Toxicological effects of bisphenol A exposure-induced cancer cells migration via activating directly integrin ß1.

Bisphenol A (BPA) is an important chemical that widely used in our life. Mounting evidences show that BPA can leak into environment, which associates with the health risks, such as initiation and metastasis of cancer, but the mechanisms still need to be interpreted. Integrin ß1 is the most known subunit in integrin family, its abnormal expression and activation are tightly linked to tumorigenesis and a number of hallmarks of cancer. Here we show that environmental concentration (10-8 M) of BPA exposure can quickly activate integrin ß1 to induce cancer cell migration, and this effect is proved as a direct interaction between BPA and integrin ß1, which is independent from classical or non-canonical estrogen receptors. The data further indicates that residues S134, D137 and E229 in integrin ß1 played important roles in the interaction as predicted by AutoDock Vina, and confirmed by spectroscopy and native-PAGE. The study is the first to show a tumorigenic mechanism of BPA on tumor metastasis by the direct activation of integrin ß1 molecule, and give rise to profound concerns about widespread use of BPA in the manufacture of plastics and human health.

Design, purification and assessment of GRP78 binding peptide-linked Subunit A of Subtilase cytotoxic for targeting cancer cells.

BACKGROUND: Targeted therapies for cancer, especially the malignant cancer, are always restricted by the deficiency of tumor-specific drug delivery methods. Subtilase cytotoxic is a virulent cytotoxin, and the subunit A (SubA) of it is able to destroy the structure of glucose-regulated protein 78 (GRP78) to induce cell apoptosis, and to be expected as anti-cancer drugs, however, the ubiquitous receptor of subunit B of Subtilase cytotoxic (SubB) restricts its application on cancer therapy. RESULTS: The present study constructed and expressed a fusion protein of GBP-SubA in E. coli Rosetta (DE3) system, in which the subunit B of Subtilase cytotoxic was replaced by GRP78 binding peptide (GBP). The fusion protein was expressed in inclusion body form. Subsequently, the denaturation/renaturation process and Ni-column purification were performed. Our data indicated the purified GBP-SubA could bind GRP78 existed on cancer cell surface specifically, internalize into cells to inactivate intracellular GRP78 and induce apoptosis. Moreover, the apoptosis induction effect of GBP-SubA was enhanced obviously along with the increased cancer cell surface GBP78. CONCLUSIONS: It indicates that the recombinant GBP-SubA possesses the dual functions of GBP and SubA to induce cancer cell apoptosis specifically, revealing that GBP-SubA holds important implications for developing as an anti-cancer peptide drug. A schematic representation of the construction and function of GBP-SubA.

Bisphenol a exposure promotes the migration of NCM460 cells via estrogen receptor-mediated integrin ß1/MMP-9 pathway.

Bisphenol A (BPA) is a widely used industrial chemical and also an environmental endocrine disruptor (EED), which serves as a monomer in the manufacture of polycarbonate plastics. BPA enters human body mainly through oral intake, and has been reported as being linked to oncogenesis in many tissues. However, the association of BPA intake with gastrointestinal cancer, such as colon cancer, has received less attention. The present study was conducted to investigate the effects of BPA on the migration of normal colon epithelial cells (NCM460 cells) and further elucidate the underlying mechanisms. Our data showed that 1 × 10(-8) M (equivalent to environmental concentration) of BPA potently promoted the migration of NCM460 cells. Interestingly, BPA treatment induced an increase of integrin ß1 expression, and the functional blocking of integrin ß1 abolished the migration-promoting effects of BPA. Moreover, the results showed that it was estrogen receptor ß but not estrogen receptor α that was involved in this migration promotion. In addition, cellular exposure of BPA stimulated the expression and activity of MMP-9, a well-known factor of cell migration. Taken together, these results indicate that environmental concentration of BPA exposure promotes cell migration through activating ERß-mediated integrin ß1/MMP-9 pathway, suggesting exposure to BPA in the colon may present a potential cancer risk. © 2014 Wiley Periodicals, Inc. Environ Toxicol 31: 799-807, 2016.

Pluronic F127 as a drug vehicle used in chick embryo chorioallantoic membrane shell-less model.

The developing vascular network is grown on the surface of chick embryo chorioallantoic membrane (CAM), so CAM is widely used as an in vivo model to study the angiogenesis. Because the CAM is hindered or wrinkled by the vehicle, the drug effect is difficult to be observed. In the present study, we firstly introduced the pluronic F127 aquogel to deliver drugs for the CAM model. The biocompatibility and advantage of this vehicle was shown by applied ranibizumab-pluronic F127 mix on the CAM. The results were showed that, the growth of blood vessels was not impaired by pluronic F127 gel, and the gel was almost imperceptible on the CAM, at the same time, the degradation of blood capillaries caused by ranibizumab was clearly visible. In conclusion, pluronic F127 was a good vehicle for angiogenesis research.

Targeted anti-colon cancer activities of a millet bran-derived peroxidase were mediated by elevated ROS generation.

Foxtail millet (Setaria italica) is the sixth most important cereal in the world. In particular, the millet-derived active components play important roles in disease prevention. In this study, we found that a peroxidase from foxtail millet bran, named FMBP, displayed profound inhibitory effects on the growth of human colon cancer cells, but not on that of the normal colon epithelial cells. Mechanistic investigations suggested that the selective anti-cancer effects of FMBP were mainly achieved by inducing more accumulation of reactive oxygen species (ROS) in colon cancer cells than normal cells. The preferential ROS accumulation in cancer cells by FMBP appears to be partially attributed to the down-regulation of NF-E2-related factor 2 (Nrf2) expression, and the reduction of catalase activities and glutathione contents. The increased ROS accumulation is speculated to block the STAT3 signaling pathway, which results in the anti-proliferative effects on colon cancer cells. Therefore, these results suggest that the millet bran-derived peroxidase has a therapeutic potential in the management of colon cancer.

Expression, purification and renaturation of truncated human integrin ß1 from inclusion bodies of Escherichia coli.

Integrins are a family of transmembrane receptors and among their members, integrin ß1 is one of the best known. It plays a very important role in cell adhesion/migration and in cancer metastasis. Preparation of integrin ß1 has a great potential value especially in studies focused on its function. To this end, recombinant plasmids were constructed containing DNA segments representing 454 amino acids of the N-terminal of integrin ß1. The recombinant plasmid was transformed into Escherichiacoli BL21 (DE3) cells and after induction by isopropyl-ß-D-thiogalactopyranoside (IPTG), the recombinant protein (molecular weight: 53 kD) was expressed, mainly in the form of inclusion bodies. The inclusion bodies were solubilized by 8M urea solution then purified by nickel affinity chromatography. The recombinant protein was renatured by a stepwise dialysis and finally dissolved in phosphate buffered saline. The final yield was approximately 5.4 mg/L of culture and the purity of the renatured recombinant protein was greater than 98% as assessed by SDS-PAGE. The integrity of the protein was shown by Western blot using monoclonal antibodies against his-tag and integrin ß1. Its secondary structure was verified as native by circular dichroism spectra and the bioactivity of the recombinant protein was displayed through the conformation switch under Mn(2+) stimulation.

Reconstructed mung bean trypsin inhibitor targeting cell surface GRP78 induces apoptosis and inhibits tumor growth in colorectal cancer.

Glucose regulated protein 78 (GRP78) has been reported to be present on cell membranes of cancer cells but not the normal cells, serving as a potential anti-cancer target. In the present study, a fusion protein containing the GRP78 binding peptide WIFPWIQL and the active fragment of mung bean trypsin inhibitor was constructed, and its targeted anti-tumor effects were investigated both in vitro and in vivo. The results showed that the fusion protein specifically inhibited growth and induced apoptosis in colorectal cancer cells but not in the normal cells. Mechanistically, these anti-tumor effects were attributed to induction of G1 phase arrest and activation of multiple apoptotic pathways. Importantly, the fusion protein could also suppress the growth of xenografted human colorectal carcinoma in vivo. Our study reveals that this fusion protein may be developed as a therapeutic agent for treatment of colon cancer, and holds important implications for developing other anti-cancer peptide drugs.

Development of microsatellite markers from tartary buckwheat.

A genomic library enriched with (gT)(n) repeats from tartary buckwheat (Fagopyrum tataricum) was constructed using 5'-anchored PCR for the development of microsattellite markers. Sequencing analysis of 5 clones from the library showed that they all contained microsatellites (totally 10 loci), and each was unique. An additional locus-specific primer was designed according to flanking sequence. Two of the microsatellite loci of 10 tartary buckwheat varieties were amplified using an anchored primer and a locus-specific primer, which revealed a clear polymorphic pattern. The data confirmed that the degenerate primer was reliably anchoring at the 5'-end of the microsatellite, and the primers developed based on this technology could be used for diversity analysis of tartary buckwheat.