Construction of Recombinant Adeno-Associated Virus Vector (AAVCMVp53) for Human Cervical Cancer Gene Therapy / 대한산부인과학회잡지

OBJECTIVE: To eliminate the potential problem of adenovirus contamination during recombinant adeno-associated virus (rAAV) vector production, we investigated new rAAV production method by a triple transfection of vector plasmid, packaging plasmid, and adenovirus helper plasmid. METHODS: This study was carried by triple transfection for the production of recombinant adeno-associated virus vector. This new production system was conducted with a plasmid construct which contained a mini-adenovirus genome capable of propagation of rAAV in the presence of adeno-assoceated viral rep and cap genes. To examine the helper functions of adenoviral plasmid on the production of rAAV vector, we carried out cotransfection with three plasmids, AAV vector, packaging construct, and adenovirus helper plasmids. The optimized plasmid quantity of transfection by calcium phosphate precipitation method was 25 micro gram of total plasmid DNA per 10 cm diameter plate of 293 cell. RESULTS: We found that rAAV yields peaked at 48 hr after Ad infection. The titer of rAAV was measured by the dot blot analysis to measure the number of particles/mL based on the quantification of viral DNA. CONCLUSION: We constructed recombinant AAVp53 without adenovirus contamination. We thought that we construct high titer rAAVp53 particle through HPLC column in the near future.

Clinical Analysis of Ovarian Pregnancy / 대한산부인과학회잡지

OBJECTIVE: The frequency of ectopic pregnancy has been recently increased by various causes. Especially in ovarian pregnancy, definitive preoperative diagnostic criteria have not been established. This study is aimed to focus on helpful clinical characterstics which can later on serve as diagnostic key points in case of suspicious ovarian pregnancy. METHODS: The study included 23 patients pathologically diagnosed with ovarian pregnancy from January 1991 to December 2002 in Kangnam St. Mary Hospital. We analyzed the age at diagnosis, pregnancy and delivery history, presence of underlying disease, clinical symptoms, therapeutic methods, human chorionic gonadotropin level, and sonographic findings for these patients. RESULTS: The average age at diagnosis was 31.6 year and 39.1% of the patients were older than the age of 35. More than half of the patients had the history of multiple pregnancy (52.2%). The common clinical symptoms were lower abdominal pain (95.6%) and vaginal bleeding (21.8%). Ipsilateral ovarian wedge resection was performed at the most frequent rate (78.3%) and ipsilateral adnexectomy was done at second most frequent rate (21.7%). HCG level was measured on 19 cases showing various individual difference. Most common sonographic findings included the fluid stasis surrounding the ovarian pregnancy lesions and ipsilateral ovary enlargement (91.3%). CONCLUSION: This study showed that ovarian pregnancy should be suspected in young woman. Of the age between 25 and 30 years, with symptom of lower abdominal pain, nulliparity, havior history of pelvic adhesion, pelvic inflammation, and showing sonographic findings of fluid collection around ovary or enlarged ovary.

The Efficacy of Lamellar Body Counts for Prediction of Perinatal Outcome / 대한산부인과학회잡지

OBJECTIVE: This study is aimed to evaluate whether lamellar body count (LBC) in amniotic fluid could be used as a predictor of neonatal morbidity as well as respiratory distress syndrome (RDS) and to determine the value of lamellar body count that maximizes sensitivity and specificity. METHODS: We conducted a prospective clinical outcome study. Amniotic fluid was obtained from 39 pregnant women at various gestational ages (29 to 36 weeks) from March 1, 2002 to February 28, 2003. They delivered within 72 hours of amniocentesis, excluding 6 cases of contaminated amniotic fluid. A LBC was performed on each specimen. The frequencies of RDS, minor and major morbidity of newborn in different LBC cutoff value were analyzed. Statistic analysis was done by Chi-square test. RESULTS: LBCs increased with gestation (r=0.533, p<0.05). The LBC cutoff value that best agreed with RDS (sensitivity 36.4%, specificity 93.8%), minor morbidity (sensitivity 100.0%, specificity 50.0%) and major morbidity (sensitivity 45.5%, specificity 81.3%) was 30,000/ l. CONCLUSION: These results indicate that LBC might be useful as a predictor of neonatal morbidity as well as respiratory distress syndrome.

Induction of Anti-tumor Immunity by Vaccination with E7 of HPV16 and IL-12 / 대한산부인과학회잡지

OBJECTIVE: Interleukin-12 is well known to induce cellular immune response materials and suppress the tumor growth. HPV infection has significant roles in cervical carcinogenesis, and HPV oncoprotein E6 and E7 are important roles in formation and maintenance of cervical cancer. E7 specific immune response was detected in cervical cancer patients, and this shows that E7 protein would be important in potential immunetherapy in cervical cancer. This study is aimed to investigate antitumor effect and E7 immune response by injection of adenovirus IL-12 and E7 in cervical cancer animal model. METHODS: In the cervical cancer animal model using C57BL/6 mice and HPV16 E7 immortalized hosts, 5 X 10(8) pfu/100 ul of PBS, AdLacZ, AdE7 and AdIL-12 were injected into the tumor mass when the tumor sized is increased to 7-8 mm. After the injection, the tumor size was caliperated every 2-3 days, and pathologic and blood studies were done on day 1, 3, 5, 7, 11, 12, and 21 days. The expression level of IL-12 and INF- and E7 specific immune response were measured by ELISA. RESULTS: After the injection of AdIL-12 into the tumor mass, 45% of tumor growth suppression was noted in comparison with control group. In the cases of combination injections of AdIL-12 and AdE7, 80% growth suppression was observed, and complete regression was shown in 40% of the study group. After injection of AdIL-12, the expression of IL-12 in the tumor mass was 9 time higher than that of control group, and 6 times higher in blood sample in comparison with control group. In the group with combined AdIL-12 and AdE7, the highest expression of INF- was noted in comparison with single injection of AdIL-12 or control group. IgGI and IgG2b isotype expression level increased 2.5 times and 2.2 times respectively 3 weeks after adenovirus injection. CONCLUSION: In cervical cancer animal model, IL-12 and E7 application using Adenovirus vector is significant antitumor effect and this demonstrates the potential immunotherapy in near future.