Mode of estrogen action on cell proliferation in CAMA‐1 cells: II. Sensitivity of G1 phase population

Abstract

The mammary cancer cell line CAMA-1 synchronized at the G1/S boundary by thymidine block or at the G1/M boundary by nocodazole was used to evaluate 1) the sensitivity of a specific cell cycle phase or phases to 17 beta-estradiol (E2), 2) the effect of E2 on cell cycle kinetics, and 3) the resultant E2 effect on cell proliferation. In synchronized G1/S cells, E2-induced 3H-thymidine uptake, which indicated a newly formed S population, was observed only when E2 was added during, but not after, thymidine synchronization. Synchronized G2/M cells, enriched by Percoll gradient centrifugation to approximately 90% mitotic cells, responded to E2 added immediately following selection; the total E2-treated population traversed the cycle faster and reached S phase approximately 4 hr earlier than cells not exposed to E2. When E2 was added during the last hour of synchronization (ie, at late G2 or G2/M), or for 1 hr during mitotic cell enrichment, a mixed response occurred: a small portion had an accelerated G1 exit, while the majority of cells behaved the same as controls not incubated with E2. When E2 addition was delayed until 2 hr, 7 hr, or 12 hr following cell selection, to allow many early G1 phase cells to miss E2 exposure, the response to E2 was again mixed. When E2 was added during the 16 hr of nocodazole synchronization, when cells were largely at S or possibly at early G2, it inhibited entry into S phase. The E2-induced increase or decrease of S phase cells in the nocodazole experiments also showed corresponding changes in mitotic index and cell number. These results showed that the early G1 phase and possibly the G2/M phase are sensitive to E2 stimulation, late G1, G1/S, or G2 are refractory; the E2 stimualtion of cell proliferation is due primarily to an increased proportion of G1 cells that traverse the cell cycle and a shortened G1 period, E2 does not facilitate faster cell division; and estrogen-induced cell proliferation or G1/S transition occurs only when very early G1 phase cells are exposed to estrogen. These results are consistent with the constant transition probability hypothesis, that is, E2 alters the probability of cells entering into DNA synthesis without significantly affecting the duration of other cell cycle phases. Results from this study provide new information for further studies aimed at elucidating E2-modulated G1 events related to tumor growth.

Modulation of the proliferative response of breast cancer cells to growth factors by oestrogen

“…Furthermore, proliferation in the presence of TGF-xc or EGF and oestradiol was actually greater than in the presence of oestradiol alone and this is not consistent with either growth factor acting as an oestrogen-induced autocrine growth factor. Experiments with transfected cell lines which express high levels of TGF-x constitutively but which retain oestrogen responsiveness , and the identification of an oestrogen responsive cell line which does not express epidermal growth factor receptor which mediates the effects of TGF-x (Leung et al, 1987), have also suggested that TGF-a plays a relatively minor role as an oestrogen induced autocrine mitogen and are consistent with our data. Although the cooperative effects of growth factors on the proliferation of cells such as fibroblasts is well documented and has given rise to the concept of competence and progression factors, the interaction of growth factors in the control of the proliferation of breast cancer cells is poorly understood.…”

Evidence for genomic and nongenomic actions of estrogen in growth plate regulation in female and male rats at the onset of sexual maturation

“…It appears that subtle changes in the fixation procedure as well as the fact that chondrocytes have a long G1 phase in relation to the S, G2 and M phase contribute to the relatively high number of PCNA-positive hypertrophic chondrocytes (Aizawa et al 1997). In addition, we speculate that E 2 may have a sexually dimorphic effect on the cell cycle G1 phase duration in chondrocytes (Leung & Potter 1987, Takahashi & Noumura 1987.…”

Estrogen-induced Activation of Cdk4 and Cdk2 during G1-S Phase Progression Is Accompanied by Increased Cyclin D1 Expression and Decreased Cyclin-dependent Kinase Inhibitor Association with Cyclin E-Cdk2

“…These include the use of hydroxymethylglutaryl-CoA reductase inhibitors (i.e. lovostatin or simvastatin (46)), isoleucine deprivation (36), or nocodazole arrest (7). The system with perhaps the most specificity is one in which cells are first growtharrested by estrogen antagonists and then rescued by estrogen, since some selective magnification of ER-regulated gene responses compared with more generalized responses to cell cycle progression might be expected.…”

“…6). Using breast cancer cells synchronized at the G 1 /S boundary or at G 2 /M to test the effect of estrogen added at different stages of the cell cycle, Leung et al (7) concluded that the sensitive cells were in early G 1 phase, immediately following mitosis. These data supported observations that both nonsteroidal (8) and steroidal antiestrogens (9,10) arrest ER-positive breast cancer cells in G 1 phase.…”