Objective:To construct recombinant human JNK3 adenovirus and study its influence on the proliferation of human neuroblatoma SH-SY5Y cells, and to study its influence on epirubicin-induced apoptosis.Methods: The full-length JNK3 cDNA fragment was amplified by PCR from the pDBLeu-JNK3 plasmid to construct the shuttle plasmid AdTrack-CMV-JNK3.Then the linearized shuttle plasmid was co-transformed into BJ5183 bacteria with backbone vector pAdEasy-1 to obtain the recombinant adenoviral plasmid Ad-JNK3 by homologous recombination.The linearized recombined adenovirus Ad-JNK3 was then transfected into HEK293 cells for packing and amplification.Viral titers were measured by endpoint dilution assay.Ad-JNK3 was identified by PCR.The expression of Ad-JNK3 in SH-SY5Y cells was detected by Western blotting assay.The influence of Ad-JNK3 on the proliferation of SH-SY5Y cells was assayed by MTT after cells were infected by 100 pfu/ml adenovirus.The cell apoptosis induced by 0.5 μg/ml epirubicin was detected by flow cytometry method and agarose gel electrophoresis.Results: The recombinant adenoviral shutter vector AdTrack-CMV-JNK3 and recombinant adenoviral vector Ad-JNK3 were successfully constructed as identified by sequence analysis.PCR assay showed that adenovirus Ad-JNK3 contained JNK3 gene.After amplification in packing cell HEK293,6.5×1010 pfu/ml titer of Ad-JNK3 was obtained.Western blotting assay showed that JNK3 protein was expressed in SH-SY5Y cells.After infected by 100 pfu/ml adenovirus,the proliferation of SH-SY5Y cells was inhibited by 28.08% with MTT method.Flow cytometry showed that Ad-JNK3 significantly promoted the apoptosis of SH-SY5Y cells induced by epirubicin.The DNA ladder of agarose gel electrophoresis was clearly seen.Conclusion: Ad-JNK3 can obviously inhibit the growth of SH-SY5Y cells and promote apoptosis induced by epirubicin,which provides a solid foundation for further studies on the function of the JNK3 gene and applying it in gene therapy for human neuroblastoma.