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Objective:To investigate the factors associated with the occurrence of coccoid forms in clinical isolates of Helicobacter pylori ( H. pylori) and its relationship with the pathological changes of gastric mucosa. Methods:A total of 66 H. pylori-infected patients admitted to People′s Hospital of Ningxia Hui Autonomous Region from January 2020 to June 2021 were included.The clinical data of the patients were collected. Immunohistochemical staining was performed on gastric mucosal biopsy specimen to observe the occurrence of coccoid forms of H. pylori and pathological changes of gastric mucosa. Chi-square test was used for statistical analysis. Results:After immunohistochemical staining of 66 gastric mucosa biopsy specimens from H. pylori-infected patients, the co-existence of helical and coccoid forms of H. pylori was found in 26(39.39%) specimens, and no simple coccoid H. pylori change was found. Among them, the patients with a history of eradication therapy had a coccoid forms rate of 52.63%(20/38), and those without a history of eradication therapy had a coccoid forms rate of 21.43%(6/28), and the difference was statistically significant ( χ2=6.57, P=0.012). There were no significant differences in the coccoid forms rates between patients with different gender, ethnicity, age and gastric mucosal pathological changes (including atrophy, gastric intestinal metaplasia, inflammation, activity)(all P>0.050). Seventeen (73.9%) of the 23 patients whose endoscopy was more than one to three months from the last eradication therapy developed coccoid forms, while three of 15 patients whose endoscopy was more than three months from the last eradication therapy had coccoid forms, and the difference was statistically significant ( χ2=10.59, P=0.002). Conclusions:The transformation of H. pylori coccoid forms is related to the previous eradication therapy. The coccoid forms of H. pylori is equally pathogenic relative to the helical forms of H. pylori.
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Objective:To compare ultrasound (US) guided versus computed tomography (CT) guided radiofrequency ablation (RFA) in treatment of early hepatocellular carcinoma (HCC).Methods:The data of 133 patients with early HCC treated by RFA in the Department of Hepatobiliary Surgery of Shandong Provincial Hospital from February 1, 2015, to January 31, 2017, was analyzed retrospectively. These patients were divided into two groups: the US-guided group and the CT-guided group. The clinical data was collected and the factors affecting prognosis were analyzed.Results:Compared with the CT-guided group, the operation time of the US-guided group was significantly shorter [(29.0±12.0)min vs. (55.0±19.0)min, P<0.05], but the number of ablation sessions per tumor was significantly less [(1.1±0.3) vs. (2.0±0.6), P<0.05]. There was no significant difference in the complete ablation rates, postoperative complication rates and postoperative length of hospital stay between the two groups ( P>0.05). The CT-guided group was superior to the US-guided group in the local tumor recurrence and progression-free survival rates ( P<0.05). On multivariate analysis, CT-guided RFA was an independent protective factor for local tumor recurrence ( HR=0.266, 95% CI: 0.073-0.967, P<0.05) and progression-free survival ( HR=0.415. 95% CI: 0.213-0.806, P<0.05), while AFP >20 ng/ml ( HR=4.821, 95% CI: 1.714-13.560, P<0.05) was an independent risk factor for progression-free survival. Conclusion:CT-guided percutaneous RFA was superior to US-guided RFA in local treatment of early HCC, probably related to more needle placements and longer ablation time under CT guidance.
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Objective To evaluate the feasibility of the combination of amplification refractory mutation system (ARMS) and quantitative real-time polymerase chain reaction (PCR) method in fast detection of clarithromycin resistance of Helicobacter pylori (H.pylori) in gastric mucosa.Methods A total of 150 gastric mucosal specimens with positive H.pylori culture were collected from the H.pylori positive patients who failed in H.pylori eradication from January to August in 2013.The drug resistant gene mutation types of H.pylori in these samples were detected by quantitative real-time PCR based on ARMS.And the accuracy was confirmed by sequencing.The clarithromycin resistance of H.pylori was determined by E-assay.Chi-square test was used for statistical analysis.Results Among 149 gastric mucosal specimens (one specimens without wild type or mutation type had been eliminated),the results of quantitative real-time PCR based on ARMS of two samples were not consistent with the results of sequencing;the consistent rate was 98.7％ (147/149).Among 149 specimens with positive H.pylori culture,104 samples (69.8％) were clarithromycin resistance.In 101 samples the clarithromycin resistance was detected by quantitative real-time PCR based on ARMS;the consistent rate was 97.1％ (101/104).Both E-assay and clarithromycin resistant rate detected by E-assay or quantitative real-time PCR based on ARMS was 69.8％ (104/149) and 67.8％ (101/149),respectively,and the difference was not significant (x2 =0.141,P=0.932).Conclusion The combination of ARMS and quantitative real-time PCR method in fast detection of clarithromycin resistance of H.pylori in gastric mucosa is strongly feasible and highly consistent has high consistent rate with sequencing and E-assay.
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Objective To evaluate the feasibility of the combination of amplification refractory mutation system (ARMS) and quantitative real-time polymerase chain reaction (PCR) method in fast detection of clarithromycin resistance of Helicobacter pylori (H.pylori) in gastric mucosa.Methods A total of 150 gastric mucosal specimens with positive H.pylori culture were collected from the H.pylori positive patients who failed in H.pylori eradication from January to August in 2013.The drug resistant gene mutation types of H.pylori in these samples were detected by quantitative real-time PCR based on ARMS.And the accuracy was confirmed by sequencing.The clarithromycin resistance of H.pylori was determined by E-assay.Chi-square test was used for statistical analysis.Results Among 149 gastric mucosal specimens (one specimens without wild type or mutation type had been eliminated),the results of quantitative real-time PCR based on ARMS of two samples were not consistent with the results of sequencing;the consistent rate was 98.7％ (147/149).Among 149 specimens with positive H.pylori culture,104 samples (69.8％) were clarithromycin resistance.In 101 samples the clarithromycin resistance was detected by quantitative real-time PCR based on ARMS;the consistent rate was 97.1％ (101/104).Both E-assay and clarithromycin resistant rate detected by E-assay or quantitative real-time PCR based on ARMS was 69.8％ (104/149) and 67.8％ (101/149),respectively,and the difference was not significant (x2 =0.141,P=0.932).Conclusion The combination of ARMS and quantitative real-time PCR method in fast detection of clarithromycin resistance of H.pylori in gastric mucosa is strongly feasible and highly consistent has high consistent rate with sequencing and E-assay.
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Background:Tissue microarray has been increasingly used in research of malignancies. It has been revealed that TGF-β signaling pathway contributes to the tumorigenesis and progress of malignancies. Aims:To determine the expressions of Runx3,Smad4,Cdk2 and p21,the key molecules in TGF-β signaling pathway by tissue microarray,and investigate their correlations with clinicopathological features and prognosis of gastric cancer. Methods:A total of 378 paraffin embedded tissue blocks,including 130 gastric cancer tissue and 248 para-cancer tissue from 130 patients undergoing radical resection of gastric cancer were obtained. Tissue microarray and immunohistochemistry were used to determine the expressions of Runx3,Smad4,Cdk2 and p21. Results:The aberrant expression rates of Runx3,Smad4, Cdk21 and p21 in gastric cancer tissue were significantly higher than those in para-cancer tissue(67. 7% ,35. 4% , 63. 8% and 70. 0% vs. 14. 1% ,12. 5% ,18. 1% and 37. 1% ,P < 0. 05,respectively). Aberrant expression of Runx3 was closely correlated with histological grade and lymph node metastasis of gastric cancer( P < 0. 05),while aberrant expressions of Smad4 and p21 were correlated with histological grade only(P < 0. 05);aberrant expression of Cdk2 was correlated with histological grade,lymph node metastasis and TNM staging(P < 0. 05). Pairwise correlations were seen among aberrant expressions of Runx3,Smad4 and p21 in gastric cancer,while Cdk2 was correlated with Runx3 only. Kaplan-Meier survival curve showed that 5-year survival rates in Runx3,Smad4 and p21 aberrant expression groups were significantly lower than those in normal expression groups(P <0. 05). Furthermore,Cox proportional hazard model indicated that Runx3 and Smad4 were independent prognostic factors for gastric cancer. Conclusions:Runx3,Smad4,Cdk2 and p21 might play pivotal roles in tumorigenesis and progression of gastric cancer. As interactions occurred among these four proteins,whether Runx3 and Smad4 could be used for predicting the prognosis of gastric cancer needs to be further studied.
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Objective To investigate the gene expression difference of IFN and their receptors in peripheral blood mononuclear cells (PBMC) of pulmonary embolism (PE) patients.Methods Twenty cases of PE patients and twenty sex and age matched controls were recruited into the study.Human cDNA microarray analysis was used to detect the gene expression difference of IFN associated genes between the two groups,and random variance model corrected t test was used to analyze the statistical data.Results In comparison with the control group, mRNA expression of type Ⅰ IFN, including IFNα5 mRNA,IFNα6 mRNA,IFNα8 mRNA,IFNα14 mRNA,IFNκ mRNA,IFNω1 mRNA,IFNε1 mRNA in PBMC of PE patients Were down-regulated (P ＜ 0.05 ).There was no significant difference in gene expression of type Ⅰ IFN receptors IFNαR1 and IFNαR2 between the PE and control groups (P ＞ 0.05 ).In comparison with the control group,mRNA expression of IFNγ gene was down-regulated ( P ＜ 0.05 ).The mRNA expression of IFNγR1 and IFNγR2 genes were upregulated compared with the control (P ＞ 0.05 ).Conclusion mRNA expression of type Ⅰ and type Ⅱ IFN in PE are significantly down-regulated,but not the IFN receptors.Reduced immune function may play an important role in the PE patients who are susceptible to virus,intracellular bacteria and parasites.
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Objectives To investigate the expression of mTOR signaling pathway in pancreatic cancer and export its signification. Methods 6 samples of pancreatic cancer and its paracancerous tissues specimens confirmed by surgery and pathologic examination were selected. RNA was extracted and expression profiles experiment was performed by using Agilent human whole genomic oligonucleotide microarrays. The expression of mTOR signaling pathway in pancreatic cancer was analyzed by bioinformatics. Results Totally 1276 differential gene were selected, and 691 were up-regulated in cancer tissue, while 585 were down-regulated.The highest score of KEGG pathway's Enrichment and gene count was hsa04150 in mTOR signaling pathway,with its Enrichment of 4.5622519 and gene count of 9, and the percentage of gene count was 1.15%, the EASE Score P value was 6.23 E-04, which had the most biological significance. Among those, there was significantly difference of expression of nine key genes including ULK2, PIK3R3, PDPK1, EIF4EBP1, PGF,VEGFB, ULK3, RICTOR and PIK3 R5 (P ＜ 0.05). Conclusions The pathogenesis of pancreatic cancer is related to the activation of the mTOR signaling pathway.
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Objective To study the expression of PIWIL1, PIWIL3 and PIWIL4 in human colon cancer and its clinical significance. Methods We collected cancerous tissues and its adjacent tissues of 106 patients with colon cancer, two tissue microarrays were constructed, with 62 and 150 points respectively. We studied the expression of PIWIL1, PIWIL3 and PIWIL4 through immunohistochemistry. Results The expression of PIWIL1, PIWIL3 and PIWIL4 were significantly higher in cancerous tissues than those in adjacent tissues (P＜0. 01). In cancerous tissues and its adjacent tissues, postive correlation were seen among PIWIL1, PIWIL3 and PIWIL4 expression (P＜0. 01). PIWIL1 expression was significant higher in low differentiation group than that in high differentiation group (t =- 2. 840, P＜0.01 ). PIWIL3 expression was higher in high clinical stage than that in low clinical stage (F= 3. 112, P＜0.05). The expression of PIWIL3 and PIWIL4 were significantly higher in patients with colon cancer with distant metastasis than those without distant metastasis (t= -3. 349, P＜0.01 ; t = - 2. 168, P＜0. 05). PIWIL3 and PIWIL4 expression were correlated with occurring of colon cancer (P＜0. 01). Conclusions The expressions of PIWIL1,PIWIL3 and PIWIL4 in colon cancer were correlated with the differentiation, clinical stage and distant metastasis of colon cancer. PIWIL3 and PIWIL4 expression were two independent related factors of occurring of colon cancer, which would be furtherly investigated to be served as novel markers for early diagnosis and promising molecular targets for colon cancer therapy.
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Objective To evaluate the impact of rosiglitazone (Ros) on liver expression of peroxisome proliferator-activated receptor γ (PPARγ),nuclear factor (NF-κB) and cyclooxygenase-2(COX-2) in treatment of nonalcoholic steatohepatitis (NASH) rats.Methods Thirty Sprague-Dawley rats were divided into normal group,model group and Ros treated group with 10 each.Except the normal group,the other two groups were given high fat diet for 12 weeks for NASH model.The rats in Ros treated group were gavaged 4 mg/kg of Ros daily at the 12th week for 8 weeks.All rats were sacrificed at the 20th week for blood sample and liver tissue.Biochemical parameters of liver function,lipid metabolism,glycometabolism and antioxidant enzyme activities were measured.The histological change of the liver were assessed with HE and Masson staining.The level of tumor necrosis factor (TNF)-α and prostaglandin E2 (PGE2) was measured using ELISA.The expression of PPARγ,NF-κB and COX-2 was detected with immunohistochemistry.The mRNA and protein expressions of COX-2 were tested by real-time PCR and Western blotting,respectively.Results In comparison with model group,Ros treated group showed significant improvement in hepatic steatosis,inflammation and fibrosis(all P value<0.05).In model group,the serum levels of fasting blood glucose,insulin and HOMA-insulin resistance index (HOMA-IRI),total cholesterol (TC),total triglyeride (TG),lowdensity lipoprotein cholesterol (LDL-C) and free fatty acids were increased,but HDL-C level was decreased.All above parameters markedly improved after Ros treatment.The levels of ALT and AST,total anti-oxidation competence,superoxide dismutase,catalase,glutathione peroxidase and malondialdehyde in Ros treated group were significantly ameliorated when compared with those in model group.Immunohistochemistry showed that the expression of NF-κB and COX-2 was significantly elevated,but PPARγ was decreased in model group.Real-time PCR and Western blot revealed that the mRNA and protein expressions of COX-2 were higher in the model group than those in normal group (0.57±0.08 vs 0.38±0.03;2.83±0.24 vs 1.00±0.03,P=0.000 and P=0.004,respectively),but significantly lower in Ros treated group (0.55±0.06 and 1.84±0.13,P<0.01).Conclusions Ros can reduce oxidative stress and insulin resistance in NASH rats by activing PPARγ expression and inhibiting expression of NF-κB and cyclooxygenases.
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Objective To investigate the microRNAs expression profile in human colorectal cancer with or without liver metastasis and try to screen miRNA associated with liver metastasis in colorectal cancer. Methods Twenty five surgical resected colorectal cancer specimens were collected and frozen in liquid nitrogen. Three without liver metastasis and three with liver metastasis were selected, from which total RNAs were isolated. The expressions of miRNAs in these two types of specimens were detected by illumine microRNA microarray, and the difference of miRNA expression was screened. The biochip results were verified with real-time RT-PCR in all colorectal caner specimens. Results The miRNA expression was significantly different in colorectal cancer with liver metastasis and without liver metastasis. Compared with colorectal cancer without liver metastasis, 28 miRNA expressions was different in colorectal cancer with liver metastasis, 4 up regulated and 24 down regulated. The quantity of miR-139-3p expression in colorectal cancer with liver metastasis was 1.75±0.40, up regulated compared with that incolorectal cancer without liver metastasis(0. 69 ±0.58,P＜0.05). The quantity of miR-19a expression in liver metastasis was 0. 39±0. 20, downregulated compare with no liver metastasis( 1.38 ± 0.98, P＜0. 05). The result of miRNA biochip was consistent with that of RT-PCR. Conclusion The difference of miRNA expression might relate to liver metastasis of colorectal cancer. The specific miRNAs expression profile might provide new target for diagnosis and treatment of colorectal cancer with liver metastasis.
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Objective To investigate microRNA expression profiles of colonic cancer without lymph node metastasis and identify the specific microRNAs associated with carcinogenesis of colon.Methods Cancerous and para-cancerous tissues (5 cm from cancer tissues of the colon) confirmed without lymph node metastasis were collected from 3 patients. The microRNAs were extracted and isolated by mirVana RNA kit. Hybridizations were made by applying the microRNAs to Agilent microRNA microarray. Data analysis was done by software of Feacture Extraction. The discovered microRNAs were confirmed by real time PCR assay. Results Twelve out of 14 microRNAs associated with colonic cancer were up-regulated in colonic tissues. They were miR-106b, miR-135b,miR-18a,miR-18b,miR-196b, miR-19a, miR-224, miR-335, miR-424, miR-20a' , miR-301b and miR-374a. The rest two of miR-378 and miR 378* were down-regulated. Two up-regulated microRNAs (miR-18a and miR-135b) were detected in 3 pairs of cancerous and para-cancerous tissues. The expression level of miR-18a and miR-135b were accordant with the results of real time PCR.Conclusions microRNA.such as miR-106b,miR-135b.miR-18a.miR-18b,miR-196b,ect,were differentially expressed between cancerous and para-cancerous tissues.Many of these target genes are supposed to participate in the process of multiple tunmorgenesis.These microRNAs play an important role in the carcinogenesis of colon.
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Objective To detect the differential expression genes(DEGs)between Barrettg esophagus(BE)and normal esophagus with oligomicroarray,and to explore the target genes related to the development of BE.Methods The total RNAs of matched BE and normal esophagus mucosa from saIne patient were isolated with one step Trizol method.Matched RNAs were qualified with 10g/L agarose gel electrophoresis.After tRNA purification,cRNAs were synthesized and labeled with fluorescence.which were tIlen hybridized with Agilent oligomicroarray containing 30,968 probes.The fluorescence intensity features were detected by Agilent scanner and quantified by software Feature Extraction.Results On average,2 biopsies by disposable jumbo biopsy forceps provided approximately 5μg RNA required for microarray.The total RNA,reverse transcription product and fluorescence labeled cRNA were all of high quality.Among 2-fold DEGs,there were 142 up-regulated genes and 284 down-regulated genes including 15 bel-2 related genes such as bel-2,MCL1,BAX,BIK and BCLAF1 Conclusion Microarray-based studies are feasible in endoscopically obtained tissues.The development of BE is a complicated process involving multi-genes,in which abnormal expression of bel-2 family related genes might be involved,but the exact mechanism needs further research.
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Objective To investigate the whole genomic expression profiles of chronic atrophic gastritis with interleukin(IL)-1β-31CC/-511TT genotype as measured by oligonucleotide microarray technique.Methods Genomic RNA was extracted from gastric biopsies of 12 patients with chronic atrophic gastritis(6 with IL-1β-31CC/-511TT and 6 with IL-1β-31TT/-511CC).The genomic profiles of IL-1β gene polymorphisms 31CC/-511TT and 31TT/-511CC were compared and tested for differential expressed genes associated with 31CC/-511TT using Agilent human whole genomic oligonucleotide microarrays.The results were further analyzed in terms of gene ontology(GO).Results There were 200 differentially expressed genes associated with IL-1β-31CC/-511TT,159 of which were up-regulated and 41 were down-regulated.These genes mainly involved in macromolecule metabolic process,post-translational protein modification,ubiquitin cycle,and protein kinase cascade.Five genes had biological activities,one of which was down-regulated gene(PCSK5)and 4 were upregulated genes(PRKCA,NPLOC4,TRIB3 and MAPKAPK3).Conclusions The chronic atrophic gastritis with IL-1β-31CC/-511TT genotype has molecular phenotypes which is associated with malignance and inflammation.These individuals are needed more intensive preemptive treatment and dynamic surveillance.
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genes related to pancreatic cancer was mainly associated with biological process,cellular location,molecular function,which suggested the development of pancreatic cancer was caused by multiple genes.
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ssociated genes,especially down-regulated expression of T cell mediated function genes,in patients with PE indicates that the etiology of PE might be related to viral infection.
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Objective To prepare polyclonal antibodies against human PIWIL and to identify their property and tissue distribution of PIWIL.Methods PIWIL polypeptide was synthesized and conjugated to Keyhole limpet hemocyanin(KLH) as an immunogen.Then PIWIL-KLH conjugations were injected into rabbits subcutaneously to produce polyclonal antibodies.The specificity and sensitivity of antibodies were identified by ELISA and Western blot after purification by affinity chromatography.PIWIL were then stained on the tissue chip to study their distribution.Results Rabbit antibodies against PIWIL were prepared after injection of PIWIL-KLH conjugation.These antibodies specially recognized PIWIL peptides.Expression of PIWIL was found in the cytoplasm of epithelia cells of varied normal tissues and tumor tissues.Conclusion The successful preparation of the polyclonal antibody against PIWIL will provide an efficient reagent for further study of its role in the pathway of miRNA and RNA interference and in the pathogenesis of human disease.
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Objective To study the effect of the recombinant vaccinia virus expressing human interleukin 2 on MKN45 gastric cancer cells in vitro. Methods Recombinant vaccinia virus expressing hIL 2 (VMJ601hIL 2) was constructed by homologous recombination using molecular virology, and VMJ601hIL 2 was detected by DNA hybridization technique and the recombinant gene product was analyzed by SDS PAGE. In addition, MKN45 gastric cancer cells was infected by VMJ601hIL 2 and the effect of VMJ601hIL 2 on the gastric cancer cells was evaluated in vitro. Results hIL 2 protein with biological activity can be secreted by MKN45 gastric cancer cells after heavily infected by VMJ601hIL 2. Conclusions It is one of the crucial steps that VMJ601hIL 2 has been constructed and identified since it forms essential prerequisite for further in vivo gene therapy of gastric cancer.
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Objective:To prepare rabbit polyclonal antibody against human argonaute 3(AGO3) protein,to identify its properties and investigate the tissue distribution of AGO3 using tissue array.Methods:AGO3 peptide was synthesized using chemical method,and then conjugated to Keyhole limpet hemocyanin(KLH) as immunogen.The AGO3-KLH conjugate were injected into rabbits subcutaneously to produce polyclonal antibodies.The specificity and sensitivity of antibodies were identified by ELISA and Western blot after purification using affinity chromatography.Then the distribution of AGO3 in tissues was examined through immuno-stainning by tissue array.Results:Rabbit polyclonal antibodies against AGO3 were raised after immunization with AGO3-KLH conjugates.The anti-serum titer after the last inoculation was up to 1∶20 000.The preparations of the antibody were confirmed to raised recognize AGO3 peptides specially by ELISA and Western blot.AGO3 protein was stained positively in the cytoplasm of tumor cells and epithelial cells in many normal tissues.Conclusion:The polyclonal antibody against AGO3 protein has been achieved successfully,and it provides an efficient tool for further studying the roles of AGO3 in the pathway of miRNA and RNA interference and in the pathogenesis of human disease.