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Aim To study the effect of G protein-coupled estrogen receptor(GPER)inhibitor G15 on the sensitivity of breast cancer tamoxifen-resistant cells to T-47DTR. Methods Experiments were carried out with 4-hydroxytamoxifen(4-OHT),the active form of tamoxifen in vivo. The sensitivity of tamoxifen-resistant breast cancer cell line T-47DTR and its parental cell line T-47D to tamoxifen was detected by MTT assay; the expression of GPER protein was analyzed by plasma separation of inhibitor G15; the effect of 4-OHT combined with G15 on the apoptosis of T-47DTR cells was analyzed by flow cytometry AnnexinV-FITC/PI double staining; the expression levels of apoptosis-related proteins Bax,Bcl-2,caspase-3,cleaved caspase-3,caspase-9,cleaved caspase-9 were analysed by Western blot. Results(1)Compared with the parental cell T-47D,the resistance of T-47DTR-resistant cells to 4-OHT was significantly enhanced.(2)When 4-OHT(2 μmol·L-1)was administered,the membrane distribution of GPER increased,indicating that GPER was activated in T-47DTR-resistant cells compared with the control group; Compared with OHT,the use of G15(5 μmol·L-1)and OHT significantly reduced the expression of GPER.(3)GPER inhibitor G15 could increase the apoptotic rate of T-47DTR-resistant cells while down-regulating the anti-apoptotic protein Bcl-2 and up-regulating the expression of pro-apoptotic proteins Bax,cleaved caspase-3,cleaved caspase-9. Conclusions The GPER inhibitor G15 increases the apoptosis of T-47DTR cells and restores the sensitivity of drug-resistant cells to tamoxifen.
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Objective:To study the influence of G protein-coupled estrogen receptor 1 (GPER1) specific agonist G1 and antagonist G2 in epilepsy susceptibility of temporal lobe epileptic rats.Methods:Sixty rats were randomly divided into control group, G1 treatment group and G15 treatment group ( n=20). Rats in the latter two groups were intraperitoneally injected with GPER1 agonist G1 (10 μg) or antagonist G15 (40 μg) for a consecutive 12 d. Lithium chloride pilocarpine epilepsy models were prepared in the 3 groups. The behavior manifestations of these rats were observed within 1 h of intraperitoneal injection of pilocarpine; Racine grading was used to evaluate the severity of epileptic seizures every 5 min; the latency of epileptic seizures (Racine grading IV) and epileptic seizure grading at different time points in the 3 groups were compared. The EEG monitoring was performed to these rats, and EEG data were recorded from 10 min before pilocarpine injection to 2 h after pilocarpine injection; EEG time-frequency was analyzed by Fast-Fourier transform (FFT); distribution of brain electrical energy and changes of θ and α wave energy during 20 min of epileptic status were compared among the 3 groups. Results:(1) As compared with that in the control group and G1 treatment group, the latency of epileptic seizures in the G15 treatment group was significantly shortened ( P<0.05); 15 and 20 min after pilocarpine injection, the epileptic seizure grading of rats in G1 treatment group was statistically lower than that in control group ( P<0.05); 15-35 min after pilocarpine injection, the epileptic seizure grading of rats in G15 treatment group was significantly higher than that in control group ( P<0.05). (2) As compared with those in the control group, rats in the G1 treatment group had smaller brain wave amplitude, while the rats in the G15 treatment group had earlier seizure time, larger brain wave amplitude and higher frequency. There were no obvious changes in the amount of brain electrical energy between the G1 treatment group and control group; while the amount of brain electrical energy in the G15 treatment group 2 h after pilocarpine injection was higher than that in the control group. As compared with those in the control group and G1 treatment group, the θ and α wave energy values of rats in the G15 treatment group were significantly increased within 20 min of epileptic status ( P<0.05). Conclusion:Activation level of GPER1 might be associated with susceptibility to epileptic seizures, and specific inhibition of GPER1 activation can enhance the susceptibility to epilepsy and increase the energy values of specific frequency bands during epilepsy.
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PURPOSE: This study was designed to identify novel fusion transcripts (FTs) and their functional significance in colorectal cancer (CRC) lines. MATERIALS AND METHODS: We performed paired-end RNA sequencing of 28 CRC cell lines. FT candidates were identified using TopHat-fusion, ChimeraScan, and FusionMap tools and further experimental validation was conducted through reverse transcription-polymerase chain reaction and Sanger sequencing. FT was depleted in human CRC line and the effects on cell proliferation, cell migration, and cell invasion were analyzed. RESULTS: One thousand three hundred eighty FT candidates were detected through bioinformatics filtering. We selected six candidate FTs, including four inter-chromosomal and two intrachromosomal FTs and each FT was found in at least one of the 28 cell lines. Moreover, when we tested 19 pairs of CRC tumor and adjacent normal tissue samples, NFATC3–PLA2G15 FT was found in two. Knockdown of NFATC3–PLA2G15 using siRNA reduced mRNA expression of epithelial–mesenchymal transition (EMT) markers such as vimentin, twist, and fibronectin and increased mesenchymal–epithelial transition markers of E-cadherin, claudin-1, and FOXC2 in colo-320 cell line harboring NFATC3–PLA2G15 FT. The NFATC3–PLA2G15 knockdown also inhibited invasion, colony formation capacity, and cell proliferation. CONCLUSION: These results suggest that that NFATC3–PLA2G15 FTs may contribute to tumor progression by enhancing invasion by EMT and proliferation.