Aberrant gene methylation in the peritoneal fluid is a risk factor predicting peritoneal recurrence in gastric cancer

AIM:To investigate whether gene methylation in the peritoneal fluid (PF) predicts peritoneal recurrence in gastric cancer patients.METHODS: The gene methylation of CHFR (checkpoint with forkhead and ring finger domains), p16, RUNX3 (runt-related transcription factor 3), E-cadherin, hMLH1 (mutL homolog 1), ABCG2 (ATP-binding cassette, sub-family G, member 2) and BNIP3 (BCL2/adenovirus E1B 19 kDa interacting protein 3) were analyzed in 80 specimens of PF by quantitative methylation-specific polymerase chain reaction (PCR). Eighty patients were divided into 3 groups; Group A (n=35):the depth of cancer invasion was less than the muscularis propria; Group B (n=31): the depth of cancer invasion was beyond the muscularis propria. Both group A and B were diagnosed as no cancer cells in peritoneal cytology and histology; Group C (n=14): disseminated nodule was histologically diagnosed or cancer cells were cytologically defi ned in the peritoneal cavity.RESULTS: The positive rates of methylation in CHFR, E-cadherin and BNIP3 were significantly different among the 3 groups and increased in order of group A, B and C (0%,0% and 21% in CHFR, P<0.05; 20%, 45% and 50% in E-cadherin, P<0.05;26%,35% and 71% in BNIP3, P<0.05). In addition, the multigene methylation rate among CHFR, E-cadherin and BNIP3 was correlated with group A, B and C (9%,19% and 57%, P<0.001). Moreover, the prognosis was analyzed in group B, excluding 3 patients who underwent a non-curative resection. Two of the 5 patients with multigene methylation showed peritoneal recurrence after surgery, while those without or with a single gene methylation did not experience recurrence (P<0.05).CONCLUSION: This study suggested that gene methylation in the PF could detect occult neoplastic cells in the peritoneum and might be a risk factor for peritoneal metastasis.

Elevated Vascular Endothelia Growth Factor-A in the Serum and Peritoneal Fluid of Patients with Endometriosis

There has been emergence of evidence suggesting that specific variants of the vascular endothelia growth factor （VEGF） family, based on their ability to regulate angiogenesis, would be pivotal in the pathogenesis of endometriosis. This study was aimed at determining whether high levels of VEGF-A could be found in the serum and peritoneal fluid （PF） of patients with endometriosis. VEGF-A levels were measured by enzyme-linked immunosorbent assay （ELISA） in serum and PF from 46 patients with surgically confirmed endometriosis, and 40 controls with no clinical evidence of the disease or detectable endometriotic lesions at the time of surgical examination. The results showed the mean VEGF-A levels were significantly higher in the serum and PF of patients with endometriosis than in the controls. The VEGF-A levels in the serum and PF of patients with severe endometriosis （stages Ⅲ-Ⅳ） were significantly higher than in those with minimal endometriosis （P〈0.001）. It was concluded that endometriosis was associated with significant modulation in the levels of circulating VEGF-A.

Levels of Lipid Perioxides and Superoxide Dismutase in Peritoneal Fluid of Patients with Endometriosis

In order to evaluate the changes of lipid perioxides and superoxide dismutase levels in peritoneal fluid in patients with endometriosis, the levels of LPO and SOD in peritoneal fluids from infertile women with endometriosis and with normal pelvis were measured. The result showed that the levels of LPO but SOD was elevated significantly in peritoneal fluid from women with endometriosis than in women with normal pelvis . It is suggested that free oxygen radicals may be involved in the pathogenesis of endometriosis associated infertility.

DETECTION OF VEGF LEVELS IN ASCITES AND PERITONEAL FLUID

Objective: The expression of vascular endothelial growth factor (VEGF) is correlated to the invasion and metastasis of tumor cells in many clinical carcinomas. In this study, we detected soluble VEGF levels in ascites and peritoneal fluid and explored its clinical significance. Methods: A total of 91 samples were collected and divided into 5 experimental groups: petitoneal fluid of patients with benign (n=10) and malignant disease (n=14), cirrhotic ascites (n=36), tuberculous ascites (n=8) and malignant ascites (n=23). Using a sandwich enzyme-linked immunoadsorbent assay, the concentration of soluble VEGF was measured in ascites (n=67) and petitoneal fluid (n=24). Results: VEGF levels in malignant ascites were 640.74264.81 pg/ml, significantly higher than those in cirrhotic ascites, tuberculous ascites and peritoneal fluid of patients with benign and malignant disease (P<0.01, separately). However, the difference of VEGF levels among the latter 4 groups had no statistic significance (P>0.05), separately). Furthermore, VEGF levels in malignant ascites from patients with ovarian cancer were higher than those with gastric and colon cancer (P<0.01, respectively), while there was no significant difference between gastric cancer and colon cancer (P>0.05). Using VEGF level of 118.96 pg/ml as a minimum cutofflimit, the sensitivity and specificity of VEGF of this assay to diagnose malignant ascites were 91.3% and 73.9% respectively. Conclusion: The elevated levels of VEGF may be useful as an index in differential diagnosis of benign and malignant ascites. It appears that VEGF plays an important role in malignant ascites formation.

Mechanism of Elevated Vascular Endothelial Growth Factor Levels in Peritoneal Fluids from Patients with Endometriosis

In order to investigate the mechanism of elevated vascular endothelial growth factor (VEGF) in peritoneal fluids from patients with endometriosis, macrophages were recovered from peritoneal fluids obtained at the time of diagnostic laparoscopy from infertile women with endometriosis (EMT group, n=20) and without endometriosis (control group, n=20). Macrophages were cultured in vitro. The VEGF levels of peritoneal fluid and the supernatant of macrophages culture were determined by enzyme linked immunoassay (ELISA). Meanwhile, the eutopic (n=20) and ectopic endometrium (n=20) from endometriosis patients, and normal edometrium (n=20) from non-endometriosis patients were obtained for the analysis of VEGF expression by labeled Streptavidin Biotin (LSAB). It was found that VEGF levels in peritoneal fluid and macrophages culture supernatant were significantly higher in EMT group than in control group (P<0.01). In normal endometrium, VEGF showed a cyclic changes and similar in eutopic and ectopic endometrium from patients with endometriosis. There was no difference in the intensity of VEGF in endometrium between two groups within each menstrual phase. It is suggested that altered VEGF production by peritoneal macrophages and ectopic endometrium secretion may contribute to the elevated VEGF levels in the peritoneal fluid of patients with endometriosis.

Effect of Monocyte Chemotactic Protein-1 on the Intraperitoneal Adhesion Formation

In order to study the role of monocyte chemotactic protein-1 (MCP-1) in the intra-peri- toneal adhesion formation, 23 infertile patients undergoing laparoscopic operation were divided into two groups: experimental group including 12 patients with intra-peritoneal adhesion and control group including 11 patients without intra-peritoneal adhesion. Peritoneal fluid (PF) and peritoneum were collected from these patients during laparoscopic examination. The expression levels of MCP-1 protein and MCP-1 mRNA were detected by using enzyme-linked immunosorbent assay (ELISA) and dot blot analysis method respectively. It was found that the levels of MCP-l protein in PF of the patients with peritoneal adhesion were significantly higher than in the control group (0.44±0. 11 ng/ ml vs 0. 19±0.09 ng/ml respectively, P<0. 01). The level of MCP-l mRNA in the peritoneum of the patients with peritoneal adhesion was significantly higher than in the control group (48. 61±3. 72 vs 19.87±2.54 respectively, P<0. 01). It was suggested that MCP-1 might play a role in the adhe- sion formation, and chemotactic cytokines expressing in the peritoneal mesothelial cells might be take part in the process.

Peritoneal Membrane Transport Characteristics in Uni-PET with Preceding Icodextrin Dwell as Compared to Classic PET with Preceding Glucose Dwell: A Prospective Study

Background: In subjects on CAPD who use icodextrin for long night dwell, it has been recommended that nocturnal exchange be replaced by dextrose dwell whenever PET is to be performed as preceding exchange with icodextrin temporarily increases peritoneal membrane permeability and therefore, gives high D/PCr and low D/D0 glucose values. Whether this temporary change is also seen with use of Uni-PET (which involves one hour dwell of 1.5% dextrose followed by 4-hour dwell of 4.25% dextrose) is not known. Methods: In this self, controlled study, subjects on CAPD, who were using icodextrin for long nocturnal dwell for at least 3 months, were screened for enrolment. Pregnancy or lactation, history of any PD related infectious complication in the past one month, present or past malignancy, and poor functional status were exclusion criteria. Enrolled subjects underwent classic PET with preceding 2.5% dextrose long nocturnal dwell on day 1 followed by Uni-PET with preceding 7.5% icodextrin long nocturnal dwell on day 2. Difference in D/PCr and D/D0 glucose between the two PETs was primary objectives. The study was approved by Institute Ethics Committee. Results: 15 out of 26 screened subjects were enrolled over a period of 18 months (July 2015-December 2016). The mean (±SD) age of study population was 60.8±9.1 years. Majority were males and diabetes was the most common cause of CKD. Mean D/PCr were 0.68 ± 0.11 and 0.64 ± 0.08 in classic PET and Uni-PET, respectively. The difference between the two values was not significant [mean difference between D/PCr (classic PET-Uni-PET): 0.040 ± 0.86;95% CI (-0.007 to 0.088);p = 0.09]. Similarly, D/D0 glucose between classic PET and Uni-PET was also similar [mean difference between D/D0 glucose (classic PET-Uni-PET): -0.02 ± 0.09;95% CI (-0.06 to 0.03);p = 0.448]. Conclusion: Peritoneal membrane small solute transport characteristics in Uni-PET with preceding icodextrin dwell are similar to classic PET with preceding glucose dwell. If Uni-PET is used, it may not be necessary to replace preceding nocturnal exchange of icodextrin with that of dextrose as is currently advised.

Role of RECK methylation in gastric cancer and its clinical significance

AIM:To investigate the relation between RECK methylation and clinicopathological characteristics of gastric cancer patients and evaluate the role of RECK methylation in peritoneal metastasis of gastric cancer.METHODS:Methylation of RECK gene in 40 paired samples of gastric cancer and its corresponding adjacent normal mucosa,lymph nodes and peritoneal irrigation fluid was detected by methylation-specific polymerase chain reaction.RESULTS:Aberrant methylation of RECK gene was detected in 27.5%(11/40)of the adjacent normal mucosa samples,in 47.5%(19/40)of gastric cancer samples,in 57.1%(12/21)of the lymph node samples,and in 35%(14/40)of peritoneal irrigation fluid samples,respectively,with a significant difference between the adjacent normal mucosa and lymph node samples(P=0.023).Presence of RECK methylation in the primary tumor samples was significantly correlated with tumor invasion(P=0.023).The accuracy of RECK methylation in peritoneal lavage fluid samples for the diagnosis of peritoneal metastasis of gastric cancer was 72.5%(26/40),with a sensitivity of 66.7%(6/9) and a specificity of 74.2%(23/31).CONCLUSION:Aberrant methylation of RECK gene may provide useful information for the early diagnosis and treatment of peritoneal metastasis of gastric cancer.

Promoter Hypermethylation of KiSS-1 Gene in Gastric Cancer

Objective： To investigate the association between KiSS-1 methylation and clinicopathological characteristics of gastric cancer and evaluate the role of peritoneal lavage fluid in detecting peritoneal metastases. Methods： The methylation status of KiSS-1 gene in 40 gastric cancer specimens, the corresponding adjacent normal mucosa, lymph nodes and peritoneal lavage fluid was investigated by methylation-spcific polymerase chain reaction（MS-PCR）. Results： Aberrant methylation of KiSS-1 gene was detected in 55%（22/40） of the adjacent normal mucosa, 82.5% （33/40）of gastric cancer specimens, 80.95%（17/21） of the lymph nodes, and 42.5%（17/40） of peritoneal lavage fluid. Methylation in gastric carcinoma and lymphonode was more frequent than in non-neoplastic gastiric mucosa. Presence of KiSS-1 methylation in peritoneal lavage fluid was significantly correlated with tumor invasion （P=0.043）. The accuracy of KiSS-1 methylation in peritoneal lavage fluid for diagnosing peritoneal metastasis was 70%, with a sensitivity of 77.8% and a specificity of 67.7%. Conclusion： Aberrant methylation of KiSS-1 gene is a common event in the occurrence and progression of gastric carcinoma, which may provide useful information for the early diagnosis of peritoneal metastases and a new therapy for gastric cancer.