Figure 4
Figure 4. AhR mediates the expansion and specification of HPs. (A) Schematic representation of pHAGE2 lentiviral reporter constructs that contain the mouse mammary tumor virus flanking the dioxin response element (MMTV-AhRRE-MMTV) driving the expression of NLS-dsRed (pHAGE2-MMTV-AhRRE-MMTV-NLS-dsRed-IRES-zsGreen) (B) Representative FACS analysis for AhRRE-dsRED in HPs infected with pHAGE2-MMTV-AhRRE-MMTV-NLS-dsRed-IRES-ZsGreen. Infected cells were untreated or treated with 5 μM of CH223191, or 0.4 μM of FICZ. (C) Representative flow cytometry dot plots of live vs dead cells (PI vs Hoechst) from day 15 HPs ± FICZ and/or CH223191. For these experiments, HPs were pretreated with the known AhR inhibitor CH223191 on day 6 before the addition of FICZ on day 7. (D) Graphical representation of experiments performed in panel C. For the live-cell gate, data are the average of 3 independent experiments ± SD: *P < .005. For the apoptotic cell gate, data are the average of 3 independent experiments ± SD: *P < .04. (E) Expression of CYP1B1 as detected by quantitative PCR of MEPs from panel C, normalized to β-actin. Data are the average of 3 independent experiments ± SD: *P < .005.

AhR mediates the expansion and specification of HPs. (A) Schematic representation of pHAGE2 lentiviral reporter constructs that contain the mouse mammary tumor virus flanking the dioxin response element (MMTV-AhRRE-MMTV) driving the expression of NLS-dsRed (pHAGE2-MMTV-AhRRE-MMTV-NLS-dsRed-IRES-zsGreen) (B) Representative FACS analysis for AhRRE-dsRED in HPs infected with pHAGE2-MMTV-AhRRE-MMTV-NLS-dsRed-IRES-ZsGreen. Infected cells were untreated or treated with 5 μM of CH223191, or 0.4 μM of FICZ. (C) Representative flow cytometry dot plots of live vs dead cells (PI vs Hoechst) from day 15 HPs ± FICZ and/or CH223191. For these experiments, HPs were pretreated with the known AhR inhibitor CH223191 on day 6 before the addition of FICZ on day 7. (D) Graphical representation of experiments performed in panel C. For the live-cell gate, data are the average of 3 independent experiments ± SD: *P < .005. For the apoptotic cell gate, data are the average of 3 independent experiments ± SD: *P < .04. (E) Expression of CYP1B1 as detected by quantitative PCR of MEPs from panel C, normalized to β-actin. Data are the average of 3 independent experiments ± SD: *P < .005.

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