H. pylori CagA activates the NLRP3 inflammasome to promote gastric cancer cell migration and invasion.

OBJECTIVE: The CagA (cytotoxin-related gene A, CagA) protein is an important factor for the pathogenicity of Helicobacter pylori (H. pylori). Although H. pylori has previously been shown to activate the NLRP3 inflammasome, it remains unclear what role CagA plays in this process. In the current study, we aimed to investigate the effect of CagA on NLRP3 activation and how it is linked to gastric cancer cell migration and invasion. METHODS: CagA positive H. pylori strain (Hp/CagA+) and CagA gene knockout mutant (Hp/ΔCagA) infected and the pcDNA3.1/CagA plasmid transfected gastric epithelial cell lines, respectively. The morphological alterations of cells under a microscope; the NLRP3 inflammasome-related markers: NLRP3, caspase-1, and ASC protein levels were detected by Western blot, IL-1ß and IL-18 levels were determined by ELISA; cell migration and invasion were determined by transwell assay; and the pyroptosis levels and intracellular ROS were determined by flow cytometry analysis. Then, pretreated with 5 mM NAC for 2 h and subsequently transfected with the pcDNA3.1/CagA plasmid for 48 h, the effects of NAC pretreatment on CagA-induced NLRP3 inflammasome-related markers expression and cell pyroptosis were examined, finally assessed the effect of CagA on migration and invasion in NLRP3-silenced cells. RESULTS: We found that Hp/CagA+ strain infection and pcDNA3.1/CagA vector transfection result in NLRP3 inflammasome activation, generation of intracellular ROS, and increased invasion and migration of gastric cancer cells. Moreover, we found that ROS inhibition via NAC effectively blocks NLRP3 activation and pyroptosis. Silencing of NLRP3 reduces the effects of CagA on gastric cancer cell migration and invasion. CONCLUSION: Our study shows that CagA can promote the invasion and migration of gastric cancer cells by activating NLRP3 inflammasome pathway. These findings provide novel insights into the mechanism of gastric cancer induction by H. pylori.

Bioinformatics analysis for the identification of differentially expressed genes and related signaling pathways in H. pylori-CagA transfected gastric cancer cells.

AIM: Helicobacter pylori cytotoxin-associated protein A (CagA) is an important virulence factor known to induce gastric cancer development. However, the cause and the underlying molecular events of CagA induction remain unclear. Here, we applied integrated bioinformatics to identify the key genes involved in the process of CagA-induced gastric epithelial cell inflammation and can ceration to comprehend the potential molecular mechanisms involved. MATERIALS AND METHODS: AGS cells were transected with pcDNA3.1 and pcDNA3.1::CagA for 24 h. The transfected cells were subjected to transcriptome sequencing to obtain the expressed genes. Differentially expressed genes (DEG) with adjusted P value < 0.05, - logFC -> 2 were screened, and the R package was applied for gene ontology (GO) enrichment and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. The differential gene protein-protein interaction (PPI) network was constructed using the STRING Cytoscape application, which conducted visual analysis to create the key function networks and identify the key genes. Next, the Kaplan-Meier plotter survival analysis tool was employed to analyze the survival of the key genes derived from the PPI network. Further analysis of the key gene expressions in gastric cancer and normal tissues were performed based on The Cancer Genome Atlas (TCGA) database and RT-qPCR verification. RESULTS: After transfection of AGS cells, the cell morphology changes in a hummingbird shape and causes the level of CagA phosphorylation to increase. Transcriptomics identified 6882 DEG, of which 4052 were upregulated and 2830 were downregulated, among which q-value < 0.05, FC > 2, and FC under the condition of ≤2. Accordingly, 1062 DEG were screened, of which 594 were upregulated and 468 were downregulated. The DEG participated in a total of 151 biological processes, 56 cell components, and 40 molecular functions. The KEGG pathway analysis revealed that the DEG were involved in 21 pathways. The PPI network analysis revealed three highly interconnected clusters. In addition, 30 DEG with the highest degree were analyzed in the TCGA database. As a result, 12 DEG were found to be highly expressed in gastric cancer, while seven DEG were related to the poor prognosis of gastric cancer. RT-qPCR verification results showed that Helicobacter pylori CagA caused up-regulation of BPTF, caspase3, CDH1, CTNNB1, and POLR2A expression. CONCLUSION: The current comprehensive analysis provides new insights for exploring the effect of CagA in human gastric cancer, which could help us understand the molecular mechanism underlying the occurrence and development of gastric cancer caused by Helicobacter pylori.

Analysis of the expression and clinical significance of ATM in gastric cancer tissues based on bioinformatics data / 中国肿瘤生物治疗杂志

@#[摘 要] 目的：探讨幽门螺杆菌（Helicobacter pylori, Hp）感染对胃癌细胞共济失调毛细血管扩张突变（ataxia-telangiectasia mutated，ATM）基因表达的影响及其临床意义。方法：从TCGA数据库中获取胃癌相关RNAseq数据，比较ATM基因的表达差异，分析ATM表达与患者临床病理参数的相关性及预后价值，用Kaplan-Meier法进行生存分析，LinkedOmics数据库分析ATM相关基因，用R语言进行GO、KEGG富集分析。选用2019年3月至2019年12月贵州医科大学附属医院12例手术切除的胃癌及癌旁组织标本，以及胃癌细胞系AGS和BGC823，用感染复数40∶1的Hp GZ7菌感染细胞，用免疫组织化学染色法检测胃癌组织中ATM蛋白的表达，qPCR法检测胃癌组织和细胞中ATM mRNA的表达。结果：TCGA数据显示胃癌和Hp感染胃癌组织中ATM miRNA表达水平均显著高于癌旁组织（均P<0.01）；胃癌组织中ATM miRNA表达与患者的T分期、AJCC分期等病理参数呈正相关（均P<0.05），ATM高表达时生存率显著降低（P<0.05）。实验检测显示，胃癌组织标本中ATM蛋白的表达水平明显高于癌旁组织（P<0.01）；Hp感染胃癌细胞中ATM miRNA表达水平显著高于未感染胃癌细胞（P<0.01）。胃癌中ATM基因与NPAT等12 461个基因呈正相关（P<0.05），与MIF等7 764个基因呈负相关（P<0.05）。GO、KEGG富集分析显示，ATM富集到DNA修复复合体、癌症中的转录失调等信号通路。结论：ATM基因在胃癌组织中高表达，患者生存率随表达水平的增高而降低，其与患者的T分期、AJCC分期等病理参数相关，且Hp感染引起ATM表达水平升高可能是Hp引起胃癌的原因之一。

Demographic and clinical characteristics of patients with type 1 diabetes mellitus: A multicenter registry study in Guangdong, China.

BACKGROUND: A lack of demographic and clinical data hinders efforts of health care providers in China to support patients with type 1 diabetes mellitus (T1D). Therefore, the aim of the present retrospective study was to provide an overview of the demographic and clinical characteristics of Chinese patients with T1D. METHODS: Hospital medical records of patients with T1D (diagnosed between January 2000 and December 2011) in 105 secondary and tertiary hospitals across Guangdong province were reviewed. Data were collected on patient age at diagnosis, presentations at onset, physical examination, and diabetes management. RESULTS: In all, 3173 patients diagnosed with T1D between January 2000 and December 2011 were included in the study (46.2% female). The median age at diagnosis was 27.5 years (interquartile range [IQR] 18.0-38.0) years and the median body mass index (BMI) at onset was 19.6 kg/m2 (IQR 17.4-21.8 kg/m2 ). Among adult patients, 0.9% were obese, 6.6% were overweight, 62.3% were normal weight, and 30.3 % were underweight. The prevalence of diabetic ketoacidosis (DKA) at onset was 50.1%. The proportion of patients with retinopathy, nephropathy, and neuropathy was 8.1%, 20.7 %, and 11.1%, respectively. CONCLUSION: The adult-onset form of T1D is not rare in China. The registry participants were characterized by older age at onset, lower BMI, and a higher prevalence of DKA at onset compared with those in regions with a high incidence of T1D, such as northern Europe. These findings contribute to a better understanding of the heterogeneity of T1D in different populations and so will help healthcare providers to develop management models that are more suitable for these patients.

[Cross-sectional study of current situation of glycemic control with type 1 diabetes in Guangdong Province].

OBJECTIVE: To investigate the glycemic control and the related factors of type 1 diabetic patients in Guangdong Province. METHODS: Medical records and blood samples of type 1 diabetic patients were collected in 89 tertiary and secondary hospitals from all of the 21 cities in Guangdong Province. The clinical data were analyzed to explore the correlates of glycemic control. HbA1c levels, measured in Guangdong Diabetes Center, were used to assess glycemic control. RESULTS: 851 patients were enrolled from August 6, 2010 to May 25, 2011. There were 408 males and 443 females. The median (interquartile range) age was 29.6 years (20.3 - 41.3 years). The onset age of diabetes was 25.3 years (15.7 - 35.5 years). The disease duration was 3.3 years (1.0 - 7.3 years). The BMI was 19.9 kg/m(2) (17.9 - 21.8 kg/m(2)). HbA1c levels were 8.6% (6.9% - 11.0%) and only 234 (27.50%) patients reached the age-specific target levels. Correlates with poorer glycemic control were 13 - 19 years old (vs 7 - 12 and ≥ 20 years old), lower household income, not on dietary intervention, never accepting diabetic education and shorter diabetic duration. CONCLUSION: The majority of Guangdong type 1 diabetic patients did not achieve target values for glycemic control, indicating an urgent need for comprehensive management to improve glycemic control.

[Relationship between different renal functional state with therapeutic effect on diabetic foot ulcers].

OBJECTIVE: To study the relationship between renal functional state and the therapeutic effect and prognosis of foot ulcers in the patients with diabetic mellitus. METHODS: The renal function was evaluated in term of glomerular filtration rate, microalbuminuria, proteinuria, blood urea nitrogen (BUN) and creatinine (Cr) levels in 126 patients with I-V class diabetic foot ulcers (according to Wagner classic standard) before the treatment, and then these patients were divided into 1-5 classes (according to Mogenson standard) and given systemic treatment and local debridement, with astragalus for topical application. The time of growth of granulation tissue (GT), the time of healing (HT), the amputation rate and mortality were observed. RESULTS: GT and HT of ulcer prolonged with worsening of diabetic nephropathy regardless of the disease phase of foot ulcers, especially the GT and HT of foot ulcers were significantly longer in IV and V phases of diabetic nephropathy than those of III phase diabetic nephropathy though the conditions of their foot ulcers were about the same. GT and HT in all the patients with the foot ulcers in the similar condition exhibited significantly positive linear correlation with the severity of diabetic nephropathy (r(1)=2.344 and r(2)=2.563, respectively, both P<0.05). The mortality of I-III phase diabetic nephropathy was significantly lower than that of IV and V phase diabetic nephropathy when the foot ulcers of these patients were of the same extent (P<0.05). CONCLUSION: A worsening of renal function would affect the treatment effect and prognosis of foot ulcers in the patients with diabetic foot ulcers, implicating that it is very important to improve the renal function in the treatment of patients with diabetic foot ulcers.