Bioaccumulation of Arsenic in Recombinant Escherichia coli Expressing Human Metallothionein

Yu-Jie Su,Jian-Qun Lin,Jian-Qiang Lin,Dong-Hui Hao 한국생물공학회 2009 Biotechnology and Bioprocess Engineering Vol.14 No.5

The recombinant Escherichia coli(E.coli) expressing human hepatic metallothionein_IA (hMT_IA) was constructed for bioaccumulation of Arsenic (As). The gene sequence of hMT_IA was modified for codon preference of E.coli and synthesized using chemical method. The vector of pGEX_4T_1 was used=and hMT_IA was expressed as the fusion protein with glutathione S-transferase (GST) tag. The bioaccumulation capability of arsenite compounds As(III) of the recombinant E.coli increased more than 3-fold from 76.3 to 319.6 μg/g dry cells compared with the control. The conditions of 50 μM of As(III) and low pHs were optimal for As(III) bioaccumulation. The heavy metals of Cd, Hg, and Zn inhibited As(III) bioaccumulation. The bioaccumulation reached 70% of the saturated value within 1 h. The recombinant E.coli will be useful in bioremediation of arsenic or other kinds of heavy metal contaminated water. The recombinant Escherichia coli(E.coli) expressing human hepatic metallothionein_IA (hMT_IA) was constructed for bioaccumulation of Arsenic (As). The gene sequence of hMT_IA was modified for codon preference of E.coli and synthesized using chemical method. The vector of pGEX_4T_1 was used=and hMT_IA was expressed as the fusion protein with glutathione S-transferase (GST) tag. The bioaccumulation capability of arsenite compounds As(III) of the recombinant E.coli increased more than 3-fold from 76.3 to 319.6 μg/g dry cells compared with the control. The conditions of 50 μM of As(III) and low pHs were optimal for As(III) bioaccumulation. The heavy metals of Cd, Hg, and Zn inhibited As(III) bioaccumulation. The bioaccumulation reached 70% of the saturated value within 1 h. The recombinant E.coli will be useful in bioremediation of arsenic or other kinds of heavy metal contaminated water.

Inclined Slice Method to Earth Pressure of Narrow Cohesionless Backfill against Rigid Walls under Various Displacement Modes

Yu-jian Lin,Fu-quan Chen,Jun-tao Yang 대한토목학회 2020 KSCE JOURNAL OF CIVIL ENGINEERING Vol.24 No.4

The displacement modes of retaining walls in engineering are complicated, and the backfill behind retaining walls is often narrow. Failure mechanisms of soils under various displacement modes of retaining walls have not been clarified yet in previous studies. According to results of the finite element limit analysis, multiple sliding surfaces were observed developing reflectively between interfaces on both sides in the active limit state. In rotational displacement modes, some layers of soil near the rotated point maintained in the non-limit state which was not considered in the previous studies. Based on numerical analysis results, failure wedges were divided into differential soil slice elements by an inclined slice method. An analytical model was established to estimate the earth pressure of the narrow backfill under various displacement modes using the limit equilibrium method. The calculated results are consistent with the data gathered from previous tests. Compared with previous studies, the proposed method can reflect failure mechanisms of the narrow backfill and be adapted to various displacement modes of the retaining wall. The accuracy of the calculation results is high in the non-limit state soil layer. In addition, the boundary criterion between the narrow and semi-limit backfills was determined. Based on extensive parametric studies, it can be concluded that increasing the soil-wall interface friction and decreasing the backfill area would be conducive to decreasing the earth pressure and the depth of resultant application point.

Scutellaria Extract Decreases the Proportion of Side Population Cells in a Myeloma Cell Line by Down-regulating the Expression of ABCG2 Protein

Lin, Mei-Gui,Liu, Li-Ping,Li, Chen-Yin,Zhang, Meng,Chen, Yuling,Qin, Jian,Gu, Yue-Yu,Li, Zhi,Wu, Xin-Lin,Mo, Sui-Lin Asian Pacific Journal of Cancer Prevention 2013 Asian Pacific journal of cancer prevention Vol.14 No.12

Background and Aims: Scutellaria is one of the most popular traditional Chinese herbal remedies against various human diseases, including cancer. In this study, we examined the active effects of Scutellaria extract and its main flavonoid constituents on the proportion of side population cells within human multiple myeloma cell line RPMI8226 in vitro and explored the potential molecular mechanisms involved. Materials and Methods: The contents of flavonoids in ethanolic extract of Scutellaria baicalensis Georgi were determined using high performance liquid chromatography. The antiproliferative effect of the ethanolic extract on RPMI-8226 was determined by CCK assay. Apoptosis was measured by annexin combining with propidium iodide in a flow cytometer. Cell cycle analysis was performed by propidium iodide staining in combination with flow cytometry analysis. Hoechst 33342 exclusion assay was used for the identification of side population within RPMI8226 cells. The expression of ABCG2 protein was assessed by Western blotting assay. Results: The content of major flavonoids constitutents of Scutellaria extract was baicalin (10.2%), wogonoside (2.50%), baicalein (2.29%), and wogonin (0.99%), respectively. The crude Scutellaria extract did not show significant anti-proliferative effect, apoptosis induction and cell cycle arrest in RPMI-8226 within the concentrations of $1-75{\mu}g/mL$. However, the ethanolic extract, baicalein, wogonin and baicalin reduced the side population cells in RPMI-8226, and data showed that baicalein and wogonin had stronger inhibitory effects. Correspondingly, they also exhibited significant effects on decreasing the expression level of ABCG2 protein in RPMI-8226 in vitro. Conclusions: Our results for the first time demonstrated a novel mechanism of action for Scutellaria extract and its main active flavonoids, namely targeting SP cells by modulating the expression of ABCG2 protein. This study provides an insight for new therapeutic strategies targeting cancer stem cells of multiple myeloma.

Taguchi Approach for Anti-heat Stress Prescription Compatibility in Mice Spleen Lymphocytes In Vitro

Xiao-yu Zhu,Gui-lin Cheng,Feng-hua Liu,Jin Yu,Yu-jie Wang,Tong-quan Yu,Jian-qin Xu,Ming Wang 대한약학회 2011 Archives of Pharmacal Research Vol.34 No.7

Heat stress (HS) may induce immunosuppression as well as inhibit the proliferation of lymphocytes. This study evaluated the effects on immune function of our prescription on splenic lymphocytes under HS as well as its compatibility. The effects of four herbal extracts from Agastache rugosa, Atractylodes lancea, Cortex Phellodendri, and Gypsum Fibrosum on heat treated splenic lymphocytes were investigated and the compatibility of the prescription was also explored by using the Taguchi method. This study revealed changes in proliferation by traditional Chinese medicines of splenic lymphocytes after HS. Proliferation in the HS group was significantly lower than the control group. Under HS, the effects of higher concentrations of Agastache rugosa (100 and 200 μg/mL), Atractylodes lancea (100 and 200 μg/mL), Cortex Phellodendri (50 and 100 μg/mL) and Gypsum Fibrosum (100 and 200 μg/mL) caused a significant increase on ConA/LPS-induced proliferation of lymphocytes than lower concentrations. We, therefore, conclude that the prescription of traditional Chinese medicines may recover splenic lymphocytes from the immunosuppression induced by HS. The Taguchi design, which allows rapid and high efficiency for the selection of the best conditions for our prescription on HS-treated splenic lymphocytes, demonstrated that Agastache rugosa (200 μg/mL), Atractylodes lancea (200 μg/mL), Cortex Phellodendri (100 μg/mL) and Gypsum Fibrosum (100 μg/mL)were the optimal conditions for the prescription. The validation experiment confirmed that our composition in optimum extraction conditions enhanced effects on ConA or LPS-stimulated lymphocytes under HS. The results showed that the Taguchi optimization approach is a suitable method for optimization of the composition of prescription.

Branched polymeric prodrug/programmed cell death 4 complexes for combinational cancer therapy

Yu‑Jing He,Lei Xing,Peng‑Fei Cui,Jia‑Liang Zhang,Jian‑Bin Qiao,Cheng‑Qiong Luo,Ge Jiang,Hu‑Lin Jiang 한국약제학회 2017 Journal of Pharmaceutical Investigation Vol.47 No.2

Here, we demonstrate a co-delivery system constructed by integrating chemotherapeutic molecules into branched polymeric prodrug which can condense nucleic acids. Demethylcantharidate (DCA) was chosen as a model drug and premodified through nucleophilic substitution reaction by its two carboxylic groups with allyl chloride. The synthesized intermediate (DCA-dially) was then used to polymerize with tris (2-aminoethyl) amine (TAEA) through progressively ammonolysis reaction. The obtained poly (DCA-alt-TAEA) (DCAT) was used to pack PDCD4 into spherical-like nanoparticles through electrostatic interaction. Gel retardation assays implied that DCAT could integrate DNA at the weight ratio of 1 and protect it from digestion by nuclease. Acid-base titration experiments showed that DCAT obtained preferable buffering capability which was beneficial for the endosomal escape of DCAT/PDCD4 complexes. Cellular tests involving gene transfection efficiency and cytotoxicity indicated that DCA and PDCD4 co-delivered by the complexes significantly and synergistically suppressed the viability of SMMC-7721 cells. These results suggest that integrating chemotherapeutic molecules into nucleic acid-packing polymeric prodrug as cationic polymer/PDCD4 complexes is a highly efficient way to co-deliver chemotherapeutic drugs and nucleic acids for cancer therapy.

Pathologic Response During Chemo-radiotherapy and Variation of Serum VEGF Levels Could Predict Effects of Chemo-Radiotherapy in Patients with Esophageal Cancer

Yu, Jing-Ping,Lu, Wen-Bin,Wang, Jian-Lin,Ni, Xin-Chu,Wang, Jian,Sun, Zhi-Qiang,Sun, Su-Ping Asian Pacific Journal of Cancer Prevention 2015 Asian Pacific journal of cancer prevention Vol.16 No.3

Background: To investigate the relationship between pathologic tumor response to concurrent chemoradiotherapy and variation of serum VEGF in patients with esophageal cancer. Materials and Methods: Forty six patients with esophageal cancer who were treated with concurrent chemo-radiotherapy were enrolled. Endoscopic and pathologic examination was conducted before and four weeks afterwards. Serum level of VEGF was documented before, four weeks later and after chemo-radiotherapy. The relationship between pathologic response and the variation of serum level of VEGF and its influence on the prognosis were investigated. Results: Serum level of VEGF decreased remarkably during and after chemo-radiotherapy in patients whose pathologic response was severe (F=5.393, 4.587, P(0.05). There were no statistical differences of serum VEGF level before, during and after chemo-radiotherapy for patients whose pathologic response was moderate or mild. There were 18 (85.7%), 7 (53.8%) and 6 patients (50.0%) whose serum VEGF level dropped in the severe, moderate and mild group, respectively, with significant differences among these groups (p=0.046). Two year survival rates of patients with severe, moderate and mild pathologic response were 61.9%, 53.8% and 33.3% respectively, and no statistically difference between severe and mild group regarding OS (p=0.245) was tested. Conclusions: Tumor pathologic response during chemo-radiotherapy and the changes of serum VEGF lever could predict curative effects of chemo-radiotherapy in patients with esophageal cancer.

Effect of different alfalfa cultivars on growth and development of the spotted alfalfa aphid, Therioaphis trifolii(Monell)

Yu Liang‐bin,Lin Ke‐Jian,Xu Lin‐bo,Wang Hui,Cui Jin,Zhang Quan‐yi,Wang Ya‐ping,Yan Li‐ying 한국곤충학회 2022 Entomological Research Vol.52 No.3

Therioaphis trifolii (Monell) is a well-known pest in alfalfa cultivation in China. Individuals of T. t r i f o l i i were allowed to feed on 10 alfalfa cultivars and their growth, reproduction, and population life table parameters were estimated. Nymphs that fed on Hunter River had the longest development time, whereas nymphs that fed on Defu had the shortest development time. Adults that fed on Debao, Zhongmu No. 2, and Defu had the shortest development time, whereas adults that fed on Golden Empress and Hunter River had the longest development times. Individuals that fed on Zhungeer produced the greatest number of offspring (13.950), and those that fed on Hunter River produced the fewest offspring (6.000). The survival rate of T. t r i f o l i i was the highest on Algonquin and lowest on Hunter River, whereas the survival time was the longest on Golden Empress and shortest on Debao. The population that fed on Algonquin had the highest intrinsic growth rate (0.560) and shortest population doubling time (1.237 days), whereas those that fed on Hunter River had the lowest intrinsic growth rate (0.217) and longest population doubling time (3.194 days). The life table parameters of the experimental T. t r i f o l i i population were used to quantify the resistance strengths of the cultivars. Results indicate that the Hunter River cultivar may effectively resist damage from T. trifolii. This study provides a theoretical basis for breeding aphid-resistant alfalfa cultivars, enabling further research into the prevention and control of aphid infestation.

Expression, Purification, and Biological Characterization of The Amino-Terminal Fragment of Urokinase in Pichia pastoris

( Jian Ping Li ),( Yu Li Lin ),( Hong Qin Zhuang ),( Zi Chun Hua ) 한국미생물 · 생명공학회 2013 Journal of microbiology and biotechnology Vol.23 No.9

Urokinase (uPA) and its receptor (uPAR) play an important role in tumor growth and metastasis. Targeting the excessive activation of this system as well as the proliferation of the tumor vascular endothelial cell would be expected to prevent tumor neovasculature and halt the tumor development. In this regard, the amino-terminal fragment (ATF) of urokinase has been confirmed as effective to inhibit the proliferation, migration, and invasiveness of cancer cells via interrupting the interaction of uPA and uPAR. Previous studies indicated that ATF expressed in Escherichia coli was mainly contained in inclusion bodies and also lacked posttranslational modifications. In this study, the biologically active and soluble ATF was cloned and expressed in Pichia pastoris. The recombinant protein was purified to be homogenous and confirmed to be biologically active. The yield of the active ATF was about 30 mg/l of the P. pastoris culture medium. The recombinant ATF (rATF) could efficiently inhibit angiogenesis, endothelial cell migration, and tumor cell invasion in vitro. Furthermore, it could inhibit in vivo xenograft tumor growth and prolong the survival of tumor-bearing mice significantly by competing with uPA for binding to cell surfaces. Therefore, P. pastoris is a highly efficient and cost-effective expression system for large-scale production of biologically active rATFs for potential therapeutic application.

Tug of War-Who is the Winner? Canker Disease Restructures the Endophytic Bacterial Community of Citrus

Jian-Yu Zhou,Yi Zhang,Wen-Ping Xu,Misbah Naz,Xiao-Meng Li,Xu Li,Bi-Ying Zhao,Shan-Shan Qi,Zhi-Cong Dai,Dao-Lin Du 한국원예학회 2023 원예과학기술지 Vol.41 No.5

High-throughput sequencing and a 16S rRNA gene clone library amplicon analysis were used to study the endophytic microbial diversity in citrus leaves in response to a Xanthomonas citri pv. Citri (Xcc) infection in an effort to develop a biocontrol scheme for citrus canker disease. There are more species of moderate citrus canker disease (MCD) than severe citrus canker disease (SCD) in terms of both Shannon and Simpson index measurements. Taxonomy has shown that the MCD Firmicutes group (25.2%) outnumber the SCD group (0.55%). Some Firmicutes can suppress Xcc, but in SCD leaves, they are in a competitive position and do not have a dominating niche; therefore, their population is greatly decreased. Sphingomonas and Methylobacterium, two genera of the phylum Proteobacteria, are growth-promoting and stress-resistant in most plants and comprise approximately 60% of the SCD group, whereas the MCD group was less abundant. In conclusion, citrus canker disease restructures bacterial communities in infected leaves, causing the endophytic community to evolve toward “weakening its spear and strengthening its shield.” This research provides support for the idea that introducing helpful bacterial strains in advance may alter the relative abundance of bacteria in a given area and hence decrease the likelihood of infection by harmful bacteria. The future of sequencing technology lies in research on microbial community functions, the elucidation of plant processes and pathogen occurrence, and support for the development of plant biological control technologies.

Adjusting optical resonance thickness to increase the conversion efficiency of polymer solar cells

Yu Sheng Tsai,Jian-Shian Lin,Wei-Ping Chu,Po-Hsun Wang,Fuh-Shyang Juang,Ming-Hua Chung,Chin-Ming Chen,Mark O. Liu 한국물리학회 2010 Current Applied Physics Vol.10 No.3

The derivatives of C60, [6,6]-phenyl C61-butyric acid methyl ester (PCBM), and 3-hexylthiophene (P3HT)were dissolved in DCB solvent, then spin coated into an active layer for polymer solar cells. The experimental parameters were studied carefully to obtain the optimum power conversion efficiency (PCE). The primary process for generation of photocurrent in an organic photovoltaic device is the generation of bound electron–hole pairs (excitons) by absorption of energy (photons) from the optical electric field. Modeling was based on the assumption that the photocurrent generation process is the result of the creation and diffusion of photogenerated species (excitons), which are dissociated by charge transfer at the active layer. Improve organic optics absorb by insert organic layer (CuPc or C60) at the active layer/Al interface. This research is divided into two components. First part, we use n-type C60 as transmission layer. When an optimum thickness of C60 is 5 nm, the Jsc of polymer solar cell can be increased from 7.26 mA/㎠ to 7.7 mA/㎠. The Voc decrease is because the energy level of C60 LUMO (lowest unoccupied molecular orbital) at 4.5 eV is higher than the 3.7 eV of PCBM. Second part, we use p-type CuPc as transmission layer. When an optimum thickness of CuPc is 3 nm, the short circuit photo-current density (Jsc) and open circuit voltage (Voc) of polymer solar cell can be increased from 7.26 mA/㎠ to 8.0 mA/㎠ and 0.56–0.58 V, respectively. The reason is the same as C60. The Voc increase is because the energy level of CuPc LUMO (lowest unoccupied molecular orbital) at 3.1 eV is lower than the 3.7 eV of PCBM. The Jsc increase is because the 3 nm of CuPc leads to a constructive interference happened in the active layer and thus optical absorption increases. In this study we used 3 nm of CuPc at the active layer/Al interface to enhance the short circuit current density, and the efficiency was increased to 2.94%.