CCR3 를 통한 염증 신호를 일으키는 NRS N 말단 부위에 대한 구조생물학적 연구

학위논문 (박사)-- 서울대학교 대학원 : 약학대학 약학과, 2019. 2. 한병우.Antisynthetase syndrome 은 autoimmune disease 의 하나로, aminoacyl-tRNA synthetase 에 대한 antibody 가 만들어진다. 그 중에서 asparaginyl-tRNA synthetase 에 대한 autoantibody 인 anti-KS 의 발생은 interstitial lung disease 와 밀접한 관련이 있는 것으로 보이는데 아직 생성 기전이 명확하게 밝혀지지 않았다. 본 연구에서는 human asparaginyl-tRNA synthetase 의 N 말단 부위의 결정구조를 규명하였고 그 구조가 Brugia malayi asparaginyl-tRNA synthetase 의 N 말단 부위와 유사하다는 것을 밝혔다. 또한 human asparaginyl-tRNA synthetase 의 N 말단 부위가 CC chemokine receptor 3 와 직접 결합하여 독립적으로 chemokine 활성을 가진다는 것을 보였고, 그 결합 방식을 NMR spectroscopy 기법을 이용하여 설명하였다. 나아가, interstitial lung disease 의 발달과 autoantibody 의 생성에 있어 human asparaginyl-tRNA synthetase 의 증가된 분비에 따른 비정상적인 N 말단 부위의 chemokine 활성이 잠재적 역할을 할 것이라 제시하였다.Asparaginyl-tRNA synthetase (NRS) is not only essential in protein translation but also associated with autoimmune diseases. Particularly, patients with antibodies that recognize NRS often develop interstitial lung disease (ILD). However, the underlying mechanism of how NRS is recognized by immune cells and provokes inflammatory responses is not well-understood. Here, I found that the crystal structure of the unique N-terminal extension domain of human NRS (named as UNE-N, where -N denotes NRS) resembles that of the chemotactic N-terminal domain of NRS from a filarial nematode, Brugia malayi, which recruits and activates specific immune cells by interacting with CXC chemokine receptor 1 and 2. UNE-N induced migration of C-C chemokine receptor 3 (CCR3)-expressing cells. The chemokine activity of UNE-N was significantly reduced by suppressing CCR3 expression with CCR3-targeting siRNA, and the loop3 region of UNE-N was shown to interact mainly with the extracellular domains of CCR3 in nuclear magnetic resonance perturbation experiments. Based on these results, evolutionarily acquired UNE-N elicits chemokine activities that would promote NRS-CCR3-mediated proinflammatory signaling in ILD.1. Introduction 1 1.1. Aminoacyl-tRNA synthetase . 1 1.2. Class 2b ARS . 10 1.3. Chemokine activities of NRS . 12 1.4. Antisynthetase syndrome . 14 1.5. Chemokine 16 1.6. Summary 18 2. Materials and Methods . 19 2.1. Cloning, protein expression, and purification . 19 2.2. Selenomethionine incorporation 28 2.3. Crystallography 28 2.4. NMR analysis . 35 2.5. Cell culture . 36 2.6. Cell migration assay 37 2.7. Secretion assay 38 2.8. Immunoblotting 39 2.9. Pull-down assay 39 2.10. Statistical analysis . 40 2.11. Data deposition 40 3. Results 41 3.1. UNE-N shares a structural scaffold with the chemotactic N-terminal domain of BmNRS but exhibits unique features 41 3.2. Th1 and Th2 cytokines induce NRS secretion from macrophages 56 3.3. UNE-N exclusively elicits chemotactic activities of NRS on Daudi and Jurkat cells 60 3.4. UNE-N directly interacts with CCR3 extracellular domains 64 3.5. Loop3 of UNE-N is an effector motif for CCR 3interactions 68 3.6. Loop3 region of UNE-N is structurally distinct from UNE-N of BmNRS targeting different receptors . 75 4. Discussion . 79 5. References 88 6. Abstract in Korean (국문 초록) 96Docto

Frey's procedure for chronic pancreatitis: a 10-year single-center experience in Korea

Purpose: Chronic pancreatitis (CP) is progressive inflammatory disease that leads to irreversible destruction of the pancreatic parenchyma. The main indications for surgical intervention in cases involving CP are intractable pain, suspicion of malignancy, and failure of other methods. However, there is no report related to Frey's procedure in Korea; hence, we aimed to investigate and analyze our institution's experience and determine the benefits of surgical treatment for CP. Methods: This was a retrospective study of 24 patients with CP who underwent Frey's procedure at Gangnam Severance Yonsei University between January 2007 and December 2017. Preoperative exocrine and endocrine pancreatic function, perioperative finding (blood loss, operation time), postoperative complications were evaluated. Statistical analytics were chi-square test, Fisher exact tests, and Wilcoxon signed-rank test and Mann-Whitney U-test. Results: Surgery was performed due to alcohol-derived CP in 12 of 24 patients (50%) and due to pancreatic stones in 15 of 24 patients (62.5%). Two patients had postoperative complications which were managed conservatively. After surgery, 7 of 24 patients were prescribed with exocrine medication. Comparison of the preoperative and postoperative conditions showed that glycated hemoglobin had no significant differences. After surgery, only 5 patients (21%) complained of intermittent abdominal pain. Conclusion: In conclusion, Frey's procedure appears to be a less burdensome surgical procedure. Thus, it could be the first option for management of patients with large pancreatic stone.ope

Clinical application of invalid foods using mealworms and evaluation of nutrition status and immune function: a study protocol for a randomized, double blind, placebo-controlled trial

Background Protein intake is important for the recovery of the immune system, physical strength, and wound healing after surgery. Sarcopenia is associated with a poor prognosis when compared to patients without sarcopenia in cancer patients. Recently, edible insects, such as mealworms, have been recognized as having a high protein content. In this study, we will evaluate the effect of nutritional status and immune function change based on a patient’s ingestion of mealworms after hepatobiliary pancreatic surgery. Methods/design This is a prospective, two-armed, phase III study investigating the effect of mealworm improving nutrition and immune status in patients after hepatobiliary pancreatic surgery. In the trial group, the patients will be provided with mealworms for 2 months after surgery. In the control group, patients will be provided with grain powder for 2 months after surgery. The target for accrual is 168 patients. We divided in to three groups according to the type of surgery. Discussion The primary endpoint is to evaluate body cell mass index 2 months postoperatively. Secondary endpoints include other body composition changes as well as nutrition index and immune function change. We expect that ingestion of mealworms can effectively improve the nutritional status and enhance the immune function. Mealworm can be used effectively for nutritional management of patients after surgery. Trial registration Clinicaltrials.gov NCT03201926 Registered June 28, 2017, retrospectively registered.ope

P16 and MGMT hypermethylation predicts surgical outcomes in curative resected mid/distal bile duct cancer

Dept. of Medicine/박사Extrahepatic bile duct cancer is a primary malignancy arising from epithelium of extrahepatic biliary tract. In patients with extrahepatic bile duct carcinoma, the 5-year survival rate was 38.3%; a very poor prognosis has been reported. In these patients, the important prognostic factors include the TNM stage, cell differentiation and histologic type. Nevertheless, we often encounter a substantial number of patients whose prognosis is not consistent with the TNM stage. Therefore, other prognostic criteria are mandatory than the TNM staging system which has been widely used at present. We aimed to evaluate the potential role of DNA promoter methylation of gene involved in a variety of celluar function including adhesion (CDH1), cell division (p16) and survival (DAPK), to predict disease free survival (DFS) and overall survival (OS) in curative resected mid/distal bile duct cancer. Sixty-five mid/distal bile duct carcinoma specimens obtained at Severance Hospital of Yonsei University College of Medicine from January 2000 to December 2006. Methylation of interest loci was confirmed using pyrosequecning.The significant methylation frequencies (MtI > 5%) of the 3 gene analyzed were 17% for P16, 54% for DAPK, 60% for E-cadherin. P16 and DAPK MtI status correlated with perineural invasion and tumor depth, respectively. In 65 patients, the 3-year and 5-year overall survival was 54.9% and 48.4%, respectively. In multivariate analysis of overall survival, presence of lymph node metastasis and P16 methylation status were identified as an independent prognostic factors for overall survival. In patient with unmethylated of P16, the 3- and 6-yeat survival rates were 60.8 % and 54.9 %, respectively. In patients with a hypermethylated of P16, the 3- and 6-yeat survival rates were 27.3 % and 0.0 %, respectively. P16 hypermethylation can predict overall survival in curative resected mid/distal bile duct cancer. Classification of mid/distal bile duct cancer by both genetic and epigenetic profiles may improve the capability of predicting prognosis and of applying tailored therapy in mid/distal cancer.ope

An Ultrasonically Powered Implantable Microprobe for Electrolytic Ablation

Electrolytic ablation (EA) is a promising nonthermal tumor ablation technique that destroys malignant cells through induction of a locoregional pH change. EA is typically performed by inserting needle electrodes inside the tumor followed by application of direct current (DC), thus inducing electrolysis and creating localized pH changes around the electrodes. In this paper, we report an ultrasonically powered implantable EA microprobe that may increase the clinical relevance of EA by allowing wireless control over device operation (capability to remotely turn the device on and off) and providing flexibility in treatment options (easier to administer fractionated doses over a longer period). The wireless EA microprobe consists of a millimeter-sized piezoelectric ultrasonic receiver, a rectifier circuit, and a pair of platinum electrodes (overall size is 9x3x2mm(3)). Once implanted through a minimally invasive procedure, the microprobe can stay within a solid tumor and be repeatedly used as needed. Ultrasonic power allows for efficient power delivery to mm-scale devices implanted deep within soft tissues of the body. The microprobe is capable of producing a direct current of 90 mu A at a voltage of 5V across the electrodes under low-intensity ultrasound (similar to 200mW/cm(2)). The DC power creates acidic (pH12.9) regions around the anode and the cathode, respectively. The pH change, measured using tissue-mimicking agarose gel, extends to 0.8cm(3) in volume within an hour at an expansion rate of 0.5mm(3)/min. The microprobe-mediated EA ablative capability is demonstrated in vitro in cancer cells and ex vivo in mouse liver.ope

Modeling Pancreatic Cancer with Patient-Derived Organoids Integrating Cancer-Associated Fibroblasts

Pancreatic cancer is a devastating disease and is highly resistant to anticancer drugs because of its complex microenvironment. Cancer-associated fibroblasts (CAFs) are an important source of extracellular matrix (ECM) components, which alter the physical and chemical properties of pancreatic tissue, thus impairing effective intratumoral drug delivery and resulting in resistance to conventional chemotherapy. The objective of this study was to develop a new cancer organoid model, including a fibrous tumor microenvironment (TME) using CAFs. The CAF-integrated pancreatic cancer organoid (CIPCO) model developed in this study histologically mimicked human pancreatic cancer and included ECM production by CAFs. The cancer cell-CAF interaction in the CIPCO promoted epithelial-mesenchymal transition of cancer cells, which was reversed by CAF inhibition using all-trans retinoic acid. Deposition of newly synthesized collagen I in the CIPCO disturbed the delivery of gemcitabine to cancer cells, and treatment with collagenase increased the cytotoxic effect of gemcitabine. This model may lead to the development of next-generation cancer organoid models recapitulating the fibrous TME.ope

Glioblastoma Cellular Origin and the Firework Pattern of Cancer Genesis from the Subventricular Zone

Glioblastoma (GBM) is a disease without any definite cure. Numerous approaches have been tested in efforts to conquer this brain disease, but patients invariably experience recurrence or develop resistance to treatment. New surgical tools, carefully chosen samples, and experimental methods are enabling discoveries at single-cell resolution. The present article reviews the cell-of-origin of isocitrate dehydrogenase (IDH)-wildtype GBM, beginning with the historical background for focusing on cellular origin and introducing the cancer genesis patterned on firework. The authors also review mutations associated with the senescence process in cells of the subventricular zone (SVZ), and biological validation of somatic mutations in a mouse SVZ model. Understanding GBM would facilitate research on the origin of other cancers and may catalyze the development of new management approaches or treatments against IDH-wildtype GBM.ope

Intraductal papillary mucinous neoplasm of the biliary tract with cardiac metastasis: A case report

Introduction: Intraductal papillary mucinous neoplasm of the biliary tract (IPNB) is a rare, low-grade neoplasm limited to the bile duct mucosa. The malignant transformation rate is low, and there have been limited reports of metastasis to other organs. Herein, we presented a rare case of a patient who was diagnosed with IPNB concurrent with invasive adenocarcinoma after surgery and was diagnosed with cardiac metastasis 6 months later. Patient concerns: A 61-year-old male patient presented with abdominal pain to a local clinic. He was diagnosed with intrahepatic cholangiocarcinoma with pancreatitis and transferred to our hospital. Diagnosis: Diagnostic testing (magnetic resonance imaging, endoscopic retrograde cholangiopancreatography, positron emission tomography-computed tomography) revealed a papillary neoplasm and invasive adenocarcinoma with papillary neoplasm in the periampullary lesion. Interventions: Pancreaticoduodenectomy, right hemihepatectomy, and left lateral sectionectomy of the liver were performed. After surgery, we planned gemcitabine-based adjuvant chemotherapy. Outcomes: Upon completion of the sixth gemcitabine chemotherapy cycle, a hyperechoic, oval-shaped mass (1.3 × 2.6 cm) was found on the outer wall of the right atrium. Resection of a cardiac tumor in the right atrium and patch repair were performed. Conclusion: To our knowledge, no other case of cardiac metastasis found during observation after surgery for an IPNB has been described. IPNBs are known to be less aggressive and to have a lower metastasis rate than intraductal papillary mucinous neoplasms; therefore, the number of case reports describing metastasis after surgery is relatively low. Our case suggests that close observation is necessary in patients diagnosed with an IPNB.ope

Practical guidelines for the surgical treatment of gallbladder cancer.

At present, surgical treatment is the only curative option for gallbladder (GB) cancer. Many efforts therefore have been made to improve resectability and the survival rate. However, GB cancer has a low incidence, and no randomized, controlled trials have been conducted to establish the optimal treatment modalities. The present guidelines include recent recommendations based on current understanding and highlight controversial issues that require further research. For T1a GB cancer, the optimal treatment modality is simple cholecystectomy, which can be carried out as either a laparotomy or a laparoscopic surgery. For T1b GB cancer, either simple or an extended cholecystectomy is appropriate. An extended cholecystectomy is generally recommended for patients with GB cancer at stage T2 or above. In extended cholecystectomy, a wedge resection of the GB bed or a segmentectomy IVb/V can be performed and the optimal extent of lymph node dissection should include the cystic duct lymph node, the common bile duct lymph node, the lymph nodes around the hepatoduodenal ligament (the hepatic artery and portal vein lymph nodes), and the posterior superior pancreaticoduodenal lymph node. Depending on patient status and disease severity, surgeons may decide to perform palliative surgeries.ope

Ivermectin and gemcitabine combination treatment induces apoptosis of pancreatic cancer cells via mitochondrial dysfunction

Pancreatic cancer is an aggressive cancer characterized by high mortality and poor prognosis, with a survival rate of less than 5 years in advanced stages. Ivermectin, an antiparasitic drug, exerts antitumor effects in various cancer types. This is the first study to evaluate the anticancer effects of the combination of ivermectin and gemcitabine in pancreatic cancer. We found that the ivermectin-gemcitabine combination treatment suppressed pancreatic cancer more effectively than gemcitabine alone treatment. The ivermectin-gemcitabine combination inhibited cell proliferation via G1 arrest of the cell cycle, as evidenced by the downregulation of cyclin D1 expression and the mammalian target of rapamycin (mTOR)/signal transducer and activator of transcription 3 (STAT-3) signaling pathway. Ivermectin-gemcitabine increased cell apoptosis by inducing mitochondrial dysfunction via the overproduction of reactive oxygen species and decreased the mitochondrial membrane potential. This combination treatment also decreased the oxygen consumption rate and inhibited mitophagy, which is important for cancer cell death. Moreover, in vivo experiments confirmed that the ivermectin-gemcitabine group had significantly suppressed tumor growth compared to the gemcitabine alone group. These results indicate that ivermectin exerts synergistic effects with gemcitabine, preventing pancreatic cancer progression, and could be a potential antitumor drug for the treatment of pancreatic cancer.ope