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丙型肝炎病毒截短型核心基因表达载体的构建与表达

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同济大学学报（医学版） 2009年第5期30卷 50-53页

作者：孙贝贝张敬石红军 hiroaki kohno 张军同济大学基础医学院生物学教研室上海200092 同济大学生命科学与技术学院上海200092 日本协和梅迪克斯株式会社

目的构建重组丙型肝炎病毒核心基因表达载体pVec-core,并进行原核表达。方法人工合成丙型肝炎病毒截短型核心基因至pUC57载体上,经双酶切后,将其克隆至原核表达质粒pVec中,测序正确后将重组质粒pVec-core转化入表达宿主菌BL21,经IPTG诱... 详细信息

目的构建重组丙型肝炎病毒核心基因表达载体pVec-core,并进行原核表达。方法人工合成丙型肝炎病毒截短型核心基因至pUC57载体上,经双酶切后,将其克隆至原核表达质粒pVec中,测序正确后将重组质粒pVec-core转化入表达宿主菌BL21,经IPTG诱导表达核心蛋白,SDS-PAGE检测目的蛋白的表达情况。采用GST亲和层析方法纯化融合蛋白并进行Western印迹法验证。结果丙型肝炎病毒核心基因表达载体构建成功,重组菌表达出相对分子质量约为40.6 kD重组融合蛋白,并以可溶形式存在于菌体中。蛋白经谷胱甘肽琼脂糖凝胶纯化,得到纯度较高的目的蛋白。经Western印迹分析,该蛋白可与HCV患者阳性血清发生特异性结合反应,具有良好的抗原活性。结论核心抗原表达载体在原核表达系统中高效表达,为利用其作为诊断抗原及研究核心蛋白的其他生物学功能奠定了基础。

关键词：丙型肝炎病毒核心蛋白原核表达纯化

Enzyme inhibition assay for pyruvate dehydrogenase complex: Clinical utility for the diagnosis of primary biliary cirrhosis

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World Journal of Gastroenterology 2005年第43期11卷 6735-6739页

作者： Katsuhisa Omagari hiroaki Hazama Shigeru kohno Second Department of Internal Medicine

Primary biliary cirrhosis (PBC) is usually diagnosed by the presence of characteristic histopathological features of the liver and/or antimitochondrial antibodies (AMA) in the serum traditionally detected by immunofluorescence. Recently, new and more accurate serological assays for the detection of AMA, such as enzyme-linked immunosorbent assay (ELISA), immunoblotting, and enzyme inhibition assay, have been developed. Of these, the enzyme inhibition assay for the detection of anti-pyruvate dehydrogenase complex (PDC) antibodies offers certain advantages such as objectivity, rapidity, simplicity, and low cost. Since this assay has almost 100% specificity, it may have particular applicability in screening the at-risk segment of the population in developing countries. Moreover, this assay could be also used for monitoring the disease course in PBC. Almost all sera of PBC-suspected patients can be confirmed for PBC or non-PBC by the combination results of immunoblotting and enzyme inhibition assay without histopathological examination. For the development of a "complete" or "gold standard" diagnostic assay for PBC, similar assays of the enzyme inhibition for anti-2-oxoglutarate dehydrogenase complex (OGDC) and anti-branched chain oxo-acid dehydrogenase complex (BCOADC) antibodies will be needed in future.

关键词： Primary biliary cirrhosis Enzyme inhibition assay Antimitochondrial antibody 2-oxo-acid dehydrogenasecomplex

Gastric mixed adenoma-neuroendocrine tumor:A case report

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World Journal of Clinical Cases 2020年第22期8卷 5744-5750页

作者： Shuzo kohno hiroaki Aoki Masayuki Kato Masaich Ogawa Kazuhiko Yoshida Department of Surgery The Jikei University Katsushika Medical CenterTokyo 125-8506Katsushika-kuJapan Department of Endoscopy The Jikei University Katsushika Medical CenterTokyo 125-8506Katsushika-kuJapan

BACKGROUND Gastric mixed adenoma-neuroendocrine tumors(NETs)are quite *** the 2019 world health organization classification of tumors of the digestive system,these were designated as a combination of grade 1 or grade 2 NETs and adenomas or tubular *** are no treatment guidelines for these tumors,and pathological and clinical studies are ***,we review previous case reports and present a case of gastric mixed *** SUMMARY A 66-year-old man underwent gastrointestinal endoscopy for the evaluation of upper abdominal *** examination of the biopsy specimen indicated the possibility of gastric cancer.A histopathological examination by endoscopic submucosal dissection showed a mixed adenoma-NET that was completely excised by endoscopic submucosal *** recurrence was observed on gastrointestinal endoscopy at the 6-mo *** Clinicians'awareness of this rare tumor is important for its timely diagnosis and treatment.

关键词： Adenoma Neuroendocrine tumor Diagnosis Pathology Stomach treatment Case report

Plasma leptin and ghrelin concentrations in patients with Crohn's disease

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World Journal of Gastroenterology 2005年第46期11卷 7314-7317页

作者： Yoshito Nishi Hajime Isomoto hiroaki Ueno Ken Ohnita Chun Yang Wen Fuminao Takeshima Ryosuke Mishima Masamitsu Nakazato Shigeru kohno Second Department of Internal Medicine Nagasaki University School of Medicine Sakamoto 1-7-1 Nagasaki Japan Department of Endoscopy Nagasaki University School of Medicine Sakamoto 1-7-1Nagasaki Japan Department of Molecular Pathology Atomic Bomb Disease Institute Nagasaki University School of MedicineSakamoto 12-4 Nagasaki Japan Third Departmentof Internal Medicine Miyazaki Medical College KiyotakeMiyazaki Japan

AIM: To determine the concentrations of leptin and ghrelin, which have opposite effects on appetite,energy expenditure, and weight control, in the plasma of patients with Crohn's disease (CD), which is often associated with weight loss and ***: Plasma leptin and ghrelin concentrations were determined in 28 outpatients with CD by radioimmunoassay. Age- and sex-matched controls with and without Helicobacter pylori(H pylori) infection (28for each) were enrolled in the study. Circulating levels of these hormones were assessed with respect to CD activity, disease localization and medical ***: There were no significant differences in ghrelin levels between CD patients and H pylorinegative controls. However, circulating ghrelin levels were significantly lower in H pylori-infected subjects than in CD patients and uninfected controls. Plasma leptin levels were comparable among the groups. Localization and medication profile had no significant impact on circulating ghrelin and leptin ***: Apart from H pyloriinfection, CD itself has no significant influence on circulating ghrelin and leptin levels in the outpatients who were mostly in inactive state.

关键词： Crohn's disease Ghrelin Leptin Helicobacter pylori

Neoadjuvant Chemotherapy for Clinical Stage II and III Thoracic Esophageal Squamous Cell Carcinoma with Curative Esophagectomy

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Journal of Cancer Therapy 2015年第15期6卷 1207-1213页

作者： Masahide Ikeguchi Yusuke kohno Kyoichi Kihara Kazunori Suzuki Kanenori Endo Seiichi Nakamura Takashi Sawada Tetsu Shimizu Tomoyuki Matsunaga Yoji Fukumoto hiroaki Saito Tottori Prefectural Central Hospital Tottori Japan Department of Surgery Division of Surgical Oncology Faculty of Medicine Tottori University Tottori Japan

Esophageal squamous cell carcinoma (ESCC) is a gastrointestinal carcinoma with a poor prognosis. To improve the survival of patients with this disease, neoadjuvant chemotherapy (NAC) has been introduced. However, the survival benefits of NAC or the correlation between NAC and postoperative complications have not been well considered. In the present study, we retrospectively investigated the clinicopathological effectiveness of NAC in patients with clinical stage II and III thoracic ESCC. This retrospective study enrolled 63 patients with clinical stage II and III thoracic ESCC, who underwent resection of the thoracic esophagus and three-field lymph node dissection between January 2007 and December 2013. NAC with cisplatin plus 5-fluorouracil (5-FU) was introduced in 38 patients. NAC did not correlate with the occurrence of postoperative complications. The 5-year disease-free survival (DFS) rate of the 38 patients with NAC (41.6%) was similar to that for the 25 patients who did not receive NAC (38.1%;P = 0.784). However, we found that the DFS of 17 patients with histopathological Grade 2 and 3 tumors who received NAC (5-year DFS rate: 58.1%) was significantly higher than that of 21 patients with low histopathological grade tumors who received NAC (5-year DFS rate: 28.6%), or than that of the 25 patients who did not receive NAC (38.1%). Moreover, we found that the effectiveness of NAC assessed macroscopically did not correlate with the effectiveness of NAC assessed microscopically. These findings may indicate that preoperative estimation of NAC effectiveness is important in avoiding unnecessary adverse drug effects caused by NAC, and in prolonging the survival of patients with thoracic ESCC.

关键词： Esophageal Squamous Cell Carcinoma Histopathological Grade Neoadjuvant Chemotherapy Prognosis

Novel serine/threonine kinase 11 gene mutations in PeutzJeghers syndrome patients and endoscopic management

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World Journal of Gastrointestinal Endoscopy 2013年第3期5卷 102-110页

作者： Hiroyuki Yajima Hajime Isomoto hiroaki Nishioka Naoyuki Yamaguchi Ken Ohnita Tatsuki Ichikawa Fuminao Takeshima Saburo Shikuwa Masahiro Ito Kazuhiko Nakao Kazuhiro Tsukamoto Shigeru kohno Department of Molecular Microbiology and Immunology Nagasaki University Graduate School of Biomedical SciencesNagasaki University School of MedicineNagasaki 852-8501Japan Department of Gastroenterology and Hepatology Nagasaki University School of MedicineNagasaki 852-8501Japan Department of Pharmacotherapeutics Nagasaki University Graduate School of Biomedical SciencesNagasaki University School of MedicineNagasaki 852-8521Japan Department of Pathology National Nagasaki Medical CenterOmuraNagasaki 856-0835Japan

AIM:To explore mutations in serine/threonine kinase 11(STK11) gene in Peutz-Jeghers syndrome(PJS) with gastrointestinal(GI) hamartomatous ***:Six Japanese PJS patients in 3 families were enrolled in this *** of the cases had hamartomatous polyposis in the gastrointestinal tract,including the small intestine,along with mucocutaneous ***-band imaging(NBI)-magnification endoscopy was employed to detect microvascular and microsurface irregularities in the GI *** magnification findings could be classified into three groups(type A,type B,or type C).Endoscopic polypectomy was performed using double-balloon enteroscopy or *** DNA was extracted from a whole blood sample from each *** of the coding exons of STK11 gene,its boundary regions,and the promoter region containing the polymorphic regions were amplified by polymerase chain reaction,and direct sequencing was performed to assess the germline ***:NBI-magnification endoscopic observation could detect the abnormalities in microvessels and microsurface structures of GI ***,we found 5 cases of type A and one case without the examination for the gastric polyps,while there were 4 cases of type B and 2 case of type A for the colorectal ***-nine small-bowel and 115 colorectal polyps over 27 sessions for each were resected endoscopically without significant *** only delayed complication included the occurrence of bleeding in a case,and this was successfully managed with *** polyps contained no malignant *** on mutation analysis,all 3 cases in Family I exhibited the +658C>T nonsense mutation in exon 5,which resulted in the production of a truncated protein(Q220X).In Family II,a case had-252C>A and-193C>A in the promoter *** Family III,a case was found to have the +1062C>G(F342L) mutation in exon ***:We found two novel mutations of STK11 in association with ***

关键词： Peutz-Jeghers syndrome Serine/threonine kinase 11 Gastrointestinal hamartomatous polyps Double-balloon enteroscopy Narrow-band imaging

Neoadjuvant Chemotherapy for Clinical Stage II and III Thoracic Esophageal Squamous Cell Carcinoma with Curative Esophagectomy

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