Targeted reversal of multidrug resistance in ovarian cancer cells using exosome­encapsulated tetramethylpyrazine.

The objective of the present study was to develop exosomes (EXOs) encapsulating tetramethylpyrazine (TMP) for the reversal of drug resistance in ovarian cancer therapy. Human A2780 cells were incubated with TMP for 48 h. Purified TMP­primed EXOs (EXOs­TMP) were isolated through ultracentrifugation. The developed EXOs­TMP were characterized using techniques such as transmission electron microscopy, nanoparticle tracking analysis, Fluorescence microscopy and western blotting. Subsequently, MTT, western blotting and flow cytometry assays were performed to evaluate the biological effects in drug­resistant A2780T cells. The results demonstrated that the incorporation of TMP into EXOs exhibited an anti­ovarian cancer effect and markedly enhanced the antitumor efficacy of paclitaxel (PTX). Furthermore, it was identified that the ability of EXO­TMP to reverse cell resistance was associated with the downregulation of multidrug resistance protein 1, multidrug resistant­associated protein 1 and glutathione S­transferase Pi protein expression. Flow cytometry analysis revealed that EXO­TMP induced apoptosis in drug­resistant cells and enhanced the apoptotic effect when combined with PTX. EXOs are naturally sourced, exhibit excellent biocompatibility and enable precise drug delivery to target sites, thereby reducing toxic side effects. Overall, EXO­TMP exhibited direct targeting capabilities towards A2780T cells and effectively reduced their drug resistance. EXOs­TMP provide a novel and effective drug delivery pathway for reversing drug resistance in ovarian cancer.

Measurement of Cumulative fission product yields on 235U induced by 2.8 MeV neutrons.

Off-line gamma-ray spectrometry was used to accurately measure the Cumulative fission product yields (CFPYs) of fission products in the 235U (n, f) reaction induced by 2.8 MeV neutrons. The 2.8 MeV quasi-monoenergetic neutron beam was produced by the CPNG-600 Cockcroft Walton accelerator at the China Institute of Atomic Energy （CIAE）and the gamma spectra were measured by the HPGe γ-ray Spectrometer. After fully considering and revising the sources of uncertainty, high-precision CFPYs of 4 fission products were obtained. This study has important applications in reactor design and operation and is conducive to the establishment of an evaluated nuclear database.

Pharmacological inhibition of the ubiquitin-specific protease 8 effectively suppresses glioblastoma cell growth.

Glioblastoma (GBM) is a malignant brain tumor. The purpose of this study is to estimate the potential effects and underlying mechanisms of a ubiquitin-specific protease 8 (USP8) small-molecule inhibitor on the phenotypic characteristics of GBM cells. The growth, migration, invasion, and stemness of GBM LN229 and T98G cells were evaluated by conducting cell proliferation, colony formation, wound healing, transwell, Ki-67 staining, spheroid formation, and ionizing radiation assays, and the results collectively showed the suppressive effects of USP8 inhibition on GBM cells. Furthermore, transcriptomic profiling of GBM cells treated with the USP8 inhibitor deubiquitinase (DUB)-IN-1 revealed significantly altered mRNA expression induced by pharmacological USP8 inhibition, from which we confirmed downregulated Aurora kinase A (AURKA) protein levels using immunoblotting assays. Our findings indicated that the proliferation, invasion, and stemness of LN229 and T98G cells were markedly suppressed by USP8 inhibition. Pharmacological USP8 suppression elicits multiple tumor-inhibitory effects, likely through dysregulating various mRNA expression events, including that of the key cell cycle regulator and oncogenic protein AURKA. Therefore, our observations corroborate the GBM-supportive roles of USP8 and suggest pharmacological USP8 inhibition is a viable therapeutic approach to target GBM. The purpose of this study was to investigate the effect and mechanism of action of the USP8 inhibitor DUB-IN-1 on GBM.

LPS preconditioning of MSC-CM improves protection against hypoxia/reoxygenation-induced damage in H9c2 cells partly via HMGB1/Bach1 signalling.

Mesenchymal stem cell-derived conditioned medium (MSC-CM) improves cardiac function after myocardial infarction; however, this cardioprotective effect is moderate and transient. Lipopolysaccharide (LPS) pretreatment partially improves MSC-CM-mediated cardioprotective effects owing to the presence of paracrine factors. However, the mechanism underlying these improved effects remains unknown. To study the effect of LPS-pretreated MSC-CM on hypoxia/reoxygenation (H/R)-induced injury, MSCs were treated with or without LPS (400 ng/mL) for 48 h, and the supernatant was collected (MSC-CM). Subsequently, H9c2 cells were co-cultured with Nor-CM (CM derived from LPS-untreated MSCs) and LPS-CM (CM derived from LPS-pretreated MSCs) for 24 h and subjected to H/R. MSC-CM inhibited the progression of H/R-induced injury in H9c2 cells, and this protective effect was enhanced via LPS pretreatment as evidenced by the improved apoptosis assessment index (i.e. caspase-3 and B-cell lymphoma-2 [Bcl-2] expression) and decreased levels of lactic dehydrogenase (LDH) and cardiac troponin (cTn). In addition, the results of haematoxylin-eosin staining (H&E), transmission electron microscopy (TEM) and TdT-mediated dUTP nick-end labelling (TUNEL) validated that MSC-CM inhibited H/R-induced injury in H9c2 cardiomyocytes. LPS pretreatment downregulated the expression of high mobility group box-1 (HMGB1) and BTB and CNC homology-1 (Bach1) proteins in MSCs but upregulated the expression of vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF) and insulin-like growth factor (IGF). HMGB1 knockdown (MSC/siHMGB1-CM) significantly decreased the expression of Bach1 and increased the expression of VEGF, HGF and IGF. Bach1 knockdown (MSC/siBach1-CM) did not alter the production of HMGB1 but increased the expression of VEGF and IGF. LPS pretreatment did not alter the expression of the paracrine factors VEGF and HGF in the MSC/siHMGB1 group but increased their expression in the MSC/siBach1 group. The myocyte anti-apoptotic effects of MSCs/siBach1-CM were similar to those of untreated MSCs, which were not enhanced by LPS. LPS-pretreated MSC-CM protects H9c2 cells against H/R-induced injury partly through the HMGB1/Bach1 signalling pathway.

Lipopolysaccharide-pretreated mesenchymal stem cell-conditioned medium optimized with 10 kDa filter attenuates the injury of H9c2 cardiomyocytes in a model of hypoxia/reoxygenation.

Mesenchymal stem cell-derived conditioned medium (MSC-CM) improves cardiac function, which is partly attributed to the released paracrine factors. Since such cardioprotection is moderate and transient, it is essential that MSC-CM's effective components are optimized to alleviate myocardial injury. To optimize MSC-CM, MSCs were treated with or without lipopolysaccharides (LPSs) for 48 h (serum-free), and the supernatant was collected. Then, LPS-CM (MSC stimulated by LPS) was further treated with LPS remover (LPS Re-CM) or was concentrated with a 10 kDa cutoff filter (10 kDa-CM). Enzyme-linked immunosorbent assay showed that all the pretreatments increased the levels of vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), and insulin growth factor (IGF) except LPS Re-CM; 10 kDa-CM was superior to the other CMs. Cell Counting Kit-8 displayed that the viability of injured H9c2 cells was enhanced with the increase in the MSC-CM concentration. We also found that the 10 kDa-CM significantly alleviated H9c2 hypoxia/reoxygenation (H/R) injury, as evidenced by the increased Bcl-2/Bax ratio, and decreased the levels of lactate dehydrogenase and cardiac troponin. Transmission electron microscopy (TEM), TdT-mediated dUTP nick-end labelling (TUNEL), and hematoxylin and eosin staining (H&E) confirmed that 10 kDa-CM inhibited H/R-induced H9c2 morphological changes. Proteomic analysis identified 41 differentially expressed proteins in 10 kDa-CM, among which anti-inflammation, proangiogenesis, and antiapoptosis were related to cardiac protection. This study indicates that 10 kDa-CM protects H9c2 cardiomyocytes from H/R injury by preserving most of the protective factors, such as VEGF, HGF, and IGF, in MSC-CM.

The optimization conditions of establishing an H9c2 cardiomyocyte hypoxia/reoxygenation injury model based on an AnaeroPack System.

Ischemia-reperfusion (I/R) injury is a major cause of cardiomyocyte apoptosis after vascular recanalization, which was mimicked by a hypoxia/reoxygenation (H/R) injury model of cardiomyocytes in vitro. In this study, we explored an optimal H/R duration procedure using the AnaeroPack System. To study the H/R procedure, cardiomyocytes were exposed to the AnaeroPack System with sugar and serum-free medium, followed by reoxygenation under normal conditions. Cell injury was detected through lactate dehydrogenase (LDH) and cardiac troponin (c-Tn) release, morphological changes, cell apoptosis, and expression of apoptosis-related proteins. The results showed that the damage to H9c2 cells increased with prolonged hypoxia time, as demonstrated by increased apoptosis rate, LDH and c-Tn release, HIF-1α expression, as well as decreased expression of Bcl-2. Furthermore, hypoxia for 10 h and reoxygenation for 6 h exhibited the highest apoptosis rate and damage and cytokine release; in addition, cells were deformed, small, and visibly round. After 12 h of hypoxia, the majority of the cells were dead. Taken together, this study showed that subjecting H9c2 cells to the AnaeroPack System for 10 h and reoxygenation for 6 h can achieve a practicable and repeatable H/R injury model.

Synthesis of folate­chitosan nanoparticles loaded with ligustrazine to target folate receptor positive cancer cells.

In addition to its vasodilatory effect, ligustrazine (LZ) improves the sensitivity of multidrug resistant cancer cells to chemotherapeutic agents. To enhance the specificity of LZ delivery to tumor cells/tissues, folate­chitosan nanoparticles (FA­CS­NPs) were synthesized by combination of folate ester with the amine group on chitosan to serve as a delivery vehicle for LZ (FA­CS­LZ­NPs). The structure of folate­chitosan and characteristics of FA­CS­LZ­NPs, including its size, encapsulation efficiency, loading capacity and release rates were analyzed. MCF­7 (folate receptor­positive) and A549 (folate receptor­negative) cells cultured with or without folate were treated with FA­CS­LZ­NPs, CS­LZ­NPs or LZ to determine cancer­targeting specificity of FA­CS­LZ­NPs. Fluorescence intensity of intracellular LZ was observed by laser scanning confocal microscopy, and concentration of intracellular LZ was detected by HPLC. The average size of FA­CS­LZ­NPs was 182.7±0.56 nm, and the encapsulation efficiency and loading capacity was 59.6±0.23 and 15.3±0.16% respectively. The cumulative release rate was about 95% at pH 5.0, which was higher than that at pH 7.4. There was higher intracellular LZ accumulation in MCF­7 than that in A549 cells and intracellular LZ concentration was not high when MCF­7 cells were cultured with folate. These results indicated that the targeting specificity of FA­CS­LZ­NPs was mediated by folate receptor. Therefore, the FA­CS­LZ­NPs may be a potential folate receptor­positive tumor cell targeting drug delivery system that could possibly overcome multidrug resistance during cancer therapy.

ARLTS1 polymorphism is associated with an increased risk of familial cancer: evidence from a meta-analysis.

Adenosine diphosphate (ADP)-ribosylation factor-like tumour suppressor gene 1(ARLTS1) might be associated with an increased risk of several types of familial cancers. However, previous studies have shown that cancer susceptibility is not completely consistent with ARLTS1 polymorphisms, and the precise mechanism remains unknown. Therefore, we conducted a meta-analysis of case-control studies by searching the PubMed, Embase, OVID, Science Direct and Chinese National Knowledge Infrastructure (CNKI) databases. In total, 12 studies met the inclusion criteria and were included in this meta-analysis. Statistical analyses were performed using STATA 11.0 software. Overall, the Cys148Arg T > C variant significantly increased cancer risk (CC vs. TT: OR = 1.27, 95% CI = 1.15-1.41, P < 0.05). The stratification indicated that the Cys148Arg variant is significantly associated with sporadic cancer (CC vs. TT: OR = 1.36, 95% CI = 1.18-1.55) and familial cancer (CC vs. TT: OR = 1.26, 95% CI = 1.12-1.43). Trp149Stop, Pro131Leu, Ser99Ser and Leu132Leu were not correlated with cancer susceptibility. Based on these results, we demonstrated that the ARLTS1 Cys148Arg polymorphism is associated with an increased risk of sporadic cancer and familial cancer, and there were no associations between the other four SNPs (i.e., Trp149Stop, Pro131Leu, Ser99Ser and Leu132Leu) and cancer risk.

[Studies on HPLC fingerprints of tanshinone microemulsion].

OBJECTIVE: To establish the HPLC fingerprints of tanshinone microemulsion. METHOD: HPLC analysis was carried on Hypersil C18(4.6 mm x 250 mm, 5 microm) analytical column; the acetonitrile and 0.5% phosphoric acid solution were used as mobile phases with gradient elution at a flow rate of 1.0 mL x min(-1). The UV detection wavelength was set at 270 nm. RESULT: Six peaks on HPLC fingerprint of Tanshinone Microemulsion are indexed. CONCLUSION: The method developed in the present study is convenient, reliable, and can be used as a quality control method for Tanshinone Microemulsion.