DEVELOPTMENT OF HYDROGEL-SOLID HYBRIDS FOR ELECTRO-MICROFLUIDICS AND SINGLE CELL ANALYSIS

학위논문 (박사) -- 서울대학교 대학원 : 공과대학 협동과정 바이오엔지니어링전공, 2020. 8. 권성훈.Agarose and other noncovalent hydrogels have good biocompatibility but their applications were restricted since they tend to have low interfacial bonding strength with other polymers or solids. Previously introduced noncovalent hydrogel-to-solid fixation strategies relied heavily on mechanical clamping which is a temporary approach and difficult to apply to kinetic parts or morphologically non-trivial adhesions. Here, we introduce a facile method that increased interfacial bonding strength of agarose hydrogel against solids via an interface-toughening hydrogel. The method showed applicability to several other noncovlanet hydrogels as well, including gelatin, alginate, agar, and chitosan. The bonding method requires no mechanical clamping, liquid glue or bulk modification of the noncovalent hydrogels polymer backbone. It is also compatible with forming micropatterns within the bonding interface. With this new bonding technique, we were able to fabricate various noncovalent hydrogel-solid integrated structures with novel functionalities for in vitro assay, soft robotics and biologically inspired systems.아가로스를 비롯한 noncovalent 하이드로겔들은 biocompatibility가 좋은 반면에 다른 고체 표면과의 접착력이 약해 그 활용성이 낮은 편이었다. 기존의 noncovalent 하이드로겔 접착 방법은 주로 기계적 고정방법에 많이 의존했는데 이는 일시적인 접착일 뿐이고, 동적 부품이나 복잡한 표면에는 적용이 어려웠다. 본 논문에선 접착면 toughness 증강을 도모하는, 그리고 활용성, 범용성이 좋은 하이드로겔 접착방법을 제시한다. 본 접착방법은 gelatin, alginate, agar, 그리고 chitosan등의 noncovalent hydrogel에 대해 적용 가능하다는 것을 보여줬다. 본 방법은 기계적 고정이 전혀 필요 없고, 액상 접착제나 하이드로겔 polymer backbone 수정을 요구하지 않는다. 또한 접착표면상에 미세구조들을 유지할 수 있다. 이 접착방법을 사용해서 전기미세유체, 단일세포전사체분석 등의 활용 예시들을 보여줬다. 본 접착방법은 이 외에도 in vitro 어세이, 소프트 로보틱스, 생체모방 등의 분야에 활용 가능할 것으로 예상한다.Chapter 1 １ Chapter 2 ６ 2.1 Fabrication of hybrid hydrogel films ７ 2.2 Fabrication of hybrid hydrogel and noncovalent hydrogel double layer structure ８ 2.3 Performing various hydrogel-to-hybrid gel bonding ８ 2.4 Agarose hydrogel to tough hydrogel bonding procedure. ９ 2.5 Preparing solids and elastomers to bond with hybrid gels. １０ Chapter 3 １１ 3.1 FTIR measurement of imine bond formation １２ 3.1.1 Sample preparation for FTIR measurement １２ 3.1.2 FTIR Measurement result １２ 3.2 13C-NMR chemical shift measurement １５ 3.2.1 Sample preparation for NMR measurement １５ 3.2.2 NMR measurement result １６ 3.3 SEM/EDS measurement of monomer diffusion layer １９ 3.3.1 Sample preparation for SEM measurement １９ 3.3.2 SEM measurement result １９ 3.3.3 EDS measurement result ２１ Chapter 4 ２５ 4.1 Bonding strength measurement ２６ 4.1.1 The effect of monomer concentration ２６ 4.1.2 The effect of agarose chain aldehyde modification ３０ 4.1.3 The effect of monomer diffusion ３１ 4.1.4 Fracture energy analysis ３３ 4.2 Noncovalent hydrogel to solid bonding ３５ 4.2.1 Noncovalent hydrogel to solid surface bonding ３５ 4.2.2 Noncovalent hydrogel to elastomer surface bonding ３７ 4.2.3 Noncovalent hydrogel to tough hydrogel bonding ４１ Chapter 5 ４５ 5.1 Zig-free hydrogel microfluidic system ４６ 5.2 Electrophoretic oligonucleotide retrieval system ５２ 5.3 Discussion ５７ Chapter 6 ５９ 6.1 Introduction of the field and the proposed approach ６０ 6.2 Device design ６２ 6.2.1 Optimization of cell assembly protocol ６６ 6.2.2 Optimization of electrophoretic mRNA capture protocol ７０ 6.2.3 Crosslinking mRNA capturing probe onto magnetic microparticles ７４ 6.2.4 Optimizing RT-PCR protocol for single cell or small number of cells using mouth pipetting ７５ 6.2.5 Critical limitation of the approach ８０ Chapter 7 ８１ 7.1 Single cell electrophoresis protocol optimization ８２ 7.1.1 Optimization of barcoded mRNA-capturing microparticle synthesis ８２ 7.1.2 Optimization of cell assembly and bead assembly ８７ 7.1.3 Optimization of electrophoretic mRNA capture protocol ９３ 7.2 Single cell RNA retrieval demonstration ９６ 7.2.1 Single cell mRNA retrieval test ９６ 7.2.2 Bead harvest and RT-PCR １０５ 7.2.3 Validation using Sanger sequencing １０７ 7.2.4 Discussion １１０ Chapter 8 : Summary １１５ Bibliography １１７ Abstract(국문초록) １２０Docto

Effect of Hysterectomy on conserved Ovarian function

Objective : To assess the impact of premenopausal total abdominal hysterectomy (TAH) on the function of remaining ovaries by reviewing the menopausal age in patients treated with total abdominal hysterectomy. Methods : We retrospectively reviewed medical records of 510 women previously treated with TAH or TAH with unilateral salpingo-oophorectomy (USO) due to benign disease at the department of Obstetric and Gynecology, Yonsei University College of Medicine, between Jan, 1989 and Dec, 1992. Out of 510 women, 94 women who were thoroughly follwed up were included in the study, and their meopausal age based on patients symptoms were compared to those of the control group. Result : Mean menopausal age in patients treated with TAH was 46.3±3.0 years and in the normal control group was 49.1±3.2 years. The mean age of menopause was significantly lowered in patients treated with TAH/TAH with USO (P<0.001). There was positive correlation between age at operation and menopausal age. Conclusion : According to this study, TAH accelerated ovarian dysfunction, and younger the patient is at the time of operation, earlier the menopause. Therefore, woman treated with TAH is at risk of early menopause and should receive adequate hormone replacement therapy.ope

Comparison of Burch Colposuspension, Pubovaginal Sling Operation and Tension-Free Vaginal Tape for Surgical Treatment of Stress Urinary Incontinence In Women

Objective : The object of this study was to eompare the cure rate and confirm the clinical efficacy of three most frequent surgical procedures for stress urinary incontinence (Burch colposuspension, pubovaginal sling operation, tension-free vaginal tape). Materials and Methods : We collected datas from the records of ninety-one patients who were diagnosed as stress urinary incontinence from Jan. 1999 to May 2001. Burch colposuspension was performed by department of gynecology, Severance hospital in thirty-three patients, pubovaginal sling operation was performed by department of urology in twenty-eight patients, and tension-free vaginal tape was performed by department of urology in thirty-one patients. We investigated the characteristics of patients, preoperative urodynamic study results, cure rates and complication rates for the result, and compared them by χ^(2)-test. Results : There were statistically no significant differences between the cure rate of each operation after 3,6 month of operation but after 12 months of follow up, the cure rate of pubovaginal sling operation was significantly higher than that Burch operation and tension-free vaginal tape. Conclusion : The cure rate of pubovaginal sling operation was significantly higher after 12 months of follow up after surgery. There was no significant difference between cure rates of Burch operation and tension- free vaginal tape. We propose randomized prospective study with larger population in the future. Key Words : Stress urinary incontinence, Burch colposuspension, Pubovaginal sling operation, Tension-free vaginal tape.ope

Translation and linguistic validation of Korean version of short form of pelvic floor distress inventory-20, pelvic floor impact questionnaire-7

The short forms of pelvic floor distress inventory (PFDI) and pelvic floor impact questionnaire (PFIQ) are useful disease specific questionnaires evaluating symptoms, quality of life for pelvic floor disorders. The purpose is to develop linguistic validation of the PFDI-20 and PFIQ-7 questionnaires. Three types of Korean version of questionnaires have been used in four locations of University Hospitals in Korea. Each version of questionnaires was developed by forward translation and back-translation by bilingual translators and was verified by the patients with pelvic floor disorder and healthy persons. For harmonization of 3 types of questionnaires, four authors reviewed, discussed all discrepancies, incorporated and produced a new version. The multi-step processes of translation and linguistic validation of the Korean version of PFDI-20 and PFIQ-7 questionnaire were completed. Further process of validation of Korean version of these questionnaires is required.ope

In Bae Yoon, an Inventor and Pioneer in Laparoscopic Surgery

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Jessie W. Hirst, a Professor Who Challenged Prejudice towards Western Medicine

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Study on endometriosis in an adolescent population

Purpose : To review diagnostic procedure, clinical stage, age distribution, treatment of endometriosis in adolescents. Material and Method : We retrospectively reviewed medical records of 39 adolescent girls(11-21) admitted to Yonsei University College of Medicine between 1990 and 1999. We identified 39 patients who underwent laparotomy or laparoscopy and was diagnosed as having endometriosis. Endometriosis was classified according to the Revised American Fertility Society Classification(AFS). The chief symptoms leading to diagnosis, clinical stage, age distribution, and treatment modality were reviewed. Results : Average age of menarche was 14.2, and the interval after the menarche was 5.9 years. The chief symptoms leading to diagnosis were chronic pelvic pain(27%), acute pelvic pain(21%), palpable pelvic mass(21%), dysmenorrhea(18%). Laparoscopy was performed in 20 patients(51%). The majority of the patients(44%) presented with stage II, 4(10%) with stage I, 11(28%) with stage Ⅲ, and 7(18%) with stage IV. GnRH agonists(64.1%), expectant managements(25.7%), OCPs(5.1%) and danazol(5.1%) were used after surgery. Conclusion : Adolescents with chronic pelvic pain have a high rate of endometriosis and should be promptly referred to a gynecologist to diagnose the etiological lesion of pelvic pain and initiate appropriate therapy.ope

Urinary Profiles of the Endogenous Steroids in Pre-Menopausal Women with Uterine Leiomyoma

목적 : 본 연구의 목적은 내인성 성호르몬이 자궁근종의 성장에 미치는 영향을 알아보고자 연구를 시행하였다. 연구대상 및 방법 : 폐경전 자궁근종 환자 27명과 같은 연령대의 정상여성 25명을 대상으로 하였다. 모든 대상군의 여성에서 24시간 소변을 모아서 소변내 estrogen, androgen 의 대사체들을 GC-MS를 이용하여 측정하였으며 두 군에서의 차이를 비교분석 하였다. 결과 : 소변내 17β-estradiol, 5-androstene-3β, 16β, 17β-triol, 11-keto-ethiocholanolone, 11β-hydroxyandrosterone, THS, THA, THE, α -cortolone, α-cortol 및 β-cortrol 가 환자군에서 의의있게 증가하였으며 17β-estradiol/estrone 및 11β-hydroxy-ethiocholanolone/11β-hydroxy-androsterone도 환자군에서 의의있게 증가하였다. 결론 : 자궁근종의 성장은 요중 estrogen과 androgen의 농도와 밀접한 관련이 있으며 이는 환자의 스테로이드 호르몬 대사 감소에 기인한 것으로 사료된다.ope

Correlation between Steroid Hormone Metabolites and Leiomyomas of Uterus

Objective: To elucidate 1) whether there are any differences in the urine concentrations of steroid hormone metabolites between patients with leiomyoma and normal controls 2) the correlation between urinary profiles of steroid hormones and leiomyomas of the uterus according to their type, location, volume, and weight. Materials of Methods: The study population consisted of 37 premenopausal patients with uterine leiomyoma and the control group consisted of 25 premenopausal normal volunteer women without uterine leiomyoma. Confirmation of the existence of uterine leiomyoma was done by ultrasonography and histopathological examination after surgery. The volume of the leiomyoma was estimated by trans-abdominal and/or trans-vaginal ultrasonography. The Leiomyomas were divided into 3 types (subserosal, intramural and submucosal). Seventeen patients had subserosal type of leiomyoma, 10 with the intramural type and 10 with the submucosal type. The locations of the leiomyoma were also divided into 3 groups (fundus, body and isthmus). Seventeen patients showed a fundus location, 10 in body, and 10 in isthmus. We compared urinary profiles of the endogenous steroids between patients with leiomyomas and normal controls, and also investigated the relationship between urinary profiles of the endogenous steroids and leiomyomas according to their type, location, volume and weight by using highly sensitive Gas Chromatography-Mass Spectrometry (GC-MS) system. Results: The mean ages of the patients with leiomyomas and the control group were 43.1 ±5.6 and 40.6 ±7.2 years, the weights were 63.4 ±7.3 and 59.4 ±8.1 ㎏, and their heights were 155.4 ±4.8 and 159.3 ±4.8 cm respectively. Seventeen patients had subserosal, 10 had intramural, and 10 had submucosal leiomyomas. There were 17 patients with leiomyoma located in fundus, 10 in body and 10 in isthmus. 17ß-estradiol, 5-AT, 11-keto ET, 11ß-hydroxy An, 11ß-hydroxy Et, THS, THA, THE, a-cortolone, a-cortol, ß-cortol, 11ß-OH Et/11ß-OH An and E2/E1 were significantly increased in patients with leiomyoma than in the control group. 17ß-estradiol was significantly increased in the intramural and the submucosal types than in the subserosal type. There was no significant difference in the concentrations of urinary steroids according to the locations of leiomyomas. There was no significant relationship between the concentration of urinary steroids and the volume of the leiomyomas. 17ß-estradiol significantly decreased as the weight of uterus increased (r=-0.322, p=0.04). Conclusion: The concentrations of steroid hormone metabolites were generally increased in patients with leiomyoma but were not significantly related to the volume and weight of the leiomyomas. Our study suggests that steroid hormones may be involved in the initiation of leiomyomas but may not be involved in their progression. In addition, the concentrations of steroid hormone metabolites are not related to the leiomyoma type and location.ope

Pregnancy Outcomes in Women with Unexplained Elevation of Maternal Serum Human Chorionic Gonadotropin Levels at Midtrimester

Objective : Our purpose was to determine the association between unexplained elevation of maternal serum human chorionic gonadotropin (hCG) in the second trimester and adverse pregnancy outcomes. Material and methods : Between February 1995 and July 1999, we evaluated 1566 pregnant women who have underwent second trimester triple marker screening tests (alpha-fetoprotein, unconjugated estriol, human chorionic gonadotropin) and delivered at Severance Hospital, Yonsei Medical Center. Multiple pregnancies, abnormal fetal karyotypes, fetal anomalies, and abortions were excluded from the study. One hundred twenty-one women with hCG levels greater than 2.0 multiples of the median (MoM) were included in the study group while 1389 women with hCG levels less than 2.0 MoM served as the control group. Pregnancy outcomes were obtained from the delivery and neonatal records in our institution. Adverse pregnancy outcomes between the two groups were compared using chi-square test and Fisher's exact test. Results : Women with unexplained elevation of human chorionic gonadotropin levels were associated with statistically significant increased risks for preeclampsia, preterm delivery, and low birth weight (p<0.05). However, there were no significant differences between the study and control groups with respect to preterm premature rupture of membranes, abnormal fetal heart rate tracing, abruptio placentae, intrauterine fetal death, and neonatal death. Conclusion : An unexplained elevation in human chorionic gonadotropin level in the second trimester may increase the risk for preeclampsia, preterm delivery, and low birth weight but not for other adverse pregnancy outcomes such as preterm premature rupture of membranes, abnormal fetal heart rate tracing, intrauterine fetal death, or neonatal death.ope