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Optimization of the human colorectal carcinoma antigen GA733‑2 production in tobacco plants
박세희,지건영,김현민,마상훈,박서영,도주희,오두병,강형식,심재성,정영희 한국식물생명공학회 2021 Plant biotechnology reports Vol.15 No.1
The colorectal carcinoma-associated protein GA733-2 is one of the representative candidate protein for the development of plant-derived colorectal cancer vaccine. Despite of its signifcant importance for colorectal vaccine development, low efciency of GA733-2 production limits its wide applications. To improve productivity of GA733-2 in plants, we here tested multiple factors that afect expression of recombinant GA733-2 (rGA733-2) and rGA733 fused to fragment crystallizable (Fc) domain (rGA733-Fc) protein. The rGA733-2 and rGA733-Fc proteins were highly expressed when the pBINPLUS vector system was used for transient expression in tobacco plants. In addition, the length of interval between rGA733-2 and left border of T-DNA afected the expression of rGA733 protein. Transient expression analysis using various combinations of Agrobacterium tumefaciens strains (C58C1, LBA4404, and GV3101) and tobacco species (Nicotiana tabacum cv. Xanthi nc and Nicotiana benthamiana) revealed that higher accumulation of rGA733-2 and rGA733-Fc proteins were obtained by combination of A. tumefaciens LBA4404 and Nicotiana benthamiana. Transgenic plants generated by introduction of the rGA733-2 and rGA733-Fc expression cassettes also signifcantly accumulated corresponding recombinant proteins. Bioactivity and stability of the plant-derived rGA733 and rGA733-Fc were evaluated by further in vitro assay, western blot and N-glycosylation analysis. Collectively, we here suggest the optimal condition for efcient production of functional rGA733-2 protein in tobacco system.
종양이식 생쥐모델에서 도두(刀豆), 우방근(牛蒡根) 추출물의 대장암 억제 효과
장지혜,지건영,최형석,양원경,김한영,김근회,강형식,이영철,김승형,Jang, Ji-Hye,Ji, Kon-Young,Choi, Hyung-Seok,Yang, Won-Kyung,Kim, Han-Young,Kim, Kun-hoae,Kang, Hyung-Sik,Lee, Young-Cheol,Kim, Seung-Hyung 대한본초학회 2017 大韓本草學會誌 Vol.32 No.5
Objective : In the present study, we examined whether Canavalia gladiata D.C. (CG) and Arctium lappa L., Redix (AL) mixture (CGAL), their components, lupeol and chicoric acid, regulate immune system and suppress the tumor in vitro and in vivo. Methods : LPS-induced reactive oxygen species (ROS) and nitric oxide (NO) were measured after treatment with CG extract (CGE), CGAL, lupeol, chicoric acid and lupeol and chicoric acid mixture (lupeol+CA) in Raw264.7 cell. To determine the effect of CGE on immune responses, immune cell population and IgG production were assessed in mice. To investigate the effect of CGAL and their component on anti-tumor activity, tumor volume and weight were measured, cell cycles and immune cell population were analyzed in MC38 injected tumor bearing mice. Also, NK cell activity was determined in splenocyte isolated from tumor bearing mice. Results : CGE, CGAL, lupeol, chicoric acid and lupeol+CA decreased the LPS-induced ROS and NO production without cell toxicity in RAW264.7 cells. CGE increased the immune cell populations of $CD4^+T$, $CD8^+T$ and macrophages in various immune organ of mice. In tumor bearing mice, CGAL, lupeol, chicoric acid and lupeol+CA suppressed tumor volume and weight. In cell cycle analysis, they decreased the percentages of S phase. In addition, CGAL, lupeol, chicoric acid and lupeol+CA immune cell populations of $CD4^+T$, $CD8^+Tcell$, NK cell and macrophage in tumor as well as NK cell activity. Conclusion : CGAL and its compounds may enhance immune responses and suppress tumor growth, and may be capable of developing health functional foods.
복분자 추출물의 Sprague-Dawley rat를 이용한단회 경구 투여 독성시험
이주영,지건영,송광훈 대한본초학회 2020 大韓本草學會誌 Vol.35 No.4
Objective : This study was performed to evaluate the toxicity after a single oral administration of black raspberry extract to male and female Sprague-Dawley (SD) rats and to determine the approximate lethal dose (ALD). Methods : We previously showed that the black raspberry extract repressed the simvastatin-mediated expression of Proprotein convertase subtilisin/kexin type 9 (PCSK9) and improved Low-Density Lipoprotein cholesterol (LDL-C) uptake by hepatocytes through the induction of the Low-Density Lipoprotein Receptor expression in hepatocytes. The groups consisted of black raspberry extract groups, as an oral dose of 2,000 ㎎/㎏ and a control group. 5 weeks SD rats were randomly assigned to 4 groups of 5 rats. Each male and female SD rats were administered orally once. For 14 days after the administration, mortality, clinical signs, changes in body weight, and necropsy findings were observed according to the “Standard for Toxicity Study of Pharmaceuticals” of Korea Food and Drug Administration (KFDA) guideline and “Acute Oral Toxicity– Fixed Dose Procedure” of OECD Test Guideline. Results : There were no cases of mortality in the group administered with 2,000 ㎎/㎏ of male and female, and no abnormalities in body weight change and clinical signs. Also, no gross abnormalities were observed at the autopsy. Conclusions : As a result of a single oral administration of the black raspberry extract to SD rats, the ALD was determined to exceed 2,000 ㎎/㎏ for both male and female SD rats.
미세먼지 유도 기도염증에 대한 배암차즈기 추출물의 호흡기 보호 효과
송형우,지건영,김복규,양원경,한창균,신한재,박양춘,황지숙,강형식,김승형,Song, Hyeongwoo,Ji, Kon Young,Kim, Bok Kyu,Yang, Won Kyung,Han, Chang Kyun,Shin, Han Jae,Park, Yang Chun,Hwang, Ji Sook,Kang, Hyung Sik,Kim, Seung Hyung 한국약용작물학회 2017 한국약용작물학회지 Vol.25 No.5
Background: Small particles increase airway inflammation upon reaching the alveoli. Here, we investigated the protective or therapeutic effects of Salvia plebeia R. Br. (SP_R) extracts on airway inflammation. Methods and Results: To investigate the anti-inflammatory activity of SP_R extracts, we measured their inhibitory effect on the production of reactive oxygen species (ROS) expression of inflammatory mediators, and immune cell infiltration in MH-S alveolar macrophage cells and in the ambient particulate matter (APM)-exposed airway inflammation mice model. The SP_R extracts inhibited the production of ROS and expression of IL-4, IL-10, IL-15, and IL-17A mRNA in APM-stimulated MH-S cells. Oral administration of SP_R extracts suppressed APM-induced inflammatory symptoms, such as high alveolar wall thickness, excess collagen fibers, decreased mRNA expression of chemokines (Ccr9, Ccl5, Ccr3), inflammatory cytokines (IL-15, TNF-${\alpha}$), and IL-4 Th2 cytokine in the lung. The SP_R extracts also inhibited ROS production, granulocyte ($CD11b^+Gr-1^+$) infiltration, IL-17A, TNF-${\alpha}$, macrophage inflammatory protein (Mip-2), and chemokine (C-X-C motif) ligand 1 (Cxcl-1) production in the airway. The specific compounds in the SR-R extracts that mediate the anti-inflammatory effects were identified. Conclusions: In this study, SP_R extracts effectively inhibited airway inflammatory responses, such as ROS production and granulocyte infiltration into the airway, by regulating the expression of chemokines and inflammatory cytokines.