scholarly journals CONTACT-INHIBITED REVERTANT CELL LINES ISOLATED FROM SV40-TRANSFORMED CELLS

1973 ◽  
Vol 56 (2) ◽  
pp. 412-428 ◽  
Author(s):  
N. Scott McNutt ◽  
Lloyd A. Culp ◽  
Paul H. Black
Keyword(s):  
Cell Lines ◽  
Transformed Cells ◽  
Cell Periphery ◽  
3T3 Cells ◽  
Heavy Meromyosin ◽  
Thin Sections ◽  
Transformed Cell Line ◽  
Close Relationship ◽  
Loss Of Contact

A comparison is made of the ultrastructure of the cell periphery in three cloned cell lines: untransformed Balb/c 3T3 cells, SV40-transformed Balb/c 3T3 cells, and revertant cells obtained from the transformed cell line by a selection technique utilizing concanavalin A. Both thin-section and surface replication techniques are used for in situ examination of the cell lines. Microfilaments, 70 Å in diameter (called alpha filaments), are abundant in untransformed and revertant cell lines, particularly in the anterior expansions of the cells, which tend to have many microvilli and small pseudopodia. Alpha filaments are diminished in the anterior expansions of transformed cells, which contain large blunt pseudopodia and relatively few microvilli. Surface replicas confirm the impression gained from thin sections that transformed cells have a greater proportion of their cell surface involved in bulging pseudopodia than either untransformed or revertant cells. Since alpha filaments are shown to bind heavy meromyosin and are similar to F-actin, these filaments are thought to be important in cell motility. These observations suggest that a close relationship exists between decreased alpha filaments, bulging pseudopodia, and loss of contact inhibition of movement in transformed cells.

scholarly journals Identification of a BALB/c-3T3 cell protein modulated by platelet-derived growth factor.

1983 ◽  
Vol 3 (1) ◽  
pp. 70-81 ◽  
Author(s):  
C D Scher ◽  
R L Dick ◽  
A P Whipple ◽  
K L Locatell
Keyword(s):  
Growth Factor ◽  
Dna Synthesis ◽  
Cell Lines ◽  
Actinomycin D ◽  
Platelet Derived Growth Factor ◽  
Cell Protein ◽  
Transformed Cells ◽  
Antigenic Determinants ◽  
3T3 Cells ◽  
Constitutive Synthesis

The platelet-derived growth factor (PDGF) stimulates density-arrested BALB/c-3T3 cells to synthesize a protein (pII; Mr, 35,000) that is constitutively synthesized by spontaneously transformed BALB/c-3T3 (ST2-3T3) cells which do not require PDGF for growth. Antisera against a major excreted protein family (MEP) of retrovirus-transformed cells quantitatively precipitated cellular pII. PDGF-stimulated pII has the same molecular weight, a similar charge, and similar antigenic determinants as authentic MEP isolated from ST2-3T3 or retrovirus-transformed cells. MEP represented about 2% of the nonnuclear proteins synthesized by ST2-3T3 cells and 0.3 to 0.6% of the proteins synthesized by PDGF-treated BALB/c-3T3 cells, a three- to sixfold increase over the background. In BALB/c-3T3 cells, less PDGF was required for pII (MEP) synthesis than for DNA synthesis. PDGF induced a selective increase in pII (MEP) within 40 min. Such preferential synthesis was inhibited by brief treatment with actinomycin D, suggesting a requirement for newly formed RNA. The constitutive synthesis of pII (MEP) by ST2-3T3 cells was not inhibited by actinomycin D. Five spontaneously or chemical carcinogen-transformed tumorigenic BALB/c-3T3 cell lines were studied; they neither required PDGF for growth nor responded to it. These cell lines became arrested at confluence with a G1 DNA content. Each of these independently isolated lines synthesized pII (MEP) constitutively. Thus, the synthesis of pII (MEP) may be required, but is not sufficient, for PDGF-modulated DNA synthesis.

scholarly journals Isolation of cellular genes differentially expressed in mouse NIH 3T3 cells and a simian virus 40-transformed derivative: growth-specific expression of VL30 genes.

1985 ◽  
Vol 5 (10) ◽  
pp. 2590-2598 ◽  
Author(s):  
K Singh ◽  
S Saragosti ◽  
M Botchan
Keyword(s):  
Cell Lines ◽  
Simian Virus 40 ◽  
Mouse Cell ◽  
Simian Virus ◽  
Differentially Expressed ◽  
Transformed Cells ◽  
3T3 Cells ◽  
Transformed Cell ◽  
Nih 3T3 ◽  
Nih 3T3 Cells

We constructed and screened a cDNA library made from simian virus 40 (SV40)-transformed NIH 3T3 cells, and we isolated cDNAs representing genes that are differentially expressed between the parental cell and its SV40-transformed derivative. We found only a small number of cDNAs representing such genes. Two isolated cDNA clones represented RNAs expressed at elevated levels in the transformed cell line in a manner relatively independent of growth conditions. The expression of two other cDNAs was growth specific because transformed cells and nonconfluent parental cells contained higher levels of the homologous RNAs than did confluent, contact-inhibited parental cells. Another cDNA was well expressed in confluent parental and confluent transformed cells, but not in nonconfluent cells. The expression of some of these cDNAs varied strikingly in different mouse cell lines. Thus the genotype or histories of different cell lines can also affect the expression of certain genes. Interestingly, the only cDNA isolated that was expressed exclusively in the transformed cell was from an SV40 message. We focused on a growth-specific cDNA which we show is derived from a mouse endogenous retrovirus-like family called VL30. We sequenced the 3' long terminal repeat (LTR) of this transcriptionally active VL30 gene. This LTR has good homology with other VL30 LTR sequences, but differences occur, particularly upstream of the VL30 promoter. We found that VL30 gene expression varied in different mouse cell lines such that C3H cell lines had very low levels of VL30 transcripts relative to NIH 3T3 cell lines. However, Southern analysis showed that both cell lines had about the same number of VL30 genes homologous to our probe and that the position of the majority of these genes was conserved. We discuss possible explanations for this difference in VL30 expression.

scholarly journals CONTACT-INHIBITED REVERTANT CELL LINES ISOLATED FROM SV40-TRANSFORMED CELLS

1971 ◽  
Vol 50 (3) ◽  
pp. 691-708 ◽  
Author(s):  
N. Scott McNutt ◽  
Lloyd A. Culp ◽  
Paul H. Black
Keyword(s):  
Cell Lines ◽  
Culture Conditions ◽  
Contact Inhibition ◽  
3T3 Cells ◽  
Nuclear Pleomorphism ◽  
Inhibition Of Growth ◽  
Cell Processes ◽  
In Situ Embedding ◽  
Peripheral Cytoplasm

The ultrastructural appearances of normal 3T3, SV40-transformed 3T3 (SV-3T3), and F1A revertant cell lines are compared. Both confluent and subconfluent cultures are described after in situ embedding of the cells for electron microscopy. There is striking nuclear pleomorphism in F1A revertant cells, with many cells having large nuclei compared to the less variable nuclear morphology of both normal 3T3 and SV-3T3 cells. Under the culture conditions used, deep infoldings of the nuclear envelope are prominent in growing cells, e.g., subconfluent normal 3T3 and confluent SV-3T3 cells. Such infoldings are infrequently seen in cultures which display contact inhibition of growth, e.g., normal 3T3 or F1A revertant cells grown just to confluence. In confluent cultures, the cytoplasmic organelles in revertant cells closely resemble those of normal 3T3 cells. In both normal and revertant cells in confluent culture, the peripheral cytoplasm (ectoplasm) has many 70 A filaments (alpha filaments), which are frequently aggregated into bundles. Alpha filaments are also abundant in the ectoplasm near regions of cell-to-cell apposition and in the motile cell processes (filopodia). The abundance and state of aggregation of alpha filaments correlates with contact inhibition of movement and growth in these cell lines since fewer bundles of alpha filaments are seen in growing cells than in contact-inhibited cells. This observation suggests that these filaments may be an important secondary component in the regulation of contact inhibition of movement and, possibly, of growth in normal and revertant cells.

scholarly journals Quantitative biochemical analysis of microtubule content in normal and transformed 3T3 cells.

1979 ◽  
Vol 82 (2) ◽  
pp. 572-576 ◽  
Author(s):  
J H Eichhorn ◽  
B Peterkofsky
Keyword(s):  
Cell Density ◽  
Cell Lines ◽  
Cell Morphology ◽  
Biochemical Analysis ◽  
Transformed Cells ◽  
Dibutyryl Camp ◽  
3T3 Cells ◽  
Normal Cells ◽  
Stages Of Growth ◽  
Tubulin Content

Microtubules in normal and transformed BALB 3T3 cells were preserved in a stabilizing medium and measured by a [3H]colchicine-binding tubulin assay, and compared to total cellular tubulin measured under nonstabilizing conditions. Essentially no change in tubulin or microtubule content was seen with changes in cell density or with changes in cellular morphology at various stages of growth of normal or transformed cells or induced by dibutyryl cAMP treatment of transformed cells. Of five cell lines transformed by a variety of agents, four had a significantly higher total tubulin content than untransformed 3T3 cells and all of them had an increased microtubule content. None of the transformed lines had a lower fraction of tubulin recoverable as sedimentable microtubules compared to untransformed cells, and in three of them this fraction was significantly higher. These results establish that microtubules are present in transformed cells to at least the extent (if not greater) than in normal cells but that there are variations in the total amount of tubulin and microtubules as well as the fraction of the total tubulin present as microtubules which are not strictly correlated with transformation or cell morphology.

Isolation of cellular genes differentially expressed in mouse NIH 3T3 cells and a simian virus 40-transformed derivative: growth-specific expression of VL30 genes

1985 ◽  
Vol 5 (10) ◽  
pp. 2590-2598
Author(s):  
K Singh ◽  
S Saragosti ◽  
M Botchan
Keyword(s):  
Cell Lines ◽  
Simian Virus 40 ◽  
Mouse Cell ◽  
Simian Virus ◽  
Differentially Expressed ◽  
Transformed Cells ◽  
3T3 Cells ◽  
Transformed Cell ◽  
Nih 3T3 ◽  
Nih 3T3 Cells

We constructed and screened a cDNA library made from simian virus 40 (SV40)-transformed NIH 3T3 cells, and we isolated cDNAs representing genes that are differentially expressed between the parental cell and its SV40-transformed derivative. We found only a small number of cDNAs representing such genes. Two isolated cDNA clones represented RNAs expressed at elevated levels in the transformed cell line in a manner relatively independent of growth conditions. The expression of two other cDNAs was growth specific because transformed cells and nonconfluent parental cells contained higher levels of the homologous RNAs than did confluent, contact-inhibited parental cells. Another cDNA was well expressed in confluent parental and confluent transformed cells, but not in nonconfluent cells. The expression of some of these cDNAs varied strikingly in different mouse cell lines. Thus the genotype or histories of different cell lines can also affect the expression of certain genes. Interestingly, the only cDNA isolated that was expressed exclusively in the transformed cell was from an SV40 message. We focused on a growth-specific cDNA which we show is derived from a mouse endogenous retrovirus-like family called VL30. We sequenced the 3' long terminal repeat (LTR) of this transcriptionally active VL30 gene. This LTR has good homology with other VL30 LTR sequences, but differences occur, particularly upstream of the VL30 promoter. We found that VL30 gene expression varied in different mouse cell lines such that C3H cell lines had very low levels of VL30 transcripts relative to NIH 3T3 cell lines. However, Southern analysis showed that both cell lines had about the same number of VL30 genes homologous to our probe and that the position of the majority of these genes was conserved. We discuss possible explanations for this difference in VL30 expression.

Identification of a BALB/c-3T3 cell protein modulated by platelet-derived growth factor

1983 ◽  
Vol 3 (1) ◽  
pp. 70-81
Author(s):  
C D Scher ◽  
R L Dick ◽  
A P Whipple ◽  
K L Locatell
Keyword(s):  
Growth Factor ◽  
Dna Synthesis ◽  
Cell Lines ◽  
Actinomycin D ◽  
Platelet Derived Growth Factor ◽  
Cell Protein ◽  
Transformed Cells ◽  
Antigenic Determinants ◽  
3T3 Cells ◽  
Constitutive Synthesis

The platelet-derived growth factor (PDGF) stimulates density-arrested BALB/c-3T3 cells to synthesize a protein (pII; Mr, 35,000) that is constitutively synthesized by spontaneously transformed BALB/c-3T3 (ST2-3T3) cells which do not require PDGF for growth. Antisera against a major excreted protein family (MEP) of retrovirus-transformed cells quantitatively precipitated cellular pII. PDGF-stimulated pII has the same molecular weight, a similar charge, and similar antigenic determinants as authentic MEP isolated from ST2-3T3 or retrovirus-transformed cells. MEP represented about 2% of the nonnuclear proteins synthesized by ST2-3T3 cells and 0.3 to 0.6% of the proteins synthesized by PDGF-treated BALB/c-3T3 cells, a three- to sixfold increase over the background. In BALB/c-3T3 cells, less PDGF was required for pII (MEP) synthesis than for DNA synthesis. PDGF induced a selective increase in pII (MEP) within 40 min. Such preferential synthesis was inhibited by brief treatment with actinomycin D, suggesting a requirement for newly formed RNA. The constitutive synthesis of pII (MEP) by ST2-3T3 cells was not inhibited by actinomycin D. Five spontaneously or chemical carcinogen-transformed tumorigenic BALB/c-3T3 cell lines were studied; they neither required PDGF for growth nor responded to it. These cell lines became arrested at confluence with a G1 DNA content. Each of these independently isolated lines synthesized pII (MEP) constitutively. Thus, the synthesis of pII (MEP) may be required, but is not sufficient, for PDGF-modulated DNA synthesis.

2-Methoxyestradiol alters cell motility, migration, and adhesion

Blood ◽  
2003 ◽  
Vol 102 (1) ◽  
pp. 289-296 ◽  
Author(s):  
Martin Sattler ◽  
Laura R. Quinnan ◽  
Yuri B. Pride ◽  
Jessica L. Gramlich ◽  
Stephanie C. Chu ◽  
...  
Keyword(s):  
Cell Growth ◽  
Cell Lines ◽  
Metastatic Potential ◽  
Resistant Cell ◽  
Model Systems ◽  
Transformed Cells ◽  
Transformed Cell Line ◽  
Spontaneous Migration ◽  
Coated Surfaces

Abstract The effect of 2-methoxyestradiol, 2ME2, an endogenous metabolite of 17β-estradiol (E2), on cell growth and cytoskeletal functions in a BCR-ABL—transformed cell line model was investigated. We determined the interaction of 2ME2 with STI571 (Gleevec, imatinib mesylate) in STI571 drug-sensitive and -resistant cell lines. In cells expressing BCR-ABL, STI571 cooperated with 2ME2 in reducing cell growth, and STI571-resistant cells were sensitive to 2ME2 treatment. 2ME2 also inhibited growth of several cancer cell lines by a mechanism independent of BCR-ABL. BCR-ABL transformation leads to altered motility, increased adhesion, and spontaneous migration in different in vitro model systems. 2ME2 was found to specifically inhibit the spontaneous motility of BCRABL—transformed Ba/F3 cells and to change the morphology and volume of treated cells. Cells attached to fibronectin-coated surfaces showed a reduced number of filipodia and lamellipodia. In addition, 2ME2 significantly reduced BCRABL—mediated adhesion to fibronectin. The spontaneous migration of BCR-ABL—transformed cells through a transwell membrane also was found to be significantly decreased by 2ME2. Cytoskeletal changes were accompanied by alteration of tubulin formation, distinct from paclitaxel treatment. These results demonstrate that 2ME2 treatment of transformed cells strongly reduces cytoskeletal functions and may also be useful for the treatment of cancers with high metastatic potential. Combination of 2ME2 with other anticancer drugs may be beneficial to treatment of drug-resistant cancers. (Blood. 2003;102:289-296)

scholarly journals CONTACT-INHIBITED REVERTANT CELL LINES ISOLATED FROM SV40-TRANSFORMED CELLS

1971 ◽  
Vol 50 (3) ◽  
pp. 682-690 ◽  
Author(s):  
Lloyd A. Culp ◽  
William J. Grimes ◽  
Paul H. Black
Keyword(s):  
Sialic Acid ◽  
Cell Lines ◽  
Acid Content ◽  
Inverse Correlation ◽  
Wild Type Virus ◽  
Transformed Cells ◽  
Type Virus ◽  
Wild Type ◽  
3T3 Cells ◽  
Sialic Acid Content

Two contact-inhibited "revertant" cell lines were isolated from an SV40-transformed mouse 3T3 cell line (SV-3T3) after exposure to 5-fluoro-2'-deoxyuridine. Revertant cells resembled 3T3 cells morphologically and grew to saturation densities which were similar to those of 3T3 cells; however, revertant cells readily formed both single and multinucleated giant cells in confluent cultures. SV40 virus was rescued from revertant cells by fusion with permissive monkey cells. The rescued virus transformed 3T3 cells with the same efficiency as wild type virus, and produced transformed colonies which were phenotypically similar to those produced by wild type virus. The revertant cells also resembled normal 3T3 cells in that they contained higher quantities of sialic acid than SV-3T3 cells. An inverse correlation was found between the saturation density of cells and their sialic acid content. Collagen content, however, of revertant cells was similar to that of SV-3T3 cells. The data presented suggest that the property of contact inhibition in revertant cells is related to the sialic acid content of the plasma membrane and that changes in sialic acid content of transformed cells are not directly specified by the viral genome.

scholarly journals Suppression of tumorigenicity in transformed cells after transfection with vinculin cDNA.

1992 ◽  
Vol 119 (2) ◽  
pp. 427-438 ◽  
Author(s):  
J L Rodríguez Fernández ◽  
B Geiger ◽  
D Salomon ◽  
I Sabanay ◽  
M Zöller ◽  
...  
Keyword(s):  
Tumor Cell ◽  
Cell Lines ◽  
Suppressive Effect ◽  
Soft Agar ◽  
Transformed Cells ◽  
Tumor Cell Lines ◽  
3T3 Cells ◽  
Apparent Increase ◽  
Cell Matrix ◽  
Endogenous Protein

Transfection of chicken vinculin cDNA into two tumor cell lines expressing diminished levels of the endogenous protein, brought about a drastic suppression of their tumorigenic ability. The SV-40-transformed Balb/c 3T3 line (SVT2) contains four times less vinculin than the parental 3T3 cells, and the rat adenocarcinoma BSp73ASML has no detectable vinculin. Restoration of vinculin in these cells, up to the levels found in 3T3 cells, resulted in an apparent increase in substrate adhesiveness, a decrease in the ability to grow in soft agar, and suppression of their capacity to develop tumors after injection into syngeneic hosts or nude mice. These results suggest that vinculin, a cytoplasmic component of cell-matrix and cell-cell adhesions, may have a major suppressive effect on the transformed phenotype.

A TEM study of the microstructure of avian eggshells

1992 ◽  
Vol 50 (2) ◽  
pp. 1102-1103
Author(s):  
S. Q. Xiao ◽  
S. Baden ◽  
A. H. Heuer
Keyword(s):  
Electron Microscope ◽  
Calcium Carbonate ◽  
Nucleation And Growth ◽  
Growth Mechanisms ◽  
Shell Surface ◽  
Thin Sections ◽  
Polycrystalline Metals ◽  
Transmission Electron ◽  
Nucleation And Growth Mechanisms

The avian eggshell is one of the most rapidly mineralizing biological systems known. In situ, 5g of calcium carbonate are crystallized in less than 20 hrs to fabricate the shell. Although there have been much work about the formation of eggshells, controversy about the nucleation and growth mechanisms of the calcite crystals, and their texture in the eggshell, still remain unclear. In this report the microstructure and microchemistry of avian eggshells have been analyzed using transmission electron microscope (TEM) and energy dispersive spectroscopy (EDS).Fresh white and dry brown eggshells were broken and fixed in Karnosky's fixative (kaltitanden) for 2 hrs, then rinsed in distilled H2O. Small speckles of the eggshells were embedded in Spurr medium and thin sections were made ultramicrotome.The crystalline part of eggshells are composed of many small plate-like calcite grains, whose plate normals are approximately parallel to the shell surface. The sizes of the grains are about 0.3×0.3×1 μm3 (Fig.l). These grains are not as closely packed as man-made polycrystalline metals and ceramics, and small gaps between adjacent grains are visible indicating the absence of conventional grain boundaries.

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