B 세포에서 Epstein-Barr virus microRNA들의 전사 및 성숙

김도년,오상택,이재면,이원근,이숙경,Kim Do Nyun,Oh Sang Taek,Lee Jae Myun,Lee Won-Keun,Lee Suk Kyeong 한국생명과학회 2005 생명과학회지 Vol.15 No.6

우리는 바이러스로는 최초로 miRNA를 생성한다고 보고된 Epstein-Barr virus (EBV)를 대상으로 EBV miRNA biogenesis를 연구했다. 먼저 EVB에 감염된 B 세포인 B95-8로부터 보고된 5 종의 EBV miRNA들인 BHRF1-1, BHRF1-2, BHRF1-3, BART1, 및 BART2 모두가 발현됨을 확인하였다. 그 다음 성숙된 EBV miRNA 서열로부터 예측되는 pri- 와 pre-miRNA들이 B95-8에서 각각 검출되어 EBV miRNA들도 이미 알려진 동물세포 miRNA와 유사한 생성 및 성숙과정을 거칠 가능성을 확인하였다 게놈 상에 2~7개씩 밀집하여 존재하는 동물세포 miRNA들과 유사하게 EBV 게놈의 2부 위에 밀집해서 존재하는 EBV miRNA들도 polycistronic하게 발현되는지 조사한 결과 B95-8에서 BHRF1-1, BHRF1-2 및 BHRF1-3를 포함하는 1,602 뉴클레오타이드의 긴 전사체가 RT-PCR로 확인되었다. 반면 BART1과 BART2는 mono cistronic하게 전사될 가능성이 확인되었다. 본 연구를 통하여 EBV miRNA들도 동물세포 miRNA들과 유사하게 poly cistronic 하게 발현될 수 있으며 pri-와 pre-miRNA 과정을 거쳐 성숙된 miRNA로 생성될 가능성을 확인하였다. We investigated microRNA (miRNA) biogenesis of Epstein-Barr virus (EBV) which is the first virus shown to produce viral miRNAs. As expected, expression of all the reported EBV miRNAs were detected by Northen blot in an EBV-infected B cell line, B95-8; BHRF1-1, BHIU1-2, BHRF1-3, BART1, and BART2. The putative EBV pri-miRWAs and pre-miRNAs predicted from the known mature EBV miRNA sequences were detected by RT-PCR in B95-8 cells. Many animal miRNA genes exist as clusters of 2-7 genes and they are expressed polycistronically. As the EBV miRNAs are clustered in two regions of the EBV genome, we examined whether these clustered EBV miRNA genes are also expressed polycistronically. A long polycistronic transcript with the expected size (1602 bp) corresponding to the BHRF1-1~BHRF1-2~BHRF1-3 was amplified. However, any polycistronic transcript containing both BART1 and BART2 was detectable in B95-8. These results suggest that EBV miRNAs may be processed in a similar way with animal miRNAs and that some of the clustered EBV miRNAs can be transcribed polycistronically.

종두상 수포증양 병변을 보인 EBV 관련성 림프구 증식증

조광현 ( Kwang Hyun Cho ),이갑석 ( Kap Sok Li ),김연경 ( Yon Kyung Kim ),전윤경 ( Yoon Kyung Jeon ),김철우 ( Chul Woo Kim ),이숙경 ( Suk Kyeong Lee ),문상은 ( Sang Eun Moon ),손숙자 ( Sook Ja Sohn ) 대한피부과학회 2004 대한피부과학회지 Vol.42 No.7

TGF-β1 Expression by Proliferated Keratinocytes in the Skin of E-Irradiated Mice

A-Ran Yoon(윤아란),Do Nyun Kim(김도년),Min Koo Seo(서민구),Sang Taek Oh(오상택),Jung Seon Seo(서정선),Se Mo Jun(전세모),Jung-Ho Cha(차정호),Seungdeok Lee(이승덕),Suk Kyeong Lee(이숙경) 한국생명과학회 2012 생명과학회지 Vol.22 No.2

우리는 방사선피부염 동물 모델을 확립하여, 이차 면역 기관에서의 면역 세포 비율 변화를 관찰하였다. 또한, 방사선 조사에 의한 병소에서 transforming growth factor-β1 (TGF-β1)과 interlukin-10 (IL-10)의 발현을 증가시킨 세포를 분석하였다. Hairless-1 (HR-1) 쥐의 posterior dorsal 부위에 6 일간 매일 10 Gy 씩 electron (E)-ray를 국부 조사하여 방사선피부염 모델을 만들었다. FACS를 이용하여 면역 세포 비율의 변화를 분석한 결과 비장과 림프절에 존재하는 항원제시세포와 T 세포 및 B 세포들의 비율이 E-irradiation에 의해 영향을 받았다. 피부에서 세포 특이적인 마커와 사이토카인들의 발현 양상은 면역형광염색법으로 확인하였다. 방사선 조사 후, TGF-β1과 interlukin-17 (IL-17)은 regulatory T 세포(Treg)보다 keratin-14 (K-14)를 발현하는 진피의 끝부분에서 높게 발현되었다. Interlukin-10 (IL-10)는 Treg 뿐만 아니라 T helper 17 (Th17) 세포, dendritic 세포, macrophage 중 어느것과도 같은 위치에서 검출되지 않았다. 우리의 데이터는 방사선피부염 동물 모델의 병소 안에서, TGF-β1이 증식된 keratinocyte에 과발현된다는 것을 나타낸다. In this study, we established a radiodermatitis animal model and investigated the change in immune cell proportions in the secondary lymphoid organs. The cells responsible for the increased transforming growth factor-β1 (TGF-β1) and interleukin-10 (IL-10) production in the lesions following irradiation were also investigated. The radiodermatitis model was constructed by locally exposing the posterior dorsal region of hairless-1 (HR-1) mice to 10 Gy electron (E)-ray/day for six consecutive days. The change in immune cell proportions was analyzed by FACS. Immunohistochemistry was carried out to detect the expression of cytokines and cell-specific markers in the skin. The proportions of antigen-presenting cells, T cells, and B cells in the lymph nodes and spleen were affected by E-irradiation. After irradiation, TGF-β1 and IL-17 were co-localized in the papillary region of the dermis with keratin-14 (K-14)-positive cells rather than with regulatory T cells (Treg). IL-10 was not co-stained with Treg, T helper 17 (Th17) cells, dendritic cells, or macrophages. Our data indicate that TGF-β1 is over-expressed mainly by proliferated keratinocytes in the lesions of a radiodermatitis animal model.

越鞠丸合補中益氣湯이 電氣刺戟스트레스를 받은 mouse의 免疫機能에 미치는 影響

權泰植,李淑慶,具炳壽 동국대학교 한의학연구소 2000 東國韓醫學硏究所論文集 Vol.9 No.-

본 실험은 越鞠丸合補中益氣湯의 항스트레효과를 조사하기 위하여 시행되었다. 越鞠丸合補中益氣湯이 전기자극으로 스트레스 상황을 유발시킨 mouse에게 끼친 B 임파구 면역계 변화를 mouse 혈청의 綿羊赤血球에 대한 적혈구 응집소가와 비장세포에서 B 세포 및 T 세포가 차지하는 백분율의 변화를 통하여 측정하였으며 이를 대조군과 비교 분석하였다. 혈청 항체가의 측정에서 스트레스 자극하에서 越鞠丸合補中益氣湯을 투여받은 그룹에서 약제를 투여받지 않은 그룹에 비하여 유의성 있는 차이를 보여(p<0.01) 면역 기능을 정상군의 수준으로 유지하는 결과를 얻었다. T 세포와 B 세포의 백분율의 측정에서도 스트레스 자극하에서 越鞠丸合補中益氣湯을 투여 받은 그룹에서 약제를 투여 받지 않은 그룹에 비하여 유의성 있는 차이를 보여(p,0.01) 면역 기능을 정상군의 수준으로 유지하는 결과를 얻었다. 실험결과들로 보아 越鞠丸合補中益氣湯이 체액성 면역 반응과 비장 내 면역 세포의 백분율을 정상의 수준으로 유지하는 효과를 보여 면역 기능의 이상에 응용할 수 있는 가능성을 제시한다고 사료된다. The present experiments were designed to study the influence of Walgookwhanhab-Bojoongikgitang on immune functions of Balb/c mice under stress condition. Walgookwhanhab-Bojoongikgitang was orally administered to the mice for 15 days. On the 10th day the mice were immunized with sheep red blood cells (SRBC) and then subjected to electric footshock for 5 days (2 sessions a day, 11 footshocks a 31 min-session). The immune responses to SRBC were determined by means of hemagglutination and B / T cell populations in the spleen were studied by FACS analysis on the 16th day. The results were as follows. 1. After electric footshock, mice became sluggish and crowded to one side of the cage. Increased antibody titer for SRBC, increased B cell population, and decreased 7 cell population in the spleen were also observed. These results confirm that electric footshock caused stress inducing immunological and behavioral changes in Balb/c mice. 2. Walgookwhanhab-Bojoongikgitang administration significantly antagonized the effect of electric footshock on the antibody titer for SRBC. As a result, antibody titers for SRBC in the mice treated with Walgookwhanga-Bojoongikgitang were maintained at the similar levels as those of control group mice even after the electric footshock. 3. Walgookwhanhab-Bojoongikgitang administration also antagonized the effect of electric footshock on the B / T populations in spleen. As a result, B and T populations in the mice treated with Walgookwhanhab-Bojoongikgitang were maintained at the similar levels as in the control group mice even after the electric footshock. Taken together, M algookwhanhab-Bojoongikgitang seem to help Balb/c mice to maintain their humoral immune response and immune cell populations at a normal range under the stress conditions, suggesting its possible therapeutic use as a immune function modulator.

HSV-TK 유전자를 암호화하는 EBV 유래 플라스미드를 이용한 유전자 치료

오상택,민경아,김종국,이숙경 한국약제학회 2003 Journal of Pharmaceutical Investigation Vol.33 No.4

Herpes simplex virus (HSV) thymidine kinase (TK) has been widely used for suicidal gene therapy in combination with nucleoside analogs such as ganciclovir (GCV). The use of HSV-TK is limited due to the side effect of GCV at high concentraions. Previous studies showed that stable transfectants of mustant HSV-TK with enhanced affinity to GCV were killed at lower GCV concentrations. In this study, we tested whether mutant HSV-TK can provide enhanced suicidal effect when trasiently transfected with Epstein-Barr virus (EBV)-based plasmid. EBV-based plasmid which contains Orip and EBNA-1 sequence is well known for a stable espisomal maintenance in human cells. Optimal transfection condition was assessed for SNU-638 gastric cancer cell line using polyethylenimine (PEI). Maximum transfection efficiency was achieved when DNA:PEI was 1:3 (w/v). Cytotoxicities of mutant and wild type HSV-TK were compared before and after partially selecting transfected cells. The were sensitive to 100?/ml hygromycin. Following GCV treatment, more cells were killed after hygromycin selection than before selection. The mutant HSV-TK showed enhanced cytotoxicity compared with the wild type HSV-TK. Our results suggest that the EBV-based plasmid encoding mutant HSV-TK may be useful to treat the diseases caused by uncontrolled cell proliferation such as cancer and rheumatoid arthritis.