Daisuke Ichikawa

Detection of gastric cancer-associated microRNAs on microRNA microarray comparing pre- and post-operative plasma.

Background:Recently, it was reported that plasma microRNAs (miRNAs) are low-invasive useful biomarkers for cancer. We attempted to isolate gastric cancer (GC)-associated miRNAs comparing pre- and post-operative paired plasma, thereby excluding the possible effects of individual variability.Methods:This study was divided into four steps: (1) microarray analysis comparing pre- and post-operative plasma; (2) validation of candidate miRNAs by quantitative RT–PCR; (3) validation study of selected miRNAs using paired plasma; and (4) comparison of the levels of selected miRNAs in plasma between healthy controls and patients.Results:From the results of microarray analysis, nine candidate miRNAs the levels of which were markedly decreased in post-operative plasma were selected for further studies. After confirmation of their post-operative marked reduction, two candidate miRNAs, miR-451 and miR-486, were selected as plasma biomarkers, considering the abundance in plasma, and marked decrease in post-operative samples. In validation, the two miRNAs were found to decrease in post-operative plasma in 90 and 93% of patients (both P<0.01). In comparison with healthy controls, the levels of both miRNAs were found to be significantly higher in patients, and the area under the curve values were high at 0.96 and 0.92.Conclusion:Plasma miR-451 and miR-486 could be useful blood-based biomarkers for screening GC.

Monoclonal antibody A7-superparamagnetic iron oxide as contrast agent of MR imaging of rectal carcinoma

Superparamagnetic iron oxide (SPIO)-based colloid has been used clinically as a tissue-specific magnetic resonance contrast agent. We coupled monoclonal antibody A7 (Mab A7), which reacts specifically with human colorectal carcinoma, to Ferumoxides (SPIO) and examined the accumulation of this conjugate in xenografted tumours in nude mice. We examined in vitro immunoreactivity of Mab A7 coupled to Ferumoxides and its in vivo distribution in nude mice with human colorectal carcinoma. Magnetic resonance imaging of tumour-bearing nude mice was performed 72 h after injection of A7-Ferumoxides. A7-Ferumoxides retained binding activities that were nearly identical to intact Mab A7. More of the radiolabelled A7-Ferumoxides accumulated in the tumour than normal mouse IgG-Ferumoxides from 12 h onwards after injection (P<0.05). Both A7-Ferumoxides and normal mouse IgG-Ferumoxides disappeared from blood linearly over time. The accumulation levels in normal tissue decreased linearly over time but were lower than levels in tumours from 6 h. In magnetic resonance T2-weighted imaging of the tumour-bearing nude mice, signal intensity was reduced at the margin of the tumour by injection of A7-Ferumoxides. Mab A7 coupled to Ferumoxides is potentially suitable as a magnetic resonance contrast agent for detecting local recurrence of rectal carcinoma.

Analysis of histological therapeutic effect, apoptosis rate and p53 status after combined treatment with radiation, hyperthermia and 5-fluorouracil suppositories for advanced rectal cancers

The tumour-suppressor gene p53 encodes a transcription factor that plays a critical role in the induction of G1 cell cycle arrest and apoptosis after DNA damage. To clarify the role of the p53 gene and apoptosis in combined hyperthermia, chemotherapy and radiation (hyperthermochemoradiotherapy, HCR therapy) for rectal cancer, we examined the histological response, rate of apoptosis, DNA fragmentation and p53 status in tumours from 28 patients undergoing HCR therapy before surgery and from 22 patients who did not have preoperative treatment. The therapeutic effect of HCR therapy was closely correlated with the rate of apoptosis; the correlation was statistically significant, suggesting that this effect occurs through apoptosis. The incidence of p53 mutations in the treated group were as follows: in tumours resistant to HCR therapy, four of seven (57.1%); intermediately sensitive, 7 of 13 (53.9%); or sensitive, three of eight (37.5%), suggesting that the therapeutic effect and apoptosis rate were related to the p53 status of the tumours to some extent, but the relation was not statistically significant. In the 22 control tumours (non-treated group), the apoptosis rate was 2.0 +/- 1.1%, and there was no significant difference in p53 status compared with the HCR group. Our study indicates that the pathological response to HCR therapy correlates with the rate of apoptosis with statistical significance and that it induces the therapeutic effect more significantly in rectal cancer cells with wild-type p53, although HCR therapy-induced apoptosis also occurs in some rectal cancers with mutated p53. Therefore, this combination therapy can induce an additive or synergistic anti-tumour effect in rectal cancers with wild-type p53 as well as in those with mutated p53 through apoptosis, offering new therapeutic opportunities and a better prognosis.

miR-509-5p and miR-1243 increase the sensitivity to gemcitabine by inhibiting epithelial-mesenchymal transition in pancreatic cancer

The epithelial-mesenchymal transition (EMT) contributes to various processes in cancer progression, such as metastasis and drug resistance. Since we have already established a cell-based reporter system for identifying EMT-suppressive microRNAs (miRNAs) in the pancreatic cancer cell line Panc1, we performed a function-based screening assay by combining this reporter system and a miRNA library composed of 1,090 miRNAs. As a result, we identified miR-509-5p and miR-1243 as EMT-suppressive miRNAs, although the mechanisms for EMT-suppression induced by these miRNAs have yet to be clarified. Herein, we demonstrated that overexpression of miR-509-5p and miR-1243 increased the expression of E-cadherin through the suppression of EMT-related gene expression and that drug sensitivity increased with a combination of each of these miRNAs and gemcitabine. Moreover, miR-509-5p was associated with worse overall survival in patients with pancreatic cancer and was identified as an independently selected predictor of mortality. Our findings suggest that miR-509-5p and miR-1243 might be novel chemotherapeutic targets and serve as biomarkers in pancreatic cancer.

Hepatic ductoplasty for iatrogenic Bismuth type 2 bile duct stricture: A case report

Highlights • The article represents recovering treatment of iatrogenic biliary tract injury by laparoscopic cholecystectomy.• Biliary tract stricture like Bismuth type 2 successfully treated by hepatic ductoplasty.• To emphasize the importance of avoiding biliary stricture is a key to prevent cholangitis and stone recurrences.

Successful laparoscopic partial gastrectomy and spleen-preserving distal pancreatectomy for gastric duplication cyst connecting with the pancreatic tail

Highlights • Gastric duplication cyst(GDC) contiguous with the stomach and pancreatic tail is extremely rare.• GDCs are usually diagnosed at a younger age. In adults, they are very rare disease and the diagnosis may be difficult.• Surgical resection is considered to be the best treatment due to the difficulty of diagnosis.• Laparoscopic surgery is less invasive and should be selected whenever possible.

Current status of laparoscopic total gastrectomy

In this article, the current state of laparoscopic total gastrectomy (LTG) was reviewed, focusing on lymph node dissection and reconstruction. Lymph node dissection in LTG is technically similar to that in laparoscopic distal gastrectomy for early gastric cancer; however, LTG for advanced gastric cancer requires extended lymph node dissections including splenic hilar lymph nodes. Although a recent randomized controlled trial clearly indicated no survival benefit in prophylactic splenectomy for lymph node dissection at the splenic hilum, some patients may receive prognostic benefit from adequate splenic hilar lymph node dissection. Considering reconstruction, there are two major esophagojejunostomy (EJS) techniques, using a circular stapler (CS) or using a linear stapler (LS). A few studies have shown that the LS method has fewer complications; however, almost all studies have reported that morbidity (such as anastomotic leakage and stricture) is not significantly different for the two methods. As for CS, we grouped various studies addressing complications in LTG into categories according to the insertion procedure of the anvil and the insertion site in the abdominal wall for the CS. We compared the rate of complications, particularly for leakage and stricture. The rate of anastomotic leakage and stricture was the lowest when inserting the CS from the upper left abdomen and was significantly the highest when inserting the CS from the midline umbilical. Scrupulous attention to EJS techniques is required by surgeons with a clear understanding of the advantages and disadvantages of each anastomotic device and approach.

Investigation of a Quality Check for Plasma Samples

Background: MicroRNA (miRNA) molecules have been detected in many body fluids and used as biomarkers. However, blood cell-derived miRNA molecules due to hemolysis have been shown to affect the amounts of plasma miRNAs. It is important to check the quality of plasma samples for clinical applications. In the present study, an objective method for quality checking plasma samples was investigated using permitted color tone level and the absorbance at 414 nm. Material and method: An ROC analysis of the macroscopic color tone and the absorbance in 1213 clinical samples was performed. The optimal cut-off absorbance value was validated using the amount of plasma miRNA. Results: AUC for detecting hemolyzed samples was very high (0.986) at a cut-off absorbance value of 1.664. A sensitivity and specificity were 99% and 92%, respectively. For validation study, 3 candidate miRNAs were selected by a miRNA microarray between fresh and hemolyzed plasma samples; the amounts of miR-16 and miR-19b increased in hemolyzed samples, whereas that of miR-223 remained unchanged. The amounts of plasma miR-16 and miR-19b were significantly increased in 5 clinical samples with higher absorbance values (p=0.0001 and p=0.0003, respectively), while the amounts of these miRNAs were not increased in samples with lower absorbance values. Conclusions: A simple method for quality checking plasma samples using color tone and absorbance is very useful and significant.