Hiroshi Kawahira

The Role of mel-18, a Mammalian Polycomb Group Gene, during IL-7–Dependent Proliferation of Lymphocyte Precursors

mel-18 is a mammalian homolog of Drosophila melanogaster Polycomb group genes. Mice lacking the mel-18 gene show a posterior transformation of the axial skeleton, severe combined immunodeficiency, and a food-passing disturbance in the lower intestine due to hypertrophy of the smooth muscle layer. In this study, the severe combined immunodeficiency observed in mel-18 mutant mice is correlated with the impaired mitotic response of lymphocyte precursors upon interleukin-7 stimulation. Strikingly, the axial skeleton and lymphoid phenotypes are identical in both mel-18 and bmi-1 mutants, indicating that the Mel-18 and Bmi-1 gene products might act in the same genetic cascade. These results suggest that mammalian Polycomb group gene products are involved in cell cycle progression in the immune system.

Preparation and characterization of phospholipid-conjugated indocyanine green as a near-infrared probe

We have rationally designed and synthesized a novel near-infrared (NIR) photoactivating probe, designated by iDOPE, in which an indocyanine green (ICG) fluorophore is covalently conjugated with a phospholipid moiety, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), to incorporate into liposome bilayers. NIR irradiation showed that iDOPE retained the optical and fluorescence properties of ICG and demonstrated photoactivator characteristics: fluorescence emission at around 820 nm in a solvent, singlet oxygen production, and concentration-dependent heat generation. Additionally, iDOPE was incorporated into liposome bilayers and maintained stable liposomally formulated iDOPE (LP-iDOPE) over 1week under physiological conditions. We also observed the tumor-specific biodistribution of LP-iDOPE of in vivo xenografts. These findings suggest that LP-iDOPE might be a promising tool for NIR optical imaging, photodynamic therapy, and photothermal therapy.

Water-filled laparoendoscopic surgery (WAFLES): feasibility study in porcine model.

PURPOSE Recent surgical techniques have been advancing under endoscopic view and insufflation of carbon dioxide gas to expand the abdominal cavity. Isotonic fluid could be one candidate for expanding cavities to facilitate surgical maneuvering. We tested the feasibility and drawbacks of replacement of irrigating materials using a porcine model (water-filled laparoendoscopic surgery [WAFLES]). MATERIALS AND METHODS Laparoscopic cholecystectomy was performed in two porcine models using instillation of sorbitol solution as irrigant. Solution irrigation was performed through one of four ports, with drainage via another port. Conventional forceps equipped with a monopolar electrode for electrocautery, laparoscope, video processor, ultrasound, and transducer for measuring intraabdominal pressure were used. RESULTS Laparoscopic cholecystectomy was successfully undertaken with the following benefits: (1) clear observation of the dissecting plane throughout maneuvering; (2) control of oozing and spilled bile by irrigation and suction; and (3) ultrasonographic and laparoscopic images can be obtained simultaneously without any restriction to probe location. However, two disadvantages should be noted: (1) difficulties in managing floating organs and (2) interruption of vision by blood. CONCLUSIONS WAFLES provides some benefits for endoscopic surgery with proper devices, including apparatuses for irrigation and suction. Efficient irrigation and selection of proper irrigant and apparatuses are required to establish an acceptable procedure.

Surgical advantages of gastric SMTs by laparoscopy and endoscopy cooperative surgery.

BACKGROUND/AIMS The treatment of gastric submucosal tumors (SMTs) is strictly surgical and enucleation of the tumor or wedge resection of the stomach is efficient to achieve R0 resection. Laparoscopic and endoscopic cooperative surgery (LECS) can be safely performed with adequate cutting lines. This study describes the initial 16 cases treated by LECS and evaluates the advantages by LECS for gastric SMTs retrospectively. METHODOLOGY Sixteen patients with gastric SMT underwent LECS from June 2007 to December 2010, their surgical data, clinical characteristics and surgical specimens of SMTs were compared. The surgical specimens of 9 gastric SMTs treated by laparoscopic wedge resection (LWR) were compared as a control. RESULTS The median (range) length of operation time, blood loss, hospital stay after surgery were minutes 172 (115- 220), <5mL (<5-115) and 10 days (6-17), respectively. The median (range) ratio of the longest diameter of the tumor divided by the longest diameter of the surgical specimen in LECS and LWR were 0.86 (0.625-1.0) and 0.69 (0.44-1.0), respectively (p=0.0189, Wilcoxon rank sum test). CONCLUSIONS LECS minimizes the surgical specimen while still providing sufficient surgical margins to successfully cure gastric SMTs.

Biologically Different Subgroups of Invasive Ductal Carcinoma of the Pancreas Dpc4 Status According to the Ratio of Intraductal Carcinoma Components

Purpose: Among invasive ductal carcinomas of the pancreas (IDCP), there is a morphologically characteristic subgroup accompanied by abundant intraductal carcinoma components (ICCs). With the aim of determining whether ICC-rich IDCP are biologically different from ICC-poor IDCP, the expression status of Dpc4 protein was analyzed. Experimental Design: A total of 43 IDCP was subdivided into two groups: (a) ICC-rich IDCP (ICCs area occupies ≥10% of the entire tumorous area); and (b) ICC-poor IDCP (with <10% of ICCs area). A total of 10 invasive carcinomas derived from intraductal papillary-mucinous neoplasms (ICs from IPMNs) were also analyzed. Each invasive and intraductal carcinoma area was then evaluated for Dpc4 protein status by immunohistochemistry. Results: In a total of 43 IDCP, there were 23 ICC-rich IDCP and 20 ICC-poor IDCP. Dpc4-positive immunostaining was observed in the invasive carcinoma component of ICC-rich IDCP, ICC-poor IDCP, and ICs from IPMN in 18 of 23 (78%), 4 of 20 (20%), and 7 of 10 (70%) cases, respectively. In the intraductal component, positive staining for Dpc4 was found in 20 of 23 (87%), 3 of 7 (41%), and 8 of 10 (80%) cases, respectively. Dpc4 expression was found in both the invasive and ICC components of ICC-rich IDCP, similar to that found in IC derived from IPMN, whereas the expression of Dpc4 was largely diminished in ICC-poor IDCP. Conclusions: Morphologically distinct subgroups of invasive ductal carcinomas of the pancreas, namely ICC-rich IDCP and ICC-poor IDCP, are also biologically distinguishable as revealed by the differential expression of Dpc4.

Salivary amylase activity is useful for assessing perioperative stress in response to pain in patients undergoing endoscopic submucosal dissection of gastric tumors under deep sedation

BackgroundAlthough endoscopic submucosal dissection (ESD) for patients with gastric tumors under the conditions of unconsciousness is considered to be minimally invasive, no objective assessment of the perioperative stress of ESD has yet been conducted. Today, stress levels can be easily and objectively assessed by monitoring salivary amylase activity (sAMY). We evaluated the perioperative changes in the sAMY in patients undergoing ESD and identified the causes of such changes.MethodsA total of 40 patients with gastric cancers/adenomas removed by ESD under general anesthesia (GA; n = 20) and under deep sedation (DS; n = 20) were enrolled. sAMY was measured using the enzyme analysis equipment, sAMY Monitor (NIPRO, Osaka, Japan) during the perioperative period of the ESD. Also, all patients were interviewed to determine their subjective stress level, using a questionnaire asking “How did you feel during ESD?”, with the choice of responses ranging from “did not wake up at all” to “I was awake and ESD was extremely stressful”.ResultsThe sAMY of the DS group increased soon after the start of ESD. Meanwhile, that of the GA group decreased just after the ESD started and was maintained at a stable level throughout the ESD. In response to the stress level questionnaire, all of the patients in the GA group and a majority of the patients in the DS group responded, “did not wake up at all”.ConclusionSympathetic agitation, expressed as an increase of sAMY, was absent in the GA group. Meanwhile, in the DS group, some patients showed high levels of sAMY which went down following the administration of an analgesic agent, thus suggesting that pain caused an elevation in the level of the stress and thereby induced an increase in sAMY. The measurement of sAMY is therefore considered to be useful for the assessment of analgesic status under DS.

Impact of vessel diameter and bandwidth of illumination in sidestream dark-field oximetry

We investigate the possibility of oxygen saturation estimation from images obtained by the sidestream dark-field (SDF) technique. The SDF technique is a method for microvascular imaging. In SDF imaging, light enters a tissue directly from illumination sources configured around a camera and then the camera captures the light scattered by the tissue. To advance the capability of the SDF imaging system, we develop a SDF oximetry method with LED illumination sources. In this paper, we evaluate some SDF oximetry methods from virtual SDF images obtained by the Monte Carlo photon propagation simulation. As a result, we verify that SDF imaging allows the estimation of oxygen saturation of the individual vessels from virtual images using the average extinction coefficients considering the bandwidth of the illumination and the effect of the integration of the camera.

Laparoscopic and luminal endoscopic cooperative surgery can be a standard treatment for submucosal tumors of the stomach: a retrospective multicenter study

Background and study aims We have developed a combined laparoscopic and luminal endoscopic surgery technique for resection of gastric submucosal tumors (SMTs) that can be performed without excessive resection of the stomach. In a multicenter retrospective study we aimed to evaluate the feasibility and safety of laparoscopic and endoscopic cooperative surgery (LECS) for gastric SMT resection. Patients and methods Between October 2007 and December 2011, 126 patients with gastric SMTs underwent LECS at 8 institutions. Patient demographics, tumor histopathologic characteristics, and operative and follow-up data were reviewed. Results 16 tumors (12.7 %) were located in the upper third of the stomach, 88 (69.8 %) in the middle third, 5 (4.0 %) in the lower third, and 17 (13.5 %) at the esophagogastric junction. The mean (standard deviation [SD]) operation time for LECS was 190.2 (66.8) min, with a mean estimated blood loss of 15.1 (38.6) mL. In 2 cases (1.6 %), the procedure was converted to open surgery because of intra-abdominal adhesions or stenosis. Morbidity was found in 6 cases (4.8 %), including 2 leakage, 2 gastric stasis, 1 fever, and 1 cystitis. Histologically, a gastrointestinal stromal tumor (GIST) was found in 86 (68.3 %) cases. The median follow-up period was 54.7 months; no local or distant tumor recurrence was observed and all patients were alive. Conclusions LECS proved to be a safe and feasible procedure for the resection of gastric SMTs, with a reasonable operation time, low bleeding, and an acceptable complication rate in this multicenter study in Japan.

Rapid diagnosis of micrometastasis of gastric cancer using reverse transcription loop-mediated isothermal amplification

Methods to detect metastases in biopsy specimens with certain rapidity and accuracy are essential to performing tailor-made surgeries for solid malignancies. Reverse transcription loop-mediated isothermal amplification (RT-LAMP) reaction is a novel technique for detecting mRNA expression of target sequences with high sensitivity and rapidity, even from crude samples without RNA purification. Applicability to detect lymph node (LN) micrometastasis of gastric cancer was tested. Total of 26 LNs were retrieved from 10 patients with primary gastric cancer. Each LN was serially sectioned, and every set of three serial sections were tested for routine histopathological (H&E) and immunohistochemical examination with anti-cytokeratin antibodies (IHC), and RT-LAMP analysis targeted cytokeratin 19 mRNA. Results from H&E/IHC and RT-LAMP analysis were compared in each set of sections. All the sections of those containing metastatic lesions equivalent to a volume of overt metastasis (maximum diameter>2 mm), 90% of those containing micrometastasis (between 2 and 0.2 mm) and 83% of those containing isolated tumor cells (<0.2 mm) were detectable using this procedure. Total analysis from lysates of clinical specimens required<75 min. This new technique is suggested to be an alternative to rapid diagnosis of micrometastasis based on conventional histopathological analysis.

Viral shedding after p53 adenoviral gene therapy in 10 cases of esophageal cancer.

We detected adenoviral DNA fragments in excretions of 10 esophageal cancer patients by DNA‐PCR after tumor injection of Ad‐CMV‐vector. A total of 220 samples consisting of feces, gargling saliva, urine, and blood plasma were assessed. A total of 29.7% of feces samples and 13.2% of gargling saliva samples were positive for adenoviral DNA fragments, but 89.7% of the positive feces samples and all of the positive gargling saliva samples turned negative on day 12 after tumor injection. Although adenoviral DNA fragments may be pathogen‐free, patients’ feces and gargling saliva contain adenoviral DNA fragments for 12 days after injection. (Cancer Sci 2010; 101: 289–291)