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ABSTRACT: To overcome difficulties that hampered widespread application of a specific delivery system in cancer gene therapy and to inhibit the growth of solid liver cancer, we utilized a strain of Bifidobacterium longum as a delivery system to transport an endostatin gene that can inhibit growth of tumor. The B. longum strain with the endostatin gene (B. longum-En) was taken orally by tumor-bearing nude mice through drencher preparation. The results showed that B. longum-En could strongly inhibit the growth of solid liver tumor in nude mice and prolong the survival time of tumor-bearing nude mice. Furthermore, tumor growth was inhibited more efficiently when the B. longum-En treatment included selenium. Enriching the B. longum-En treatment with selenium improves the activity of NK and T cells and stimulates the activity of IL-2 and TNF-alpha in BALB/c mice. These results suggest that B. longum may be a highly specific and efficient vector for transporting anticancer genes in cancer gene therapy.
Cancer Gene Therapy 03/2005; 12(2):133-40. · 2.80 Impact Factor
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ABSTRACT: To obtain an efficient delivery system for transporting endostatin gene to mouse liver tumor xenografts by administration of aerosol.
Recombinant plasmid pcDNA3.0/endostatin containing human endostatin gene together with signal peptide from alkaline phosphatase were transferred into human umbilical vein endothelial cell (HUVEC) by transferrin(TF)-liposome-endostatin complex. Western blot was used to detect the expression of human endostatin in transfected HUVEC cells and its medium. After the tumor-bearing mice were administrated with TF-liposome-endostatin complex, the lung tissue was analyzed by immunohistochemical method for expression of endostatin and the tumors were treated with CD-31 antibody to detect the density of microvessels in tumor tissues. The inhibition of tumor growth was estimated by the weight of tumors from groups treated with different doses of TF-liposome-endostatin complex. DNA fragmentation assay was used to detect the apoptosis of the cells from primary liver tumor.
Western blot analysis and immunohistochemical method confirmed the expression of endostatin protein in vitro and in vivo. After the tumor sections were treated with CD-31 antibody, the positive reaction cells appeared brown while the negative cells were colorless. The positively stained area of the TF-liposome-endostatin treated group was significantly smaller (P<0.01, 645.8+/-55.2 microm(2)) than that of the control group (1 325.4+/-198.5 microm(2)). The data showed a significant inhibition of angiogenesis. After administration of TF-liposome-endostatin, comparing with the control group administrated with TF-liposome-pcDNA3.0, liver tumor growth in the mice treated with 50, 250 and 500 mg DNA/kg was inhibited by 36.6 %, 40.8 %, and 72.8 %, respectively (P<0.01). And a typical DNA fragmentation of apoptosis was found in the cells from tumor tissues of the mice treated with TF-liposome-endostatin but none in the control group.
Endostatin gene could be efficiently transported into the mice with TF-liposome-DNA delivery system by administration of aerosol. TF-liposome-mediated endostatin gene therapy strongly inhibited angiogenesis and the growth of mouse xenograft liver tumors. It also could promote the development of apoptosis of tumors without direct influence on tumor cells.
World Journal of Gastroenterology 03/2003; 9(2):262-6. · 2.47 Impact Factor
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ABSTRACT: In order to overcome difficulties that hampered widespread application of antiangiogenesis in cancer therapy, a highly specific delivery system may be engaged in vivo to deliver and express antiangiogenic genes. We selected a strain of Bifidobacterium adolescentis (B. adolescentis) as the delivery system to transport endostatin gene to solid tumors. B. adolescentis with endostatin gene were injected into tumor-bearing mice through the tail vein. After the mice were sacrificed, the tumor and some normal tissues of the mice were examined. B. adolescentis were only found in the tumors and no bacilli were found in other normal tissues. Also, a strong inhibition of angiogenesis had been shown to inhibit local tumor growth in the administrated group. These results suggested that B. adolescentis only germinated and proliferated in solid tumors and might be a highly specific and efficient vector for transporting anticancer genes into target tumor in cancer gene therapy.
Cancer Gene Therapy 03/2003; 10(2):105-11. · 2.80 Impact Factor
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ABSTRACT: To study the effect of human endostatin mediated by retroviral gene transfer on the growth of human hepatocarcinoma cell line SMMC7721 in nude mice.
Human endostatin gene together with rat serum albumin signal peptide was transferred into human liver carcinoma SMMC7721 cells by retroviral vector pLncx to build a stable transfectant (SMMC-endo). PCR and Western blot analysis were used to verify the transfection and secretion of human endostatin gene in SMMC7721 cells. The endothelial cell proliferation assay in vitro was conducted to test the biological activity of the expressed human endostatin. The inhibitory effect of endostatin expressed by transfected SMMC7721 on the growth rates of tumor cells in vivo was observed. The mean microvessel density in the specimen was also counted.
PCR amplification proved that the genome of SMMC-endo cells contained a 550bp specific fragment of endostatin gene. Western blot analysis confirmed the secretion of human endostatin gene in the conditioned medium of transfected SMMC-endo cells. The endothelial proliferation assay showed that the conditioned medium of SMMC-endo cells significantly inhibited the proliferation of human umbilical vein endothelial cells by 48 %, significantly higher than that of SMMC-pLncx (10.2 %, P<0.01). In vitro experiments revealed that only in 3 out of 5 mice tumors were formed and the mean size of flank tumors from SMMC-endo cells was 94.5 % smaller than that from the control SMMC-pLncx cells 22 days after tumor inoculation (P<0.001). The mean microvessel density in tumor samples from SMMC-endo cells was only 8.6+/-1.1, much fewer than that of 22.6+/-4.5 from SMMC-pLncx cells (P<0.01).
Human endostatin mediated by retroviral gene transfer can inhibit human liver carcinoma cell SMMC7721 growth in nude mice.
World Journal of Gastroenterology 12/2002; 8(6):1045-9. · 2.47 Impact Factor
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ABSTRACT: To explore the inhibitory effect of human endostatin gene mediated by retroviral vector on the growth of human liver carcinoma.
A recombinant retroviral plasmid containing human endostatin gene and signal peptide was engineered and transferred into PA317 cells to produce retrovirus. Human liver carcinoma cells (SMMC7721) were infected with the above retrovirus to build a stable endostatin-transfected liver carcinoma cell line (SMMC-endo). The control liver carcinoma cell line (SMMC-pLncx) was developed in a similar way except that the plasmid was replaced by an empty retroviral vector. Immunohistochemistry and Western blot were used to test the expression and secretion of human endostatin. The biological activity of the expressed human endostatin was assessed by endothelial cell proliferation assay. The growth rates of SMMC-endo and control SMMC-pLncx cells in vivo and in vitro were also observed.
The expression and secretion of human endostatin by endostatin-transfected SMMC-endo cells were confirmed by immunohistochemistry and Western blot. Compared with the control group, concentrated supernatant of SMMC-endo cells remarkably inhibited the proliferation of human umbilical vein endothelial cells by 48%, significantly higher than the inhibition by the control (10.2%; P < 0.01). The endostatin-transfected SMMC-endo cells had similar in vitro growth rates to SMMC-pLncx cells. The in vivo experiment showed that the growth rate of SMMC-endo cells was slowed. Only in 3 out of 5 mice were tumors formed and flank tumors of SMMC-endo cells were 94.5% smaller than those of control cells 22 days after inoculation into nude mice (P < 0.001).
Gene transfer of human endostatin mediated by retroviral vector is an effective form of cancer therapy.
Chinese medical journal 11/2002; 115(11):1664-9. · 0.86 Impact Factor
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ABSTRACT: To explore the effect of human endostatin expressed by host cells on the growth of human liver carcinoma in vivo.
Human endostain gene was transferred into SMMC7721 cells by retroviral pLncx to build endostatin-transfected cell line. PCR, immunohistochemistry and Western blot analysis were applied to examine the transfection, expression and secretion of endostatin. Endothelial cell proliferation assay was used to determine the biological activity of expressed endostatin. The in vivo and in vitro growth rates of the endostatin-transfected and control SMMC7721 cells were also observed.
PCR proved that the genome of endostatin-transfected SMMC7721 cells contained a 550 bp specific fragment of endostatin. The expression and secretion of human endostatin from endostatin-transfected SMMC7721 cells were confirmed by immunohistochemistry and Western blot analysis. Endostatin expressed by host cells could inhibit the proliferation of human umbilical vein endothelial cells by 48% (P < 0.01). In vitro proliferation assay showed that endostatin-transfected SMMC7721 cells had no change in proliferation rate compared to control SMMC7721 cells. In comparison with control group, however, tumor growth rate in vivo from endostatin-transfected SMMC7721 cells was inhibited greatly by 94.5%, 22 days after inoculation into nude mice (P < 0.01).
Human endostatin mediated by retroviral gene transfer can inhibit greatly the growth of human liver carcinoma SMMC7721 in vivo.
Zhonghua wai ke za zhi [Chinese journal of surgery] 09/2002; 40(9):692-5.
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ABSTRACT: To construct a stable transfectant of human liver carcinoma cell line SMMC7721 that could secret human endostatin and to explore the effect of human endostatin expressed by the transfectant on endothelial cell proliferation.
Recombinant retroviral plasmid pLncx-Endo containing the cDNA for human endostatin gene together with rat albumin signal peptide was engineered and transferred into SMMC7721 cell by lipofectamine. After selection with G418, endostatin-transfected SMMC7721 cells were chosen and expanded. Immunohistochemical staining and Western blot were used to detect the expression of human endostatin in transfected SMMC7721 cells and its medium. The conditioned medium of endostatin-transfected and control SMMC7721 cells were collected to cultivate with human umbilical vein endothelial cells for 72 hours. The inhibitory effect of endostatin, expressed by transfected SMMC7721 cells, on endothelial proliferation in vitro was observed by using MTT assay.
A 550 bp specific fragment of endostatin gene was detected from the PCR product of endostatin-transfected SMMC7721 cells. Immunohistochemistry and Western blot analysis confirmed the expression and secretion of foreign human endostatin protein by endostatin-transfected SMMC7721 cells. In vitro endothelial proliferation assay showed that 72 hours after cultivation with human umbilical vein endothelial cells, the optical density (OD) in group using the medium from endostatin-transfected SMMC7721 cells was 0.51 +/- 0.06, lower than that from RPMI 1640 group (0.98 +/- 0.09) or that from control plasmid pLncx-transfected SMMC7721 cells (0.88 +/- 0.11). The inhibitory rate for medium from endostatin-transfected SMMC7721 cells was 48%, significantly higher than that from empty plasmid pLncx-transfected SMMC7721 cells (10.2%, P<0.01).
Human endostatin can be stably expressed by SMMC7721 cell transferred with human endostatin gene and its product can significantly inhibit the proliferation of human umbilical vein endothelial cell in vitro.
World Journal of Gastroenterology 04/2002; 8(2):253-7. · 2.47 Impact Factor
