HCAP1 Gene Expression And Effects Of Exogenous HCAP1 Gene On Cell Apoptosis And Bax/Bcl-2 Expressionin Hematopoietic Malignancies

Objective: The HCAP1 is a novel gene that cloned from the region of loss ofheterzygosity at human chromosome 17pl3.3. It has been supposed HCAP1 is a candidate tumor suppressor gene. Our previous studies showed overexpression of exogenous HCAP could suppress the cell viability, inhibit the colony formation on agar and induce the apoptosis to some extent in leukemia/ lymphoma cell lines. In this study, the expression of HCAP1 gene was examined in the normal peripheral leukocytes from the healthy individuals and in the leukemic cells from the patients with leukemia, apoptosis in T leukemia/ lymphoma cell line Jurket transfected with exogenous HCAP1 was measured using more sensitive TUNEL method labeling with fluorescence, the regulation of overexpression on apoptotic related protein Bax and Bcl-2 in Jurkat cells and Raji cells was also investigated in order to investigate the possible molecular mechanism of HCAP1 on inducing of apoptosis.Methods: (1) The expressions of HCAP1 protein were examined withimmunocytochemical method and Western blot in the normal peripheral leukocytes and the tumor cells from the patients with hematopoietic malignancies. (2) The HCAP1 gene was transfected into the Jurkat and Raji cells by medium with liposome, the overexpression of exogenous HCAP1 gene was detected with Western blot technique.(3) Apoptosis induced by overexpression of HCAP1 in Jurkat cells was measured with TUNEL Assay labelling with fluorescence and mounted with DAPI.〦ffects of overexpression of HCAP1 on apoptotic related protein Bax and Bcl-2 in the Jurkat and Raji cells were examined with Western blot technique.Results: (1)Using imrmmocytochemical method, it was showed HCAP1 protein wasexpressed in both normal peripheral leukocytes from healthy individuals and the leukemic cells from leukemia patients and localized in nuclear.(2)Using Western blot technique, no difference in the levels of HCAP1 protein was detected between the normal peripheral leukocytes from 18 healthy individuals and the leukemic cells from 20 cases of patients with leukemia (P>0.05). (3) Using Western blot technique, overexpression of HCAP1 protein was detected in the Jurkat and Raji cells transfected with exogenous HCAP1 gene, compared with the control cells transfected with empty vectors or without any transfection.(4) At 24 hour after transfection, using TUNEL assay labelling with fluorescence, the percentage of apoptotic cells of Jurkat cells transfected with exogenous HCAP1 gene was increased to 26.67 + 3.88%, compared to 4.83 + 2.08% in control cells tranfected with empty vectors (P<0.01). (5) Using Western blot technique, the increase of the expression level of Bax protein was detected in the Jurkat and Raji cells transfected with exogenous HCAP1 gene, compared with the control cells transfected with empty vectors or the control cells without any transfection. Whereas, the decrease of the expression level of Bcl-2 protein was detected in the Jurkat and Raji cells transfected with exogenous HCAP1 gene, compared with the control cells transfected with empty vectors or the control cells without any transfection.Conclusion: (1)The HCAP1 protein was expressed in both normal peripheralleukocytes from healthy individuals and in the leukemic cells from patients with leukemia, localized in nuclear. However, no significantly difference in the levels of HCAP1 protein between them was detected. (2) Overexpression of the HCAP1 gene in Jurkat cells can induce cell apoptosis. (3) Overexpression of HCAP1 gene in the Jurkat and Raji cells can upregulate the expression of pro-apoptotic protein Bax, anddownregulate the expression of anti-apoptotic gene Bcl-2. The regulation of Bax/Bcl-2 protein might be involved in the molecular pathway of apoptosis induced by the HCAP1 protein.