In vitro interaction of human pancreatic cancer cells and rat dorsal root ganglia: a co-culture model / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To establish an in vitro model of perineural invasion (PNI) with co-culture of human pancreatic cancer cells and rat root ganglion, to observe the neurite outgrowth and pancreatic cancer cell proliferation and migration, and to explore the molecular basis of perineural invasion (PNI) of pancreatic cancer.</p><p><b>METHODS</b>Human pancreatic cancer cell line (MIA PaCa-2) and rat dorsal root ganglion (DRG) were co-cultured in Matrigel matrix to generate the PNI model. The neurite outgrowth, pancreatic cancer cell colony formation, neurite-colony contact and retrograde migration were observed under an inverted microscope. The data were analyzed with the Image-Pro Plus 5.0 system. The proliferative index (PI) was measured by immunohistochemical staining with the Ki-67 antibody. In order to determine the absorbance (A) of the pancreatic cancer cells, MTT assay was used. The apoptotic index (AI) was evaluated by flow cytometry.</p><p><b>RESULTS</b>Neurite outgrowth was stimulated in the presence of pancreatic cancer cells. After 72 hours of the co-culture, MIA PaCa colonies co-cultured with DRG exhibited a significantly larger colony area (242.83 ± 4.92) than that of the control (182.50 ± 5.39, P < 0.001). In the MIA PaCa-2/DRG co-culture system, the neurites exhibited a trend of growing towards the pancreatic cancer cell colony. However, the pancreatic cancer cells showed a trend of retrogradely migrating to the DRG along the neurite outgrowth, when MIA PaCa-2 colonies touched the DRG. The positive rate of Ki-67 nuclear antigen was significantly higher than in the co-culture group. The PI value was higher in the experimental group (12.80%) than that in the control group (6.81%, P < 0.01). The MTT assay showed that proliferation of the pancreatic cancer cells was more active than that in the control group. Flow cytometry analysis showed that the apoptosis rate of the pancreatic cancer cell was 2.46%, significantly lower than that of the control group (4.89%, P < 0.001).</p><p><b>CONCLUSIONS</b>An in vitro co-culture model of rat dorsal root ganglion and human pancreatic cancer cell line is successfully established in this study. This MIA PaCa-2/DRG co-culture system demonstrates that the neural-pancreatic carcinoma cell interaction is a mutually beneficial process for the growth of neurites and pancreatic carcinoma cells. The pancreatic cancer cells show a trend of migrating to the DRG along the neurite outgrowth.</p>

Study on gene control region of mitochondrial DNA in familial breast cancer / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To study on mutations in D-loop region which is gene control region of mitochondrial genome in patients with familial breast cancer.</p><p><b>METHODS</b>Twenty-three breast cancer patients came from twenty-one families of breast cancer, and eighteen healthy controls participated in the study. PCR amplification of D-loop region in mitochondrial DNA was performed and then the product was sequenced to analyze mutations.</p><p><b>RESULTS</b>One hundred and twenty-six mutations in D-loop region were found in twenty-three patients with familial breast cancer, and four mutations were new. In all of twenty-three patients, thirty-seven mutations were found in D310 which was hot spot of D-loop region in mitochondrial DNA. In these mutations, T>C in 310, TC insert in 311-312, CA deletion in 522-523 and C>G in 527 were multi-presentation mutations in patients with familial breast cancer. Mutations had no difference in the same family member of breast cancer family except that occurrence in the region of D310. In the same family, mutations in D310 of patients were different from controls.</p><p><b>CONCLUSION</b>Mutations in D310 of familial breast cancer patients may enhance their susceptibility to breast cancer.</p>

Study on the effects of mitochondrial pathways on apoptosis in colon carcinoma cells induced by tumor necrosis factor related apoptosis inducing ligand / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To explore the effects of mitochondrial pathways on apoptosis in colon carcinoma cells induced by Tumor necrosis factor related apoptosis inducing ligand and offer evidences for TRAIL application in clinic.</p><p><b>METHODS</b>Apoptosis, integration of mitochondria (including DeltaPsim, cardiolipin), activity of Caspase-9 and release of cytochrome c in colon carcinoma cells SW1116 treated with TRAIL, were detected by means of flowcytometry, fluorometer method and western-blot at the different time point.</p><p><b>RESULTS</b>After treated with TRAIL for 4 hours, the apoptosis index was 32.98%, and the damage of mitochondria occurred with DeltaPsim, cardiolipin decreased, and the activity of Caspase-9 and cytochrome c increased. The Caspase-9 activity at 24 hour was (48.12+/-2.21)micromol.L(-1).h(-1).mg(-1) protein. Mitochondrial damage induced by TRAIL could be inhibited by Caspase inhibitor Z-VAD. fmk.</p><p><b>CONCLUSION</b>Mitochondrial pathways involved in the apoptosis of colon carcinoma cell induced by TRAIL. Cytochrome c was released and Caspase-9 was activated in the Caspase-dependent manner after the damage of mitochondrial.</p>

Study on the potential mechanisms of leukemia cell resistance to TRAIL-induced apoptosis / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the potential mechanisms of leukemia cell resistance to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) -induced apoptosis.</p><p><b>METHODS</b>Cells apoptosis, changes of mitochondrial membrane potential, activity of NF-kappaB, activity of caspase-8 and expressions of apoptosis-related proteins in TRAIL treated K562 and CEM cells, were detected by flowcytometry, ELISA and Western blotting methods, respectively.</p><p><b>RESULTS</b>After treated with TRAIL, the apoptosis indexes were 29.98% and 14.1%, and mitochondrial membrane potential were decreased to 73.25% and 25.4% in K562 and CEM cells respectively. Constitutive level of caspase-8 expression in CEM was lower than that in K562 cells. Both cells became over-expressed Bcl-xL and down-regulated Bax. The ratio of Bcl-xL/Bax in CEM cells was higher than that in K562 cells. Compared with that in K562, the NF-kappaB activity increased significantly in CEM after treatment with TRAIL in early stage.</p><p><b>CONCLUSION</b>CEM cells were more resistant to TRAIL-induced apoptosis than K562 cells did. The potential mechanisms associated with CEM drug resistance might be the lower expression of the constitutive level of caspase-8, lower sensitivity of mitochondrial inner membrane, early increase in NF-KB activity and altered expression of Bcl-2 proteins family.</p>

Study on inhibitory effect of antisense VEGF RNA on the growth of hepatocellular in vitro in vivo / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To determine the effects of PCMV-FGEV transfection on the profile of SMMC-7721 hepatocellular in vitro in vivo.</p><p><b>METHODS</b>SMMC-7721 hepatocellular was transfected with PCMV-FGEV antisense, PCMV-VEGF sense and empty vector plasmid encapsulated by lipofectamine. The positive cell clones were selected with G418. The stable transfection and expression of VEGF in the SMMC-7721 hepatocellular were determined by in situ hybridization and immunochemical analysis. The effect of PCMV-FGEV transfection on SMMC-7721 hepatocellular proliferation was observed by MTT colorimetric assay. Flow cytometry was used to determine the effects of PCMV-FGEV transfection on cell apoptosis of SMMC-7721. The growth of transfected cells was also observed in nude mice.</p><p><b>RESULTS</b>There was reduced VEGF expression in SMMC-7721 transfected with PCMV-FGEV confirmed by in situ hybridization and immunohistochemical analysis. There was no effect of PCMV-FGEV transfection on cell proliferation and cell apoptosis of SMMC-7721 in vitro. The growth of cell with PCMV-FGEV transfected was slow in nude mice (vivo) and accompanied with obvious apoptosis. The latent time of tumor in the antisense mice group was 25.0+/-1.8 days, which was longer than that in sense and control group significantly (F=19.455, P<0.01). On the other hand, the average tumor weight in antisense group (0.96 g+/-0.28 g) was the smallest among the three groups (F=21.501, P<0.01).</p><p><b>CONCLUSIONS</b>The expression of VEGF was inhibited by PCMV-FGEV. There was no effect on cell proliferation and cell apoptosis of SMMC-7721 by transferring PCMV-FGEV gene into SMMC-7721 cells in vitro. But in vivo it can inhibit tumor growth and induce cell apoptosis.</p>