Abstract
IntroductionIt has been demonstrated that leukemia inhibitory factor (LIF) induces epithelium apoptosis through Stat3 activation during mouse mammary gland involution. In contrast, it has been shown that this transcription factor is commonly activated in breast cancer cells, although what causes this effect remains unknown. Here we have tested the hypothesis that locally produced LIF can be responsible for Stat3 activation in mouse mammary tumors.MethodsThe studies were performed in different tumorigenic and non-tumorigenic mammary cells. The expression of LIF and LIF receptor was tested by RT-PCR analysis. In tumors, LIF and Stat3 proteins were analyzed by immunohistochemistry, whereas Stat3 and extracellular signal-regulated kinase (ERK)1/2 expression and phosphorylation were studied by Western blot analysis. A LIF-specific blocking antibody was used to determine whether this cytokine was responsible for Stat3 phosphorylation induced by conditioned medium. Specific pharmacological inhibitors (PD98059 and Stat3ip) that affect ERK1/2 and Stat3 activation were used to study their involvement in LIF-induced effects. To analyze cell survival, assays with crystal violet were performed.ResultsHigh levels of LIF expression and activated Stat3 were found in mammary tumors growing in vivo and in their primary cultures. We found a single mouse mammary tumor cell line, LM3, that showed low levels of activated Stat3. Incidentally, these cells also showed very little expression of LIF receptor. This suggested that autocrine/paracrine LIF would be responsible for Stat3 activation in mouse mammary tumors. This hypothesis was confirmed by the ability of conditioned medium of mammary tumor primary cultures to induce Stat3 phosphorylation, activity that was prevented by pretreatment with LIF-blocking antibody. Besides, we found that LIF increased tumor cell viability. Interestingly, blocking Stat3 activation enhanced this effect in mammary tumor cells.ConclusionLIF is overexpressed in mouse mammary tumors, where it acts as the main Stat3 activator. Interestingly, the positive LIF effect on tumor cell viability is not dependent on Stat3 activation, which inhibits tumor cell survival as it does in normal mammary epithelium.
Highlights
It has been demonstrated that leukemia inhibitory factor (LIF) induces epithelium apoptosis through Stat3 activation during mouse mammary gland involution
These cells showed very little expression of LIF receptor. This suggested that autocrine/paracrine LIF would be responsible for Stat3 activation in mouse mammary tumors
This hypothesis was confirmed by the ability of conditioned medium of mammary tumor primary cultures to induce Stat3 phosphorylation, activity that was prevented by pretreatment with LIF-blocking antibody
Summary
It has been demonstrated that leukemia inhibitory factor (LIF) induces epithelium apoptosis through Stat activation during mouse mammary gland involution. The pleiotropic cytokine leukemia inhibitory factor (LIF) is a secreted 38 to 67 kDa glycoprotein first named for its ability to induce macrophage differentiation in the murine myeloid leukemic cell line M1 [1] This factor has been detected in a variety of adult mouse tissues and displays different biological activities, including effects on bone metabolism, inflammation, neural development and embryogenesis [2]. Be modulated by progestins and antiprogestins [5], and by its capacity to induce the proliferation of several estrogendependent (MCF-7 and T47D) and estrogen-independent (SK-BR3 and BT20) breast cancer cell lines as well as fresh breast carcinoma cells [4,6] In spite of these data, little is known about the relevance of LIF for mammary tumor development in vivo. It has been reported that among the seven members of the Stat family, Stat is the major mediator of gp130 signals [10,11]
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