Wen-Liang Fang

Epigenetic regulation of miR-34b and miR-129 expression in gastric cancer.

MicroRNAs (miRNAs) are small noncoding RNAs that play fundamental roles in diverse biological and pathological processes by targeting the expression of specific genes. Here, we identified 38 methylation‐associated miRNAs, the expression of which could be epigenetically restored by cotreatment with 5‐aza‐2′‐deoxycytidine and trichostatin A. Among these 38 miRNAs, we further analyzed miR‐34b, miR‐127‐3p, miR‐129‐3p and miR‐409 because CpG islands are predicted adjacent to them. The methylation‐silenced expression of these miRNAs could be reactivated in gastric cancer cells by treatment with demethylating drugs in a time‐dependent manner. Analysis of the methylation status of these miRNAs showed that the upstream CpG‐rich regions of mir‐34b and mir‐129‐2 are frequently methylated in gastric cancer tissues compared to adjacent normal tissues, and their methylation status correlated inversely with their expression patterns. The expression of miR‐34b and miR‐129‐3p was downregulated by DNA hypermethylation in primary gastric cancers, and the low expression was associated with poor clinicopathological features. In summary, our study shows that tumor‐specific methylation silences miR‐34b and miR‐129 in gastric cancer cells.

Aberrant hypermethylation of miR-9 genes in gastric cancer

Carcinogenesis of the stomach involves multiple steps including genetic mutation or epigenetic alteration of tumor suppressor genes or oncogenes. Recently, tumor suppressive miRNAs have been shown to be deregulated by aberrant hypermethylation during gastric cancer progression. In this study, we demonstrate that three independent genetic loci encoding for miR-9 (miR-9-1, miR-9-2 and miR-9-3) are simultaneously modified by DNA methylation in gastric cancer cells. Methylation-mediated silencing of these three miR-9 genes can be reactivated in gastric cancer cells through 5-Aza-dC treatment. Subsequent analysis of the expression levels of miR-9 showed that it was significantly down-regulated in gastric cancers compared with adjacent normal tissues (P value < 0.005). A similar tendency toward a tumor-specific DNA methylation pattern was shown for miR-9-1, miR-9-2 and miR-9-3 in 72 primary human gastric cancer specimens. Ectopic expression of miR-9 inhibited cell proliferation, migration and invasion, suggesting its tumor suppressive potential in gastric cancer progression.

Epigenetic regulation of miR-196b expression in gastric cancer

MicroRNAs (miRNAs) are short noncoding RNAs that play important roles in cellular processes and disease pathogenesis via the control of specific targeted gene expression. The miR‐196s miRNA is encoded at three paralogous loci in three HOX clusters and acts as an oncogenic miRNA in cancer progression. Recent studies have demonstrated that the expression of miR‐196b increases cell proliferation and survival in leukemic cells. Here, we used a sequential methylation analysis to reveal that the methylation status correlated well with miR‐196b expression in different cell lines. Treatment with the demethylating drug 5‐Aza‐dC reactivated miR‐196b transcription in methylation‐silenced cells. Using in vitro methylation approach, we further provide evidences that promoter hypermethylation represses miR‐196b transcriptional activation tightly in human cancer cell lines. We also demonstrate that the expression of miR‐196b is significantly elevated in gastric cancer and that hypomethylation status of miR‐196b CpG islands frequently is observed in primary gastric tumors. Our results provide important information on miR‐196s regulation and demonstrate that abnormal DNA hypomethylation induces overexpression of miR‐196b in gastric cancer.

Aberrant expression of miR-196a in gastric cancers and correlation with recurrence

MicroRNAs (miRNAs) are short noncoding RNAs (˜22 nt) that play important roles in the pathogenesis of human diseases by negatively regulating gene expression. Here, we examined the relationship between miR‐196a and gastric cancer. By the analysis of 72 gastric cancer samples, we found that the expression level of miR‐196a microRNA significantly increased in primary gastric cancer tissues versus adjacent normal tissues. In addition, extracellular miR‐196a detected in conditioned medium was strongly correlated with its cellular expression status and increased circulating miR‐196a in patient serum was associated with gastric cancer disease status and relapse. Furthermore, ectopic expression of miR‐196a microRNA promoted the epithelial‐mesenchymal transition and migration/invasion capabilities of transfected cells, suggesting its oncogenic potential in gastric cancer progression. Altogether, our data demonstrate that miR‐196a exerts an oncogenic role in gastric cancer and miR‐196a may be a novel biomarker for detecting gastric cancer and for monitoring disease recurrence.

Functional and morphological changes in pancreatic remnant after pancreaticoduodenectomy.

Objectives: Pancreatic exocrine insufficiency has been reported to be more common in pancreaticogastrostomy (PG) than in pancreaticojejunostomy (PJ) after pancreaticoduodenectomy (PD). This study aimed to evaluate the long-term outcome after PD between these 2 groups. Methods: We evaluated the long-term functional status of 42 surviving patients diagnosed with periampullary lesions who underwent PJ or PG after PD and followed up for more than 1 year. Among these, 23 patients underwent PJ and 19 patients underwent PG. To compare the 2 groups, we analyzed the (1) pancreatic exocrine insufficiency by questioning the presence or absence of steatorrhea, (2) pancreatic endocrine function by measuring glycohemoglobin A1c, fasting blood glucose, and history of new-onset diabetes, (3) nutritional status by measuring serum total protein, albumin, cholesterol, and triglyceride, (4) gastric emptying time, (5) panendoscopic findings, (6) changes of pancreatic duct diameter by computed tomography, and (7) relaparotomy rate. Results: The mean follow-up time for PG and PJ were 37 ± 23 and 103 ± 52 months, respectively (P < 0.05). A total of 52.4% patients developed pancreatic exocrine insufficiency, and 11.9% had new-onset diabetes. There was no significant difference between PJ and PG groups. A significantly improved postoperative nutritional state regarding serum total protein and albumin were noticed in both groups. There was no significant difference in terms of gastric emptying time, positive panendoscopic findings, and changes in pancreatic duct diameter. The pancreatic remnant-related relaparotomy rate was higher in the PJ group as compared with the PG group (17.4% vs 0%; P = 0.056). Conclusions: There is no significant difference in pancreatic exocrine or endocrine insufficiency, gastric emptying time, and positive panendoscopic findings between PJ and PG. Pancreaticojejunostomy was associated with a higher pancreatic remnant-related relaparotomy rate; however, because of a shorter follow-up in the PG group, a continuous long-term follow-up is still needed.

Transcriptional regulation of miR-196b by ETS2 in gastric cancer cells

E26 transformation-specific sequence (ETS)-2 is a transcriptional modulator located on chromosome 21, alterations in its expression have been implicated with a reduced incidence of solid tumors in Down syndrome patients. MicroRNAs (miRNAs) are thought to participate in diverse biological functions; however, the regulation of miRNAs is not well characterized. Recently, we reported that miR-196b is highly expressed in gastric cancers. Herein, we demonstrate that miR-196b expression was significantly repressed by ETS2 during gastric cancer oncogenesis. We demonstrate that knockdown of endogenous ETS2 expression increases miR-196b expression. A genomic region between −751 and −824 bp upstream of the miR-196b transcriptional start site was found to be critical for the repression activity. This putative regulatory promoter region contains three potential ETS2-binding motifs. Mutations within the ETS2 binding sites blocked the repression activity of ETS2. Furthermore, knockdown of ETS2 or overexpression of miR-196b significantly induced migration and invasion in gastric cancer cells. In addition, alterations in ETS2 and miR-196b expression in gastric cancer cell lines affected the expression of epithelial–mesenchymal transition-related genes. The levels of vimentin, matrix metalloproteinase (MMP)-2 and MMP9 were drastically induced, but levels of E-cadherin were decreased in shETS2- or miR-196b-transfected cells. Our data indicate that ETS2 plays a key role in controlling the expression of miR-196b, and miR-196b may mediate the tumor suppressor effects of ETS2. We demonstrated that miR-196b was transcriptionally regulated by ETS2 and there was an inverse expression profile between miR-196b and ETS2 in clinical samples. This finding could be beneficial for the development of effective cancer diagnostic and alternative therapeutic strategies.

Comparison Between Pancreaticojejunostomy and Pancreaticogastrostomy After Pancreaticoduodenectomy

BACKGROUND/PURPOSE Pancreatic leakage is a leading cause of morbidity and mortality after pancreaticoduodenectomy (PD). Pancreaticogastrostomy (PG) has been reported to be associated with a lower pancreatic leakage rate and morbidity rate than pancreaticojejunostomy (PJ). This study compared the preoperative characteristics, surgical risk factors, intraoperative parameters, and postoperative outcome between PJ and PG. METHODS From March 1992 to March 2005, a comparative study between PJ and PG for patients with periampullary lesions undergoing PD was conducted. A total of 377 consecutive patients underwent PD. Among them, 188 patients underwent PJ and 189 underwent PG. RESULTS The overall mortality, morbidity and pancreatic leakage following PD were 5%, 45.1% and 10.6%, respectively. The mortality, morbidity and pancreatic leakage were 8.9%, 56.4% and 17.6% in the PJ group, and 2.1%, 33.9% and 3.7% in the PG group (p < 0.001). Mean operative time was 9.3 hours versus 6.7 hours (p < 0.001), mean blood loss was 1032 mL versus 891 mL (p = 0.064) and mean hospital stay was 34.8 days versus 26.1 days (p < 0.001) in the PJ and PG groups, respectively. PJ, soft pancreas, pancreatic duct stenting, low surgical volume (< 20) and age (> 65 years) were identified as risk factors for pancreatic leakage, while PJ, soft pancreas, pancreatic duct stenting and low surgical volume (< 20) were four significant risk factors for surgical morbidity. Further, PJ, pancreatic leakage, low surgical volume (< 20) and age (> 65 years) were identified to be surgical risk factors for mortality. CONCLUSION PG is a safer method than PJ following PD as a significantly lower rate of pancreatic leakage, surgical morbidity and mortality, shorter operation time, and shorter postoperative hospital stay are reported.

Comparison of the Operative Outcomes and Learning Curves between Laparoscopic and Robotic Gastrectomy for Gastric Cancer

Background Minimally invasive surgery, including laparoscopic and robotic gastrectomy, has become more popular in the treatment of gastric cancer. However, few studies have compared the learning curves between laparoscopic and robotic gastrectomy for gastric cancer. Methods Data were prospectively collected between July 2008 and Aug 2014. A total of 145 patients underwent minimally invasive gastrectomy for gastric cancer by a single surgeon, including 73 laparoscopic and 72 robotic gastrectomies. The clinicopathologic characteristics, operative outcomes and learning curves were compared between the two groups. Results Compared with the laparoscopic group, the robotic group was associated with less blood loss and longer operative time. After the surgeon learning curves were overcome for each technique, the operative outcomes became similar between the two groups except longer operative time in the robotic group. After accumulating more cases of robotic gastrectomy, the operative time in the laparoscopic group decreased dramatically. Conclusions After overcoming the learning curves, the operative outcomes became similar between laparoscopic and robotic gastrectomy. The experience of robotic gastrectomy could affect the learning process of laparoscopic gastrectomy.

The management of high‐risk patients with primary hyperparathyroidism – minimally invasive parathyroidectomy vs. medical treatment

Objective  Parathyroidectomy (PTx) for high‐risk primary hyperparathyroidism (PHPT) patients poses a surgical challenge. We hypothesize that a minimally invasive parathyroidectomy (MIP) under local anaesthesia may minimize the perioperative risks and facilitate easier clinical care than medical treatment for these patients.

Mutations in PI3K/AKT pathway genes and amplifications of PIK3CA are associated with patterns of recurrence in gastric cancers

Mutations in genes involved in the PI3K/AKT pathway and amplifications of the PIK3CA gene in gastric cancer and their associations with clinicopathological characteristics and EBV infection were analyzed in this study. A total of 431 patients with gastric adenocarcinomas were enrolled, and 39 mutation hotspots were evaluated in these patients using MALDI-TOF mass spectrometry were analyzed. PIK3CA amplifications were analyzed using real-time quantitative PCR. Regarding patients with intestinal-type gastric cancer, those with mutations in PI3K/AKT pathway genes were also more likely to have tumors located in the lower-third of the stomach than were those without mutations. Regarding patients with diffuse-type gastric cancer, those with PI3K/AKT pathway mutations were more likely to have tumors located in the upper-third of the stomach and to have more hematogenous metastases, particularly in the liver and lungs, than were patients without such mutations (22.2% vs. 4.5%). No significant survival difference was observed between patients with vs. without PI3K/AKT pathway mutations. Mutations in PI3K/AKT pathway genes were associated with hematogenous metastasis in patients with diffuse-type gastric cancer. Only when the tumors were located in the middle-third of stomach, tumor with mutations of the PIK3CA gene or mutations of the PI3K/AKT pathway genes were associated with more EBV infection than those without mutations. Patients with PIK3CA amplifications were more likely to have diffuse-type and poorly differentiated gastric cancers and were more likely to experience peritoneal recurrence compared with those without PIK3CA amplifications. Even upon subgroup analysis, PI3KCA amplifications were found to not affect the patients’ outcomes.