than mock-immunized mice. These results suggest that UspA1 and UspA2 serve different virulence functions and that both are promising vaccine candidates.

scholarly journals Characterization of proteins from the cytoskeleton of Giardia lamblia

1983 ◽  
Vol 59 (1) ◽  
pp. 81-103 ◽  
Author(s):  
R. Crossley ◽  
D.V. Holberton
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulphate ◽  
Giardia Lamblia ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Molecular Size ◽  
Molecular Weights

Proteins from the axonemes and disc cytoskeleton of Giardia lamblia have been examined by sodium dodecyl sulphate/polyacrylamide gel electrophoresis. In addition to tubulin and the 30 X 10(3) molecular weight disc protein, at least 18 minor components copurify with the two major proteins in Triton-insoluble structures. The most prominent minor bands have the apparent molecular weights of 110 X 10(3), 95 X 10(3) and 81 X 10(3). Protein of 30 X 10(3) molecular weight accounts for about 20% of organelle protein on gels. In continuous 25 mM-Tris-glycine buffer it migrates mostly as a close-spaced doublet of polypeptides, which are here given the name giardins. Giardia tubulin and giardin have been purified by gel filtration chromatography in the presence of sodium dodecyl sulphate. Well-separated fractions were obtained that could be further characterized. Both proteins are heterogeneous when examined by isoelectric focusing. Five tubulin chains were detected by PAGE Blue 83 dye-binding after focusing in a broad-range ampholyte gel. Giardin is slightly less acidic than tubulin. On gels it splits into four major and four minor chains with isoelectric points in the pI range from 5.8 to 6.2. The amino acid composition of the giardin fraction has been determined, and compared to Giardia tubulin and a rat brain tubulin standard. Giardins are rich in helix-forming residues, particularly leucine. They have a low content of proline and glycine; therefore they may have extensive alpha-helical regions and be rod-shaped. As integral proteins of disc microribbons, giardins in vivo associate closely with tubulin. The properties of giardins indicate that in a number of respects - molecular size, charge, stoichiometry - their structural interaction with tubulin assemblies will be different from other tubulin-accessory protein copolymers studied in vitro.

Involvement of tyrosyl-tRNA synthetase in splicing of group I introns in Neurospora crassa mitochondria: biochemical and immunochemical analyses of splicing activity

1989 ◽  
Vol 9 (5) ◽  
pp. 2089-2104
Author(s):  
A L Majumder ◽  
R A Akins ◽  
J G Wilkinson ◽  
R L Kelley ◽  
A J Snook ◽  
...  
Keyword(s):  
Neurospora Crassa ◽  
Molecular Mass ◽  
Mitochondrial Protein ◽  
Gel Filtration ◽  
Nuclear Gene ◽  
Trna Synthetase ◽  
Synthetase Activity ◽  
Group I Introns ◽  
Group I

We reported previously that mitochondrial tyrosyl-tRNA synthetase, which is encoded by the nuclear gene cyt-18 in Neurospora crassa, functions in splicing several group I introns in N. crassa mitochondria (R. A. Akins and A. M. Lambowitz, Cell 50:331-345, 1987). Two mutants in the cyt-18 gene (cyt-18-1 and cyt-18-2) are defective in both mitochondrial protein synthesis and splicing, and an activity that splices the mitochondrial large rRNA intron copurifies with a component of mitochondrial tyrosyl-tRNA synthetase. Here, we used antibodies against different trpE-cyt-18 fusion proteins to identify the cyt-18 gene product as a basic protein having an apparent molecular mass of 67 kilodaltons (kDa). Both the cyt-18-1 and cyt-18-2 mutants contain relatively high amounts of inactive cyt-18 protein detected immunochemically. Biochemical experiments show that the 67-kDa cyt-18 protein copurifies with splicing and synthetase activity through a number of different column chromatographic procedures. Some fractions having splicing activity contain only one or two prominent polypeptide bands, and the cyt-18 protein is among the few, if not only, major bands in common between the different fractions that have splicing activity. Phosphocellulose columns resolve three different forms or complexes of the cyt-18 protein that have splicing or synthetase activity or both. Gel filtration experiments show that splicing activity has a relatively small molecular mass (peak at 150 kDa with activity trailing to lower molecular masses) and could correspond simply to dimers or monomers, or both, of the cyt-18 protein. Finally, antibodies against different segments of the cyt-18 protein inhibit splicing of the large rRNA intron in vitro. Our results indicate that both splicing and tyrosyl-tRNA synthetase activity are associated with the same 67-kDa protein encoded by the cyt-18 gene. This protein is a key constituent of splicing activity; it functions directly in splicing, and few, if any, additional components are required for splicing the large rRNA intron.

scholarly journals High Molecular Weight Factor VIII Coagulant Activity in Cryoprecipitate and Polyethylene Glycol Precipitates

1977 ◽  
Author(s):  
K. A. Rickard ◽  
T. Exner ◽  
H. Kronenberg
Keyword(s):  
Molecular Weight ◽  
Polyethylene Glycol ◽  
Factor Viii ◽  
High Molecular Weight ◽  
Gel Filtration ◽  
Lower Molecular Weight ◽  
Human Antibody ◽  
High Molecular Weight Material ◽  
Coagulant Activity ◽  
Molecular Weight Form

Gel filtration of human plasma cryoprecipitate on Sepharose 2B indicated the molecular weight of factor VIII coagulant activity (VIIIc) to be significantly greater than that found in antihaemophilic concentrate. Polyethylene glycol at 3% concentration precipitated approximately half of the VIIIc from cryoprecipitate. This activity eluted as high molecular weight material on gel filtration. The addition of more polyethylene glycol to a concentration of 8% precipitated most of the remaining VIIIc from cryoprecipitate. This activity appeared to be of significantly lower molecular weight, approximately corresponding in elution volume to that observed for antihaemophilic concentrate. The possibility that an antibody to VIIIc generated in a patient treated with cryoprecipitate might be directed against the higher molecular weight form of factor VIII was investigated. However, no significant differences between the higher and lower molecular weight forms of factor VIII either in stability or in reactivity with human antibody to factor VIII were found.

Gel Filtration of Plasma from Rabbits Injected with Honomeric DES-AB 125I-Fibrin and 131I–Fibrinogen

1979 ◽  
Author(s):  
I. Kröhnke ◽  
I. Hahn ◽  
W. Krell ◽  
G. Müller-Berghaus
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Gel Filtration ◽  
Void Volume ◽  
Molecular Configuration ◽  
Chromatographic Behaviour ◽  
Fibrin Monomer ◽  
Deutsche Forschungsgemeinschaft

We intended to study the chromatographic behaviour of soluble des-AB fibrin prepared in vitro and injected into rabbits. To prepare des-AB 1251-fibrin, purified rabbit 125I-fibrinogen was clotted by thrombin and the formed clot dissolved in Tris-buffered 3 M urea. Gel filtration of des-AB fibrin in urea through sepharose-CL 6B columns equilibrated with buffered 3 M. urea revealed monomeric fibrin. Rabbits received 131I-fibrinogen and 5 min later monomeric des-AB 125I-fibrin in urea. 30 min after injection blood was drawn and the plasma obtained filtered through sepharose-CL 6B columns eguilibrated with buffered plasma. At 20°C as well as at 37°C des-AB 125I-fibrin was eluted in the void volume in front of the 131I-fibrinogen peak. The data demonstrate that monomeric des-AB 125I-fibrin in urea injected into rabbits remains soluble in plasma. Possibly, monomerJ fibrin is converted to circulating soluble high-molecular weight fibrin aggregates or fibrin monomer changes its molecular configuration, thus showing a different gel filtration behaviour.(Supported by the Deutsche Forschungsgemeinschaft).

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