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Copyright of Journal of Henan Agricultural Sciences is the property of Editorial Board of Journal of Henan Agricultural Sciences and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Lung cancer is the leading cause of morbidity and mortality of malignant diseases in China. Approximately 57% lung cancer patients harbored distant metastases at initial diagnosis which is relevant to poor outcomes. The research strategy of anti-lung cancer metastasis now has became the new treatment directions and thoughts for lung cancer treatment. Previous studies have shown that changes in the corresponding driving genes on different signaling pathways may be related to the transfer of different organs, and the biological alteration of tumor to some extent can affect the metastatic behavior and metastatic pattern of tumor. However, current clinical and basic studies have not elucidated the molecular mechanism of the specific distant organ metastasis in the pathway of lung cancer related signal transduction, clinical research on the correlation between gene mutation and organ transfer specificity is also relatively rare. This review aims to summarize the characteristics of the expression of epidermal growth factor receptor (EGFR), anaplastic lymphoma kinase (ALK), V-Ki-ras2 Kirsten rat sarcoma viral oncogene homologue (KRS) in non-small cell lung cancer, and the correlation between the distribution of metastatic organs. [ABSTRACT FROM AUTHOR]

Background and objective It has been proven that CRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-associated 9) system was the modern gene-editing technology through the constitutive expression of nucleases Cas9 in the mammalian, which binds to the specific site in the genome mediated by single-guide RNA (sgRNA) at desired genomic loci. The aim of this study is that the animal model of EZH2 gene knockout was constructed using CRISPR/Cas9 technology. Methods In this study, we designed two single-guide RNAs targeting the Exon3 and Exon4 of EZH2 gene. Then, their gene-targeting efficiency were detected by SURVEYOR assay. The lentivirus was perfused into the lungs of mice by using a bronchial tube and detected by immunohistochemistry and qRT-PCR. Results The experimental results of NIH-3T3 cells verify that the designed sgEZH2 can efficiently effect the cleavage of target DNA by Cas9 in vitro. The immunohistochemistry and qRT-PCR results showed that the EZH2 expression in experimental group was significantly decreased in the mouse lung tissue. Conclusion The study successfully designed two sgRNA which can play a knock-out EZH2 function. An EZH2 knockout animal model was successfully constructed by CRISPR/Cas9 system, and it will be an effective animal model for studying the functions and mechanisms of EZH2. [ABSTRACT FROM AUTHOR]

Copyright of Chinese Journal of Lung Cancer is the property of Chinese Journal of Lung Cancer and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Objective: Constructing retroviral vector of small hairpin RNA (shRNA ) targeting human transducer and activator of transcription (STAT3) , establishing stable infected HPV 16 positive cervical cancer cell lines, to further explore the role of STAT3 in the development of HPV 16 positive cervical cancer. Methods: Design and synthetise three siRNA sequences targeting STAT3, then select one with best gene interference effect, and construct STAT3-shRNA vector, pSUPERretro-puro-STAT3. The recombinant plasmid was transfected into packaging cell line 293FT and then infected into HPV16 positive cervical cancer cell line SiHa and CaSki cells. Cells were selected with puromycin for 2 weeks, passaged around every 3 days , and named with SiHa/STAT3-sh and CaSki/STAT3-sh. Expression of STAT3 was analyzed by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western blotting. Results: The knockdown of the expression of STAT3 mRNA was over 99% (P<0.01) , that of the expression of STAT3 total protein was over 70% (P<0.01) and that of the expression of STAT3 phosphorylated protein was over 78% (P<0.01) in both SiHa/STAT3-sh and CaSki/STAT3-sh cells. Conclusions: HPV 16 positive cervical cancer cell lines with stable silence of STAT3 were successfully established. [ABSTRACT FROM AUTHOR]

Objective To investigate the expression of differentially expressed gene PAPSS2 in Kashin-Beck disease during the differentiation process of osteoblast-like cells MC3T3-E1 osteoblast and its effect on the cell mineralization and alkaline phosphatase (ALP) activity. Methods We cultured MC3T3-E1 cells and induced osteogenic differentiation, and then observed PAPSS2 expression during osteoblast differentiation process at the gene and the protein levels. PAPSS2 low expression shRNA lentivirus packaging and PAPSS2 retroviral vector were constructed, tittered, and checked for cell transfection efficiency. MC3T3-E1 was transfected and blank control group was set up. We observed changes of ALP activity and mineralization of osteoblasts after silence or high expression of PAPSS2. High expression vector was transfected with lentivirus-mediated RNA interference (RNAi) technology or retrovirus. Results The mRNA and protein expressions of PAPSS2 increased during differentiation of osteoblasts as the mineralization progressed. Our lentivirus-mediated RNAi technology successfully reduced the expression of PAPSS2 in MC3T3-E1 osteoblasts. And the retroviral transfection high expression vector significantly increased PAPSS2 gene expression. Silence PAPSS2 expression significantly decreased ALP activity and cell mineralization, contrary to its role in the high expression. Conclusion PAPSS2 can regulate ALP activity of osteoblasts and cell mineralization. Its abnormal expression may be related to bone lesion in Kashin-Beck disease. [ABSTRACT FROM AUTHOR]

PURPOSE: To establish immortalized oral mucosal epithelial (OME) cell line by transfection with human telomerase reverse transcriptase (hTERT). METHODS: Oral mucosal epithelial cells were infected with retroviral vector encoding hTERT. The infected cells were selected with G418 and checked by immunocytochemistry (ICC) for in vitro proliferation and the ability of tumorigenesis. RT-PCR and Western blot were used to detect the expression of hTERT. The data was processed with SPSS11.0 software package. RESULTS: Both infected and uninfected cells showed "flagstone" appearance; ICC confirmed the epithelial origin of the infected cells based on positive pan -cytokeratin and negative vimentin expression; Compared with uninfected cells, which arrested at the population doublings (PDL) of 9, the infected cells were more active by proliferated and reached at 80 to date; RT-PCR and Western blot showed hTERT expression. CONCLUSIONS: The infected oral mucosal epithelial cells were immortalized after transfection with hTERT and can serve as a genetically defined model of oral epithelial tumor study. Supported by National Natural Science Foundation of China(81072227) and Science and Technology Planning Project of Guangdong Province(2008B050100008). [ABSTRACT FROM AUTHOR]

Background and objective CD147, a type of transmembrane glycoprotein embedded on the surface of tumor cells, can promote tumor invasion and metastasis. This aim of this study is to construct a CD147 lentiviral expression vector, establish its stably transfected A549 cell line, and observe the effect of CD147 on MMP-9 proliferation as well as on the invasive ability of human lung adenocarcinoma cells. Methods Full-length CD147 gene was amplified by real-time polymerase chain reaction (RT-PCR), inserted into a pEGFP vector to construct pEGFP-CD147 and pEGFP vectors, and then transfected into 293FT cells to precede the lentivirus equipment package. Subsequently, we collected the lentivirus venom to infect the A549 cells and establish a stable, overexpressed cell line named A549-CD147. The mRNA expression of MMP-9 was examined by RT-PCR. The proliferation and invasive ability of the human lung cancer cells before and after transfection were examined by the CCK-8 and Transwell methods. Results A CD147 lentiviral expression vector (pEGFP-CD147) was successfully constructed by restrictive enzyme digestion and plasmid sequencing. RT-PCR and Western blot analyses revealed increased mRNA and protein expression of CD147 gene in cells transfected with pEGFP-CD147 compared with the control groups. Therefore, the A549-CD147 cell line was successfully established through the experiment. The mRNA expression of MMP-9 also significantly increased after the upregulation of CD147 expression. Meanwhile, CCK-8 and Transwell assays indicated that the proliferation and invasive ability significantly increased in the A549-CD147 cells. Conclusion A lentiviral CD147 expression vector and its A549 cell line (A549-CD14) were successfully constructed. CD147 overexpression upregulated the protein expression of MMP-9, and strengthened the proliferation and invasive ability of human lung adenocarcinoma cells. [ABSTRACT FROM AUTHOR]

PURPOSE: To investigate the influence of HPVI6 E5 on E6 and E7 gene in human oral immortalized epithelial cells (HIOEC). METHODS: The pLEGFP-E5 was transferred into HIOEC cell; Deletion mutation expression vectors were constructed and transferred into HIOEC; RT-PCR was used to detect the gene expression in HIOEC; The expression of E6 anti E7 mRNA was detected by real time-PCR;Cell proliferation of HIOEC was evaluated by MIT. All statistical analysis was performed by using SPSS 13.0 software package. RESULTS: The deletion mutation expression vector of E5 was successfully constructed and applied for further experiments; E5 and the deletion mutation genes were transferred into HIOEC successfully; HPVI6 E5 promoted cell proliferation of HIOEC and made the increased expression raised of E6 and E7 genes; However, the deletion mutation had no significant effect on HIOEC. CONCLUSIONS: HPVI6 E5 is transferred into HIOEC successfully. E5 may promote cell proliferation and upregulate the mRNA expression of E6 and E7 genes. Supported by Natural Science Foundation of Anhui Province (070413086). [ABSTRACT FROM AUTHOR]

PURPOSE: To investigate the effect of Dlx2 overexpression on osteogenic differentiation of MC3T3-E1 cells in vitro. METHODS: Dlx2-expression retrovirus vector was constructed by subcloning and verified by sequencing. MC3T3-E1 cells were transfected with pMSCV-Dlx2 and selected with puromycin for stable clones. The expression of Dlx2 was determined by RT-PCR and Western blot. The level of osteogenetic biomarkers ALP, OCN, Runx2 and Msx2 was quantified by real-time PCR. ALP detection and Alizarin red staining were conducted to evaluate the effect of Dlx2 overexpression on osteogenic differentiation. The data were analyzed for ANOVA with SNK method using SAS 6.04 software package. RESULTS: pMSCV-Dlx2 was successfully constructed and verified by direct sequencing and Dlx2 overexpression in vitro. Enhanced ALP activity and Alizarin red staining were observed in MC3T3-E1-Dlx2 cells as compared with the control. During osteogenetic induction, Dlx2 overexpression up-regulated ALP and Msx2 in the early stage, while enhanced OCN expression in the late stage. Runx2 expression was different but not reached statistical significance. CONCLUSION: Dlx2 overexpression induces osteogenic differentiation of MC3T3-E1 cells via up - regulating bone formation-related genes. Supported by Shanghai Leading Academic Discipline Project (S30206), Research Fund of Science and Technology Commission of Shanghai Municipality (10JC1408700), Research Fund of Shenkang Company (SHDC12010205), Research Fund of Shanghai Municipal Health Bureau (2009077) and Combined Engineering and Medicine Project of Shanghai Jiao Tong University (YG2010MS55). [ABSTRACT FROM AUTHOR]

Objective To construct the lentiviral vector carrying miR - 122 ( Lv - miRl 22 ) and investigate the function of stably expressed miR - 122 in hepatocellular carcinoma ( HCC) cells. Methods Pre - miR - 122 was cloned into lentiviral vector pLUNIG to obtain the Lv - miR122 that had stable expression of miR - 122 ; Lv - miR122 was packaged in 293T cells along with other three vectors pRRE, pRSV -REV, and pCMV - VSVG, and virus supernatant was obtained by concentration; the virus supernatant was used to infect HCC cell line HepG2. The expression of miR - 122 was measured by Q - RT - PCR. The proliferation of HepG2 cells was analyzed by MTT assay and col-ony formation assay. Blank lentiviral vector ( Lv - Ctrl) - infected HepG2 cells and uninfected HepG2 cells were used as controls. Results The expression of miR - 122 was significantly higher in Lv - miR122 - infected HepG2 cells than in Lv - Ctrl - infected HepG2 cells ( t -- 10. 745, P < 0. 01 ). The Lv - miR122 - infected HepG2 cells ( with stable expression of miR - 122 ) had significantly suppressed prolifera-tion compared with Lv - Ctrl - infected and uninfected HepG2 cells (P <0. 05 for both comparisons) ; moreover, miR - 122 significantly in-hibited the colony formation of HepG2 cells ( t -- 5. 256, P < 0. 01 ) . Conclusion The Lv - miR122 can be successfully constructed, and the carried miR -122 can be stably expressed and can inhibit the proliferation of HepG2 cells. [ABSTRACT FROM AUTHOR]