| Primary liver cancer, a common malignancy in China, is highly malignant and proceeding rapidly, which makes it difficult to treat. Although surgical operation is a treatment of choice in all therapeutic methods today, its effect is still limited. Gene therapy, a new therapeutic hot spot at present, is regarded as one of the most hopeful therapy projects. F1t3 ligand is a recently cloned primitive hematopoietic growth factor, which can stimulate hematopoisis in vivo and enhance inimulogic function. Using retroviral vetor-mediated, we introduced mFlt3L into hepatocellular cell line for studying its biology functions, in order to provide basic research of clinical utilizations.This study includes: 1. Retroviral vector carrier pBabe/Flt3L was established using retroviral vector pBabe Puro with gene expression. The cell strain 'ir -cre pack with transfection monophilic retrovirus and the vector w -crip transfected with bi-ophilic retroviral in viral supernant were moncloned after screening with 3.0 ii glml puromycin for 4 weeks. It was termed ~i' -crip/Flt3L. The highest virus titre determined by NIH3T3 was 2.0 X 1O6CFU/ml. 2. The retroviral vector DNA was transfected into Hepal -6 cell line. We get resistant colonies using puromycin selection. It was termed Hepal-6IFIt3L. The mRNA of F1t3L was detected in the Hepal-6/F1t3L by RT-PCR, and the Flt3L protein was 11 .5ngIl ~6 I4Shr in the Hepal -6IF1t3L cell culture supernatant. 3. The in vivo experimental studies showed, transduction of Hepal-6 with retroviral vectors expressing F1t3L did not influence in vitro growth but could reduce tumorigenicity in syngeneic mice, and the ininiunogenicity was augmented. Administration of FIt3L as a tumor vaccine could protect mice from a subsequent challenge with untransduced Hepal-6 in 100% of mice, compared to 0% of controls. To simulate minimal disease, parental Hepal -6 cells were injected into the3contralateral chest wall 5 days prior to treatment with Hepal-6/pBabe or Hepal6IF1t3L. Hepal-6IF1t3L treatment decreased contralateral parental tumor formation(80% tumor free) compared with Hepal -6/pBabe treatment(O% tumor free). Pathological staining showed that there were significantly more intratumor inflammatory infiltration and CD4~. CD8~ T lymphocytes cell infiltration than that of control group. |
