탄소-수소 활성화 및 설페이트 클릭 반응의 화학생물학 응용

학위논문 (박사)-- 서울대학교 대학원 : 자연과학대학 화학부, 2018. 2. 김병문.Factors affecting cellular phenomena comprise a series of complicated and elaborate signal transductions. Due to the complexity of biomolecules, direct observation of a specific biomolecule is extremely difficult. Therefore conjugation of functionality onto the biomolecule is often implemented for the purpose of monitoring a bio-target. Since hydrophobic interaction between a biomolecule and a ligand is usually not strong enough to be monitored by common analytical tools, covalent bond formation of an artificial tag molecule has been developed. After Huisgens discovery of 1,3-dipolar cycloaddition between an azide and an alkyne, Sharpless group reported that the reaction can be catalyzed by a Cu(I) species. Furthermore, introduction of azidophenylalanine into proteins through expanded genetic code renders possible the conjugation of an artificial functionality such as fluorescence, and anti-cancer activity, etc. It is of particular note that the fluorescence attachment makes the spatiotemporal observation of bio-target possible. As a consequence, both the development of a new bioconjugation method, and the synthesis of functional molecules to be ligated have become significant research subjects in chemical biology field. This dissertation includes research results of two parts, 1) developing synthetic method of fluorescent molecules which can be used as a biosensor, and 2) discovery of novel tyrosine bioconjugation chemistry. The first part shows one of our approaches in utilizing novel metal assisted 1,3-cycloaddition and Pd-mediated C-H activation for efficient syntheses of indolizinone fluorophores containing a small molecule library. Since the photophysical property of indolizinone is regulated by the electronic characteristics of C7 and C9 substituents, diversification of those substituents was crucial for the construction of a fluorescent molecule library. Through the new synthetic route, synthesis of a number of fluorescent molecules was accomplished, as diversification of C9 substituent occurs at the late stage of the synthesis. Moreover, several biosensors were synthesized, which had been previously infeasible. In part 2, tyrosine selective conjugation method through sulfate click reaction is described. Since SuFEx (sulfur(VI) fluoride exchange) chemistry was known as the reaction between aryl fluorosulfate and aryl silyl ether, aryloxy anion has scarecely been recognized as a useful reaction partner for SuFEx. In this part, a reaction condition to allow for the reaction of aryl fluorosulfate with tyrosine in the presence of several nucleophilic amino acid residues was carefully selected, and a feasibility test of a new site-specific biomolecule modification was successfully carried out on a model peptide, TAT 47-57, and a physiologically important protein, erythropoietin.SuFEx 2 1.4 References 3 Chapter 2. Developing Synthetic Method of Indolizin-one based Fluorescent Molecules. 5 2.1 Introduction. 5 2.2 Results and Discussion. 7 2.2.1 Synthesis of Fluorophore through 1,3-Dipolar Cycloaddition 7 2.2.2 Coupling Reaction of the Fluorophore and Aryl Iodide through Pd-mediated C-H Activation 8 2.2.3 Fluorescence Quenching through PeT. 11 2.2.4 Synthesis of Hydrogen Peroxide Sensor 16 2.2.5 Heck type Coupling Reaction of the Fluorophore and Styrene Derivatives. 18 2.3 Conclusion. 21 2.4 Experimental Section 22 2.4.1 Materials and Methods 22 2.4.2 Preparations of Compounds 24 2.4.3 Cell Culture and Imaging 40 2.5 References 41 Chapter 3. Discovery of Tyrosine Selective Bioconjugation. 43 3.1 Introduction. 43 3.2 Results and Discussion. 45 3.2.1 SuFEx Reaction of Aryloxy Anion and Aryl Fluorosulfate. 45 3.2.2 Chemoselectivity of SuFEx Reaction among Various Nucleophilic Amino Acid Model Compounds. 47 3.2.3 Fluorescence Tagging on TAT 47-57 Peptide 49 3.2.4 PEGylation of Erythropoietin 52 3.3 Conclusion. 58 3.4 Experimental Section 59 3.4.1 Materials and Methods 59 3.4.2 Preparations of Compounds, Peptides and Proteins. 60 3.4.3 Cell Culture and Imaging 70 3.4.4 In Vivo Mice Experiment 71 3.5 References 78 Abstract in Korean 75Cahpter 1. Introduction. 1 1.1 Bioconjugation of Protein 1 1.2 Fluorescencetagging and biosensor 2 1.3 Sulfur (VI) Fluoride ExchangeDocto

중환자실 간호사의 자기효능감과 자기통제력이 공감피로와 공감만족에 미치는 영향

학위논문 (석사)-- 서울대학교 대학원 : 간호학과 성인간호학 전공, 2016. 2. 고진강.중환자실 간호사들은 일반부서의 간호사에 비해 높은 공감피로를 보이고 있고, 공감피로는 소진을 증가시키며 간호업무성과를 저하시키는 요인으로 작용한다. 따라서 중환자실 간호사의 공감피로를 감소시키고 공감만족을 증진시키기 위한 중재방법을 개발하려는 노력이 필요하다. 본 연구의 목적은 중환자실 간호사의 공감피로와 공감만족에 미치는 자기효능감과 자기통제력의 영향을 조사하는 것이다. 본 연구는 자가보고식 설문지를 활용한 횡단면적 조사연구이다. 서울시내 소재 일 종합병원의 성인 중환자실 간호사를 대상으로 2015년 8월부터 9월에 자료를 수집하여 총 148부를 자료분석에 사용하였다. 연구 도구는 일반적 특성 25문항, 공감피로 10문항, 공감만족 10문항, 자기효능감 17문항, 자기통제력 28문항의 총 90문항으로 구성된 설문지를 사용하였다. 연구대상자의 자기효능감은 평균 3.27점 이었고, 자기통제력은 평균 3.6점이었다. 연구대상자의 공감피로는 30.82점이었고, 공감만족은 30.16점 이었다. 연구대상자들은 현재부서 경력이 짧을수록, 외상경험이 있는 경우, 스트레스 정도가 높은 경우, 우울증상을 경험한 경우에 공감피로 정도가 높았다. 연구대상자들은 연령이 높을수록, 심혈관내과 중환자실과 외과 중환자실에 근무하는 경우, 업무만족도가 높을 수록, 친구, 가족, 동료로부터 사회적 지지를 많이 받을 수록, 의료진과의 의사소통이 더 원활할 수록, 보호자와의 관계가 좋을 수록, 외상경험이 있는 경우, 자기계발 활동을 하는 경우, 스트레스 정도가 낮을 수록, 최근 2주간 병의원에서 치료받은 경험이 있는 경우에 공감만족이 높은 것으로 나타났다. 본 연구에서 자기효능감은 공감만족과 강한 양의 상관관계를 보였으나 공감피로와는 유의한 상관관계를 보이지 않았고, 자기통제력은 공감피로와 공감만족 정도에 유의한 상관관계를 보이지 않았다. 공감피로와 공감만족은 약한 양의 상관관계를 보였다. 연구대상자의 공감피로에 영향을 미치는 요인은 우울증상의 경험과 공감만족이었다. 공감만족에 영향을 미치는 요인은 업무만족, 공감피로, 자기효능감, 의료진과의 의사소통, 스트레스 정도, 치료경험 유무, 보호자와 관계, 외상경험이 있었다. 본 연구를 통해 중환자실 간호사의 자기통제력은 공감피로와 공감만족에 직접적인 영향을 미치지 않는 것으로 조사되었으나 중환자실 간호사의 자기효능감은 공감만족과 양의 상관관계에 있으며 유의하게 영향을 미치는 요인인 것으로 나타났다. 본 연구의 결과에 따라, 중환자실 간호사의 자기효능감을 증진시킬 수 있는 방안들을 개발하고 적용하는 것이 중환자실 간호사의 공감만족을 향상시키는데 도움이 될 것이라고 제언한다.Ⅰ.서론 1 1.연구의 필요성 1 2.연구의 목적 3 3.용어정의 4 Ⅱ.문헌고찰 6 1.공감피로 6 2.공감만족 10 3.자기효능감 13 4.자기통제력 18 Ⅲ.연구방법 20 1.연구설계 20 2.연구대상자 20 3.연구 도구 21 4.자료수집방법 24 5.자료분석방법 25 6.윤리적 고려 26 Ⅳ.연구결과 27 1.대상자의 일반적인 특성 27 2.대상자의 자기효능감, 자기통제력, 공감피로 및 공감만족 정도 36 3.대상자의 특성에 따른 공감피로와 공감만족 정도 및 각 개념 간 상관관계 41 4.대상자의 공감피로와 공감만족에 영향을 미치는 요인 49 Ⅴ.논의 61 Ⅵ.결론 및 제언 72 1.결론 72 2.제언 75 참고문헌 77 부록 87 1.부록 Ⅰ 87 2.부록 Ⅱ 98 3.부록 Ⅲ 99 Abstract 101Maste

임산부의 치주질환 활성도와 조산과의 상관관계에 관한 연구

Purpose We designed this study for the purpose of determining the relationship between periodontal disease activity and PLBW, using the evaluation of probing pocket depth, loss of attachment, gingival index, gingival crevicular fluid amount and subgingival microflora. Methods A total of 100 volunteer mothers(mean age 30.44) at the Department of Obstetrics and Gynecology Seoul National University Hospital were selected for this study.Pregnancy outcomes were categorized into cases and controls in two ways. our definition was based on the following; Group 1 : Any PLBW cases Vs. All NBW controls Group 2 : PLBW cases Vs. NBW controls A periodontal exam was performed on the Ramfjord( #16, 21, 24, 36, 41, 44) teeth and Clinical evaluation consisted of probing pocket depth, loss of attachment, gingival index and gingival crevicular fluid amount. Subgingival plaque samples were collected by three sterile #35 paper points. The total number of anaerobic colonies and aerobic bacteria were enumerated after incubation. Antisera to P. gingivalis, P. intermedia, A. actinomycetemcomitans were produced in white rabbits with live whole cells suspensions. The specific fluorescent bacteria obtained by immunofluorescence and total cell counts obtained by dark-field microscopy were counted on four fields. The percent of each specific microorganism in the total cell count was determined. Results Any PLBW and PLBW cases showed significantly greater probing depth and attachment loss than all NBW and NBW controls. Cases group had significantly increased anaerobic bacterial counts compared with control group and no differences in the other microbes. This study confirmed that periodontal disease is a statistically significant risk factor for PLBW by investigating clinical parameters and subgingival plaque analysis.이 연구는 1996년도 서울대학교 병원지정연구비 지원에 의한 결과입니다

Combined Intrathoracic and Intraperitoneal Splenosis after Splenic Injury: Case Report and Review of the Literature

Splenosis is defined as an autotransplantation of the splenic tissue after splenic rupture or splenectomy, and occurs most frequently in the peritoneal cavity. Splenosis is usually asymptomatic and is found incidentally. We report a case of combined intrathoracic and intraperitoneal splenosis in a 54-year-old male who worked as a miner for 10 years in his twenties, and was a current smoker. He was referred to our hospital for further evaluation of an incidental left diaphragmatic mass. Positron emission tomography-computed tomography and bronchoscopy were performed to evaluate the possibility of malignancy. There was no evidence of malignancy, but the spleen was not visualized. Reviewing his medical history revealed previous splenectomy, following a dynamite explosion injury. Therefore, splenosis was suspected and technetium-99m-labeled heat-damaged red blood cell scan confirmed the diagnosis. Radionuclide imaging is a useful diagnostic tool for splenosis, which could avoid unnecessary invasive procedures.ope

용융방사공정에서 다이팽윤현상의 1차원 모델링

Thesis (master`s)--서울대학교 대학원 :섬유고분자공학과,1998.Maste

Effects of rehabilitation program based on self-efficacy on poststroke patients

학위논문(박사)--서울대학교 대학원 :간호학과 간호학전공,2002.Docto

Strategic Plan of Okryu-dong Stream Park in Seo-Chon, Seoul

학위논문 (석사)-- 서울대학교 대학원 : 환경조경학과, 2011.2. 조경진.Maste

Targeting EGFR Signaling Pathway in NSCLC and Malignant Glioma Cells for Radiosensitization

학위논문 (박사)-- 서울대학교 대학원 : 협동과정 종양생물학 전공, 2013. 2. 김인아.Epidermal growth factor receptor (EGFR) signalings play important roles in the pathogenesis of lung cancer and malignant glioma, and therefore, molecular inhibition of the EGFR signaling pathway represents a promising strategy to enhance the anti-tumor activity of radiation. However, therapeutic resistance has emerged as an important clinical issue. Therefore, we investigated whether inhibitors targeting downstream molecules in pathways associated with EGFR signaling would radiosensitize a panel of non-small cell lung cancer (NSCLC) and malignant glioma cell lines showing activated EGFR. In the first part, we evaluated the efficacy of targeting EGFR-associated downstream signaling in NSCLC with activated EGFR, mutant K-RAS, or both. Specific inhibition of K-RAS attenuated downstream signaling and increased radiosensitivity of A549 and H460 cells, while inhibition of EGFR did not. A549 cells having a K-RAS mutation at codon V12 were radiosensitized by small interfering RNA (siRNA) targeting this codon. H460 cells harboring mutation at codon V61 were radiosensitized by siRNA targeting this codon. K-RAS siRNA did not radiosensitize H1299 cells possessing wild-type K-RAS. Inhibition of the phosphoinositide 3-kinase (PI3K)-AKT-mammalian target of rapamycin pathway led to significant radiosensitization of the two cell lines, while selective inhibition of extracellular signal-regulated kinase signaling did not. Inhibitors targeting PI3K-AKT-mTOR pathway also abrogated G2 arrest following irradiation and induced delay of γH2AX foci formation. A dual inhibitor of Class I PI3K and mTOR increased radiosensitivity of A549 and H460 cells effectively. Inhibition of PI3K-AKT signaling was associated with down-regulation of p-DNA-PK, respectively. While apoptosis was the major mode of cell death when the cells were pretreated with LY294002 or AKT inhibitor VIII, the cells were pretreated by rapamycin or PI103 showed mixed mode of cell death including autophagy. In the second part, we evaluated whether targeting EGFR-associated signaling would radiosensitize EGFR-activated glioma cells and further increase the radiosensitizing effect of temozolomide (TMZ). Although the result of radiotherapy (RT) for high grade glioma has significantly improved by concurrent and adjuvant TMZ, glioblastoma multiform (GBM) still has a dismal prognosis. Therefore, we tried to identify the effective targeting strategies to improve the therapeutic outcome of concurrent RT and TMZ. We first investigated the efficacy of targeting downstream molecules in EGFR-associated signaling pathway, such as Src, PI3K-AKT-mTOR, and STAT3, in glioma cell lines U251 and T98G. Next, we tested the effect of ligand-independent modulation using an HSP90 inhibitor and epigenetic modulation using a histone deacetylase (HDAC) inhibitor. U251 cells showing a high proportion of methylated methyl guanine transferase (MGMT) were highly responsive to the radiosensitizing effect of TMZ. Treatment with a Src inhibitor, PP2a dual inhibitor of Class I PI3K and mTOR, PI103a STAT3 inhibitor, Cpd188a HSP90 inhibitor, 17-DMAGor a HDAC inhibitor, LBH589, further increased the cytotoxic effect of RT plus TMZ in this cell line. Conversely, T98G cells showing a high proportion of unmethylated MGMT had a lower response to the radiosensitizing effect of TMZ although treatment with PI103, 17-DMAG, or LBH-589 increased the cytotoxic effect of radiotherapy plus TMZ. The mechanism of enhanced radiosensitizing effects of TMZ was multifactorial, involving impaired DNA damage repair and induction of autophagy or apoptosis. Taken together, these results suggest possible mechanisms for counteracting EGFR prosurvival signaling implicated in radioresistance of NSCLC and malignant glioma cells, and offer a potential strategy for overcoming resistance to EGFR inhibitors combined with irradiation.CONTENTS Abstract……………………………………….…………………………………………i Contents……………………………………………………………………………iv List of Tables……………………………………………………vii List of Figures……………………………………………………… viii General Introduction ………………………………………………..1 1. EGFR in cancer …. ……………………………………………………………………...1 2. Mechanisms of Resistance to EGFR Targeted Therapy…………………………………3 2-1. Alternative mechanisms for activating downstream signaling……………………….3 2-2. Nuclear EGFR and resistance to radiation therapy…………………………………...5 Purpose…………………………………………………………………8 PartI………………………………………………………………………9 Targeting Epidermal Growth Factor Receptor-Associated Signaling Pathways in Non -Small Cell Lung Cancer Cells: Implication in Radiation Response 1.Introduction………………………………………………………… 9 2.Materials and Methods……………………………………………… 11 2-1.Cell culture ………………………………………………………… 11 2-2.Pharmacologic inhibitors ………………………………………………. 11 2 - 3. RNA interference ..……………………………………………… …………………………11 2-4.Clonogenic assays………………. ……………………………………………… 12 2-5.Western analysis……………… ……………………………………………… 13 2- 6. Flow cytometric analysis ………………………………………………………………….13 2 - 7. Immunocytochemistry …………………………………………………………………….. 14 3. Results …………………………………………………………………………………15 3-1. siRNA inhibition of K-RAS increased A549 radiosensitivity with activated EGFR- KRAS signaling…………………………………………………………………………………………..15 3-2. Pharmacologic inhibition of ERK signaling did not increase A549 radiosensitivity………..18 3-3. Pharmacologic inhibition of PI3K-AKT-mTOR signaling increased A549 radiosensitivity …………………………………………………………………………………………………...18 3-4. Effects of combined inhibitor and radiation treatment on cell cycle progression…………...20 3-5. Inhibition of PI3K-AKT-mTOR signaling induced prolongation of γH2AX foci…………..20 3-6. A dual inhibitor of Class I PI3K and mTOR signaling led to effective radiosensitization in A549 cells………………………………………………………………………………………...22 3-7. Marker for DNA damage repair……………………………………………………………..24 3-8. Mode of cell death………………………………………………………………………….. 26 3-9.Toxicity towards normal human fibroblasts……………………………………………….... 29 4. Discussion……………………………………………………………………………………31 PartII............................................................................................................35 Enhancement of the Radiosensitizing Effect of Temozolomide: Targeting EGFR-associated Signaling in Malignant Glioma Cells 1.Introduction…………………………………………………………………………. 35 2.Materials and Methods………………………………………………………. 37 2-1.Cell culture …………………………………………………………………. 37 2-2. Pharmacologic inhibitors ……………………………………………… 37 2-3. Clonogenic assays ……….……………………………………………… 38 2-4. Western analysis ………………………………………………………… 38 2-5. Immunocytochemistry ………………………………………………… 39 2-6. Caspase-3/7assay……….……………………………………………… 39 2-7. Annexin V-FITC/Propidium Iodide (PI) double-staining……………….39 2-8. Cellular Senescence-Associated β-Galactosidase Assay……………39 2-9. Statistical Analysis …………………………………………………..40 3.Results …………………………………………………………………… 41 3-1. Specific inhibition of EGFR using RNA interference……………………41 3-2. Targeting downstream signaling: Src, PI3K/mTOR, and STAT3…………44 3-3. Ligand-independent modulation using HSP90 inhibitor…………….. 47 3-4. Epigenetic modulation using HDAC inhibitor …………………………47 3-5. Impairment of DNA damage repair following irradiation…………..50 3-6. Mode of cell death……………………………………………………53 3-7. Toxicity towards normal human astrocytes……………………. 59 4.Discussion………………………………………………………………..…62 General discussion………………………………………………………………..…66 References………………………………………………………………70 Abstract in Korean…………..………………………………………83Docto

Clinical study on therapeutic effects of expanded polytetrafluoroethylene membrane and cacium carbonate in infrabony defects

치의학과/박사[한글] 치주질환으로 인해 상실된 치주조직의 재생을 위해 이용되는 치주조직유도재생술과 골이식재를 이용한 치료효과를 비교하고자 expanded polytetrafluoroethylene (e-PTFE) 차단막과 골이식재인 porous resorbable calcium carbonate를 함께 사용하여 술 후 6개월간 치료에 대한 임상적 효과를 54명의 만성 치주염으로 진단된 환자의 71개의 골내낭을 대상으로 치은박리수술을 시행한 20개의 골내낭을 대조군으로, e-PTFE 차단막을 사용하여 치은박리수술을 시행한 20개의 골내낭을 실험 Ⅰ군으로, e-PTFE 차단막과 골이식재인 porous resorbable calcium carbonate를 함께 사용한 16개의 골내낭을 실험 Ⅱ군으로, 골이식재인 porous resorbable calcium carbonate만 이식한 15개의 골내낭을 실험 Ⅲ군으로 설정하였다. 술 전과 술 후 6 개월 후의 치료효과를 치주낭탐침 깊이, 부착상실, 탐침골 깊이, 치은퇴축의 변화로 비교하였으며 이러한 변화에 미치는 임상지수들간의 상관관계를 알아보아 다음과 같은 결과를 얻었다. 1. 치주낭탐침 깊이의 변화는 대조군과 실험군 모두 술 후 6개월 후에 유의성 있는 감소를 보였다(p<0.01). 대조군에 비하여 실험 Ⅰ군(p<0.01), 실험 Ⅱ군(p<0.05)에서 유의성 있는 더 많은 감소를 보였다. 2. 부착상실의 변화는 대조군과 실험군 모두 술 후 6개월 후에 유의성 있는 감소를 보였으며 (p<0.01) 대조군에 비하여 실험 Ⅰ, Ⅱ군에서 유의성 있는 더 많은 감소를 보였다(p<0.01). 3. 탐침골 깊이의 변화는 대조군에서는 유의성 있는 변화를 보이지 않았으나 실험 Ⅰ, Ⅱ, Ⅲ군에서는 술 후 6개월후에 유의성 있는 감소를 보였다(p<0.01). 4. 치은퇴축의 변화는 대조군과 실험 Ⅰ, Ⅲ군에서는 술 후 6개월후에 유의성 있는 증가를 보였다(p<0.01). 실험 Ⅱ군에서는 유의성 있는 증가를 보이지 않았다. 5. 실험 Ⅰ, Ⅱ, Ⅲ군간의 비교에서는 치주낭탐침 깊이, 부착상실, 탐침골 깊이, 치은퇴축의 변화에서 통계학적으로 차이가 없었다. 6. 각 임상지수간의 상관관계에서는 초기 치주낭탐침 깊이에 대한 치주낭탐침 깊이의 변화량및 부착상실의 변화량은 대조군과 실험군 모두 상관관계가 있었으며 (p<0.05), 치주낭탐침 깊이의 변화와 부착상실의 변화(p<0.05), 부착상실의 변화와 탐침골 깊이의 변화는 대조군과 실험군 모두에서 상관관계를 보였다(p<0.05). 이상의 결과로 치주염으로 인한 골내낭 치료시 e-PTFE차단막을 사용한 군에서는 치주낭탐침 깊이와 부착상실의 변화에, e-PTFE 차단막, 골이식재 그리고 이들을 함께 사용한 군에서는 탐침골 깊이의 변화에 좋은 효과를 보였으므로 치주조직의 재생을 위한 치료효과 를 높이기 위해서는 e-PTFE 차단막을 사용하는 것이 좋으리라 사료된다. [영문] The ultimate goal of periodontal therapy is promoting the regeneration of lost periodontal tissue. The purpose of this study is to compare the effect of treatment using expanded polytetrafluoroethylene (e-PTFE) as a barrier membrane in conjunction with porous resorbable calcium carbonate as a bone graft material. 71 infrabony defects from 54 patients with clinical diagnosis of chronic periodontitis were selected among those 20 defects were treated via flap operation only and designated as the control group, another 20 defects were treated with e-PTFE barrier membrane covering via flap operation and designated as the experimental group 1, 16 defects were treated with e-PTPE barrier membrane covering and porous resorbable calcium carbonate grading, and designated as the experimental group Ⅱ, 15 defects were treated with porous resorbable calcium carbonate grafting via flap operation. Clinical parameters including probing depth, loss of attachment, probing bone level and gingival recession have been recorded at 6th months, and the significance of the changes has been analyzed. Correlation coefficients among these clinical parameters were analyzed. The results are as follows : 1. Probing depth were reduced with statistically significance (p<0.01). Experimental group Ⅰ (p<0.01) and experimental group Ⅱ (p<0.05) showed statistically significant higher decrease compared to the control group. 2. Loss of attachment showed statistically significant decrease in both control and experimental groups Ⅰ, Ⅱ, Ⅲ(p<0.01) and experimental group Ⅰ and Ⅱ showed statistica 3. Probing bone level showed insignificant decrease in the control group, but experimental group Ⅰ, Ⅱ, Ⅲ showed statistically significant decrease. (p<0.01) 4. Gingival recession showed statistically significant increase in the control group and experimental group Ⅰ, Ⅲ (p<0.01), but no statistically significant mental group Ⅱ. 5. Comparisons among the experimental groups showed no statistically significant changes in all parameters. 6. Correlations among the changes of probing depth and loss of attachment to the initial probing depth skewed statistically significant in all groups (p<0.05), and the clinical parameters showed statistical significance between probing depth and loss of attachment (p<0.75), and between loss of attachment and probing bone level in all groups. (p<0.05) On the bases of these results, treatments using expand ed polytetrafluoroethylene( e-PTFE) membrane promote changes on probing depth and loss of attachment in infra-bony defects, and using e-PTFE membrane, bone graft material, and e-PTFE membrane in conjunction with bone graft material changes on probing bone level is promoted. Therefore using e-PTFE membrane increases the efficiency on therapeutic effects for periodontal regeneration.restrictio