염증성 관절염에서 NO 의 역할에 관한 연구

배상철(Sang Cheol Bae),김동욱(Dong Yook Kim),김태환(Tae Hwan Kim),전재범(Jae Bum Jun),정성수(Sung Soo Jung),이인홍(In Hong Lee),유대현(Dae Hyun Yoo),김성윤(Seong Yoon Kim),이은영(Eun Young Lee),장성렬(Sung Yeoul Chang) 대한내과학회 1997 대한내과학회지 Vol.52 No.1

N/A Objectives: Nitric Oxide(NO) is a toxic, inorganic, gaseous free radical produced during the metabolism of L-Arginine by NO synthase(NOS). It has been implicated in a rapidly growing number of physiological and pathophysiological processes such as cytotoxic effects against microbes and tumor cells, blood vessel dilation and neurotransmitter. Recently there is growing evidence implicating NO in immune regulation, inflammation, autoimmunity, and arthritis. We performed this study to determine a role for nitric oxide in inflammatory arthritis especially rheumatoid arthritis(RA). Methods: We measured (D the concentrations of nitrite, a breakdown product of nitric oxide, in serum and synovial fluid from patients with RA and osteoarthritis(OA) and in the serum of controls ② the concentrations of nitrite in the supernatant of cultured synovial tissue with RA and OA and ③ determined whether human chondrocytes and synoviocytes can synthesize nitric oxide and if so, how production is regulated by cytokines and antirheumatic drugs. Results: 1) Serum nitrite concentrations in patients with RA and OA were higher than in controls. In both disease groups synovial fluid nitrite was higher than serum nitrite. Serum and synovial fluid nitrite concenrations in RA were higher than those in OA. However, those findings are not statistically significant. 2) Although these findings are not statistically significant, the concentration of nitrite in the supernatant of cultured synavial tissue with RA was higher than that in OA. 3) IL-l5 and TNF-a induced the biosynthesis of NO by chondrocytes and synoviocytes. IGF-1 and TGF-5 failed to provoke the production of NO. The biosynthesis of NO required an induction period of approximately 6 hours and was inhibited by L- NMMA and cycloheximide. Dexamethasone, indomethacin, gold sodium thiomalate and methotrexate had no effect on the induction of NO biosynthesis. Conclusion: These results suggest a role for nitric oxide as an inflommatory mediator in inflammatory arthritis.

부인암 세포주에서 항암제 투여에 따른 신생혈관 형성 관련 유전자 전령 리보핵산 및 단백질 표현 양상에 관한 연구

김영재 ( Young Jae Kim ),공미숙 ( Mi Sook Kong ),이영미 ( Young Me Lee ),장성렬 ( Sung Yeoul Chang ),김승룡 ( Seung Ryong Kim ),김경태 ( Kyung Tai Kim ),황윤영 ( Youn Yeung Hwang ),조삼현 ( Sam Hyun Cho ) 대한산부인과학회 2007 Obstetrics & Gynecology Science Vol.50 No.3

연구목적: 본 연구자는 확립된 세포주에서 세포독성제 및 면역 조정제에 의한 항신혈관 형성효과를 확인하고자 하였다. 유방암 세포주 MCF-7, 자궁 경부 편평 상피암 세포주 SiHa, 난소암 세포주 A2780, 그리고 cisplatin 내성 난소암 세포주 A2780-CDDP에서 cisplatin, paclitaxe l(taxol), 및 thalidomide로 처리한 후에 vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), 그리고 thrombospondin-1,2 (TSP-1,2)의 전령 리보 핵산 및 단백질의 표현 양상의 변화를 규명하고자하였다. 연구대상 및 방법: 유방암 세포주 MCF-7, 자궁 경부 편평 상피암 세포주 SiHa, 난소암 세포주 A2780, 그리고 cisplatin 내성 난소암 세포주 A2780-CDDP에서 cisplatin, paclitaxel (taxol), 및 thalidomide로 각각 처리한 후에 vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), 그리고 thrombospondin-1,2 (TSP-1,2)의 전령 리보 핵산 및 단백질의 표현 양상의 변화를 규명하고자 RT-PCR, 단백 추출 및 Western blot assay를 사용하였다. 결과: 난소암 세포주 A2780과 동일 세포주에서 분리한 cisplatin 내성 주 A2780-CDDP, 유방암 세포주 MCF-7 및 자궁경부 편평 상피암 세포주인 SiHa에서 VEGF gene은 212 bp, bFGF는 238 bp, 그리고 TSP-1은 492 bp로 합성되었으나 TSP-2 는 합성되지 않았으며 VEGF 및 bFGF 단백질은 각각 21 KDa와 17 KDa로 합성되었으나 TSP-1 단백질은 합성되지 않았다. 각 세포주의 VEGF gene의 발현 정도는 cisplatin, taxol 및 thalidomide에 별 큰 영향을 받지 않았다. bFGF gene의 발현 정도는 taxol처리 후 유방암 세포주에서 만 감소하였으며 TSP-1 gene의 발현도는 taxol 처리 후 유방암 세포주에서 자궁경부암 세포주 보다 큰 폭의 감소가 있었고 난소암 세포주에서 thalidomide처리 후 뚜렷한 감소를 보였다. Cisplatin, taxol, 그리고 thalidomide에 VEGF 및 bFGF 단백질 발현도는 아무런 영향을 받지 않았다. 결론: 난소암 세포주 A2780, cisplatin 내성 난소암 세포주 A2780-CDDP, 유방암 세포주 MCF-7, 그리고 자궁경부 편평 상피암 세포주 SiHa에서 VEGF, bFGF 그리고 TSP-1 gene들 및 단백질의 발현 양상은 cisplatin 처리에 의해 변화되지는 않는 것으로 확인되어 난소암, 유방암 그리고 자궁경부암에서 cisplatin의 신혈관 형성 억제 효과 판정에는 VEGF, bFGF 및 TSP-1 등의 직접적인 단백 발현 양상만을 표적으로 삼기 보다는 기저층 파괴를 담당하는 신호 전달체등의 발현 변화를 동시에 분석하여야 할 것으로 사료되며, 통계적으로 유의하지는 않았지만 taxol 처리에 따른 신혈관 형성 단백 유전자들의 발현 양상에 있어 bFGF가 유방암 세포주에서 약간 감소되었고, TSP-1은 유방암 세포주에서 자궁경부암 세포주보다 약간 더 많은 감소를 보여 bFGF와 TSP-1은 유방암 및 자궁경부암에서 taxol의 신혈관 형성 저해 연구의 평가 기준으로 재연구될 필요가 있을 것으로 생각된다. 역시 통계적으로 유의한 정도는 아니었으나 Thalidomide 처리에 따른 전기 단백질들의 표현 양상에는 난소암 세포주 A2780에서 TSP-1 단백 발현의 감소가 다른 세포주보다 많은 것으로 나타나, 실험 방법에 따라 같은 세포 독성제라도 신혈관 형성 저해 효과 판정이 다르다는 것을 확인시켜 줌과 동시에 향후 thalidomide를 난소암의 보조적 치료제로 이용하는 연구의 필요성을 제시하였다. Objective: Angiogenesis is central role to both the proliferation and metastasis of malignant tumor. The intense interest in angiogenesis has also lead to a re-examination of the activity of established cytotoxic agents which are known to be an antiangiogenic effect anecdotally. In this study, anti-angiogenic effect of cisplatin, paclitaxel and thalidomide was evaluated in human ovarian cancer cell lines and cervical cancer cell line. Methods: Human ovarian cancer cell line A2780, cisplatin resistant human ovarian cancer cell line A2780-CDDP, human breast cancer cell line MCF-7, and squamous cell uterine cervical carcinoma cell line SiHa were used to evaluate the level of mRNA and protein expression of VEGF, bFGF and TSP-1, 2 before and after the treatment with cisplatin, paclitaxel, and thalidomide using RT-PCR, protein extraction, and Western blot. The results were analyzed with Wilcoxon signed rank test in the SAS ver 8.1. Results: Targeted mRNAs were synthesized as 212 bp VEGF, 238 bp bFGF, and 492 bp band sized except mRNA of TSP-2 via RT-PCR. The protein of VEGF and bFGF were appeared as 21KDa and 17 KDa size, however, the protein of TSP-1 was not appeared through western blot. No effect of cisplatin on protein expression was measured in these cell lines, but paclitaxel influenced the expression of bFGF in MCF-7 cell line and the expression of TSP-1 in MCF-7 and SiHa cell lines. TSP-1 expression was influenced by thalidomide in A2780 cell line. The protein expression of VEGF and bFGF were not influenced following treatment with cisplatin, paclitaxel, and thalidomide. Conclusion: These results were suggested that bFGF and TSP-1 will be used as a target gene for the assay of antiangiogenic effect of paclitaxel in breast and uterine cervical cancer tissue and TSP-1 will be used as that of thalidomide in ovarian cancer. Furthermore, thalidomide will be tried as an adjunctive agent for the improvement of the survival in the case of the patient with ovarian cancer.

퇴행성관절염 환자에서 제2형 콜라겐 유잔자의 점돌연변이에 관하 연구

김태환 ( Tae Hwan Kim ),홍관표 ( Kwan Pyo Hong ),전재범 ( Jae Bum Jun ),정성수 ( Sung Soo Jung ),이인홍 ( In Hong Lee ),배상철 ( Sang Cheol Bae ),유대현 ( Dae Hyun Yoo ),이은영 ( Eun Young Lee ),장성렬 ( Sung Yeoul Chang ),김성윤 대한류마티스학회 1997 대한류마티스학회지 Vol.4 No.1

목적: 퇴행성관절염(Osteoarthritis)은 관절연골의 퇴행성 변화를 특징으로 하는 가장 흔한 류마티스 질환의 하나로 전 인구의 약 15%를 차지한다. 이 질환의 원인으로는 비만, 직업으로 인한 관절의 빈번한 사용, 외상, 대사 장애, 선천적 질환과 동반된 경우 등이 밝혀져 있지만 아직까지 정확한 원인은 밝혀지지 않은 상태이다. 제2형 콜라겐은 연골에 가장 풍부하게 존재하는 물질로 유전적 이상이 퇴행성관절염을 일으킨다고 알려져 있다. 이에 전자는 비교적 이른 나이에 발생하고, 가족력이 있는 수부관절염 환자에서 제2형 콜라겐 exon 31 유전자의 점돌연변이를 관찰하고자 하였다. 방법: 환자의 말초혈액에서 genomic DNA를 얻고 분리된 genomic DNA를 주형으로 하고 제2형 콜라겐의 exon 31번의 시발체를 이용하여 PCR을 시행하였다. PCR산물을 제한효소 Dsa I와 반응시켜 C가 T로 변이된 위치를 2% agarose gel상에 전기영동하여 확인하였다. 결과: 퇴행성관절염 환자를 대상으로 제2형 콜라겐의 exon31번의 점돌연변이를 관찰하였는데 점돌연변이는 관찰되지 않았다. 결론: 제2형 콜라겐의 점돌연변이가 밝혀진 exon11, 48번에 대해 연구를 시행할 예정이며 특히 척추골단이형성증이나 연골이형성증 변화가 있는 환자로 범위를 넓혀갈 예정이다. Objectives: Human osteoarthritis is a heterogeneous and multifactorial disease characterized by the progressive deterioration of the cartilage of diarthrodial joints. In some instances, there is an identifiable cause, such as trauma or congenital malformation. but, mostly the etiology remains unknown. Since familial aggregation is seen, genetic factors may be important, particularly in osteoarthritis of the hand. Methods: Blood samples from patients and controls were obtained for DNA analysis. Exon 31 of type II procollagen gene was amplified by polymerase chain reaction. and screening for the mutation was undertaken using a restriction enzyme digestion (Dsa I). Results: The patients` phenotype represented typical. but early-onset and family history, osteoarthritis. No mutation in exon 31 of type II procollagen gene could be identified. Conclusion: Screening of the 31 exon did not. however, reveal any mutation. It needs further evaluation in other sites of type II procollagen genes.

난소암 세포주와 cisplatin 내성주에서의 MDR 관련 유전자의 발현에 관한 연구

문영진(Young Jin Moon),황윤영(Youn Yeung Hwang),이재규(Jai Kyu Lee),나영정(Young Jeong Na),이영미(Young Mi Lee),장성렬(Sung Yeoul Chang),조삼현(Sam Hyun Cho),김경태(Kyung Tai Kim) 대한산부인과학회 2001 Obstetrics & Gynecology Science Vol.44 No.9

N/A Objective : Expressions of P-glycoprotein, the multidrug resistance-associated protein (MRP) and the lung resistance protein (LRP) with the MDR phenotype widely divergent in human cancer cell lines. This study focused on the altered gene expression related drug transport. Methods : To examine correlations between MDR-associated genes and PKC isozyme with cisplatin resistance on the level of the mRNA expression, we analyzed MDR-associated gene (LRP, MDR1/P-gp and MRP) expression and PKC isozyme, topoisomerase II alpha and beta in cisplatin-sensitive ovarian cancer cell line A2780 and cisplatin - resistant cell line A2780cp using cDNA-PCR approach. Results : LRP mRNA levels were significantly increased in A2780cp compared to the drug sensitive variant. In contrast, MRP mRNA levels were not significantly correlated with drug sensitivity. A modest increase in PKCη and MDR1/P-gp mRNA expression activity was also observed in ovarian cancer A2780cp cell lines that were resistant to CDDP. The level of topoisomerase II alpha and beta were not affected. Conclusion : These results showed that MDR1/P-gp expression may be an important determinant of the MDR phenotype in cell lines intrinsically resistant to cancer chemotherapeutic agents and a multifactorial emergence of MDR phenotype of tumor with a possible involvement of the PKC isozymes may be associated with CDDP resistant ovarian cancer cell line.

활액세포에서 유도된 apoptosis에 관한 연구

채영규,백승연,박재찬,정성수,김명희,김성윤,최영길,장성렬 漢陽大學校 自然科學硏究所 1997 自然科學論文集 Vol.16 No.-

류마티스 관절염환자의 활액세포에서 스테로이드에 의한 apoptosis가 일어나는지를 DO11.00 세포주와 함께 DNA ladder 형성유무로 검증하였다. DNA ladder 형성은 apoptosis의 특징이나 단지 DO11.00 세포주에서만 확인되었다. 이 결과는 류마티슴 환자의 활액세포는 스테로이드에 의한 apoptosis를 생성시키지 못함을 입증하였다. To detect the apoptotic cell death in rheumatoid synoviocytes, we examined DNA ladder formation with the treatment of steroids, together with DO11.00 cell lines. The DNA ladder formation that is characteristic of apoptosis was detected only in samples from DO11.00 cell lines. The findings indicate that rheumatoid synoviocytes do not undergo steroid-mediated apoptosis.

천포창 환자에서의 혈청 IgA 자가항체의 출현 빈도에 관한 연구

이창우,전재홍,장성렬 대한피부과학회 2000 大韓皮膚科學會誌 Vol.38 No.1

Background:Pemphigus is an autoimmune bullous disease with circulating desmosomal autoantibodies of IgG. In direct IF studies with perilesional tissue, IgA or IgM antibodies can be seen in addition to IgG. Objective:We examined sera of patients with pemphigus for the presence/frequency of IgA autoantibodies as well as IgG by indirect IF and immunoblot assay. Patients:Twenty patients of pemphigus (PV 10, PF 10) who showed positive findings in indirect IF examinations. Methods:Indirect IF study with normal human skin substrates and immunoblot analysis using A431 cell extracts (with multi-step immunostaining) were performed with patients sera. Results:In indirect IF, IgA autoantibodies that bind to the epidermal keratinocyte antigens were detected in 4 cases among the 20 patients (PV 2 and PF 2). In immunoblot analysis IgA bands reacting to PV/PF antigens were observed in 7 cases from the 20 patients with pemphigus (PV 3, PF 4). The serum titers of IgA autoantibodies were lower than those of IgG in every single case. Conclusion:In patients with pemphigus (PV/PF), 35% of cases have serum IgA autoantibodies as well as IgG autoantibodies specific to the pemphigus antigens (Dsg 1/Dsg 3). However, pathogenic roles of the associated IgA autoantibody are not clear.