Isolation, Identification, and Performance Studies of a Novel Paraffin-degrading Bacterium of Gordonia amicalis LH3

Dong-Hui Hao,Xin Song,Jian-Qiang Lin,Yu-Jie Su,Yin-Bo Qu,Jian-Qun Lin 한국생물공학회 2008 Biotechnology and Bioprocess Engineering Vol.13 No.1

In this study, we describe the isolation and identification of a novel long-chain n-alkane degrading strain, Gordonia amicalis LH3. Under aerobic conditions, it utilized approximately 18.0% of paraffin (2% w/v) after 10 day of incubation, and the paraffin compositions of C18C24 alkalines were utilized preferentially. Under anaerobic conditions, paraffin utilization was approximately 1/8 that seen under aerobic conditions, and the compositions of C34 and C36 alkalines were utilized preferentially. The effects of salinity, temperature, and biosurfactants on paraffin degradation were also evaluated. The strain was also demonstrated to grow on oil, and decreased oil viscosity by 44.7% and degraded oil by 10.4% under aerobic conditions. Our results indicated that G. amicalis LH3 has potential applications in paraffin control, microbial enhanced oil recovery (MEOR), and the bioremediation of hydrocarbon-polluted environments.

Effect of 5-aza-2'-deoxycytidine on Cell Proliferation of Non-small Cell Lung Cancer Cell Line A549 Cells and Expression of the TFPI-2 Gene

Dong, Yong-Qiang,Liang, Jiang-Shui,Zhu, Shui-Bo,Zhang, Xiao-Ming,Ji, Tao,Xu, Jia-Hang,Yin, Gui-Lin Asian Pacific Journal of Cancer Prevention 2013 Asian Pacific journal of cancer prevention Vol.14 No.7

Objective: The present study employed 5-aza-2'-deoxycytidine (5-Aza-CdR) to treat non-small cell lung cancer (NSCLC) cell line A549 to investigate the effects on proliferation and expression of the TFPI-2 gene. Methods: Proliferation was assessed by MTT assay after A549 cells were treated with 0, 1, 5, 10 ${\mu}mol/L$ 5-Aza-CdR, a specific demethylating agent, for 24, 48 and 72h. At the last time point cells were also analyzed by flow cytometry (FCM) to identify any change in their cell cycle profiles. Methylation-specific polymerase chain reaction (MSPCR), real time polymerase chain reaction(real-time PCR) and western blotting were carried out to determine TFPI-2 gene methylation status, mRNA expression and protein expression. Results: MTT assay showed that the growth of A549 cells which were treated with 5-Aza-CdR was significantly suppressed as compared with the control group (0 ${\mu}mol/L$ 5-Aza-CdR). After treatment with 0, 1, 5, 10 ${\mu}mol/L$ 5-Aza-CdR for 72h, FCM showed their proportion in G0/G1 was $69.7{\pm}0.99%$, $76.1{\pm}0.83%$, $83.8{\pm}0.35%$, $95.5{\pm}0.55%$ respectively (P<0.05), and the proportion in S was $29.8{\pm}0.43%$, $23.7{\pm}0.96%$, $15.7{\pm}0.75%$, $1.73{\pm}0.45%$, respectively (P<0.05), suggesting 5-Aza-CdR treatment induced G0/G1 phase arrest. MSPCR showed that hypermethylation in the promoter region of TFPI-2 gene was detected in control group (0 ${\mu}mol/L$ 5-Aza-CdR), and demethylation appeared after treatment with 1, 5, 10 ${\mu}mol/L$ 5-Aza-CdR for 72h. Real-time PCR showed that the expression levels of TFPI-2 gene mRNA were $1{\pm}0$, $1.49{\pm}0.14$, $1.86{\pm}0.09$ and $5.80{\pm}0.15$ (P<0.05) respectively. Western blotting analysis showed the relative expression levels of TFPI-2 protein were $0.12{\pm}0.01$, $0.23{\pm}0.02$, $0.31{\pm}0.02$, $0.62{\pm}0.03$ (P<0.05). TFPI-2 protein expression in A549 cells was gradually increased significantly with increase in the 5-Aza-CdR concentration. Conclusions: TFPI-2 gene promoter methylation results in the loss of TFPI-2 mRNA and protein expression in the non-small cell lung cancer cell line A549, and 5-Aza-CdR treatment could induce the demethylation of TFPI-2 gene promoter and restore TFPI-2 gene expression. These findings provide theoretic evidence for clinical treatment of advanced non-small cell lung cancer with the demethylation agent 5-Aza-CdR. TFPI-2 may be one molecular marker for effective treatment of advanced non-small cell lung cancer with 5-Aza-CdR.

Allogeneic Hemopietic Stem Cell Transplants for the Treatment of B Cell Acute Lymphocytic Leukemia

Dong, Wei-Min,Cao, Xiang-Shan,Wang, Biao,Lin, Yun,Hua, Xiao-Ying,Qiu, Guo-Qiang,Gu, Wei-Ying,Xie, Xiao-Bao Asian Pacific Journal of Cancer Prevention 2014 Asian Pacific journal of cancer prevention Vol.15 No.15

Objective: Explore the feasibility of allo-hemopietic stem cell transplants in treating patients with B cell acute lymphocytic leukemia. Methods: Between september 2006 and February 2011, fifteen patients with B cell acute lymphocytic leukemia (ALL) were treated by allo-hemopietic stem cell transplants (HSCT). Stem cell sources were peripheral blood. Six patients were conditioned by busulfan (BU) and cyclophosphamide (CY) and nine patients were conditioned with TBI and cyclophosphamide (CY). Graft versus host disease (GVHD) prophylaxis regimen consisted of cyclosporine A (CSA), methotrex ate (MTX) and mycophenolatemofetil (MMF). Results: Patients received a median of $7.98{\times}10^8{\cdot}kg^{-1}$ ($5.36-12.30{\times}10^8{\cdot}kg^{-1}$) mononuclear cells (MNC). The median time of ANC> $0.5{\times}10^9/L$ was day 12 (10-15), and PLT> $20.0{\times}10^9/L$ was day 13 (11-16). Extensive acute GVHD occurred in 6 (40.0%) patients, and extensive chronic GVHD was recorded in 6 (40.0%) patients. Nine patients were alive after 2.5-65 months follow-up. Conclusion: Allogeneic stem cell transplant could be effective in treating patients with B cell acute lymphocytic leukemia.

Synthesis, Characterization and pH-Responsive Self-Assembly Behavior of Amphiphilic Multiarm Star Triblock Copolymers Based on PCL, PDEAEMA, and PEG

You Qiang Yang,Wen Jing Lin,Li Juan Zhang,Cheng Zhi Cai,Wei Jiang,Xin Dong Guo,Yu Qian 한국고분자학회 2013 Macromolecular Research Vol.21 No.9

A series of amphiphilic 4- and 6-armed star triblock copolymers based on poly(ε-caprolactone) (PCL),poly(2-(diethylamino)ethyl methacrylate) (PDEAEMA), and poly(poly(ethylene glycol) methyl ether methacrylate)(PPEGMA) were designed and synthesized by a combination of ring opening polymerization (ROP) and continuous activators regenerated by electron transfer atom transfer radical polymerization (ARGET ATRP). The continuous ARGET ATRP of DEAEMA and PEGMA was in situ monitored by react infrared spectroscopy (ReactIR) and showed good first-order kinetic characteristics. The molecular weights and chemical structures of the copolymers and their precursors were confirmed by gel permeation chromatography (GPC) and 1H NMR. The critical micelle concentration (CMC) values of the star copolymers in aqueous solution were extremely low (2.2-4.0 mg/L), depending on the architecture of the copolymers. The pH-responsive self-assembly behavior of the star copolymers in aqueous solution was investigated by a combination of dynamic light scattering (DLS), UV-vis spectrometry and scanning electron microscopy (SEM). When the pH values decreased from 10 to 3, no obvious fluctuation of the visible light transmittance of the micelle solutions was observed for lower polymer concentrations of 0.1 and 1 mg/mL, while sharp increase occurred at higher concentration of 10 mg/mL. The hydrodynamic diameters (Dh) of the micelle solutions appeared slight increase with the increase of concentration, and increased rapidly as the pH decreased from 10 to 4 followed by a slight decrease at pH 3. The effects of pH value on the zeta potentials exhibited almost the same tendency with the Dh. This may due to the fact that the protonation of tertiary amine groups in DEAEMA can induce the swelling of micelles. The PCL and PDEAEMA contents and the topological structures (4-or 6-arm) showed significant influences on the pH-sensitivity of the micelles. Overall, the results demonstrated that the structures and pH-sensitivity of these amphiphilic copolymers could be well-controlled and their self-assembled micelles are promising carriers for delivery of anticancer hydrophobic drugs.

Knockdown of Radixin by RNA interference Suppresses the Growth of Human Pancreatic Cancer Cells in Vitro and in Vivo

Chen, Shu-Dong,Song, Mao-Min,Zhong, Zhi-Qiang,Li, Na,Wang, Pi-Lin,Cheng, Shi,Bai, Ri-Xing,Yuan, Hui-Sheng Asian Pacific Journal of Cancer Prevention 2012 Asian Pacific journal of cancer prevention Vol.13 No.3

Radixin, encoded by a gene on chromosome 11, plays important roles in cell motility, invasion and tumor progression. However, its function in pancreatic cancer remains elusive. In this study, radixin gene expression was suppressed with a lentivirus-mediated short-hairpin RNA (shRNA) method. We found that radixin shRNA caused down-regulation of radixin in PANC-1 cells, associated with inhibition of pancreatic cancer cell proliferation, survival, adhesion and invasive potential in vitro. When radixin-silenced cells were implanted in nude mice, tumor growth and microvessel density were significantly inhibited as compared to blank control cells or nonsense shRNA control cells. Thrombospondin-1 (TSP-1) and E-cadherin were up-regulated in radixin-silenced PANC-1 cells. Our results suggest that radixin might play a critical role in pancreatic cancer progression, possibly through invvolvement of down-regulation of TSP-1 and E-cadherin expression.

New Hydrophobic Charge-induction Resin with 2-mercaptoimidazole as the Ligand and Its Separation Characteristics for Porcine IgG

Wimonrat Phottraithip,Dong-Qiang Lin,Fei Shi,Shan-Jing Yao 한국생물공학회 2013 Biotechnology and Bioprocess Engineering Vol.18 No.6

New hydrophobic charge-induction chromatography(HCIC) resin coupled with 2-mercaptoimidazole(MI) as the ligand was prepared and used to purify IgGfrom porcine plasma. The cellulose matrix was activatedby divinylsulfone (DVS), and was then coupled with MI asthe functional ligand. The reaction conditions wereoptimized as pH: 11, volume ratio of DVS to matrix: 1.0,reaction time: 4 h. The ligand density reached about 100μmol/mL gel. The adsorption isotherms of porcine IgGwas determined at different pH values, and high saturatedadsorption capacities of 78.02 mg IgG/mL gel were foundat pH 8. The adsorption of IgG showed a typical pHdependentproperty of HCIC, and the adsorption capacitydecreased significantly in acidic conditions. The preparedresin was used to separate IgG from porcine plasma. Afterprecipitating the fibrinogen by salting-out, the supernatantwas loaded onto the column at pH 7 and the elution pHwas optimized. The results indicated that acidic elution pHwas necessary to recover the IgG. The purity of IgG in theelution fractions was in the range of 81 ~ 90%, whichdemonstrated that HCIC with the new ligand showed theexcited separation performs and is a potential effectivetechnique to purify IgG from the complex feedstock.

Galectin-9 Acts as a Prognostic Factor with Antimetastatic Potential in Hepatocellular Carcinoma

Zhang, Zhao-Yang,Dong, Jia-Hong,Chen, Yong-Wei,Wang, Xian-Qiang,Li, Chong-Hui,Wang, Jian,Wang, Guo-Qiang,Li, Hai-Lin,Wang, Xue-Dong Asian Pacific Journal of Cancer Prevention 2012 Asian Pacific journal of cancer prevention Vol.13 No.6

Considerable research has been conducted concerning galectin-9 and carcinomas, but little information is available about any relation with the hepatocellular carcinoma. In this study, we employed a small interfering RNA (siRNA) targeting galectin-9 to down-regulate the expression in HepG2 cells. As a result, after galectin-9 expression was reduced, cell aggregation was suppressed, while other behaviour such as the proliferation, adhesion and invasion to ECM, cell-endothelial adhesion and transendothelial invasion of the cells were markedly enhanced. When tumors of 200 patients with hepatocellular carcinoma were tested for galectin-9 expression by immunohistochemistry, binding levels demonstrated intimate correlations with the histopathologic grade, lymph node metastasis, vascular invasion and intrahepatic metastasis (P<0.05). Moreover, survival analysis indicated that patients with galectin-9 expression had much longer survival time than those with negative lesions, and the Log-rank test indicated that this difference was statistical significant (P<0.0001). The Cox proportional hazards model suggested that negative galectin-9 expression in hepatocellular carcinoma represented a significant risk factor for patient survival. We propose that galectin-9 might be a new prognostic factor with antimetastatic potential in patients with hepatocellular carcinoma.

Separation of Nattokinase from Bacillus subtilis Fermentation Broth by Expanded Bed Adsorption with Mixed-mode Adsorbent

Lu Miao-Hua,Lin Dong-Qiang,Wu Yuan-Chun,Yun Jun-Xian,Mei Le-He,Yao Shan-Jing The Korean Society for Biotechnology and Bioengine 2005 Biotechnology and Bioprocess Engineering Vol.10 No.2

Mixed-mode hydrophobic/ionic matrices exhibit a salt-tolerant property for adsorbing target protein from high-ionic strength feedstock, which allows the application of undiluted feedstock via an expanded bed process. In the present work, a new type of mixed-mode adsorbent designed for expanded bed adsorption, Fastline $PRO^{\circledR}$, was challenged for the capture of nattokinase from the high ionic fermentation broth of Bacillus subtilis. Two important factors, pH and ion concentration, were investigated with regard to the performance of nattokinase ad-sorption. Under initial fermentation broth conditions (pH 6.6 and conductivity of 10 mS/cm) the adsorption capacity of nattokinase with Fastline PRO was high, with a maximum capacity of 5,350 U/mL adsorbent. The elution behaviors were investigated using packed bed adsorption experiments, which demonstrated that the effective desorption of nattokinase could be achieved by effecting a pH of 9.5. The biomass pulse response experiments were carried out in order to evaluate the biomass/adsorbent interactions between Bacillus subtilis cells and Fastline PRO, and to demonstrate a stable expanded bed in the feedstock containing Bacillus subtilis cells. Finally, an EBA process, utilizing mixed-mode Fastline PRO adsorbent, was optimized to capture nattokinase directly from the fermentation broth. The purification factor reached 12.3, thereby demonstrating the advantages of the mixed-mode EBA in enzyme separation.

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.

Effect of Heat Treatment on the Lipophillic Pigments of Fresh Green Tea Liquor

Jian-Liang Lu,Zhan-Bo Dong,Shun-Shun Pan,Chen Lin,Xin-Qiang Zheng,Borthakur Devajit,Yue-Rong Liang 한국식품과학회 2009 Food Science and Biotechnology Vol.18 No.3

Changes in lipophillic pigments concentration and its relation to color of fresh green tea liquor during heat treatment were studied. The results showed liquor greenness decreased markedly with extension of incubation time at 55℃, while the brightness and yellowness changed a little. Significant increase in ‘a’ and ‘b’ values of tea liquor was observed at 95℃. Color change of liquor at 55℃ was accompanied by a decrease in the level of chlorophylls, lutein and neoxanthin, and an increase in the pheophytins and β-carotene levels. However, all pigments except β-carotene decreased with time extension at 95℃. Significant correlation was found between pigments and color difference index. The browning of fresh green tea liquor was attributed to vicissitudes of lipophillic pigments during heat treatment, especially to the change of chlorophylls/pheophytins ratio. Result also showed addition of Zn²? at 1.6 μ㏖/ℓ could partially alleviate the decrease in greenness during heat treatment.