Inactivation of recombinant human brain-type creatine kinase during denaturation by guanidine hydrochloride in a macromolecular crowding system.

Fan, Yong-Qiang,Liu, Hong-Jian,Li, Chang,Luan, Yu-Shi,Yang, Jun-Mo,Wang, Yu-Long Humana Press 2013 Applied biochemistry and biotechnology Vol.169 No.1

<P>In this study, we quantitatively examined the effects of the macromolecular crowding agents, polyethylene glycol 2000 (PEG 2000) and dextran 70, on guanidine hydrochloride (GdnHCl)-induced denaturation of recombinant human brain-type creatine kinase (rHBCK). Our results showed that both PEG 2000 and dextran 70 had a protective effect on the inactivation of rHBCK induced by 0.5 M GdnHCl at 25 C. The presence of 200 g/L PEG 2000 resulted in the retention of 35.33 % of rHBCK activity after 4 h of inactivation, while no rHBCK activity was observed after denaturation in the absence of macromolecular crowding agents. The presence of PEG 2000 and dextran 70 at a concentration of 100 g/L could decelerate the k (2) value of the slow track to 21 and 33 %, respectively, in comparison to values obtained in the absence of crowding agents. Interestingly, inactivation of rHBCK in the presence of 200 g/L PEG 2000 followed first-order monophasic kinetics, with an apparent rate constant of 8??10(-5)?s(-1). The intrinsic fluorescence results showed that PEG 2000 was better than dextran 70 at stabilizing rHBCK conformation. In addition, the results of the phase diagram indicate that more intermediates may be captured when rHBCK is denatured in a macromolecular crowding system. Mixed crowding agents did not produce better results than single crowding agents, but the protective effects of PEG 2000 on the inactivation and unfolding of rHBCK tended to increase as the ratio of PEG 2000 increased in the mixed crowding agent solution. Though it is not clear which crowding agents more accurately simulated the intracellular environment, this study could lead to a better understanding of protein unfolding in the intracellular environment.</P>

Optimized Transformation of Streptomyces sp. ATCC 39366 Producing Leptomycin by Electroporation

Yong-Qiang Fan,Hong-Jian Liu,Li Yan,Yu-Shi Luan,Hai-Meng Zhou,Jun-Mo Yang,Shang-Jun Yin,Yu-Long Wang 한국미생물학회 2013 The journal of microbiology Vol.51 No.3

Streptomyces sp. ATCC 39366 produces leptomycin derivatives. Leptomycin B, a potent and specific inhibitor against the export of nuclear proteins, is the main product; however, the introduction of DNA into this strain is almost impossible, which has impeded its further use. We developed a Streptomyces sp. ATCC 39366 transformation protocol to introduce foreign DNA via electroporation. Various conditions were examined, including treatments of the cell wall with weakening agents, electroporation parameters, and DNA content. We found that only plasmid DNA isolated from a dam- ET12567 strain resulted in successful transformation. The mycelium growing in a yeast-peptone-dextrose medium supplemented with 1% glycine at 28°C on a rotary shaker (220 rpm) was more dispersed than those without supplementation and prone to electroporation. The maximum transformation efficiency of 8×102 CFU/μg plasmid DNA was obtained at a field strength of 13 kV/cm with a time constant of 13 ms (25-μF capacitor; parallel resistance, 600 Ω) using 1-mm electrocuvettes. The results of the transformations of two other Streptomyces species indicated that the optimized conditions established in this study might only be applicable to Streptomyces sp. ATCC 39366. However, this is the first report of successful transformation of Streptomyces sp. ATCC 39366, and will facilitate the construction of a gene knockout mutant in Streptomyces sp. ATCC 39366 to produce series of new leptomycin derivatives.

Association of MDR1 Gene Polymorphisms with Susceptibility to Hepatocellular Carcinoma in the Chinese Population

Ren, Yong-Qiang,Han, Ju-Qiang,Cao, Jian-Biao,Li, Shao-Xiang,Fan, Gong-Ren Asian Pacific Journal of Cancer Prevention 2012 Asian Pacific journal of cancer prevention Vol.13 No.11

Objective: The objective of this study was to evaluate the association of MDR1 gene polymorphisms with susceptibility to hepatocellular carcinoma (HCC). Methods: A total of 689 HCC patients and 680 cancer-free subjects were enrolled. Human MDR1 gene polymorphisms were investigated by created restriction site-polymerase chain reaction (CRS-PCR) and DNA sequencing methods. Multiple logistic regression models were applied to estimate the association between MDR1 gene polymorphisms and susceptibility to HCC. Results: We detected a novel c.4125A>C polymorphism and our findings suggested that this variant was significantly associated with susceptibility to HCC. A significantly increased susceptibility to HCC was noted in the homozygote comparison (CC versus AA: OR=1.621, 95% CI 1.143-2.300, ${\chi}^2$=7.4095, P=0.0065), recessive model (CC versus AC+AA: OR=1.625, 95% CI 1.167-2.264, ${\chi}^2$=8.3544, P=0.0039) and allele contrast (C versus A: OR=1.185, 95% CI 1.011-1.389, ${\chi}^2$=4.4046, P=0.0358). However, no significant increase was observed in the heterozygote comparison (AC versus AA: OR=0.995, 95% CI 0.794-1.248, ${\chi}^2$=0.0017, P=0.9672) and dominant model (CC+AC versus AA: OR=1.106, 95% CI 0.894-1.369, ${\chi}^2$=0.8560, P=0.3549). Conclusions: These findings suggest that the c.4125A>C polymorphism of the MDR1 gene might contribute to susceptibility to HCC in the Chinese population. Further work will be necessary to clarify the relationship between the c.4125A>C polymorphism and susceptibility to HCC on larger populations of diverse ethnicity.

Attributable Causes of Liver Cancer Mortality and Incidence in China

Fan, Jin-Hu,Wang, Jian-Bing,Jiang, Yong,Xiang, Wang,Liang, Hao,Wei, Wen-Qiang,Qiao, You-Lin,Boffetta, Paolo Asian Pacific Journal of Cancer Prevention 2013 Asian Pacific journal of cancer prevention Vol.14 No.12

Objectives: To estimate the proportion of liver cancer cases and deaths due to infection with hepatitis B virus (HBV), hepatitis C virus (HCV), aflatoxin exposure, alcohol drinking and smoking in China in 2005. Study design: Systemic assessment of the burden of five modifiable risk factors on the occurrence of liver cancer in China using the population attributable fraction. Methods: We estimated the population attributable fraction of liver cancer caused by five modifiable risk factors using the prevalence data around 1990 and data on relative risks from meta-analyses, and large-scale observational studies. Liver cancer mortality data were from the 3rd National Death Causes Survey, and data on liver cancer incidence were estimated from the mortality data from cancer registries in China and a mortality/incidence ratio calculated. Results: We estimated that HBV infection was responsible for 65.9% of liver cancer deaths in men and 58.4% in women, while HCV was responsible for 27.3% and 28.6% respectively. The fraction of liver cancer deaths attributable to aflatoxin was estimated to be 25.0% for both men and women. Alcohol drinking was responsible for 23.4% of liver cancer deaths in men and 2.2% in women. Smoking was responsible for 18.7% and 1.0%. Overall, 86% of liver cancer mortality and incidence (88% in men and 78% in women) was attributable to these five modifiable risk factors. Conclusions: HBV, HCV, aflatoxin, alcohol drinking and tobacco smoking were responsible for 86% of liver cancer mortality and incidence in China in 2005. Our findings provide useful data for developing guidelines for liver cancer prevention and control in China and other developing countries.

Antiproliferative Effects of Celecoxib in Hep-2 Cells through Telomerase Inhibition and Induction of Apoptosis

Zhao, Yong-Qiang,Feng, Hui-Wei,Jia, Tao,Chen, Xue-Mei,Zhang, Hui,Xu, An-Ting,Zhang, Hai-Ling,Fan, Xian-Liang Asian Pacific Journal of Cancer Prevention 2014 Asian Pacific journal of cancer prevention Vol.15 No.12

Background: To investigate the effect of celecoxib on telomerase activity and apoptosis in a human laryngeal squamous carcinoma cell line (Hep-2 cells). Materials and Methods: The growth inhibition rate of Hep-2 cells in vitro was measured by MTT assay, and apoptosis by TUNEL assay and flow cytometry (FCM). The TRAP-ELISA method was used to determine telomerase activity in Hep-2 cells. The mRNA expression of human telomerase RNA component(hTR), human telomerase reverse transcriptase (hTERT) and human telomerase-associated protein(hTEP1) was determined by RT-PCR assay. Expression of Bax and Bcl-2 proteins was assessed by Western blotting. Results: Celecoxib can inhibit proliferation and induce apoptosis in a dose- and time-dependent manner, repress telomerase activity, decrease hTERT mRNA and Bcl-2 protein expression and increase Bax protein expression, PGE2 had no effect on telomerase. Conclusions: Celecoxib had the antiproliferative and pro-apoptotic effect in Hep-2 cells. Apoptosis was accompanied by a decrease in telomerase activity which was directly correlated with hTERT mRNA and up-regulation of Bax/Bcl-2. Bcl-2 may thus play an important role in telomerase activity as well as apoptosis.

Antitumor effects of dammarane-type saponins from steamed Notoginseng

He, Fan,Ding, Yan,Liang, Chun,Song, Seok Bean,Dou, De-Qiang,Song, Gyu Yong,Kim, Young Ho Medknow PublicationsMedia Pvt Ltd 2014 Pharmacognosy magazine Vol.10 No.39

<P>Six dammarane-type saponins were extracted from steamed <I>Panax notoginseng</I>. Their chemical structures were identified spectroscopically as ginsenosides Rh<SUB>1</SUB> (1), Rg<SUB>1</SUB> (2), 20 (S)-Rg<SUB>3</SUB> (3), 20 (R)-Rg<SUB>3</SUB> (4), Rb<SUB>3</SUB> (5), and Rb<SUB>1</SUB> (6). Compounds (0.1-10 μM) were tested for inhibition of tumor necrosis factor-α (TNF)-induced nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) luciferase reporter activity using a human kidney 293T cell-based assay. Ginsenoside Rb<SUB>3</SUB> (5) showed the most significant activity with an IC<SUB>50</SUB> of 8.2 μM. This compound also inhibited the induction of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) messenger Ribonucleic acid (mRNA) in a dose-dependent manner after HepG2 cells had been treated with TNF-α (10 ng/mL).</P>

Potential Therapeutic Targets for the Primary Gallbladder Carcinoma: Estrogen Receptors

Zhang, Ling-Qiang,Zhang, Xiu-De,Xu, Jia,Wan, Yong,Qu, Kai,Zhang, Jing-Yao,Wang, Zhi-Xin,Wei, Ji-Chao,Meng, Fan-Di,Tai, Ming-Hui,Zhou, Lei,Liu, Chang Asian Pacific Journal of Cancer Prevention 2013 Asian Pacific journal of cancer prevention Vol.14 No.4

Gallbladder carcinoma, the most frequent malignant neoplasm of the biliary tract system, has always been considered to feature late clinical presentation and diagnosis, limited treatment options and an extremely poor prognosis. In recent years, while the incidence of gallbladder cancer has appeared to be on the increase, the available treatment methods have not greatly improved survival of the affected patients. Thus, exploring new therapeutic targets for this devastating disease is an urgent matter at present. Epidemical studies have demonstrated that the incidence of gallbladder carcinoma exhibits a distinct gender bias, affecting females two to three times more than males, pointing to crucial roles of estrogen. It is well known that estrogen acts on target tissues by binding to estrogen receptors (ERs), which are mainly divided into three subtypes, $ER{\alpha}$, $ER{\beta}$ and $ER{\gamma}$. $ER{\alpha}$ and $ER{\beta}$ appear to have overlapping but also unique even opposite biological effects. As important pathogenic mediators, ERs have been considered to relate to several kinds of tumors. In gallbladder carcinoma tissue, ERs have been shown to be positively expressed, and ERs expression levels are associated with differentiation and prognosis of this cancer. Nevertheless, the exact mechanisms of estrogen inducing growth of gallbladder carcinoma remain poorly understood. On the base of the current investigations, we deduce that estrogen participates in promotion of gallbladder carcinoma by influencing the formation of gallstones, stimulating angiogenesis, and promoting abnormal proliferation. Since ERs mediate the carcinogenic actions of estrogen in gallbladder, and therapy targeting ERs may provide new directions for gallbladder carcinoma. Therefore, it should be stressed that ERs are potential therapeutic targets for gallbladder carcinoma.

rs10505474 and rs7837328 at 8q24 Cumulatively Confer Risk of Prostate Cancer in Northern Han Chinese

Zhang, Lin-Lin,Sun, Liang,Zhu, Xiao-Quan,Xu, Yong,Yang, Kuo,Yang, Fan,Yang, Yi-Ge,Chen, Guo-Qiang,Fu, Ji-Cheng,Zheng, Chen-Guang,Li, Ying,Mu, Xiao-Qiu,Shi, Xiao-Hong,Zhao, Fan,Wang, Fei,Yang, Ze,Wang, Asian Pacific Journal of Cancer Prevention 2014 Asian Pacific journal of cancer prevention Vol.15 No.7

Aims: Genome-wide association studies (GWAS) have identified several risk variants for prostate cancer (pCa) mainly in Europeans, which need to be further verified in other racial groups. We selected six previously identified variants as candidates and to define the association with PCa in Northern Han Chinese. Methods: 749 subjects from Beijing and Tianjin in Northern China were included. Six variants (rs10505474, rs7837328, rs4242384, rs7813, rs486907 and rs1058205) were genotyped by high resolution melting (HRM) assays. The individual and cumulative contribution for of the risk of PCa and clinical covariates were analyzed. Results: Among the six candidate variants, onlyrs10505474, and rs7837328, both locating at 8q24 region, were associated with PCa in our population.rs10505474 (A) was associated with PCa ($OR_{recessive}=1.56$, p=0.006); and rs7837328 (A) was associated with PCa ($OR_{dominant}=1.38$, p=0.042/$OR_{recessive}=1.99$, p=0.003). Moreover, we observed a cumulative effects between them ($p_{trend}=2.58{\times}10^{-5}$). The joint population attributable risk showed the two variants might account for 71.85% of PCa risk. In addition, we found the homozygotes of rs10505474 (A) and rs7837328 (A) were associated with PCa clinical covariants (age at onset, tumor stage, respectively) ($p_{age}=0.046$, $P_{tumorstage}=0.048$). Conclusion: rs10505474 (A) and rs7387328 (A) at 8q24 are associated with PCa and cumulatively confer risk, suggesting the two variations could determine susceptibility to PCa in the Northern Chinese Han population.