Control of cell proliferation in the human embryonic cornea: an autoradiographic analysis of the effect of growth factors on DNA synthesis in endothelial and stromal cells in organ culture and after explantation in vitro

1986 ◽  
Vol 83 (1) ◽  
pp. 1-21
Author(s):  
L. Hyldahl
Keyword(s):  
Endothelial Cells ◽  
Growth Factors ◽  
Growth Factor ◽  
Organ Culture ◽  
Dna Synthesis ◽  
Stromal Cells ◽  
Free Access ◽  
Corneal Endothelial Cells ◽  
Igf I ◽  
Autoradiographic Analysis

A novel technique for studying the growth properties of human embryonic corneal endothelial and stromal cells in organ culture was devised. Human embryonic eye globes were microdissected so that a passage was opened between the outer environment and the anterior chamber, which rendered free access of tissue culture medium to the endothelial cell monolayer. The dissected eye globes were maintained in organ culture for 24 h in the continuous presence of tritiated thymidine. Cross-sections were cut through the whole eye globes and subjected to autoradiographic analysis in order to estimate the mitogenic response of each corneal cell type to externally supplied growth factors and hormones. It was found that the corneal endothelial cells could be stimulated to initiate DNA synthesis by exposure to epidermal growth factor (EGF). The stimulatory effect of this growth factor could be enhanced if either insulin-like growth factor I (IGF I) or a combination of insulin, transferrin and high density lipoprotein (HDL) was simultaneously added. Similar results were obtained by adding growth factors and hormones to primary cell cultures from human embryonic corneas. It was also found that the stromal cell could be stimulated to initiate DNA synthesis by the addition of EGF and IGF I or a combination of insulin, transferrin and HDL. Taken together, these results suggest that the proliferation of corneal endothelial cells and stromal cells is dependent on EGF-like factors as well as on some insulin-like substance during embryogenesis.

scholarly journals Stimulatory effects of insulin-like growth factors on DNA synthesis in the human embryonic cornea

Development ◽  
1986 ◽  
Vol 98 (1) ◽  
pp. 71-83
Author(s):  
Louise Hyldahl ◽  
Wilhelm Engström ◽  
Paul N. Schofield
Keyword(s):  
Endothelial Cells ◽  
Growth Factors ◽  
Organ Culture ◽  
Dna Synthesis ◽  
Tritiated Thymidine ◽  
Cell Monolayer ◽  
Free Access ◽  
Corneal Endothelial Cells ◽  
Anterior Portion ◽  
Igf I

10- to 12-week-old human embryonic eye globes were microdissected so that a passage was opened between the outer environment and the anterior chamber which rendered free access of tissue culture medium to the endothelial cell monolayer. The dissected eye globes were maintained in organ culture for 24 h in the continuous presence of tritiated thymidine. Cross sections were cut through the whole eye globes and subjected to autoradiographic analysis in order to estimate the mitogenic response of human embryonic corneal endothelial cells to externally supplied growth factors and hormones. It was found that the corneal endothelial cells could be stimulated to initiate DNA synthesis by exposure to insulin-like growth factor I (IGF-I). The thymidine-labelling index doubled after IGF-I supplementation. Northern blot analysis revealed the abundant presence of IGF-II transcripts in the posterior eye. In contrast, the anterior portion of the eye, including the cornea, contains barely detectable levels of IGF-II transcripts. IGF-I transcripts were detected in both parts of the eye at much lower concentrations than those for IGF-II. No insulin transcripts were found. These results demonstrate that mRNA for both IGF-I and IGF-II is present in the late first trimester eye. The observed stimulatory effects of IGF-I in organ culture suggest that local production of IGF-I and IGF-II may stimulate cell proliferation in vivo.

Stimulatory effects of basic fibroblast growth factor on DNA synthesis in the human embryonic cornea

Development ◽  
1990 ◽  
Vol 109 (3) ◽  
pp. 605-611
Author(s):  
L. Hyldahl ◽  
P.N. Schofield ◽  
W. Engstrom
Keyword(s):  
Endothelial Cells ◽  
Growth Factor ◽  
Dna Synthesis ◽  
Fibroblast Growth Factor ◽  
Basic Fibroblast Growth Factor ◽  
First Trimester ◽  
Free Access ◽  
Fibroblast Growth ◽  
Corneal Endothelial Cells ◽  
Topographical Distribution

First trimester human embryonic eye globes were micro-dissected so that a passage was opened between the outer environment and the anterior chamber, which rendered free access of tissue culture medium to the endothelial cell monolayer. The dissected eye globes were maintained in organ culture for 24h in the continuous presence of tritiated thymidine. Sections were cut through the whole eye globes and were subject to autoradiographic analysis in order to estimate the mitogenic response of human corneal endothelial cells to externally supplied growth factors and hormones. It was found that the corneal endothelial cells could be stimulated to initiate DNA synthesis by exposure to basic fibroblast growth factor (bFGF). The thymidine labelling index nearly doubled after bFGF addition. Northern blot analysis revealed the presence of bFGF transcripts in the embryonic eye. In contrast we were unable to trace any bFGF transcripts in other first trimester human embryonic organs. In an attempt to determine the topographical distribution of bFGF mRNA within the eye, we found that transcript levels were higher in the posterior regions of the eye globe. Immunostaining with the appropriate antibody showed conclusively that bFGF protein was present in both the anterior and posterior human eye. These results suggest that local production of bFGF may stimulate cell proliferation in vivo.

Post-transcriptional control of the onset of DNA synthesis by an insulin-like growth factor

1984 ◽  
Vol 4 (9) ◽  
pp. 1807-1814
Author(s):  
J Campisi ◽  
A B Pardee
Keyword(s):  
Cell Cycle ◽  
Growth Factors ◽  
Growth Factor ◽  
Dna Synthesis ◽  
Transcriptional Control ◽  
S Phase ◽  
G1 Phase ◽  
Insulin Like Growth Factor ◽  
Igf I ◽  
Translational Inhibition

The control of eucaryotic cell proliferation is governed largely by a series of regulatory events which occur in the G1 phase of the cell cycle. When stimulated to proliferate, quiescent (G0) 3T3 fibroblasts require transcription, rapid translation, and three growth factors for the growth state transition. We examined exponentially growing 3T3 cells to relate the requirements for G1 transit to those necessary for the transition from the G0 to the S phase. Cycling cells in the G1 phase required transcription, rapid translation, and a single growth factor (insulin-like growth factor [IGF] I) to initiate DNA synthesis. IGF I acted post-transcriptionally at a late G1 step. All cells in the G1 phase entered the S phase on schedule if either insulin (hyperphysiological concentration) or IGF I (subnanomolar concentration) was provided as the sole growth factor. In medium lacking all growth factors, only cells within 2 to 3 h of the S phase were able to initiate DNA synthesis. Similarly, cells within 2 to 3 h of the S phase were less dependent on transcription and translation for entry into the S phase. Cells responded very differently to inhibited translation than to growth factor deprivation. Cells in the early and mid-G1 phases did not progress toward the S phase during transcriptional or translational inhibition, and during translational inhibition they actually regressed from the S phase. In the absence of growth factors, however, these cells continued progressing toward the S phase, but still required IGF at a terminal step before initiating DNA synthesis. We conclude that a suboptimal condition causes cells to either progress or regress in the cell cycle rather than freezing them at their initial position. By using synchronized cultures, we also show that in contrast to earlier events, this final, IGF-dependent step did not require new transcription. This result is in contrast to findings that other growth factors induce new transcription. We examined the requirements for G1 transit by using a chemically transformed 3T3 cell line (BPA31 cells) which has lost some but not all ability to regulate its growth. Early- and mid-G1-phase BPA31 cells required transcription and translation to initiate DNA synthesis, although they did not regress from the S phase during translational inhibition. However, these cells did not need IGF for entry into the S phase.

scholarly journals Insulin Growth Factor-I and Epidermal Growth Factor Receptors Recruit Distinct Upstream Signaling Molecules to Enhance AKT Activation in Mammary Epithelial Cells

Endocrinology ◽  
2006 ◽  
Vol 147 (12) ◽  
pp. 6027-6035 ◽  
Author(s):  
Jodie M. Fleming ◽  
Gwenaëlle Desury ◽  
Tiffany A. Polanco ◽  
Wendie S. Cohick
Keyword(s):  
Growth Factors ◽  
Growth Factor ◽  
Dna Synthesis ◽  
Egf Receptor ◽  
Mammary Epithelial ◽  
Insulin Receptor Substrate ◽  
Phosphatidylinositol 3 Kinase ◽  
Mapk Activation ◽  
Igf I ◽  
Epidermal Growth

IGF-I and epidermal growth factor (EGF) stimulate both normal mammary epithelial cell (MEC) growth and tumorigenesis. Whereas both growth factors increase DNA synthesis in MECs, how they evoke a greater response in combination when they activate similar signaling pathways remains unknown. In the present study, we investigated the signaling pathways by which these mitogens act in concert to increase DNA synthesis. Only EGF activated the MAPK pathway, and no further increase in MAPK activation was observed when both mitogens were added together. Both growth factors activated the phosphatidylinositol-3 kinase pathway, and simultaneous treatment enhanced phosphorylation of both AKT and its downstream target, p70S6K. The enhanced activation of AKT was observed at multiple time points (5 and 15 min) and growth factor concentrations (2.5–100 ng/ml). IGF-I activated AKT via insulin receptor substrate-1 and p85, the regulatory subunit of phosphatidylinositol-3 kinase. Treatment with EGF had no effect on insulin receptor substrate-1; however, it activated the EGF receptor, SHC, and c-Src. EGF treatment caused the association of SHC with Grb2 and Gab2 with phospho-SHC, phospho-Gab1, Grb2, and p85. Interestingly, inhibition of Src activation blocked the ability of EGF, but not IGF-I, to activate AKT. This corresponded with a decrease in phosphorylation of the EGF receptor and its association with phospho-SHC as well as downstream signaling. Unexpectedly, inhibition of Src increased basal MAPK activation. This is the first study to show that EGF and IGF-I use separate upstream components within a given MEC line to enhance AKT phosphorylation, contributing to increased DNA synthesis.

Growth factor stimulated cell proliferation is accompanied by an elevated labile intracellular pool of zinc in 3T3 cells

2002 ◽  
Vol 80 (8) ◽  
pp. 790-795 ◽  
Author(s):  
Shirley C Paski ◽  
Zhaoming Xu
Keyword(s):  
Cell Proliferation ◽  
Growth Factors ◽  
Growth Factor ◽  
Dna Synthesis ◽  
Cell Number ◽  
3T3 Cells ◽  
Intracellular Pool ◽  
Igf I ◽  
Epidermal Growth ◽  
Number Counts

Growth factors such as platelet-derived growth factor (PDGF), epidermal growth factor (EGF), and insulin-like growth factor-I (IGF-I) are required for quiescent 3T3 cells to proliferate, but zinc deprivation impairs IGF-I-induced DNA synthesis. We recently showed that labile intracellular pool of zinc is involved in cell proliferation. Our objective was to determine whether the labile intracellular pool of zinc plays a role in growth factor (PDGF, EGF, and IGF-I) - stimulated proliferation of 3T3 cells. Quiescent 3T3 cells were cultured in DMEM with or without growth factors. Labile intracellular pool of zinc, DNA synthesis, and cell proliferation were assessed using fluorescence microscopy, 3H-thymidine incorporation, and total cell number counts, respectively. After 24 h, growth factors stimulated DNA synthesis (24%) but not cell proliferation. After 48 h, growth factors stimulated both DNA synthesis (37%) and cell proliferation (89%). In response to growth factor stimulation, the labile intracellular pool of zinc was also elevated after 24 or 48 h of treatment. In summary, growth factor (PDGF, EGF, and IGF-I) - stimulated increase in DNA synthesis and cell proliferation were accompanied by an elevated labile intracellular pool of zinc in 3T3 cells. Since elevation of the labile intracellular pool of zinc occurred along with increased DNA synthesis, but cell proliferation remained unchanged, the elevation of the labile intracellular pool of zinc likely occurred during the S phase to provide the zinc needed to support DNA synthesis and ultimately cell proliferation.Key words: PDGF, EGF, IGF-I, labile intracellular pool of zinc, cell proliferation, DNA synthesis, 3T3 cells.

Growth Factors and Receptors in Juvenile Nasopharyngeal Angiofibroma and Nasal Polyps: An Immunohistochemical Study

2003 ◽  
Vol 127 (11) ◽  
pp. 1480-1484
Author(s):  
Paul J. Zhang ◽  
Randal Weber ◽  
Ho-Hi Liang ◽  
Teresa L. Pasha ◽  
Virginia A. LiVolsi
Keyword(s):  
Endothelial Cells ◽  
Growth Factors ◽  
Growth Factor ◽  
Signaling Pathways ◽  
Stromal Cells ◽  
Nasal Polyps ◽  
Bone Morphogenic Protein ◽  
Juvenile Nasopharyngeal Angiofibroma ◽  
Nasopharyngeal Angiofibroma ◽  
Bone Morphogenic Protein 4

Abstract Background.—Juvenile nasopharyngeal angiofibroma is a rare nasopharyngeal tumor that occurs exclusively in adolescent boys. It is a histologically benign but locally persistent growth of stromal and vascular tissue. Although male hormones and some growth factors, such as transforming growth factor β1 (TGF-β1), insulin-like growth factor II (IGF-II), and, lately, the proto-oncogene β-catenin, have been implicated in the histogenesis of the tumor, the biologic signaling pathways that drive this peculiar fibrovascular proliferation are still nuclear. Objective.—To evaluate immunoexpressions of β-catenin, c-Kit, p130Cas, TGF-β3, bone morphogenic protein 4, nerve growth factor (NGF), and the IGF receptor (IGF-1R) in a series of juvenile nasopharyngeal angiofibromas and to compare to that of a group of nasal polyps. Design.—A standard immunohistochemical technique was used on paraffin sections of 12 sporadic juvenile nasopharyngeal angiofibromas and 15 nasal polyps with microwave or steam antigen retrieval. Immunoreactivity was analyzed semiquantitatively in stromal cells and endothelial cells of each case. Results.—The expressions of β-catenin (nuclear), c-Kit (cytoplasmic), and NGF (cytoplasmic) were higher and more frequent in stromal cells of juvenile nasopharyngeal angiofibromas than those of nasal polyps. Both juvenile nasopharyngeal angiofibromas and nasal polyps showed similarly frequent and strong immunoreactivity for p130Cas and TGF-β3 and weak immunoreactivity for bone morphogenic protein 4 in both stromal cells and endothelial cells. No IGF-1R immunoreactivity was detected in any case of either group. Conclusions.—Our results support the role of β-catenin in juvenile nasopharyngeal angiofibromas and suggest a potential involvement of c-Kit and NGF signaling pathways in the juvenile nasopharyngeal angiofibromas. Although the biologic significance of c-Kit in juvenile nasopharyngeal angiofibromas has yet to be defined, the finding of frequent and high c-Kit expression might have therapeutic importance for patients with juvenile nasopharyngeal angiofibromas.

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