Notch-1 inhibits apoptosis in murine erythroleukemia cells and is necessary for differentiation induced by hybrid polar compounds

Strikingly increased expression of notch-1 has been demonstrated in several human malignancies and pre-neoplastic lesions. However, the functional consequences of notch-1 overexpression in transformed cells remain unclear. We investigated whether endogenously expressed notch-1 controls cell fate determination in mouse erythroleukemia (MEL) cells during pharmacologically induced differentiation. We found that notch-1 expression is modulated during MEL cell differentiation. Premature downregulation of notch-1 during differentiation, by antisense S-oligonucleotides or by enforced expression of antisense notch-1 mRNA, causes MEL cells to abort the differentiation program and undergo apoptosis. Downregulation of notch-1 expression in the absence of differentiation inducer increases the likelihood of spontaneous apoptosis. We conclude that in MEL cells, endogenous notch-1 expression controls the apoptotic threshold during differentiation and growth. In these cells, notch-1 allows differentiation by preventing apoptosis of pre-committed cells. This novel function of notch-1 may play a role in regulating apoptosis susceptibility in notch-1 expressing tumor cells.     

Growth inhibition and differentiation induction in murine erythroleukemia cells by 4-hydroxynonenal

4-Hydroxynonenal (HNE) is one of the major end products of lipid peroxidation. Here we show that the exposure of murine erythroleukemia (MEL) cells to 1 μM HNE, for 10.5 h over 2 days, induces a differentiation comparable with that observed in cells exposed to DMSO for the whole experiment (7 days). The exposure of MEL cells for the same length of time demonstrates a higher degree of differentiation in HNE-treated than in DMSO-treated MEL cells. The protooncogene c-myc is down-modulated early, in HNE-induced MEL cells as well as in DMSO-treated cells. However, ornithine decarboxylase gene expression first increases and then decreases, during the lowering of the proliferation rate. These findings indicate that HNE, at a concentration physiologically found in many normal tissues and in the plasma, induces MEL cell differentiation by modulation of specific gene expression.     

Growth inhibition and differentiation induction in murine erythroleukemia cells by 4-hydroxynonenal

4-Hydroxynonenal (HNE) is one of the major end products of lipid peroxidation. Here we show that the exposure of murine erythroleukemia (MEL) cells to 1 μM HNE, for 10.5 h over 2 days, induces a differentiation comparable with that observed in cells exposed to DMSO for the whole experiment (7 days). The exposure of MEL cells for the same length of time demonstrates a higher degree of differentiation in HNE-treated than in DMSO-treated MEL cells. The protooncogene c-myc is down-modulated early, in HNE-induced MEL cells as well as in DMSO-treated cells. However, ornithine decarboxylase gene expression first increases and then decreases, during the lowering of the proliferation rate. These findings indicate that HNE, at a concentration physiologically found in many normal tissues and in the plasma, induces MEL cell differentiation by modulation of specific gene expression.     

Inhibition of proliferation and differentiation and promotion of apoptosis by cyclin L2 in mouse embryonic carcinoma P19 cells

Cyclin L2 (CCNL2) is a novel member of the cyclin gene family. In a previous study, we demonstrated that CCNL2 expression was upregulated in ventricular septum tissues from patients with ventricular septal defect compared to healthy controls. In the present study, we established a stable CCNL2-overexpressing P19 cell line that can differentiate to myocardial cells when treated with 1% dimethyl sulfoxide (DMSO). Our data showed that stable CCNL2-overexpressing P19 cells were less differentiated after treatment with 1% DMSO and that expression of myocardial cell differentiation-related genes (such as cardiac actin, GATA4, Mef2C, Nkx2.5, and BNP) were reduced compared to vector-only transfected P19. Moreover, P19 cells overexpressing the CCNL2 gene had a reduced growth rate and a remarkably decreased S phase. We also found that these cells underwent apoptosis, as detected by two different apoptosis assays. The anti-apoptotic Bcl-2 protein was also downregulated in these cells. In addition, real-time PCR analysis revealed that expression of Wnt and β-catenin was suppressed and GSK3β was induced in the CCNL2-overexpressing P19 cells. These data suggest that overexpression of CCNL2 inhibited proliferation and differentiation of mouse embryonic carcinoma P19 cells and induced them to undergo apoptosis, possibly through the Wnt signal transduction pathway.