In-vitro activation of cytotoxic T lymphocytes by fusion of mouse hepatocellular carcinoma cells and lymphotactin gene-modified dendritic cells.
OBJECTIVE
To investigate the in-vitro activation of cytotoxic T lymphocytes (CTLs) by fusion of mouse hepatocellular carcinoma (HCC) cells and lymphotactin gene-modified dendritic cells (DCs).


METHODS
Lymphotactin gene modified DCs (DCLptn) were prepared by lymphotactin recombinant adenovirus transduction of mature DCs which differentiated from mouse bone marrow cells by stimulation with granulocyte/macrophage colony-stimulating factor (GM-CSF), interleukin-4 (IL-4) and tumor necrosis factor alpha (TNF-alpha). DCLptn and H22 fusion was prepared using 50% PEG. Lymphotactin gene and protein expression levels were measured by RT-PCR and ELISA, respectively. Lymphotactin chemotactic responses were examined by in-vitro chemotaxis assay. In-vitro activation of CTLs by DCLptn/H22 fusion was measured by detecting CD25 expression and cytokine production after autologous T cell stimulation. Cytotoxic function of activated T lymphocytes stimulated with DCLptn/H22 cells was determined by LDH cytotoxicity assay.


RESULTS
Lymphotactin gene could be efficiently transduced to DCs by adenovirus vector and showed an effective biological activity. After fusion, the hybrid DCLptn/H22 cells acquired the phenotypes of both DCLptn and H22 cells. In T cell proliferation assay, flow cytometry showed a very high CD25 expression, and cytokine release assay showed a significantly higher concentration of IFN-gamma and IL-2 in DCLptn/H22 group than in DCLptn, DCLptn+H22, DC/H22 or H22 groups. Cytotoxicity assay revealed that T cells derived from DCLptn/H22 group had much higher anti-tumor activity than those derived from DCLptn, H22, DCLptn+H22, DC/H22 groups.


CONCLUSIONS
Lymphotactin gene-modified dendritoma induces T-cell proliferation and strong CTL reaction against allogenic HCC cells. Immunization-engineered fusion hybrid vaccine is an attractive strategy in prevention and treatment of HCC metastases.
Sheng
Xi-Ling
XL
Department of Intensive Care Unit, the Red Cross Hospital, Hangzhou 310003, Zhejiang Province, China.
Zhang
Hao
H
D000256:Adenoviridae; D000818:Animals; D006528:Carcinoma, Hepatocellular / Q000276:immunology / Q000378:metabolism / Q000473:pathology*; D002459:Cell Fusion; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002478:Cells, Cultured; D019744:Chemokines, C / Q000235:genetics* / Q000378:metabolism; D016207:Cytokines / Q000378:metabolism; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism / Q000473:pathology*; D005260:Female; D053645:Interleukin-2 Receptor alpha Subunit / Q000378:metabolism; D008114:Liver Neoplasms, Experimental / Q000276:immunology / Q000378:metabolism / Q000473:pathology*; D008213:Lymphocyte Activation; D051379:Mice; D008807:Mice, Inbred BALB C; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology / Q000473:pathology / Q000502:physiology*; D014161:Transduction, Genetic
D019744:Chemokines, C; D016207:Cytokines; D053645:Interleukin-2 Receptor alpha Subunit; C486249:Xcl1 protein, mouse