Antitumor immunity by a dendritic cell vaccine encoding secondary lymphoid chemokine and tumor lysate on murine prostate cancer / 亚洲男科学杂志(英文版)

<p><b>AIM</b>To investigate the antitumor immunity by a dendritic cell (DC) vaccine encoding secondary lymphoid chemokine gene and tumor lysate on murine prostate cancer.</p><p><b>METHODS</b>DC from bone marrow of C57BL/6 were transfected with a plasmid vector expressing secondary lymphoid chemokine (SLC) cDNA by Lipofectamine 2,000 liposome and tumor lysate. Total RNA extracted from SLC+lysate-DC was used to verify the expression of SLC by reverse transcriptase-polymerase chain reaction (RT-PCR). The immunotherapeutic effect of DC vaccine on murine prostate cancer was assessed.</p><p><b>RESULTS</b>We found that in the prostate tumor model of C57BL/6 mice, the administration of SLC+lysate-DC inhibited tumor growth most significantly when compared with SLC-DC, lysate-DC, DC or phosphate buffer solution (PBS) counterparts (P < 0.01). Immunohistochemical fluorescent staining analysis showed the infiltration of more CD4(+), CD8(+) T cell and CD11c(+) DC within established tumor treated by SLC+lysate-DC vaccine than other DC vaccines (P < 0.01).</p><p><b>CONCLUSION</b>DC vaccine encoding secondary lymphoid chemokine and tumor lysate can elicit significant antitumor immunity by infiltration of CD4(+), CD8(+) T cell and DC, which might provide a potential immunotherapy method for prostate cancer.</p>

Functional localization of metastasis suppressor genes for HCC on human chromosome 8 / 中华肝脏病杂志

<p><b>OBJECTIVE</b>We previously showed that introduction of a normal, neomycin-tagged human chromosome 8 reduced the metastatic capacity of C5F rat liver cancer cell line, which had high metastatic potential without affecting tumorigenicity, suggesting the presence of one or more metastasis suppressor genes encoded on human chromosome 8. We proceeded to define further the region harboring the metastasis suppressor gene(s) and to determine the random loss of human chromosome 8 by PCR amplification of sequence tag site (STS) markers.</p><p><b>METHODS</b>The national Center for Biotechnology Information (NCBI) databases were used as references of the relative genetic distances of the STS markers. C5F genomic DNA and A9/neo8 genomic DNA were used as negative and positive controls for chromosome 8 amplification, respectively. Genomic DNA was isolated and quantified from cultured hybrid clones (A9/C5F-1 and A9/C5F-2 microcell hybrid clones served as metastasis-unsuppressed groups; A9/C5F-4, A9/C5F-8 and A9/C5F-10 microcell hybrid clones served as metastasis suppressed groups). STS-PCR products were separated by electrophoresis through 2% agarose gel.</p><p><b>RESULTS</b>Metastasis-suppressed microcell hybrid clones (A9/C5F-4, A9/C5F-8 and A9/C5F-10) conserved STS markers between D8S542 --> D8S1973 (8p21.1-23.1). In contrast, metastasis-unsuppressed clones (A9/C5F-1 and A9/C5F-2) lacked several markers in this region. In attempts to refine the region retained in the microcell suppressed clones, more densely spaced STS markers in the human chromosome 8p21.1-23.1 were used. We found that the metastasis-suppressed clones retained 18cM region between D8S542 and D8S1973 (8P21.1-23.1), where as the metastasis-unsuppressed clones lacked the region.</p><p><b>CONCLUSION</b>Our results suggest that a metastasis suppressor gene is located within the interval between D8S542 and D8S1973 on human chromosome 8p21.1-23.1.</p>

A preliminary study of the killing function in vitro by T lymphocytes activated by dendritic cells loaded with exosomes secreted by hepatic cancer cell lines with high or low metastatic potentials / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To study the tumor cell killing function of T lymphocytes stimulated by dendritic cells (DC) and to analyze the differences of protein contents of exosomes in each type of cell.</p><p><b>METHODS</b>The exosomes of hepatic cell lines with high (P group) or low (F group) metastatic potentials were isolated by a process of four-step centrifugation and the collected exosomes were observed under an electron microscope (EM). The tumor cell killing experiment was performed by adding T lymphocytes activated by DC loaded with exosomes from corresponding P and F group cells and was studied using 3H-TdR experiments. The proteomic analysis was performed by surface-enhanced laser desorption/ ionization time of flight mass spectrometry (SELDI-TOF-MS ) on the exosomes of P and F group cells.</p><p><b>RESULTS</b>The density distribution and content of exosomes in the P group were not equal to those in the F group observed by EM. The CD80, CD86, MHC-I and MHC-II in the P group were 64.27+5.00, 44.89+10.11, 84.35+19.89 and 59.03+19.37, and those in the F group were 71.53+4.85, 50.01+9.50, 80.68+29.87 and 58.86+21.11, respectively (P>0.05, compared with the control group). The counts per minute value in the P group was 528.40+179.06 and 78.80+24.44 in the F group after being loaded with exosomes (P<0.01, compared with the control group). There were significant differences between the proteins in the exosomes of hepatic cancer cell lines with high or low metastatic potentials.</p><p><b>CONCLUSION</b>Exosomes have potential values of application in immunotherapy and in biotherapy for recurrences and metastases of hepatic carcinomas.</p>

The relationship between lymphangiogenesis and lymphatic metastasis in murine hepatic carcinoma of high and low metastatic potentialities / 中华肝脏病杂志

<p><b>OBJECTIVES</b>To study the relationship between lymphangiogenesis and lymphatic metastasis in mice bearing hepatic carcinoma and analyze the mechanism of the lymphatic metastasis.</p><p><b>METHODS</b>Hepatic carcinoma cell lines of high and low potentialities of lymphatic metastasis were injected into the footpads of Balb/c mice. Their metastases to lymph nodes were examined. The tumor tissues of each group were stained with 5'-nucleotidase-ALP to observe the lymphoangiogenesis. The total RNA of high and low metastatic potential cell lines were extracted for metastasis gene DNA array. The vascular endothelial cell growth factor C (VEGF-C) and VEGF-D of each cell line were detected using semi-quantitative RT-PCR and were further quantatively analyzed using real time PCR.</p><p><b>RESULTS</b>The para-common iliac a. and renal hilar lymph nodes metastases of the high metastatic potential cells were significantly higher than in the controls (P>0.05). The quantity of lymphatic vessels in the high metastasis group was significantly larger than that of the control group (P<0.05). The expressions of CD44, E-cadherin, HER2/neu, H-Ras and VEGF-C in the high metastasis group were higher than those in the low metastasis group shown by the cDNA micro array experiment but the expressions of nm23A, nm23-E4, p16ink4a, CD61 were lower. The VEGF-C expression was higher and the VEGF-D was lower in the high metastasis group compared to those of the low metastasis group shown by semi-quantitative RT-PCR. The secretion of VEGF-D was significantly lower and the ratio of VEGF-C/VEGF-D was significantly higher in the high metastasis group than the low metastasis group (P<0.05).</p><p><b>CONCLUSIONS</b>The lymphatic metastasis of hepatic carcinoma is related to lymphoangiogenesis. The changes of VEGF-C and VEGF-D expressions might be a cause influencing the lymphoangiogenesis. VEGF-C/VEGF-D might be an effective parameter in affecting lymphatic metastases.</p>

Relationship between DLC-1 expressions and metastasis in hepatocellular carcinoma / 中华肝脏病杂志

<p><b>OBJECTIVES</b>To study the relationship between the expression level of DLC-1 mRNA (located in 8p) and the invasion/metastasis of human hepatocellular carcinoma (HCC).</p><p><b>METHODS</b>Fifty-one surgical specimens of human HCC were divided into high-invasive and low invasive groups according to their clinicopathological features. DLC-1 mRNA expression was studied in the 51 HCC specimens as well as 5 different metastasis potential cell lines using real-time quantitative PCR (RQ-PCR).</p><p><b>RESULTS</b>The expression level of DLC-1 mRNA in HCC specimens with high invasiveness was significantly lower than that with low invasiveness (P < 0.05). The expression levels of DLC-1 mRNA were significantly different between non-metastatic (Hep3B and HepG2) and metastatic (MHCC97-H, MHCC97-L and HCCLM3) cell lines (P < 0.05). From MHCC97-L to HCCLM3, with an increase of invasiveness and metastatic potentials, the expression level of DLC-1 decreased correspondingly, and its expression level in HCCLM3 was significantly lower than that in MHCC97-L (P < 0.01).</p><p><b>CONCLUSION</b>The expression of DLC-1 mRNA may play an important role in inhibiting the invasiveness and metastasis of HCC.</p>

Influence of reconstruction of immunological functions of T lymphocytes on mouse hepatocarcinoma metastasis / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the effectiveness of reconstruction of immunological functions of T cells on the degree of metastases of mouse hepatocarcinoma and the mechanisms of their functioning.</p><p><b>METHODS</b>The T cell model of immunological functions in Balb/c nu/nu mice was established and the effectiveness of the model was evaluated. The mice were divided into 4 groups. The immunological functions of T cells in experiment groups of Balb/c nu/nu mice were reconstructed. Metastases of the cancer in lymph nodes in each group were examined histologically. The formation time and growth rate of the tumors were calculated. The expression of MHCI and II of the tumor cell line and the difference of expression of immune associated gene were detected by Th1-Th2-Th3 gene array.</p><p><b>RESULTS</b>The ratio of CD3, CD4, CD8 and CD4/CD8 in the reconstructed group was higher than that in the control group. The average formation time was 7.7+/-0.6 days in Balb/c nu/nu mice and 11.5+/-1.3 days in Balb/c mice. The extent of metastases of the experiment group was lower than that of the control group (P < 0.05). The expression of MHCI of the high metastasis cell line was lower than that of the low metastasis cell line (P < 0.05). The expressions of Th1/Th2 associated genes in lymphocytes of high metastasis mice were lower than those of the low metastasis mice.</p><p><b>CONCLUSION</b>Reconstruction of the immunological function of T cells can influence the metastasis of mouse hepatocarcinoma. The alteration of MHC molecule and low expression of Th1/Th2 correlated genes in lymphocytes may be a factor influencing the metastasis of liver cancer.</p>

Effects of hepatectomized rat serum on the transdifferentiation of adult rat bone marrow cells into hepatocyte-like cells / 中华肝脏病杂志

<p><b>OBJECTIVES</b>To study the effects of hepatectomized rat serum and hepatocyte growth factor (HGF) on the transdifferentiation of adult rat bone marrow stem cells (ABMSCs) into hepatic parenchymal cells.</p><p><b>METHODS</b>The serum was collected from the rats 24 hours after being subjected to subtotal hepatectomy. ABMSCs were collected and cultured in DMEM/F12 (1:1) containing the hepaetectomized rat serum or HGF. The differentiated hepatocyte-like cells were labeled with CM-DiI and administrated by tail vein injection into the isogeneic rats. The cultured and injected cells were both identified by immunocytochemistry and cultured cells were assayed using RT-PCR and Western blot.</p><p><b>RESULTS</b>Hepatectomized rat serum and HGF were demonstrated to have the effect of inducing transdifferentiation of ABMSCs into hepatocyte-like cells in vitro. The differentiated cells expressed albumin mRNA and albumin after 7 days++'s co-incubation. Albumin-expressing and CM-DiI positive hepatocyte-like cells were characterized in livers and spleens of the rats injected with the cultivated cells.</p><p><b>CONCLUSION</b>ABMSCs could transdifferentiate into hepatic parenchymal cells by hepatectomized rat serum or HGF.</p>