In this study, DHT and E2 respectively up-regulate p66Shc protein level in PCa and OVa cells.
The ovalbumin (OVA -expressing B16-OVA -expressingas generously provided by Dr. Edith M. Lord and Dr. John Frelinger (University of Rochester Medical Center, Rochester, NY) [ 36].
When the long-term survivors were challenged with ovalbumin negative EL4 tumor cells (which are the parent cells to the ovalbumin positive E.G7-Ova cell line), the inoculated EL4 tumors were rejected by all long-term survivors.
The E.G7-OVA cell line was originally derived from the murine thymoma line, EL-4, by transfection with a neomycin-selectable vector expressing full-length chicken ovalbumin (Moore et al, 1988).
Flow cytometry showed that an earlier passage of the E.G7-OVA cell line (nominal passage number 10) expressed significantly (P<0.01) higher levels of ovalbumin (842±95, mean±s.e., n=3) when compared to a later passage (402±51, mean±s.e., n=3) (nominal passage number 13).
As shown in Figure 6A B16-OVA cells did not secrete α-MSH in vitro.
These results were further confirmed by immunofluorescence stainings of tumor tissue at d22 after inoculation of B16-OVA cells.
Thus, we quantified α-MSH in culture supernatants from B16-OVA cells at d1, d3, d5 and d7 of cell culture by radioimmunoassay.
Mice were challenged with 2.5×105 B16-Ova cells and vaccinated on days 6, 9, and 12 - cytokine production was analyzed on day 14.
Lung metastases were induced by intravenous injection of 3×105 B16F1-OVA cells and mice were sacrificed after 12 days to quantify lung tumor numbers.
B16F1-OVA cells (provided by Dr. C. Parrish [18]) were maintained in RPMI supplemented with 10% FBS, penicillin, streptomycin and L-glutamine.
