Suppression of Aurora-A by RNA interference inhibits laryngeal cancer Hep-2 cell growth / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To investigate the effects of knockdown of Aurora-A by RNA interference on laryngeal cancer Hep-2 cell growth in vitro and in vivo.</p><p><b>METHODS</b>A plasmid containing siRNA against Aurora-A was constructed and transfected into human laryngeal cancer cell line Hep-2. Measurements included the CCK-8 assay for viability and proliferation, Transwell assay for invasion, colony formation assay for cell anchorage-independent growth. Western blot and immunohistochemistry assay for protein expression. Tumorigenicity was observed in vivo.</p><p><b>RESULTS</b>In Hep-2 cells transfected by Aurora-A siRNA (designated as siRNA-3), protein expression of Aurora-A was suppressed by 52%. In CCK-8 assay, absorbance value of siRNA-3 cells (3.268 ± 0.106, (x(-) ± s)) was lower than that of Hep-2 cells (3.722 ± 0.152, F = 17.634, P < 0.001). In Transwell assay, the average invasive cells per field in siRNA-3 cells (110.0 ± 18.0) was less than that in Hep-2 cells (236.0 ± 26.0, F = 26.462, P < 0.01). In colony formation assay, the average colony number of siRNA-3 cells (31.0 ± 6.6) was lower than that of Hep-2 cells (104.0 ± 14.0). The average tumor size in siRNA-3 group was (127.77 ± 174.83) mm(3), which was less than Hep-2 cell group (837.26 ± 101.80) mm(3), (F = 28.187, P < 0.001). Silencing of Aurora-A decreased the expression of focal adhesion kinase (FAK) and matrix metalloproteinase-2 (MMP-2), key regulators in cell adhesion and invasion.</p><p><b>CONCLUSIONS</b>The knockdown of Aurora-A inhibits the growth and invasiveness of Hep-2 cells in vitro and in vivo, which may be a promising therapeutic strategy for LSCC.</p>

Etiology and antimicrobial resistance of community-acquired pneumonia in adult patients in China / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Appropriate antimicrobial therapy of community-acquired pneumonia (CAP) is mainly based on the distribution of etiology and antimicrobial resistance of major pathogens. We performed a prospective observational study of adult with CAP in 36 hospitals in China.</p><p><b>METHODS</b>Etiological pathogens were isolated in each of the centers, and all of the isolated pathogens were sent to Zhongshan Hospital for antimicrobial susceptibility tests using agar dilution.</p><p><b>RESULTS</b>A total of 593 patients were enrolled in this study, and 242 strains of bacteria were isolated from 225 patients. Streptococcus pneumoniae (79/242, 32.6%) was the most frequently isolated pathogen, followed by Haemophilus influenzae (55/242, 22.7%) and Klebsiella pneumoniae (25/242, 10.3%). Totally 527 patients underwent serological tests for atypical pathogens; Mycoplasma pneumoniae and Chlamydia pneumoniae infections were identified in 205 (38.9%) and 60 (11.4%) patients respectively. Legionella pneumophila infections were identified in 4.0% (13/324) of patients. The non-susceptibility rate of isolated Streptococcus pneumoniae to erythromycin and penicillin was 63.2% and 19.1% respectively. Six patients died from the disease, the 30-day mortality rate was 1.1% (6/533).</p><p><b>CONCLUSIONS</b>The top three bacteria responsible for CAP in Chinese adults were Streptococcus pneumonia, Haemophilus influenza and Klebsiella pneumonia. There was also a high prevalence of atypical pathogens and mixed pathogens. The resistance rates of the major isolated pathogens were relatively low except for the high prevalence of macrolide resistance in Streptococcus pneumoniae.</p>

Interaction of SerpinB5 and MAFbx in gastric cancer cell and its action site / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To explore the interaction between SerpinB5 and MAFbx in gastric cancer cell and to identify the interaction sites.</p><p><b>METHODS</b>The interaction between SerpinB5 and MAFbx was screened and validated by yeast two-hybrid screening and co-immunoprecipitation. The expression of MAFbx was analyzed after SerpinB5 expression being modified by RNA interference and pGBKT7-SerpinB5 transfection. The impact of SerpinB5 on the expression of MAFbx was studied in gastric cancer cell line SUN-16. A model of MAFbx was constructed by homology modeling. The related residues for interaction were analyzed by Autodock4.0.</p><p><b>RESULTS</b>The interaction between SerpinB5 and MAFbx was validated. The expression of MAFbx changed along with SerpinB5 expression. Amino acids including PRO261, ASN361, and LYS362 were key residue in the interaction of SerpinB5 and MAFbx.</p><p><b>CONCLUSION</b>SerpinB5 interacts with MAFbx in gastric cancer cell. Amino acids including PRO261, ASN361, and LYS362 are potential binding sites.</p>

Influence of silencing Polo-like kinase 1 on migration and invasion of colorectal cancer cells / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To examine the role of Polo-like kinase 1(PLK1) in the migration and invasiveness of human colorectal cancer cells.</p><p><b>METHODS</b>Nine colorectal cancer cell lines were cultured. Cell lines with the highest level of PLK1 expression was selected by PCR and Western blot. Three siRNA oligo segments targeting PLK1 were designed and selected cell lines transfected. Successful transfection was confirmed using real-time PCR and Western blot. Changes in migration and invasiveness of the selected cell line were evaluated by Transwell test.</p><p><b>RESULTS</b>Colorectal cancer cell line SW1116 was selected with the highest expression of PLK1 at both mRNA level and protein level. The expression of PLK1 in SW1116 was reduced by the three siRNA oligo segments to varying degrees, and the No.1 siRNA oligo segment was the most efficient. In migration test, the number of cells crossing through chambers in PLK1-siRNA group was 44 ± 14, which was lower than that in the negative control group (242 ± 40) and in blank control group(240 ± 38). In invasion test, the number of cells crossing through chambers in PLK1-siRNA group was 62 ± 3, which was lower than that in negative control group (207 ± 12) and in blank control group (211 ± 15). These differences were statistically significant(P<0.01).</p><p><b>CONCLUSION</b>PLK1 silencing by siRNA may inhibit the migration and invasiveness of colorectal cancer cells, suggesting that PLK1 might play an important role in the infiltration and metastasis of colorectal cancer.</p>

Establishment of experimental angiogenic models with applications of quantitative digital image analysis / 中华病理学杂志

<p><b>OBJECTIVE</b>To establish experimental models for tumor neovascularization and to apply quantitative digital imaging analysis in the study.</p><p><b>METHODS</b>An endothelial tube formation model was established by human umbilical vein endothelial cells (HUVECs). A vasculogenic mimicry model was established by SGC-7901 gastric cancer cell line. Fertilized eggs were used to establish a chorioallantoic membrane angiogenesis model. Using gene transfection experiment, IRX1 tumor suppressor gene was chosen as a therapeutic target. Image Pro Plus (IPP) analysis software was used for digital vascular images analysis with parameters including points, lines, angles and integral absorbance (IA) for the tubular formation or vasculogenic mimicry.</p><p><b>RESULTS</b>Digital image analysis by IPP showed that HUVEC tubular formation was significantly inhibited in IRX1 transfectant, compared with controls. The tubular numbers in three groups were 12.80 +/- 3.83, 29.00 +/- 5.34 and 28.20 +/- 4.32 (P<0.01). The connection points of tubules in three groups were 13.20 +/- 2.59, 25.00 +/- 2.24 and 24.60 +/- 3.21 (P<0.01). The tubular lengths of three groups were (821.5 +/- 12.5), (930.9 +/- 13.5) and (948.4 +/- 18.1) microm (P=0.022). The IA values of PAS stain in three groups were 3606 +/- 363, 14 200 +/- 1251 and 15 043 +/- 1220 (P<0.01). In chick chorioallantoic membrane model, the angular numbers of tubules in three groups were 6.41 +/- 2.60, 10.27 +/- 2.65 and 9.18 +/- 1.99 (P<0.01).</p><p><b>CONCLUSIONS</b>The endothelial tube formation model, vasculogenic mimicry model and chorioallantoic membrane angiogenesis model are useful for gene therapy and drug screening with targeting neoplastic vascularization. Professional image analysis software may greatly facilitate the quantitative analysis of tumor neovascularization.</p>

Silencing invariant chains of dendritic cells enhances anti-tumor immunity using small-interfering RNA / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Genetic modification of dendritic cells (DCs) has been used as an effective approach to enhance anti-tumor immunity. RNA interference (RNAi), which can cause the degradation of any RNA in a sequence-specific manner, is a post-transcriptional gene silencing mechanism. In this study, small-interfering RNA (siRNA) specific for the Ii gene was transfected into DCs, and the anti-tumor immunity of Ii-silenced DCs was assessed.</p><p><b>METHODS</b>The silencing effect of siRNA was evaluated by Western blotting and real-time PCR analyses. In vitro cytotoxic activity of T cells was evaluated using a Cytotox 96(®) non-radioactive cytotoxicity assay kit. The time to tumor onset and the tumor volumes were used as reliable indices to assess the anti-tumor immunity in vivo. To further examine the mechanisms underlying the anti-tumor immunity, flow cytometry analysis was used.</p><p><b>RESULTS</b>The Ii expression of DCs was significantly reduced after Ii siRNA transfection. Significant in vitro anti-tumor ability was exhibited when DCs were co-transfected with Ii siRNA plus endogenous tumor antigen (P < 0.05). Furthermore, tumor growth was greatly inhibited when mice were immunized with DCs transfected with Ii siRNA plus tumor antigen prior to or subsequent to tumor implantation. Flow cytometry analysis in vitro and in vivo indicated that both CD4(+) and CD8(+) T cells were significantly activated in the Ii siRNA group (P < 0.05).</p><p><b>CONCLUSION</b>Silencing of the Ii gene of DCs may offer a potential approach to enhance DC-based anti-tumor immunity.</p>

Demethylation of the gamma-synuclein gene CpG island in colorectal cancer and its clinical significance / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To explore the relationship between gamma-synuclein gene expression and CpG island demethylation in colorectal cancer(CRC), and the relationship between the demethylation and clinicopathological factors of CRC.</p><p><b>METHODS</b>The expression of gamma-synuclein mRNA was examined in 30 pairs of tumor tissues and tumor-matched non-neoplastic adjacent tissues(NNAT) by RT-PCR. CRC cell lines including COLO205, LoVo, and SW480 were used and treated with a demethylating agent, 5-aza-2'-deoxycytidine(5-aza-C). Before and after the treatment, the expression of gamma-synuclein mRNA in the cells was determined by RT-PCR, and bisulfite sequencing PCR was also used to analyze methylation status of CpG island. The methylation status of gamma-synuclein was then examined in 67 CRC samples and 30 NNAT samples by nested methylation-specific PCR (NMSP) and real time methylation-specific PCR(real-time MSP). The relationship between the demethylation of gamma-synuclein in CRC and clinicopathological factors was analyzed.</p><p><b>RESULTS</b>The mean gamma-synuclein mRNA expression was 0.66+/-0.34 in CRC samples, which was much higher than 0.45+/-0.26 in NNAT samples(P=0.011). 5-aza-C could induce expression and demethylation of gamma-synuclein in COLO205, LoVo and SW480 cells. gamma-Synuclein gene was demethylated in 80.0%(24/30) of the CRC samples and 50.0%(15/30) of the NNAT samples. The demethylated status of gamma-synuclein was much higher in CRC samples than that in NNAT samples(P=0.030), and was significantly correlated with clinical stage, lymph node involvement, and distant metastasis of CRC(P<0.05).</p><p><b>CONCLUSION</b>The upregulation of gamma-synuclein expression in CRC is primarily attributed to the demethylation of CpG island, which may be used as a marker for prognosis.</p>

Analysis of human gastric cancer by transcriptome and proteome profiling / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To screen differential expression genes and proteins at transcriptome and proteome levels between human gastric cancer tissue and corresponding normal mucosa.</p><p><b>METHODS</b>Fresh-frozen gastric cancers were collected from patients treated at Ruijin Hospital. A total of 22 pairs of gastric cancer tissues and the corresponding noncancerous mucosa were analyzed. Commercially available cDNA microarray with 14 592 genes/ESTs was used. Genes were considered to be up-or down-regulated when the intensity ratio Cy3/Cy5 was > or = 2 or < or = 0.5 in over 50% samples (P<0.05). Immobilized pH gradient(IPG)-based 2-DE was applied to separate the total proteins of gastric cancer tissue and paired normal tissue. After staining and analysis by software,the differential expression proteins were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS) or MALDI-TOF-TOF-MS.</p><p><b>RESULTS</b>As compared with corresponding noncancerous tissue, there were totally 149 up-regulating genes/ESTs and 238 down-regulating genes/ESTs in gastric cancer, including 29 genes with 3-fold over-expression ratio and 21 genes with 5-fold under-expression. Fifteen protein spots were identified successfully, among whom there were ten over-expressed and five under-expressed proteins in gastric cancer tissue compared with normal tissue. Most of over-expressed genes and proteins were related to cell motility, cell proliferation, signal transduction, while those under-expressed genes and proteins were related to defense response, toxoid metabolism.</p><p><b>CONCLUSION</b>Studying gastric cancer at transcriptome and proteome levels can help demonstrate tumorigenesis and biological characteristics of gastric cancer comprehensively and provide powerful tools to find new biomarkers associated with gastric cancer and therapy targets.</p>

Hedgehog signaling pathway activates in gastric carcinoma and promotes the proliferation through GLI1 in MKN28 cell / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To investigate the effect of Hedgehog (HH) pathway on proliferation and in vitro tumorigenicity of gastric cancer cell lines.</p><p><b>METHODS</b>The expression of SHH, PTCH, SMO, SUFU and GLI1 in seven cell lines were tested by RT-PCR. siRNA targeting GLI1 mRNA was transfected into MKN28 cells. Cell proliferation and in vitro tumorigenicity were examined by CCK8 and soft agar colony formation test.</p><p><b>RESULTS</b>SHH in six gastric cancer cell lines was up-regulated. Expression of PTCH in KATOIII cell lines and expression of SUFU in MKN28 and KATOIII were reduced. GLI1 siRNA significantly inhibited the expression of GLI1 in MKN28 cell line. Growth rate and colony formation rate of MKN28 cells treated with GLI1 siRNA were significantly lower than those of control cells (all P <0.001).</p><p><b>CONCLUSIONS</b>HH signaling pathway is widely activated in gastric cancer cell lines. The activation of HH signaling pathway promotes the growth of MKN28 cells.</p>

Vascular endothelial growth factor D promoting the growth of lymphatic vessel in gastric carcinoma / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To investigate the relationship between vascular endothelial growth factor D (VEGF-D) and metastasis of lymphatic vessel in gastric carcinoma.</p><p><b>METHODS</b>The human VEGF-D cDNA was amplified from total RNA isolated from human normal gastric tissue, then it was inserted into T-A clone plasmid and subcloned into pEGFP eukaryotic expression vector. After the full-length sequence expected was confirmed by enzymatic digestion and sequencing,the human gastric carcinoma cell line SGC7901, which expressed a low level of VEGF-D, was transfected with the pEGFP/VEGF-D expression vector. Stable SGC7901 clones with high expression of VEGF-D were selected in vitro with G418, which were then combined and subcutaneously injected into nude mice to observe the density and morphology of lymphatic vessel. The outcomes were later compared with those of SGC7901 cells transfected with null vector(pEGFP) by immunostaining with a specific antibody LYVE-1.</p><p><b>RESULTS</b>The average weight of tumors in the pEGFP group (1.13+/-0.40) g at day 35, was significantly lower than that in the pEGFP/VEGF-D group (2.24+/-0.82)g (P<0.05). The lymphatic vessel density (LVD) in the pEGFP group (2.89+/-1.32) was significantly lower than that in the pEGFP/VEGF-D group (5.74+/-1.30)(P<0.01). There were dilated functional lymphatic vessels around the tumor margin.</p><p><b>CONCLUSION</b>VEGF-D may promote the growth and metastasis of tumor in gastric carcinoma by increasing the growth of lymphatic vessels.</p>

Effects of N(omega)-nitro-L-arginine methyl ester and aminoguanidine on lipopolysaccharide-induced airway hyperresponsiveness in guinea pigs / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>The down-regulation of constitutive nitric oxide synthase (cNOS) and up-regulation of inducible nitric oxide synthase (iNOS) are associated with the allergen-provocated airway hyperresponsiveness (AHR). This study aimed to determine whether their alteration also plays an important role in the AHR induced by lipopolysaccharide (LPS).</p><p><b>METHODS</b>Hartley male guinea pigs, weighing between 250 g and 350 g, were injected with LPS at a dose of 1 mg/kg every 24 hours for three days. A non-selective NOS inhibitor, N(omega)-nitro-L-arginine methyl ester (L-NAME), or a selective inducible NOS inhibitor, aminoguanidine (AG), were used thirty minutes before each injection of LPS. Airway reactions, nitric oxide (NO) production and inflammatory changes were detected 24 hours after the last dose of LPS.</p><p><b>RESULTS</b>AG significantly decreased the NO production in the bronchoalveolar lavage fluid (BALF) and sharply reduced the intensity of bronchoconstriction to histamine challenge. L-NAME also significantly decreased the NO production in the BALF, but had no effect on airway reactions or, perhaps, a tendency to enhance the intensity of AHR.</p><p><b>CONCLUSIONS</b>The data suggest that inducible NOS contributes to the AHR induced by repetitive intraperitoneal LPS, and constitutive NOS was also involved.</p>

Cloning and functional analysis of the tumor metastasis related gene tropomyosin2 / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To construct an eukaryotic expression vector of tumor associated gene tropomyosin 2 (TM2) tagged with green fluorescence protein and study its biological role in tumorigenesis. METHODS; Whole length of TM2 cDNA sequence was amplified from human gastric cancer cell line AGS by reverse transcript PCR. The recombinant TM2 was inserted into eukaryotic expression vector pIRES2 tagged with enhanced green fluorescence protein. pIRES2-EGFP was transfected into human hepatoma cell line QGY-7703 cells by liposome technique and detected by fluorescence microscopy and flow cytometry. TM2 protein level was determined by Western blot. The in vitro invasion and motility were further studied. Its tumorigenesis was assessed on nude mice.</p><p><b>RESULTS</b>The recombinant TM2 was shown to be expressed in QGY-7703 cells by fluorescence microscopy and flow cytometry. The protein level of TM2 was increased. The invasion ability of TM2- transfected cells was increased(45.6 +/- 9.9) than that in controls(21.6 +/- 3.3, P < 0.05), as well as mobility(41.4 +/- 11.8 vs. 16.7 +/- 3.7). The tumor volume was (241.5 +/- 95.1) mm3, significantly higher than that in blank-vector controls (100.6 +/- 85.4) mm3 and negative-controls (123.7 +/- 92.3) mm3.</p><p><b>CONCLUSION</b>TM2 plays a role in growth and metastasis of hepatic tumor.</p>

Inhibition of the proliferation of human gastric cancer cells SGC-7901 in vitro and in vivo using Bcl-2 siRNA / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Bcl-2, the anti-apoptotic protein is overexpressed in the majority of gastric cancers and associated with its pathogenesis. To better understanding of the role of Bcl-2, RNA interference (RNAi) was used to inhibit Bcl-2 expression in the human gastric cancer cells in vitro and in vivo.</p><p><b>METHODS</b>Bcl-2 small interfering RNA (siRNA) was transfected into human gastric cancer cells SGC-7901, and Bcl-2 expression was monitored by real-time polymerase chain reaction (PCR) and Western blot. Cell proliferation, apoptosis, and telomerase activity were examined by MTT, flow cytometry, and TRAP assay, respectively. Gastric cancer cells treated with 100 nmol/L Bcl-2 siRNA were subcutaneously transplanted into nude mice and tumor growth was assessed.</p><p><b>RESULTS</b>Bcl-2 siRNA significantly inhibited the expression of Bcl-2 in human gastric cancer cells at both mRNA and protein levels in a time- and dose-dependent manner. Bcl-2 siRNA also decreased telomerase activity (by 78.76%) and increased the rate of apoptosis (by 37.47%). SGC-7901 cell growth was also significantly suppressed in vivo and in vitro.</p><p><b>CONCLUSIONS</b>Bcl-2 expression knockdown suppressed the growth of gastric cancer cells. Thus, Bcl-2 may play a very important role in carcinogenesis of gastric cancer and its knockdown may offer a new potential gene therapy approach for human gastric cancer in future.</p>

Polo like kinase 1 expression and prognostic value in gastric carcinomas / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To identify the expression of polo like kinase 1 (plk1) and to discuss its relationship with the clinicopathological parameters and prognosis in gastric carcinoma.</p><p><b>METHODS</b>Plk1 protein expression levels in 89 cases of resected gastric carcinomas were detected by immunohistochemistry method, the relations between plk1 expression levels and the survival periods were estimated by Kaplan-Meier curve.</p><p><b>RESULTS</b>The positive rate of plk1 expression in gastric cancer tissues was 42.7% (38/89), significantly higher than that (13.5%) in the adjacent noncancerous tissues (12/89) (P<0.01). The expression levels of plk1 were closely related to tumor differentiation, invasion and TNM stage (P<0.05). Patients with plk1-positive expression had worse prognosis than those with plk1-negative expression in gastric cancer patients (P<0.05).</p><p><b>CONCLUSIONS</b>Plk1 may promote carcinogenesis and gastric cancer development, its overexpression can be a novel marker for diagnosing certain biological behaviours and predicting prognosis in gastric cancer.</p>

Chromosomal alterations analyzed by comparative genomic hybridization in primary gastric carcinoma / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To identify genetic abnormalities in primary gastric carcinoma.</p><p><b>METHODS</b>Comparative genomic hybridization (CGH) was used in screening DNA copy number changes along all chromosomes in 23 cases of primary gastric cancer.</p><p><b>RESULTS</b>Twenty-one out of 23 cases showed chromosomal losses and gains for at least one of the chromosomal arms in primary gastric cancer. The mean number of chromosomal alterations was 7.52. Chromosomal gains predominated over chromosomal losses in a ratio of 5.38:2.14. The most often involved chromosomal gains were observed in 8q (9/21, 42.9%), 20q (9/21, 42.9%), 17q (8/21, 38.1%), 3q (7/21, 33.3%), 7q (7/21, 33.3%), 11q (6/21, 28.6%), 13q (6/21, 28.6%), 1q (5/21, 23.8%) and 20p (5/21, 23.8%). The chromosomal arms with frequent losses were 17p (7/21, 33.3%), 18q (6/21, 28.6%), 5q (5/21, 23.8%), 8p (5/21, 23.8%), and 9p (5/21, 23.8%).</p><p><b>CONCLUSIONS</b>The phenomenon of gain and loss of chromosomal regions is observed in primary gastric cancer, which may induce the amplification of oncogenes and the loss of tumor suppressor genes to regulate the development and progression of gastric cancer.</p>

Mitosis arrest caused by inhibition of PLK1 expression in gastric cancer MKN45 cells / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To observe the effect of polo-like kinase 1 (PLK1) gene depletion on mitosis phenotype and elucidate its vital role in gastric cancer cell line (MKN45) mitosis.</p><p><b>METHODS</b>The PLK1 expression in MKN45 cells was blocked by RNA interference (RNAi), the expression level of PLK1 mRNA and protein were measured by real-time quantitative PCR and Western blot, respectively. The morphological change of microtubules and mitosis phenotype in MKN45 cells were observed by immunofluorescence staining and laser confocal microscopy, the morphological changes of cells were observed by reverse microscopy, the variation of cell cycle distribution was detected by flow-cytometry.</p><p><b>RESULTS</b>After RNAi targeting PLK1, PLK1 mRNA and protein level decreased obviously, the cell microtubules became obscure and lost cohesiveness, the mitosis phenotype also varied substantially (P < 0.05), more gastric cancer cells became rounded and showed G(2) phase cell DNA content (P < 0.05).</p><p><b>CONCLUSION</b>PLK1 gene plays a key role in mitosis and its inhibition can lead to mitosis arrest in MKN45 cells.</p>

Mitotic arrest of gastric cancer cells induced by silencing of STK15 gene / 中华病理学杂志

<p><b>OBJECTIVE</b>To investigate the role of STK15 in regulating mitosis of gastric cancer cells (MKN45) by gene silencing through RNA interference mechanism.</p><p><b>METHODS</b>RNA interference technique was used to inhibit STK15 expression in MKN45 cells. The expression levels of STK15 mRNA and protein were measured by real-time quantitative RT-PCR and Western blot respectively and cell morphological changes were investigated by reverse microscopy. In addition, cell cycle distribution and cellular proliferation were determined by flow-cytometry and MTT assay respectively. Finally, the mitotic phenotype of MKN45 cells was studied by immunofluorescence staining and confocal microscopy.</p><p><b>RESULTS</b>Silencing of STK15 gene by RNA interference was confirmed by marked decrease of STK15 mRNA and protein levels in the treated MKN45 cells. This silencing correlated with rounding of the cells, decreasing of DNA content in G(2) phase (P < 0.05) and a lowered proliferation index (P < 0.05), along with alterations of mitotic phenotype of MKN45 (P < 0.05).</p><p><b>CONCLUSION</b>STK15 gene may play a key role in regulating cellular mitosis and its inhibition by RNA interference leading to mitosis arrest in MKN45 cells.</p>

Immunotherapeutic efficacy of both helper T lymphocytes and cytotoxic T lymphocytes epitopes augmented dendritic cells tumor vaccine on gastric cancer / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To investigate the immunotherapy efficacy of both helper T lymphocytes (Th) and cytotoxic T lymphocytes (CTL) epitopes augmented dendritic cells (DCs) tumor vaccine on gastric cancer.</p><p><b>METHODS</b>Naïve spleen T cells were stimulated by mixed peptides (a mixture of Th epitope MAGE-3 (22-36)) primed DCs per week in vitro. After 4 cycles of restimulation, peptide specific T cells were harvested and subgroups of which were determined with flow cytometry. Cytokines secreting profiles by CD4+ T cells and cytotoxicities of CD8+ T cells on tumor cells were assessed. The protective immunity by referred DCs tumor vaccines was also monitored.</p><p><b>RESULTS</b>Both Th and CTL epitopes primed DCs could elicit both CD4+ T cells and CD8+ T cells in vitro,of which CD4+ T cells released high amount of Th1 type cytokines (IFN-gamma, IL-2) on recognizing specific antigen, as well as CD8+ T cells exhibited efficient tumor-killing capacity. The effects induced by DCs pulsed with single epitope (Th or CTL epitope) in vivo were less effective than those induced by DCs pulsed with mixture epitopes.</p><p><b>CONCLUSIONS</b>Both Th and CTL epitopes augmented DCs tumor vaccine can induce CD4+ Th1 and CD8+ CTL mediated immune responses to eradicate gastric cancer cells.</p>

Identification of biomarkers in the serum of the patients with poorly differentiated gastric cancer / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To investigate the application of proteomics in the field of serology,and to screen the differential expression proteins related with poorly differentiated gastric carcinoma.</p><p><b>METHODS</b>Two-dimensional electrophoresis (2-DE) was applied to segregate the total proteins in the serum form gastric cancer patients and health volunteers. After staining,the differential expression proteins were analyzed using PDQuest software,and identified using matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS).</p><p><b>RESULTS</b>Electrophoresis figures with high resolution and reproducibility were obtained. Six differential expression proteins were found only in the serum from gastric cancer patients, while four other proteins from healthy volunteers.</p><p><b>CONCLUSIONS</b>Protein expression is differential in the serum from the gastric cancer patients and health volunteers. It is hopeful to find the biomarkers related with poorly differentiated gastric carcinoma using proteomics.</p>

Inhibition of serine/threonine kinase 15 gene expression induces apoptosis of gastric cancer cells / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To observe the effect of inhibition of serine/threonine kinase15 (STK15) gene expression on apoptosis induction in gastric cancer cell line-MKN45 and discuss the role of STK15 in viability of gastric cancer cells.</p><p><b>METHODS</b>The STK15 expression was inhibited by chemically synthesized siRNA. The STK15 mRNA and protein level were respectively measured by real-time quantitative PCR and western blotting,the change of cell cycle distribution and apoptosis rate were detected by flow-cytometry, cell morphological change was observed by Hoechst staining,and pro-caspase 3 level was also detected by western blot.</p><p><b>RESULTS</b>After treatment by siRNA targeting STK15 after 48 h, STK15 mRNA and protein level decreased obviously. More MKN45 cells accumulated at G(2)/M phase (P< 0.05). The apoptosis rate of STK15 siRNA treated MKN45 cells was higher than that of control cells(P< 0.05) with the pro-caspase 3 level decreased.</p><p><b>CONCLUSIONS</b>Inhibition of STK15 gene expression may induce apoptosis in MKN45 cells through the pathway of caspase3. STK15 gene play a key role in proliferation and viability of MKN45 cells.</p>