Publications by authors named "Pei Cui"

25 Publications

  • Page 1 of 1

Requirement of the transcription factor YB-1 for maintaining the stemness of cancer stem cells and reverting differentiated cancer cells into cancer stem cells.

Stem Cell Res Ther 2019 08 2;10(1):233. Epub 2019 Aug 2.

College of Life Sciences and Laboratory for Marine Biology and Biotechnology of Qingdao National Laboratory for Marine Science and Technology, Zhejiang University, Hangzhou, 310058, People's Republic of China.

Background: Cancer stem cells always express high levels of stemness-associated transcription factors to maintain their features. However, the regulatory mechanism of the stemness of cancer stem cells mediated by transcription factors has not been extensively explored.

Methods: The YB-1 gene in cancer stem cells was knocked out by the CRISPR/Cas9 system. The YB-1 knockout cancer stem cells were transfected with a vector expressing YB-1 to rescue YB-1, and then the cell proliferation, cell cycle, apoptosis, and stemness, as well as tumorigenesis in nude mice, were assessed to examine the effect of YB-1 in cancer stem cells. The target genes of YB-1 were confirmed by CHIP-seq. The totipotency or pluripotency of differentiated cancer stem cells were detected by tumorsphere formation assay and quantitative real-time PCR.

Results: The deletion of YB-1 gene inhibited the proliferation of breast cancer stem cells and melanoma stem cells, leading to cell cycle arrest and apoptosis, and induced irreversible differentiation of cancer stem cells. The tumorigenicity ability of YB-1-deleted cancer stem cells was significantly reduced in vitro and in vivo. The results of ChIP-seq showed that YB-1 maintained the stemness of cancer stem cells by promoting the expressions of stemness-associated genes (FZD-1, p21, GLP-1, GINS1, and Notch2). Furthermore, simultaneous expressions of YB-1 and the other four (SOX2, POU3F2, OCT-4, and OLIG1) or five (SOX2, SALL2, OCT-4, POU3F2, and Bmi-1) transcription factors in YB-1 knockout cancer stem cells restored the stemness of YB-1 knockout cancer stem cells.

Conclusions: Our study indicated that YB-1 was required for maintaining the stemness of cancer stem cells and reverting the differentiated tumor cells into cancer stem cells.
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http://dx.doi.org/10.1186/s13287-019-1360-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6679460PMC
August 2019

Investigating the potential to assess severe lung inhalation injuries using computed tomography.

Burns 2019 03 21;45(2):310-316. Epub 2018 Dec 21.

Research Laboratory of Burns and Trauma, The 181st Hospital of Chinese PLA, Guilin 541002, People's Republic of China; Department of Burns, Plastic and Wound Repair Surgery, The 181st Hospital of Chinese PLA, Guilin 541002, People's Republic of China. Electronic address:

Purpose: The purpose of this study is to investigate the potential use of computed tomography (CT) in assessing inhalation injuries at various levels by studying the changes in lung imaging of rabbits with severe inhalation injury.

Methods: The sham, serious, critical, and extremely critical lung inhalation injury models were established by the New Zealand white rabbits' inhalation of steam for 0s, 0.25s, 0.50s, and 1.00s, respectively. Lung CT scans were performed at 1, 4, and 12h after the administration of steam and a radiologist's scores (RADS) were collected for each CT scan. Lung tissues were later collected to measure the lung wet/dry weight (W/D) ratio and to determine pathological scores. The correlation of the RADS with the lung-tissue pathological scores and W/D changes was investigated.

Results: The RADS and lung-tissue pathological scores are dependent on the time after injury and the level of injury. W/D ratios are dependent on the level of injury. The W/D ratio showed an increasing trend from 1h to 4h for the 0.25s, 0.50s, and 1s inhalation injury groups, while the W/D ratio decreased from 4h to 12h for the 0.25s and 0.50s inhalation injury groups. Further analysis indicates that, at the same time point, the lung RADS positively correlates with both the lung pathological scores and W/D ratios.

Conclusion: A lung CT scan is able to reflect the early-stage lung injuries of rabbits with different levels of severe inhalation injury.
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http://dx.doi.org/10.1016/j.burns.2018.11.012DOI Listing
March 2019

Acute Respiratory Distress Syndrome Induced by White Smoke Inhalation: a Potential Animal Model For Evaluating Pathological Changes and Underlying Mechanisms.

Cell Physiol Biochem 2018 10;47(6):2396-2406. Epub 2018 Jul 10.

Department of Burn surgery, Changhai Hospital, Naval Military Medical University, Shanghai, China.

Background/aims: White smoke inhalation (WSI) is an uncommon but potentially deadly cause of acute respiratory distress syndrome (ARDS). However, no clinical treatment protocol has been established for the treatment of WSI-induced ARDS. Therefore, it is necessary to investigate the effects of WSI in ARDS and the mechanisms underlying the effects of WSI to determine a novel therapeutic target.

Methods: On the basis of the duration of continued inhalation of white smoke (3 min, 5 min, and 7 min), rats were divided into three groups (WSI-3 min, WSI-5 min, and WSI-7 min). The survival rate, pathological change, and computed tomography (CT) score were evaluated to determine the modeling conditions. In the established WSI-5 min models, evaluations were performed to evaluate the following: arterial blood gas levels, lung wet/dry weight ratio, the expression of inflammatory cytokines, and the effect of NF-κB signaling pathway.

Results: The survival rate of rats at 72 h post-WSI in the WSI-3 min, WSI-5 min, and WSI-7 min groups was 83.33%, 75%, and 25%, respectively. Results from evaluation of H&E staining, CT scan, arterial blood gas levels, and lung wet/dry weight ratio suggest that the pathological changes in the rat in the WSI-5 min and WSI-7 min groups are very similar to those in patients with ARDS induced by WSI. Additionally, the expression of INF-γ, TGF-β1, TNF-α, and IL-1β were increased, and the NF-κB signaling pathway was activated in the WSI-5 min group.

Conclusion: The rat model of WSI-5 min can be used as a WSI-induced ALI model for further experiments. The NF-κB signaling pathway may be a potential therapeutic target for the treatment of WSI- induced ARDS.
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http://dx.doi.org/10.1159/000491614DOI Listing
August 2018

Human amnion-derived mesenchymal stem cells alleviate lung injury induced by white smoke inhalation in rats.

Stem Cell Res Ther 2018 04 12;9(1):101. Epub 2018 Apr 12.

Department of Burn surgery, Changhai Hospital, Naval Military Medical University, Shanghai, 200433, China.

Background: White smoke inhalation (WSI) is an uncommon but potentially deadly cause of acute lung injury and acute respiratory distress syndrome for which no effective pharmaceutical treatment has been developed. This study aimed to determine the protective effects of human amnion-derived mesenchymal stem cells (hAMSCs) against WSI-induced lung injury in rats.

Methods: hAMSCs were injected into rats via the tail vein 4 h after WSI. At 1, 3, 7, 14, and 28 days after cell injection, hAMSCs labeled with PKH26 in lung, heart, liver, and kidney tissues were observed by fluorescence microscopy. The lung injury score was determined by hematoxylin and eosin staining. Lung fibrosis was assessed by Masson's trichrome staining. The computed tomography (CT) score was assessed by CT scanning. The wet/dry weight ratio was calculated. The levels of interleukin (IL)-1β, IL-6, and IL-10 were determined by enzyme-linked immunosorbent assays. The expression of surfactant protein (SP)-A, SP-C, and SP-D was measured by Western blotting.

Results: The injected hAMSCs were primarily distributed in the lung tissues in WSI-induced rats. Compared with the model and phosphate-buffered saline (PBS) group, hAMSC treatment led to reduced lung injury, lung fibrosis, CT score, and inflammation levels in WSI-induced mice. hAMSC treatment also resulted in increased cell retention in the lung, partial pressure of oxygen (PaO), and PaO/fraction of inspired oxygen (FiO) levels, and pulmonary SP-A, SP-C, and SP-D expression compared with that in the model and PBS group.

Conclusions: hAMSCs are a potential cell-based therapy for WSI-induced lung injury.
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http://dx.doi.org/10.1186/s13287-018-0856-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5898065PMC
April 2018

Effect of an upstream bulge configuration on film cooling with and without mist injection.

J Environ Manage 2017 Dec 29;203(Pt 3):1072-1079. Epub 2017 Jun 29.

School of Energy and Environmental Engineering, Hebei University of Technology, Tianjin 300401, China.

To meet the economic requirements of power output, the increased inlet temperature of modern gas turbines is above the melting point of the material. Therefore, high-efficient cooling technology is needed to protect the blades from the hot mainstream. In this study, film cooling was investigated in a simplified channel. A bulge located upstream of the film hole was numerically investigated by analysis of the film cooling effectiveness distribution downstream of the wall. The flow distribution in the plate channel is first presented. Comparing with a case without bulge, different cases with bulge heights of 0.1d, 0.3d and 0.5d were examined with blowing ratios of 0.5 and 1.0. Cases with 1% mist injection were also included in order to obtain better cooling performance. Results show that the bulge configuration located upstream the film hole makes the cooling film more uniform, and enhanceslateral cooling effectiveness. Unlike other cases, the configuration with a 0.3d-height bulge shows a good balance in improving the downstream and lateral cooling effectiveness. Compared with the case without mist at M = 0.5, the 0.3d-height bulge with 1% mist injection increases lateral average effectiveness by 559% at x/d = 55. In addition, a reduction of the thermal stress concentration can be obtained by increasing the height of the bulge configuration.
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http://dx.doi.org/10.1016/j.jenvman.2017.06.055DOI Listing
December 2017

Activation of the pro-migratory bone morphogenetic protein receptor 1B gene in human MDA-MB-468 triple-negative breast cancer cells that over-express CYP2J2.

Int J Biochem Cell Biol 2016 11 5;80:173-178. Epub 2016 Oct 5.

Pharmacogenomics and Drug Development Group, Discipline of Pharmacology, School of Medical Sciences, Sydney Medical School, University of Sydney, NSW 2006, Australia. Electronic address:

Secondary metastases are the leading cause of mortality in patients with breast cancer. Cytochrome P450 (CYP) 2J2 (CYP2J2) is upregulated in many human tumors and generates epoxyeicosanoids from arachidonic acid that promote tumorigenesis and metastasis, but at present there is little information on the genes that mediate these actions. In this study MDA-MB-468 breast cancer cells were stably transfected with CYP2J2 (MDA-2J2 cells) and Affymetrix microarray profiling was undertaken. We identified 182 genes that were differentially expressed in MDA-2J2 cells relative to control (MDA-CTL) cells (log[fold of control] ≥2). From gene ontology pathway analysis bone morphogenetic protein (BMP) receptor 1B (BMPR1B) emerged as an important upregulated gene in MDA-2J2 cells. Addition of the BMPR1B ligand BMP2 stimulated the migration of MDA-2J2 cells, but not MDA-CTL cells, from 3D-matrigel droplets. Migration of MDA-2J2 cells was prevented by the BMPR antagonist dorsomorphin. These findings indicate that over-expression of CYP2J2 in MDA-MB-468-derived breast cancer cells activates BMPR1B expression that may contribute to increased migration. Targeting BMPR1B may be a novel approach to inhibit the metastatic activity of breast cancers that contain high levels of CYP2J2.
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http://dx.doi.org/10.1016/j.biocel.2016.10.004DOI Listing
November 2016

Effect of estrogen on the expression of GnRH and kisspeptin in the hypothalamus of rats during puberty.

Theriogenology 2015 Dec 12;84(9):1556-64. Epub 2015 Aug 12.

Anhui Provincial Laboratory of Animal Genetic Resources Protection and Breeding, Department of Animal Science, College of Animal Science and Technology, Anhui Agricultural University, Hefei, Anhui, China; Anhui Provincial Laboratory for Local Livestock and Poultry Genetic Resource Conservation and Bio-Breeding, Department of Animal Science, College of Animal Science and Technology, Anhui Agricultural University, Hefei, Anhui, China; Department of Animal Veterinary Science, College of Animal Science and Technology, Anhui Agricultural University, Hefei, Anhui, China. Electronic address:

The aim of this study was to assess whether changes in kisspeptin and GnRH levels could be attributed to sex steroids at puberty onset. We used the ovariectomy (OVX) model in rats treated with 17β-estradiol (E2; OVX + E2), or oil (OVX + oil), and in intact rats treated with E2 (intact + E2) or oil only (intact + oil) to determine gene expression changes of Kiss1 and Gnrh1 in the hypothalamus and protein expression of kisspeptin and GnRH in the different areas of the hypothalamus. In the intact + E2 and OVX + E2 rats on the day of the onset of puberty, GnRH-immunoreactive (ir) cell numbers decreased (P < 0.05) in the arcuate nucleus but were increased in the preoptic area; Kisspeptin-ir cells increased (P < 0.05) in the arcuate nucleus, periventricular nucleus, and preoptic area; no difference (P > 0.05) was found in the paraventricularis nucleus for GnRH-ir or kisspeptin-ir cells. Additionally, levels of Kiss1 and Gnrh1 messenger RNA in the hypothalamus were significantly higher (P < 0.05) in the OVX + E2 or intact + E2 rats than in the OVX + oil or intact + oil animals, respectively. In the OVX + oil rats, OVX significantly increased (P < 0.05) levels of Gnrh1 and Kiss1 messenger RNA and the expression of GnRH and kisspeptin in the hypothalamus compared to intact + oil animals. These results suggest that kisspeptin and GnRH play major roles in modulating the activity of estrogen circuits at the onset of puberty.
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http://dx.doi.org/10.1016/j.theriogenology.2015.08.004DOI Listing
December 2015

Tetracyclic triterpenoids in herbal medicines and their activities in diabetes and its complications.

Curr Top Med Chem 2015 ;15(23):2406-30

Faculty of Pharmacy, the University of Sydney, NSW 2006, Australia.

Tetracyclic triterpenoids, including the dammarane, cucurbitane, cycloartane, lanostane and protostane groups, is a class of triterpenoids widely distributed in various medicinal plants, particularly those commonly used for the treatment of diabetes and its complications, such as Panax ginseng, Panax quinquefolium, Panax notoginseng, Gynostemma pentaphyllum, Astragalus membranaceus, Momordica charantia, and Ganoderma lucidum. This review highlights recent findings on the chemistry and bioactivities of tetracyclic triterpenoids from these plants and other popular herbal medicines.
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http://dx.doi.org/10.2174/1568026615666150619141940DOI Listing
June 2016

Effect of active immunization against GnRH-I on the reproductive function in cat.

Anim Sci J 2015 Aug 19;86(8):747-54. Epub 2015 Jan 19.

Anhui Provincial Laboratory of Animal Genetic Resources Protection and Breeding, College of Animal Sciences and Technology, Anhui Agricultural University, Hefei, Anhui, China.

This study was designed to explore the effect of active immunization against maltose binding protein-gonadotropin releasing hormone I hexamer (MBP-GnRH-I6) on the reproductive function in cats. Each immunized cat was administered twice intramuscularly in the neck at 16 and 20 weeks old. The concentrations of the testosterone and estradiol and the level of anti-GnRH-I antibody in the serum were measured by radioimmunoassay and ELISA, respectively. The results showed that the weight and size of testicles and ovaries, and the concentrations of serum testosterone and estradiol in the immunized animals were lower than those of the control cats (P < 0.05), but that the levels of anti-GnRH-I antibody were significant higher compared to control animals (P < 0.05). Testicular tissues from the immunized male cats showed that seminiferous tubules were depauperate with the lumen relatively empty and that the differentiation of spermatogonia was not obvious. Tissues from the immunized female cats showed that the ovaries had many primordial follicles and primary follicles, but no secondary follicle was observed. These results showed active immunization against MBP-GnRH-I6 could make the gonads atrophy and reduce the concentrations of gonadal hormones, which suggested that MBP-GnRH-I6 was a very effective immunogen in the cat.
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http://dx.doi.org/10.1111/asj.12355DOI Listing
August 2015

Analysis of the serum concentrations of kisspeptin and neurokinin B in the geese during reproductive cycle and their localisation in the ovary.

Anim Reprod Sci 2014 Dec 28;151(1-2):78-84. Epub 2014 Sep 28.

Anhui Provincial Laboratory of Animal Genetic Resources Protection and Breeding, College of Animal Sciences and Technology, Anhui Agricultural University, No. 130 of Changjiang West Road, Hefei 230036, Anhui, China; Anhui Provincial Laboratory for Local Livestock and Poultry Genetic Resource Conservation and Bio-Breeding, No. 130 of Changjiang West Road, Hefei 230036, Anhui, China; Department of Animal Veterinary Science, College of Animal Science and Technology, Anhui Agricultural University, No. 130 of Changjiang West Road, Hefei 230036, Anhui, China. Electronic address:

Kisspeptin and neurokinin B (NKB) have various functions. Their expression has been demonstrated in rat and human ovary, and estrogen affects their activity, suggesting a role for these molecules in the control of ovary function. However, whether these signaling systems are present in geese ovary, and the associated with serum estrogen remains largely unexplored. In this study we investigated the expression of kisspeptin and NKB in the ovary, and analysed their changes in the serum during the reproductive cycle and their association with serum estradiol in the geese. The results showed both kisspeptin and NKB immunoreactivity was found in the ovary, with marked expression in the granular layer and theca of the follicle, where intense coexpression of kisspeptin and NKB was also detected. The serum concentrations of kisspeptin and NKB in geese were significantly higher (P<0.05) in broody period than in laying period and laying cessation stage. However, the level of estradiol was markedly higher (P<0.05) in laying period than in broody and laying cessation stage. Serum kisspeptin was positively correlated with NKB (r=0.866, P<0.001), serum estradiol was negatively correlated with kisspeptin (r=-0.977, P<0.05) and NKB (r=-0.887, P<0.05). Overall, the existence of kisspeptin and NKB in geese ovary, and the difference of their serum concentrations during reproductive cycle and the inverse correlation with serum estradiol are highly suggestive of a role for kisspeptin and NKB in the regulation of geese reproductive function.
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http://dx.doi.org/10.1016/j.anireprosci.2014.09.014DOI Listing
December 2014

Functional analysis of novel polymorphisms in the human SLCO1A2 gene that encodes the transporter OATP1A2.

AAPS J 2013 Oct 6;15(4):1099-108. Epub 2013 Aug 6.

Faculty of Pharmacy, The University of Sydney, Sydney, NSW, 2006, Australia,

The solute carrier organic anion transporting polypeptide 1A2 (OATP1A2, SLCO1A2) is implicated in the cellular influx of a number of drugs. We identified five novel single nucleotide polymorphisms (SNPs) in coding exons of the SLCO1A2 gene in a cohort of subjects: G550A, G553A, G673A, A775C, and G862A, that encoded the OATP1A2 variants E184K, D185N, V255I, T259P, and D288N, respectively. The function and expression of these variant transporters were assessed in HEK-293 cells. We found that the novel variants, E184K, D185N, T259P, and D288N, were associated with impaired estrone-3-sulfate, imatinib, and methotrexate transport (∼20-50% of wild-type control); function was retained by OATP1A2-V255I. From biotinylation assays, the decreased function of these variants was due, at least in part, to impaired plasma membrane expression. The four loss-of-function variants were studied further using mutagenesis to produce variants that encode residues with different charges or steric properties. From immunoblotting, the replacement of negatively charged residues at amino acid positions 184 and 185 impaired membrane expression, while either a positive or negative charge at residue 288 supported the correct membrane targeting of OATP1A2. Replacement of T259 with bulky residues disrupted transporter stability. From molecular models, E184, D185, and D288 were located near several charged residues such that intramolecular ionic interactions may stabilize the transporter structure. Individuals who carry these novel SNPs in the SLCO1A2 gene may be at risk from impaired efficacy or enhanced toxicity during treatment with drugs that are substrates for OATP1A2.
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http://dx.doi.org/10.1208/s12248-013-9515-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3787238PMC
October 2013

Anti-proliferative actions of N'-desmethylsorafenib in human breast cancer cells.

Biochem Pharmacol 2013 Aug 31;86(3):419-27. Epub 2013 May 31.

Pharmacogenomics and Drug Development Group, Discipline of Pharmacology, University of Sydney, NSW 2006, Australia.

The multi-kinase inhibitor sorafenib is used for the treatment of renal and hepatic carcinomas and is undergoing evaluation for treatment of breast cancer in combination with other agents. Cytochrome P450 (CYP) 3A4 converts sorafenib to multiple metabolites that have been detected in patient plasma. However, recent clinical findings suggest that combination therapy may elicit inhibitory pharmacokinetic interactions involving sorafenib that increase toxicity. While sorafenib N-oxide is an active metabolite, information on the anti-tumor actions of other metabolites is unavailable. The present study evaluated the actions of sorafenib and its five major metabolites in human breast cancer cell lines. All agents, with the exception of N'-hydroxymethylsorafenib N-oxide, decreased ATP formation in four breast cancer cell lines (MDA-MB-231, MDA-MB-468, MCF-7 and T-47D). Prolonged treatment of MDA-MB-231 cells with N'-desmethylsorafenib, N'-desmethylsorafenib N-oxide and sorafenib (10 μM, 72 h) produced small increases in caspase-3 activity to 128-139% of control. Sorafenib and its metabolites, again with the exception of N'-hydroxymethylsorafenib N-oxide, impaired MEK/ERK signaling in MDA-MB-231 cells and modulated the expression of cyclin D1 and myeloid cell leukemia sequence-1, which regulate cell viability. When coadministered with doxorubicin (0.5 or 1 μM), sorafenib and N'-desmethylsorafenib (25 μM) produced greater effects on ATP production than either treatment alone. Thus, it emerges that, by targeting the MEK/ERK pathway, multiple sorafenib metabolites may contribute to the actions of sorafenib in breast cancer. Because N'-desmethylsorafenib is not extensively metabolized and does not inhibit major hepatic CYPs, this metabolite may have a lower propensity to precipitate pharmacokinetic drug interactions than sorafenib.
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http://dx.doi.org/10.1016/j.bcp.2013.05.014DOI Listing
August 2013

Genomic sequence analysis of a new reassortant infectious bursal disease virus from commercial broiler flocks in Central China.

Arch Virol 2013 Sep 31;158(9):1973-8. Epub 2013 Mar 31.

Henan Center for Animal Disease Control and Prevention, Animal Husbandry Bureau of Henan Province, Zhengzhou, 450008, Henan Province, People's Republic of China.

We report the complete nucleotide sequence of a reassortant infectious bursal disease (IBD) virus (IBDV) HN isolate from commercial broiler flocks in central China. The genome consisted of 3,232 and 2,652 nucleotides in the coding regions of segments A and B, respectively. Alignment of both nucleotide and deduced amino acid sequences and phylogenetic analysis revealed that the genome segments A and B of HN were derived from the attenuated strain B87 and the VV strain OKYM. This is a new reassortant IBDV strain that has emerged in nature, involving segment A of a cell-culture-adapted attenuated vaccine strain B87.
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http://dx.doi.org/10.1007/s00705-013-1682-yDOI Listing
September 2013

Antiproliferative and antimigratory actions of synthetic long chain n-3 monounsaturated fatty acids in breast cancer cells that overexpress cyclooxygenase-2.

J Med Chem 2012 Aug 2;55(16):7163-72. Epub 2012 Aug 2.

Pharmacogenomics and Drug Development Group, Faculty of Pharmacy, University of Sydney, NSW 2006, Australia.

Cyclooxygenase-2 (COX-2) is overexpressed in many human cancers and converts the n-6 polyunsaturated fatty acid (PUFA) arachidonic acid to prostaglandin E(2) (PGE(2)), which drives tumorigenesis; in contrast, n-3 PUFA inhibit tumorigenesis. We tested the hypothesis that these antitumor actions of n-3 PUFA may involve the n-3 olefinic bond. n-3 Monounsaturated fatty acids (MUFAs) of chain length C16-C22 were synthesized and evaluated in MDA-MB-468 breast cancer cells that stably overexpressed COX-2 (MDA-COX-2 cells). Longer chain (C19-C22) n-3 MUFAs inhibited proliferation, activated apoptosis, decreased PGE(2) formation, and decreased cell invasion; C16-C18 analogues were less active. Molecular modeling showed that interactions of Arg120, Tyr355, and several hydrophobic amino acid residues in the COX-2 active site with C19-C22 MUFA analogues were favored. Thus, longer-chain n-3 MUFAs may be prototypes of novel anticancer agents that decrease the formation of PGE(2) in tumor cells that contain high levels of COX-2.
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http://dx.doi.org/10.1021/jm300673zDOI Listing
August 2012

Role of human CYP3A4 in the biotransformation of sorafenib to its major oxidized metabolites.

Biochem Pharmacol 2012 Jul 10;84(2):215-23. Epub 2012 Apr 10.

Pharmacogenomics and Drug Development Group, University of Sydney, Sydney, NSW 2006, Australia.

The tyrosine kinase inhibitor drug sorafenib is used in the treatment of liver and renal cancers but adverse effects may necessitate dose interruption and under-dosage may lead to therapeutic failure. Sorafenib also undergoes cytochrome P450 (CYP)-dependent biotransformation to the N-oxide and other metabolites. However, although CYPs are major determinants of efficacy and toxicity the roles of these enzymes in the formation of multiple sorafenib metabolites are unclear. In the present study CYP-mediated pathways of sorafenib oxidation in human liver were evaluated. cDNA-expressed CYP3A4 was the major catalyst in the formation of the principal N-oxide and N-hydroxymethyl metabolites of sorafenib, as well as the minor N-desmethyl metabolite. In contrast, CYP3A5 exhibited only ~5% of the activity of CYP3A4 and eleven other CYPs and three flavin-containing monooxygenases were inactive. In human hepatic microsomes metabolite formation was correlated with CYP3A4-mediated midazolam 1'-hydroxylation, but not with other CYP-specific substrate oxidations. In accord with these findings the CYP3A4 inhibitor ketoconazole selectively inhibited microsomal sorafenib oxidation pathways. From computational modeling studies atoms in the structure of sorafenib that undergo biotransformation were within ~5.4 Å of the CYP3A4 heme. Important hydrogen bonding interactions between sorafenib and amino acids Ser-119 and Glu-374 in the active center of CYP3A4 were identified. These findings indicate that sorafenib is oxidized selectively by human CYP3A4. This information could be adapted in individualized approaches to optimize sorafenib safety and efficacy in cancer patients.
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http://dx.doi.org/10.1016/j.bcp.2012.04.001DOI Listing
July 2012

Immune responses of chickens inoculated with a recombinant fowlpox vaccine coexpressing glycoprotein B of infectious laryngotracheitis virus and chicken IL-18.

FEMS Immunol Med Microbiol 2011 Nov;63(2):289-95

College of Animal Science and Veterinary Medicine, Henan Agricultural University, Zhengzhou, Henan Province, China.

Infectious laryngotracheitis virus (ILTV) is an alphaherpesvirus that causes severe and economically significant respiratory disease in poultry worldwide. Herein, the immunogenicity of two recombinant fowlpox viruses (rFPV-gB and rFPV-gB/IL18) containing ILTV glycoprotein B (gB) and chicken interleukin-18 (IL-18) were investigated in a challenge model. One-day-old specific-pathogen-free chickens were vaccinated by wing-web puncture with the two rFPVs and challenged with the virulent ILTV CG strain. There were differences in antibody levels elicited by either rFPV-gB/IL18 or rFPV-gB as determined using ELISA. The ratios of CD4(+) to CD8(+) in chickens immunized with rFPV-gB/IL18 were higher (P < 0.05) than in those immunized with rFPV-gB, and the level of proliferative response of the T cells in the rFPV-gB/IL18-vaccinated group was higher (P < 0.05) than that in the rFPV-gB group. All chickens immunized with rFPV-gB/IL18 were protected (10/10), whereas only eight of 10 of the chickens immunized with the rFPV-gB were protected. The results showed that the protective efficacy of the rFPV-gB vaccine could be enhanced by simultaneous expression of chicken IL-18.
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http://dx.doi.org/10.1111/j.1574-695X.2011.00850.xDOI Listing
November 2011

Roles of mitogen-activated protein kinases in the regulation of CYP genes.

Curr Drug Metab 2010 Dec;11(10):850-8

Pharmacogenomics and Drug Development Group, Faculty of Pharmacy, University of Sydney, NSW 2006, Australia.

Cells undergo phenotypic changes after exposure to a wide range of exogenous stimuli that include growth factors, proinflammatory cytokines and environmental chemicals. Such stimuli may arise as components of disease pathogenesis and cellular injury, or as a result of exposure to environmental chemicals and radiation. These stimuli modulate the proliferation and differentiation of cells by altering the regulation of genes that control homeostasis. A generalized response appears to be a decline in the expression and function of many cytochrome P450 (CYP) genes in liver and other tissues. Thus, individuals who have been exposed to such exogenous stimuli often exhibit a decreased capacity for drug clearance, which has important consequences for concurrent drug therapy. Several signaling pathways transduce exogenous stimuli within cells, with the mitogen-activated protein kinases (MAPKs) being one of the most important. Evidence is increasing that MAPKs may impair the expression of multiple CYP genes by modulating the activity of transcription factors, including nuclear receptors, the aryl hydrocarbon receptor, and the activator protein-1 complex. MAPKs catalyze the phosphorylation of transcription complexes that incorporate these factors, which modulates their capacity to transactivate target genes, including CYPs. An understanding of the mechanisms that account for the regulatory impact of MAPKs on the transcriptional factors that regulate CYP genes will provide critical insight into the consequences from exposure to injurious stresses that impact cellular function.
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http://dx.doi.org/10.2174/138920010794479664DOI Listing
December 2010

The ω-3 epoxide of eicosapentaenoic acid inhibits endothelial cell proliferation by p38 MAP kinase activation and cyclin D1/CDK4 down-regulation.

Br J Pharmacol 2011 Mar;162(5):1143-55

Pharmacogenomics and Drug Development Group, Faculty of Pharmacy, University of Sydney, New South Wales, Australia.

Background And Purpose: Dietary intake of ω-3 polyunsaturated fatty acids (ω-3 PUFAs) like eicosapentaenoic acid (EPA) decreases cancer risk, while arachidonic acid and other ω-6 PUFAs increase risk, but the underlying mechanisms are unclear. Cytochrome P450 (CYP)-derived epoxides contribute to enhanced tumourigenesis due to ω-6 PUFA intake. Thus, ω-6 arachidonic acid epoxides (EETs) inhibit apoptosis and stimulate proliferation by up-regulating cyclin D1 expression in cells. The present study evaluated the corresponding ω-3 PUFA epoxides and assessed their role in the regulation of cell proliferation.

Experimental Approach: Four chemically stable EPA epoxides (formed at the 8,9-, 11,12-, 14,15- and 17,18-olefinic bonds) were synthesized and tested against growth-related signalling pathways in brain microvascular endothelial bEND.3 cells. Cell cycle distribution was determined by flow cytometry and cyclin gene expression by immunoblotting and real-time PCR. The role of the p38 mitogen-activated protein (MAP) kinase in cyclin D1 dysregulation was assessed using specific inhibitors and dominant-negative expression plasmids.

Key Results: The ω-3 17,18-epoxide of EPA decreased cell proliferation, interrupted the cell cycle in S-phase and down-regulated the cyclin D1/cyclin-dependent kinase (CDK)-4 complex, whereas the 8,9-, 11,12- and 14,15-epoxides were either inactive or enhanced proliferation. Cyclin D1 down-regulation by 17,18-epoxy-EPA was mediated by activation of the growth-suppressing p38 MAP kinase, but the alternate EPA-epoxides were inactive.

Conclusions And Implications: The present findings suggest that the epoxide formed by CYP enzymes at the ω-3 olefinic bond may contribute to the beneficial effects of ω-3 PUFA by down-regulating cyclin D1 and suppressing cell proliferation.
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http://dx.doi.org/10.1111/j.1476-5381.2010.01113.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3051386PMC
March 2011

Construction and immunogenicity of a recombinant fowlpox vaccine coexpressing S1 glycoprotein of infectious bronchitis virus and chicken IL-18.

Vaccine 2010 Nov 14;28(51):8112-9. Epub 2010 Oct 14.

College of Animal Science and Veterinary Medicine, Henan Agricultural University, Wenhua Road 95#, 450002 Zhengzhou, Henan Province, People's Republic of China.

Infectious bronchitis virus (IBV) poses a major threat to the chicken industry worldwide. In this study, we developed a recombinant fowlpox virus (rFPV) vaccine expressing the IBV S1 gene and chicken interleukin-18 gene (IL-18), rFPV-S1/IL18. Recombinant plasmid pSY-S1/IL18 was constructed by cloning chicken IL-18 into fowlpox virus transfer plasmid containing S1 gene and transfected into the chicken embryo fibroblasts cell pre-infected with S-FPV-017 to generate rFPV-S1/IL18. Expression of the recombinant proteins was confirmed by RT-PCR and IFA. We also constructed the recombinant fowlpox virus rFPV-S1 without IL-18. One-day-old chickens were vaccinated by wing-web puncture with the two rFPVs, and the induced humoral and cellular responses were evaluated. There was a significant difference in ELISA antibody levels (P<0.05) elicited by either rFPV-S1 or rFPV-S1/IL18. The ratios of CD4(+) to CD8(+) in chickens immunized with rFPV-S1/IL18 were significantly higher (P<0.05) than in those immunized with rFPV-S1. All chickens immunized with rFPV-S1/IL18 were completely protected (20/20) after challenge with the virulent IBV HN99 strain 43 days after immunization, while only 15 out of 20 of the chickens immunized with the rFPV-S1 were protected. Our results show that the protective efficacy of the rFPV-S1 vaccine could be enhanced significantly by simultaneous expression of IL-18.
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http://dx.doi.org/10.1016/j.vaccine.2010.09.106DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7115522PMC
November 2010

Impaired transactivation of the human CYP2J2 arachidonic acid epoxygenase gene in HepG2 cells subjected to nitrative stress.

Br J Pharmacol 2010 Apr 24;159(7):1440-9. Epub 2010 Feb 24.

Pharmacogenomics and Drug Development Group, Faculty of Pharmacy, University of Sydney, NSW, Australia.

Background And Purpose: Human cytochrome P450 2J2 (CYP2J2) generates epoxyfatty acids that modulate cellular apoptosis and proliferation. CYP2J2 regulation has not been intensively studied but induction of the activator protein-1 (AP-1) subunit c-fos mediates CYP2J2 down-regulation in hypoxia, a component of ischaemic injury. Decreased CYP2J2 expression may contribute to tissue injury.

Experimental Approach: HepG2 cells were treated with sodium nitroprusside (SNP) to induce nitrative stress, which has been associated with inflammation and infection in liver and other tissues. CYP2J2 protein and mRNA expression were evaluated by immunoblotting and real-time PCR respectively. The role of mitogen-activated protein (MAP) kinases in CYP2J2 dysregulation was assessed using specific inhibitors and dominant negative MAP kinase expression plasmids. CYP2J2-luciferase reporter constructs and electromobility shift assays (EMSAs) were used to identify SNP-regulated regions in the CYP2J2 gene.

Key Results: Cytochrome P450 2J2 was down-regulated by SNP while the AP-1 proteins c-jun and c-fos were up-regulated; inhibition of p38 and ERK MAP kinases normalized their expression. The gene elements at -105/-95 and -56/-63 were required for the down-regulation of CYP2J2 induced by nitrative stress.

Conclusions And Implications: p38 and ERK MAP kinases transduce stress stimuli that down-regulate CYP2J2. Targeting these kinases may prevent the loss of CYP2J2 and epoxy-fatty acids that protect cells against deleterious stresses.
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http://dx.doi.org/10.1111/j.1476-5381.2009.00628.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2850401PMC
April 2010

Facile and stereoselective synthesis of (Z)-15-octadecenoic acid and (Z)-16-nonadecenoic acid: monounsaturated omega-3 fatty acids.

Lipids 2010 Feb;45(2):159-65

Faculty of Pharmacy, University of Sydney, Sydney, NSW 2006, Australia.

Facile syntheses of the monounsaturated omega-3 fatty acids, (Z)-15-octadecenoic acid and (Z)-16-nonadecenoic acid, are presented. Commercially available hydroxy fatty acids were esterified and oxidised, followed by the Wittig reaction to introduce the omega-3 olefinic bond; hydrolysis yielded the omega-3 fatty acids in high purity. An examination of different reaction conditions for the Wittig step found that THF as solvent and coupling temperatures of -78 degrees C gave optimal stereoselectivity, affording the omega-3 olefins in Z:E ratios >or= 97:3. The syntheses have overall yields of approximately 43%, and utilise straightforward, robust chemistry, that may be readily scaled up and reproduced. Also presented is a method for accurately determining the double bond geometry and isomeric purity of the fatty acid products using 1H-13C-HSQC NMR and GC-MS, respectively.
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http://dx.doi.org/10.1007/s11745-009-3378-3DOI Listing
February 2010

Functional characterization of nonsynonymous single nucleotide polymorphisms in the human organic anion transporter 4 (hOAT4).

Br J Pharmacol 2010 Jan 10;159(2):419-27. Epub 2009 Dec 10.

Pharmacogenomics and Drug Development Laboratory, Faculty of Pharmacy, The University of Sydney, Sydney, NSW, Australia.

Background And Purpose: The human organic anion transporter (hOAT) family of transmembrane carrier proteins mediate the cellular flux of anionic substances, including certain hormones and anti-cancer drugs. hOAT4 is highly expressed at the apical membrane of the renal tubular cell and facilitates drug re-absorption in the kidney. In the present study, the impact of 10 nonsynonymous single nucleotide polymorphisms (SNPs) of hOAT4 on transport function in COS-7 cells was characterized.

Experimental Approach: Transport uptake assay was used to assess the function of the variant transporters. Cell surface biotinylation and western blot analysis were used to investigate the expression characteristics of the transporter proteins. Comparative modelling was used to interpret the influence of nonsynonymous changes in terms of hOAT4 structure.

Key Results: Four naturally occurring hOAT4 variants (L29P, R48Y, V155G and T392I) exhibited a significant loss of function. Substitution of leucine-29, which is a conserved residue in OATs, with a proline residue, impaired the synthesis or the apparent stability of the transporter and membrane insertion was disrupted in the R48Y variant. In the case of the V155G and T392I variants, impaired function was due to decreased affinity of the transporter for oestrone sulphate and impaired transporter-substrate turnover respectively. The T392I variant was inhibited more extensively than the wild-type transporter by the cationic substrate tetraethyl ammonium.

Conclusions And Implications: Several naturally occurring SNPs encode variant hOAT4s that may impair the renal tubular re-absorption of important drug substrates.
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http://dx.doi.org/10.1111/j.1476-5381.2009.00545.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2825363PMC
January 2010

Synthesis and NMR characterization of the methyl esters of eicosapentaenoic acid monoepoxides.

Chem Phys Lipids 2009 May 27;159(1):30-7. Epub 2009 Feb 27.

Faculty of Pharmacy, University of Sydney, NSW 2006, Australia.

The activities of cytochrome P450-derived epoxide metabolites of omega-6 polyunsaturated fatty acids (PUFAs) in cellular homeostasis have generated considerable topical interest, but there is less information on the effects of omega-3 PUFA epoxides. Mass spectroscopic data on the epoxides of the omega-3 PUFA eicosapentaenoic acid (EPA) have been reported but the absence of corresponding NMR data currently hinders their biological assessment. In the present study five monoepoxy derivatives of EPA methyl ester were synthesized by treating EPA methyl ester with m-chloroperbenzoic acid. The individual regioisomers were purified by normal-phase chromatography and characterized by LC-MS/MS and a combination of NMR approaches including (1)H-, (13)C-, (1)H-(1)H-COSY, (1)H-(13)C-HSQC, and (1)H-(13)C-HMBC. The chromatographic properties for these monoepoxides were studied in normal-phase and reversephase-HPLC systems and the MS/MS fragmentation patterns using electrospray ionization were established. This paper also focuses on the NMR characterization of epoxide, olefinic and methylenic moieties and the complete assignments of the isomers.
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http://dx.doi.org/10.1016/j.chemphyslip.2009.02.005DOI Listing
May 2009

Monoepoxy octadecadienoates and monoepoxy octadecatrienoates 1: NMR spectral characterization.

Chem Phys Lipids 2008 Apr 11;152(2):122-30. Epub 2008 Mar 11.

Pharmacogenomics, Faculty of Pharmacy, University of Sydney, NSW 2006, Australia.

Methyl esters of gamma-linolenic acid, alpha-linolenic acid and stearidonic acid were epoxidised using m-chloroperbenzoic acid to achieve nine cis-monoepoxy-C18 fatty acid methyl esters (FAMEs), including novel methyl cis-monoepoxy derivatives of stearidonic acid and a cis-6,7-epoxy derivative of gamma-linolenic acid. These nine monoepoxy FAMEs were purified by normal-phase HPLC, identified by LC-MS, 1H and 13C NMR, and characterized by mass spectrometry and NMR spectroscopy. This study is focused on structural characterization of these C18 monoepoxy FAMEs using techniques in NMR spectroscopy including 1H, 13C, 1H-1H correlated spectroscopy (COSY) and 1H-13C heteronuclear correlation (HETCOR). The proton and carbon NMR chemical shifts of the epoxide, the double bonds, and the interrupted methylenes are assigned. Also discussed is an interpretation of the 1H and 13C NMR spectra of these monoepoxides including the changes in the 13C resonance of the olefinic carbons on the neighboring double bonds resulting from epoxide formation.
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http://dx.doi.org/10.1016/j.chemphyslip.2008.02.003DOI Listing
April 2008

Monoepoxy octadecadienoates and monoepoxy octadecatrienoates 2: mass spectral characterization.

Chem Phys Lipids 2008 Apr 11;152(2):65-70. Epub 2008 Mar 11.

Pharmacogenomics, Faculty of Pharmacy, University of Sydney, NSW 2006, Australia.

Methyl esters of C18 polyunsaturated fatty acids, including gamma-linolenic acid, alpha-linolenic acid and stearidonic acid, were epoxidised using m-chloroperbenzoic acid. Nine monoepoxides were obtained by normal-phase HPLC, identified using LC-MS and NMR, and characterized by NMR spectroscopy and mass spectrometry. This study is focused on structural characterization using LC-MS and LC/APCI/MS/MS. The elution profiles of these monoepoxides in RP-HPLC are determined as 12,13->9,10->6,7-epoxy, 9,10->15,16->12,13-epoxy and 15,16->12,13->9,10-epoxy derivatives of gamma-linolenic, alpha-linolenic and stearidonic acid methyl esters, respectively. The major diagnostic fragmentations in MS/MS identified are postulated to be induced by cleavages of the epoxide ring and alpha-bond cleavage to the epoxy group from [M+H]+ and/or [M+H-MeOH]+.
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http://dx.doi.org/10.1016/j.chemphyslip.2008.02.004DOI Listing
April 2008