Construction of Modified Bacillus thuringiensis cry1Ac Genes for Transgenic Crop Through Multi Site-directed Mutagenesis

Xu, Hong Guang,Roh, Jong-Yul,Wang, Yong,Choi, Jae-Young,Shim, Hee-Jin,Liu, Qin,Tao, Xueying,Woo, Soo-Dong,Jin, Byung-Rae,Je, Yeon-Ho Korean Society of Sericultural Science 2009 International Journal of Industrial Entomology Vol.19 No.1

The newly cloned Bacillus thuringiensis cry1-5 gene showed high activity to both Plutella xylostella and Spodoptera exigua, while cry1Ac only showed high activity against P. xylostella but low to S. exigua. Through the alignment of amino acid sequences between Cry1Ac and Cry1-5, we found 12 different residues in domain I (6 residues) and domain II (6 residues). In this study, the modified cry1Ac gene, which is constructed according to a crop-preferring codon usage, was used as a template to construct mutant B. thuringiensis cry1Ac genes based on cry1-5 gene through multi site-directed mutagenesis. Total 63 various mutant cry genes were obtained at 12 positions randomly. Among them, ten mutant cry genes, whose domain I was totally converted and domain II was randomly, were selected to express in baculovirus expression system as a polyhedrin fusion form. The recombinant proteins were 95 kDa in size and were stably activated as 65 kDa by trypsin. The expressed mutant Cry proteins were applied to bioassays against P. xylostella and S. exigua. All mutants showed high insecticidal activity both to P. xylostella and S. exigua similar to cry1-5. These results suggest that these mutant cry genes might be expected of desirable cry genes for introduction to transgenic crops.

Construction of Modified Bacillus thuringiensis cry1Ac Genes for Transgenic Crop Through Multi Site-directed Mutagenesis

Hong Guang Xu,Jong Yul Roh,Yong Wang,Jae Young Choi,Hee Jin Shim,Qin Liu,Xue Ying Tao,Soo Dong Woo,Byung Rae Jin,Yeon Ho Je 한국잠사학회 2009 International Journal of Industrial Entomology Vol.19 No.1

The newly cloned Bacillus thuringiensis cry1-5 gene showed high activity to both Plutella xylostella and Spodoptera exigua, while cry1Ac only showed high activity against P. xylostella but low to S. exigua. Through the alignment of amino acid sequences between Cry1Ac and Cry1-5, we found 12 different residues in domain I (6 residues) and domain II (6 residues). In this study, the modified cry1Ac gene, which is constructed according to a crop-preferring codon usage, was used as a template to construct mutant B. thuringiensis cry1Ac genes based on cry1-5 gene through multi site-directed mutagenesis. Total 63 various mutant cry genes were obtained at 12 positions randomly. Among them, ten mutant cry genes, whose domain I was totally converted and domain II was randomly, were selected to express in baculovirus expression system as a polyhedrin fusion form. The recombinant proteins were 95 kDa in size and were stably activated as 65 kDa by trypsin. The expressed mutant Cry proteins were applied to bioassays against P. xylostella and S. exigua. All mutants showed high insecticidal activity both to P. xylostella and S. exigua similar to cry1-5. These results suggest that these mutant cry genes might be expected of desirable cry genes for introduction to transgenic crops.

Construction of modified Bacillus thuringiensis cry1AC genes based on cry1-5 genes through multi site-directed mutagenesis

Hong Guang Xu,Jong Yul Roh,Jae Young Choi,Hee Jin Shim,Yong Wang,Qin Liu,Byung Rae Jin,Yeon Ho Je 한국응용곤충학회 2008 한국응용곤충학회 학술대회논문집 Vol.2008 No.10

Bt crystal proteins, encoded by cry genes, are a group of insecticidal proteins unique in the Gram-positive and spore-forming bacterium, Bacillus thuringiensis. These cry genes are widely applied as one of the most successful candidates for constructing transgenic crops resistant to pest insects. In our previous report, we found Cry1-5 had high insecticidal activity against Spodoptera larvae although its amino acid sequences showed high similarity (95.6%) to those of Cry1Ac which had low activity. In comparison with Cry1Ac, Cry1-5 had 12 different residues on domain I and II. In order to convert these residues to Cry1-5 randomly, 10 mutagenic primers were designed. Through multi site-directed mutagenesis, we mutated the modified cry1Ac gene by plant codon usage in pOBI-Modcry1Ac based on cry1-5 and constructed 63 mutant cry genes. Among them, 10 mutant cry genes on domain II were selected and their recombinant proteins were expressed by baculovirus expression system. From bioassay results to P. xylostella and S. exigua, we found some mutants have high insecticidal activities to be applicable to transgenic crops.

Ginsenoside Ro, an oleanolic saponin of Panax ginseng, exerts antiinflammatory effect by direct inhibiting toll like receptor 4 signaling pathway

Hong-Lin Xu,Guang-Hong Chen,Yu-Ting Wu,Ling-Peng Xie,Zhang-Bin Tan,Bin Liu,Hui-Jie Fan,Hong-Mei Chen,Gui-Qiong Huang,Min Liu,Ying-Chun Zhou 고려인삼학회 2022 Journal of Ginseng Research Vol.46 No.1

Background: Panax ginseng Meyer (P. ginseng), a herb distributed in Korea, China and Japan, exerts benefits on diverse inflammatory conditions. However, the underlying mechanism and active ingredients remains largely unclear. Herein, we aimed to explore the active ingredients of P. ginseng against inflammation and elucidate underlying mechanisms. Methods: Inflammation model was constructed by lipopolysaccharide (LPS) in C57BL/6 mice and RAW264.7 macrophages. Molecular docking, molecular dynamics, surface plasmon resonance imaging (SPRi) and immunofluorescence were utilized to predict active component. Results: P. ginseng significantly inhibited LPS-induced lung injury and the expression of proinflammatory factors, including TNF-a, IL-6 and IL-1b. Additionally, P. ginseng blocked fluorescence-labeled LPS (LPS488) binding to the membranes of RAW264.7 macrophages, the phosphorylation of nuclear factor-kB (NF-kB) and mitogen-activated protein kinases (MAPKs). Furthermore, molecular docking demonstrated that ginsenoside Ro (GRo) docked into the LPS binding site of toll like receptor 4 (TLR4)/myeloid differentiation factor 2 (MD2) complex. Molecular dynamic simulations showed that the MD2-GRo binding conformation was stable. SPRi demonstrated an excellent interaction between TLR4/MD2 complex and GRo (KD value of 1.16 × 10<SUP>-9</SUP> M). GRo significantly inhibited LPS488 binding to cell membranes. Further studies showed that GRo markedly suppressed LPS-triggered lung injury, the transcription and secretion levels of TNF-α, IL-6 and IL-1β. Moreover, the phosphorylation of NF-kB and MAPKs as well as the p65 subunit nuclear translocation were inhibited by GRo dose-dependently. Conclusion: Our results suggest that GRo exerts anti-inflammation actions by direct inhibition of TLR4 signaling pathway.

Construction of modified Bacillus thuringiensis cry1AC genes based on cry1-5 genes through multi site-directed mutagenesis

Hong Guang Xu,Jong Yul Roh,Jae Young Choi,Hee Jin Shim,Yong Wang,Qin Liu,Byung Rae Jin,Yeon Ho Je 한국응용곤충학회 2008 한국응용곤충학회 학술대회논문집 Vol.2008 No.05

Bt crystal proteins, encoded by cry genes, are a group of insecticidal proteins unique in the Gram-positive and spore-forming bacterium, Bacillus thuringiensis. These cry genes are widely applied as one of the most successful candidates for constructing transgenic plants resistant to pest insects. In our previous report, we found Cry1-5 had high insecticidal activity against Spodoptera larvae although its amino acid sequences showed high similarity (97.9%) to those of Cry1Ab which had low activity. In comparison with Cry1Ac, Cry1-5 had 12 different residues in domain Ⅰ and domain Ⅱ, and we focused on domain Ⅰand domain Ⅱ regions and designed 10 mutagenic primers to change 12 residues. Through multi site-directed mutagenesis, we mutated the modified cry1Ac gene by plant codon usage in pOBⅠ-Mod-cry1Ac based on cry1-5 and constructed 63 various mutant cry genes. In the further study, we will express those mutant proteins as a fusion form with polyhedrin using baculovirus expression system and subsequently do bioassay to Spodoptera larvae.

Surgical treatment for hepatocellular carcinoma with bile duct invasion

Xu-guang Hu,Wei Mao,Sung Yeon Hong,Bong-Wan Kim,Wei-guang Xu,Hee-Jung Wang 대한외과학회 2016 Annals of Surgical Treatment and Research(ASRT) Vol.90 No.3

Purpose: There is still some debate on surgical procedures for hepatocellular carcinoma (HCC) patients with bile duct tumor thrombi (BDTT, Ueda type 3 or 4). What is adequate extent of liver resection for curative treatment? Is extrahepatic bile duct resection mandatory for cure? The aim of this study is to answer these questions. Methods: Between February 1994 and December 2012, 877 consecutive HCC patients underwent hepatic resection at Ajou University Hospital. Thirty HCC patients (3.4%) with BDTT (Ueda type 3 or 4) were retrospective reviewed in this study. Results: In total, 20 patients enrolled in this study were divided into 2 groups: patients who underwent hemihepatectomy with extrahepatic bile duct resection (group 1, n = 10) and with only removal of BDTT (group 2, n = 10). The 1-, 3- and 5-year overall survival rates were 75.0%, 50.0%, and 27.8%, respectively. The 1-, 3-, and 5-year survival rates of group 1 were 100.0%, 80.0%, and 45.7%, and those of group 2 were 50.0%, 20.0%, and 10.0%, respectively (P = 0.014). The 1-, 3-, and 5-year recurrences free survival rates of group 1 were 90.0%, 70.0%, and 42.0%, and those of group 2 were 36.0%, 36.0%, and 0%, respectively (P = 0.014). Thrombectomy and infiltrative growth type (Ig) were found as independent prognostic factors for recurrence free survival by multivariate analysis. Thrombectomy, Ig, and high indocyanine green retention rate at 15 minutes were found as independent prognostic factors for overall survival by multivariate analysis. Conclusion: We suggest that the appropriate surgical procedure for icteric HCC patients should be comprised of ipsilateral hemihepatectomy with caudate lobectomy and extrahepatic bile duct resection.

Corrigendum to "Ginsenoside Ro, an oleanolic saponin of Panax ginseng, exerts anti-inflammatory effect by direct inhibiting toll like receptor 4 signaling pathway" [J Ginseng Res 46 (2022) 156-166]

Xu, Hong-Lin,Chen, Guang-Hong,Wu, Yu-Ting,Xie, Ling-Peng,Tan, Zhang-Bin,Liu, Bin,Fan, Hui-Jie,Chen, Hong-Mei,Huang, Gui-Qiong,Liu, Min,Zhou, Ying-Chun The Korean Society of Ginseng 2022 Journal of Ginseng Research Vol.46 No.2

18F-fluorodeoxyglucose PET/CT as an independent predictor for patients with hepatocellular carcinoma combined with major portal vein tumor thrombus

Xu-Guang Hu,Xue-Yin Shen,Jin-Niang Nan,In-Gyu Kim,Joon-Kee Yoon,Sung-Yeon Hong,Mi-Na Kim,Bong-Wan Kim,Hee-Jung Wang 대한외과학회 2020 Annals of Surgical Treatment and Research(ASRT) Vol.99 No.1

Purpose: Hepatocellular carcinoma (HCC) patients with major portal vein tumor thrombosis (mPVTT) complications were generally characterized by extremely poor prognoses. The aim of this study was to explore the role of 18F-fluorodeoxyglucose (18F-FDG) PET/CT imaging in predicting HCC complicated by mPVTT. Methods: Five hundred one HCC patients received surgery in our hospital during November 2008 to December 2014, among which 32 patients (6.4%) were diagnosed as HCC complicated by mPVTT. Six cases were excluded for reasons of complex medical conditions, including 2 cases of salvage liver transplantation, 2 cases of re-resection, 1 case of mPVTT combined with inferior vina cava tumor thrombosis, and 1 case of residual portal vein tumor thrombosis. Ultimately, 26 cases were enrolled in this study. The maximal tumor standardized uptake value (SUVmax) was identified as a predictive factor and detected. The univariate and multivariate regression analyses were performed to identify the prognostic factors for recurrence-free survival (RFS) and overall survival (OS) of HCC patients complicated by mPVTT. Results: Our results showed that the median OS was 16 months. The 1-, 3-, and 5-year cumulative OS was 55.6%, 31.7%, and 31.7%, respectively. The multivariate regression analysis revealed that SUVmax ≥ 4.65 was the only independent risk factor for RFS and OS. Conclusion: SUVmax was an independent predictor for RFS and OS of patients suffering from both HCC and mPVTT. L ow SUVmax could serve as an effective factor for selecting candidates with low recurrence risks and for helping with improving patient survival after surgical resection.

Growth Inhibition and G1 Cell Cycle Arrest Mediated by 25-Methoxyhispidol A, a Novel Triterpenoid, Isolated from the Fruit of Poncirus trifoliata in Human Hepatocellular Carcinoma Cells

Hong, ji-Young,Min, Hye-Young,Guang Hua Xu,Lee, Jong-Gu,Lee, Seung-Ho,Kim, Young Shik,Kang, Sam Sik,Lee, Sang Kook 이화여자대학교 약학연구소 2009 藥學硏究論文集 Vol.- No.19

Poncirus trifoliata (Rutaceae) extracts have been known to possess anti-allergic, anti-inflammatory and antiviral activities. However, other biological activities, especially, the anticancer potential of extracts of P. trifoliata or its constituents, have not been fully investigated yet. In this study, we have evaluated the antiproliferative effects of a novel triterpenoid, 25-methoxyhispidol A, isolated from the fruit of P. trifoliata against SK-HEP-1 human hepatocellular carcinoma cells. Flow cytometric analysis indicated that 25-methoxyhispidol A arrests the cell cycle in the G1 phase at the earlier time and subsequently induces apoptosis of the cancer cells. Further study revealed that the cell cycle arrest in the G1 phase by 25-methoxyhispidol A correlated well with the inhibition of phosphorylation of the retinoblastoma (Rb) protein, and with the down-regulation of cyclin D1 and cyclin-dependent kinase cdk4 and the induction of cdk inhibitor p21^(WAF1/Cip1) protein. These findings suggest the potential of 25-methoxyhispidol A isolated from the fructus of P. trifoliata as an antitumor agent against human hepatocarcinoma cells by arresting the cell cycle and inducing apoptosis.

Novel Classification for Right Posterior Portal Vein (RPPV) Evaluated by SYNAPSE VINCENT

( Xu-guang Hu ),( Ingyu Kim ),( Sung Yeon Hong ),( Xue-yin Shen ),( Mina Kim ),( Bong-wan Kim ),( Hee-jung Wang ) 대한간학회 2018 춘·추계 학술대회 (KASL) Vol.2018 No.1

Aims: There was no consensus on right posterior section (RPS) graft was a routine option for adult living donor liver transplantation (LDLT). The main reason for it was that the vascular pedicles of the RPS graft which was the second order branches were complicated. And there was no deep and detailed description of the patterns of the 2^nd, 3^rd portal branch in RPS. The present study is an attempt to describe the patterns of the 2^nd, 3^rd portal branch in RPPV. Methods: Between November 2008 and January 2017, a total of 106 preoperative liver multidetector-row computed tomography (MDCT) images were obtained from the donor and the 3D images were reconstructed by SYNAPSE VINCENT medical imaging system. The patterns of the 2^nd, 3^rd portal branch of the RPPV were investigated using SYNAPSE VINCENT medical imaging system. Results: We classified the RPPV into four types. Type A: the RPPV have a common trunk (more than 5mm) with two main 3^rd order branch (36 cases, 34%); Type B: there was no common trunk with two main 2^nd order branch (16 cases, 15%); Type C: there was one major common trunk with multiple 3^rd order branch (48 cases, 45.3%); Type D: the right posterior section have sliding branches to/from anterior portal vein (6 cases, 5.7%). The median length of the type A portal trunk was 12.35mm (range 5.0mm-28.7mm). In the present study, there were 16 cases of right posterior sector (RPS) graft which were selected by volumetry-Based criteria. There was only one RPS graft has multiple portal veins opening. Conclusions: The Couinaud segment was not always supplied with one inflow portal pedicle. In the present study, there were 52 cases (Type A and B, 49%) which one segment has only one inflow portal pedicle. Others, 54 cases which one segment have two or more 3rd inflow portal pedicle. RPPV type C and D could not apply with anatomic segmentectomy, only the systematic resection is possible.