Journal: Oxidative Medicine and Cellular Longevity

Article Title: HSP90AB1 as the Druggable Target of Maggot Extract Reverses Cisplatin Resistance in Ovarian Cancer

doi: 10.1155/2023/9335440

Figure Lengend Snippet: ME enhances the sensitivity of A2780/CDDP and SKOV3/CDDP cells to cisplatin (Cis). Cells were treated with different concentrations of cisplatin with and without ME (6 mg/ml) for 48 h. (a) The viability of A2780 and A2780/CDDP cells was assayed using CCK8 kits. (b) The viability of SKOV3 and SKOV3/CDDP cells was assayed. SKOV3/CDDP cells were infected with lentiviral particles to express luciferase, and then cells were treated with cisplatin (3.2 μ g/ml) and/or ME (4 mg/ml or 6 mg/ml) for 48 h followed by 150 μ g/ml D-luciferin for 10 min. (c) The luciferase-positive SKOV3/CDDP cells were analyzed using Living Image software and a GloMax® 96 Microplate Luminometer. The quantification of luciferase-positive cells is shown on the right. ∗ p ≤ 0.05 vs. blank; # p ≤ 0.05 vs. cisplatin; (d) A2780/CDDP and SKOV3/CDDP cells were double stained with Annexin V-FITC and PI then analyzed using flow cytometry, and the quantitative analysis was located in the right. ∗ p ≤ 0.05, ∗∗ p ≤ 0.01, ∗∗∗ p ≤ 0.001. (e) Levels of p-p53 in A2780/DDP cells were measured by immunofluorescence staining (400x magnification). The panel under the staining shows the quantitative analysis of p-p53 in A2780/CDDP cells. The nuclei are stained with DAPI. ∗ p ≤ 0.05 vs. blank; # p ≤ 0.05 vs. cisplatin. (f) Representative images of the wound healing assays in A2780/CDDP and SKOV3/CDDP cells are shown, and the panels under the images show the wound closure rate. ∗ p ≤ 0.05 vs. 0 h; # p ≤ 0.05 vs. blank; ns means no significance vs. 0 h. (g) The transwell assay was used to analyze the invasion of A2780 and A2780/CDDP cells. The invasive cell number was quantified using ImageJ and located under the images. ∗ p ≤ 0.05. Three independent experiments were performed with similar results. Data are shown as the mean ± SEM.

Article Snippet: The human ovarian cancer cell line A2780 (RRID: CVCL_0134) was purchased from Shanghai Pituo Biological Technology Co., Ltd. (China), and the cell line SKOV3 (RRID: CVCL_0532) was purchased from Solarbio Science & Technology Co., Ltd. (China).

Techniques: Infection, Luciferase, Software, Staining, Flow Cytometry, Immunofluorescence, Transwell Assay

Journal: Oxidative Medicine and Cellular Longevity

Article Title: HSP90AB1 as the Druggable Target of Maggot Extract Reverses Cisplatin Resistance in Ovarian Cancer

doi: 10.1155/2023/9335440

Figure Lengend Snippet: Enrichment analysis of DEGs in A2780/CDDP cells based on high-throughput RNA sequencing. The total RNA from A2780 and A2780/DDP cells was extracted using RNA Miniprep kit reagents. Next-generation sequencing analysis was performed on the BGISEQ-500 platform by BGI Genomic Services. (a) Volcano plot showing significantly upregulated genes (red dots) and downregulated genes (blue dots). (b) A2780 vs. A2780/CDDP Gene Ontology (GO) analysis on a cellular component of DEGs. (c) A2780 vs. A2780/CDDP KEGG pathway enrichment analysis of DEGs. (d) The heat map shows the relative transcript levels of the DEGs in A2780 and A2780/CDDP cells. (e) The protein–protein interaction network shows the upregulated DEGs from A2780/CDDP cells compared with A2780 cells. (f) qRT-PCR analyses of the mRNA levels of MYC in A2780 cells and A2780/CDDP cells. (g) qRT-PCR analyses of the mRNA levels of HSP90AB1 in A2780 cells and A2780/CDDP cells. Three independent experiments were performed with similar results. Data are shown as the mean ± SEM. ∗ p ≤ 0.05, ∗∗∗ p ≤ 0.001.

Article Snippet: The human ovarian cancer cell line A2780 (RRID: CVCL_0134) was purchased from Shanghai Pituo Biological Technology Co., Ltd. (China), and the cell line SKOV3 (RRID: CVCL_0532) was purchased from Solarbio Science & Technology Co., Ltd. (China).

Techniques: High Throughput Screening Assay, RNA Sequencing, Next-Generation Sequencing, Quantitative RT-PCR

Journal: Oxidative Medicine and Cellular Longevity

Article Title: HSP90AB1 as the Druggable Target of Maggot Extract Reverses Cisplatin Resistance in Ovarian Cancer

doi: 10.1155/2023/9335440

Figure Lengend Snippet: ME induces cisplatin to promote apoptosis by suppressing the expression of HSP90AB1/IGF1R in cisplatin-resistant ovarian cancer cells. (a) The expression of HSP90AB1, IGF1R, and IGFBP2 in A2780 cells and A2780/CDDP cells was assessed by western blot. (b) The lower panel shows the quantitative analysis. ∗∗ p ≤ 0.01, ∗∗∗ p ≤ 0.001. (c) The expression of HSP90AB1, IGF1R, and IGFBP2 in SKOV3 cells and SKOV3/CDDP cells was assessed by western blot. (d) The lower panel shows the quantitative analysis. ∗ p ≤ 0.05, ∗∗ p ≤ 0.01. The GSH level in A2780 cells (e) and SKOV3 cells (f) was analyzed using a reduced GSH assay kit. ∗ p ≤ 0.05, ns means no significance. Relative mRNA expression of MYC (g), HSP90AB1 (h), and IGF1R (i) in A2780/CDDP cells was determined by real-time PCR. ∗ p ≤ 0.05, ∗∗∗ p ≤ 0.001, ns means no significance. A2780/CDDP and SKOV3/CDDP cells were treated with cisplatin (3.2 μ g/ml) and ME (6 mg/ml) for 48 h. The protein was extracted for further analysis. (j–m) The protein expression of HSP90AB1, IGF1R, MYC, PTEN, p-H2AX, and BCL2 in A2780/CDDP cells (j) and SKOV3/CDDP cells (l) was measured by western blot. The lower panel shows the quantitative analysis of the western blot in A2780/CDDP cells (K) and SKOV3/CDDP cells (m). ∗ p ≤ 0.05, ns means no significance. Drug-resistant cells were treated with cisplatin, ME, or both. The amount of GSH in A2780/CDDP cells (n) and SKOV3/CDDP cells (o) was analyzed using a reduced GSH assay kit. ∗ p ≤ 0.05, ns means no significance. Three independent experiments were performed with similar results. Data are shown as the mean ± SEM.

Article Snippet: The human ovarian cancer cell line A2780 (RRID: CVCL_0134) was purchased from Shanghai Pituo Biological Technology Co., Ltd. (China), and the cell line SKOV3 (RRID: CVCL_0532) was purchased from Solarbio Science & Technology Co., Ltd. (China).

Techniques: Expressing, Western Blot, GSH Assay, Real-time Polymerase Chain Reaction

Journal: Oxidative Medicine and Cellular Longevity

Article Title: HSP90AB1 as the Druggable Target of Maggot Extract Reverses Cisplatin Resistance in Ovarian Cancer

doi: 10.1155/2023/9335440

Figure Lengend Snippet: Inhibition of HSP90 ATPase activity with geldanamycin promotes apoptosis and suppresses migration in cisplatin-resistant ovarian cancer cells. A2780/CDDP cells and SKOV3/CDDP cells were pretreated with 5 μ M geldanamycin (In-HSP) for 1 h and then treated with cisplatin (3.2 μ g/ml) and ME (6 mg/ml) for 72 h. (a) A2780/CDDP cells and SKOV3/CDDP cells were double stained with Annexin V-FITC and PI and then analyzed by flow cytometry, and the quantitative analysis was located on the right. ∗ p ≤ 0.05, ∗∗ p ≤ 0.01, ∗∗∗ p ≤ 0.001. (b, c) A2780/CDDP cells and SKOV3/CDDP cells expressing luciferase were pretreated with 150 μ g/ml D-luciferin for 10 min. The luciferase-positive A2780/CDDP cells (b) and SKOV3/CDDP cells (c) were analyzed using Living Image software and a GloMax® 96 Microplate Luminometer. The quantification of luciferase-positive cells is shown on the right. ∗∗∗ p ≤ 0.001, ∗∗∗∗ p ≤ 0.0001. Representative images of the wound healing assays of A2780/CDDP cells (d) and SKOV3/CDDP cells (e) are shown, and the quantification of the wound closure rate is on the right. ∗ p ≤ 0.05 vs. 0 h; ns mean no significance vs. 0 h; ## p ≤ 0.01. (f) The expression of HSP90AB1, IGF1R, p-H2AX, p-p53, and BAX in A2780/CDDP cells was analyzed by western blot. (g) The panel on the right shows the quantitative analysis. ∗ p ≤ 0.05. (h) The expression of HSP90AB1, IGF1R, p-H2AX, p-p53, and BCL2 in SKOV3/CDDP cells was measured by western blot. (i) The panel on the right shows the quantitative analysis. ∗ p ≤ 0.05. (j–m) The coimmunoprecipitation assay. SKOV3/CDDP cells were treated with geldanamycin for 24 h. The expression levels of HSP90AB1 and IGF1R were measured (j), and the panel on the right shows the quantitative analysis (k). ∗ p ≤ 0.05. The same cell lysates were immunoprecipitated with IGF1R, HSP90AB1, and isoform-matched immunoglobulin (IgG). (l) Western blot assays of SKOV3/CDDP cells using site-specific antibodies against HSP90AB1 and IGF1R, and the panel on the right shows the quantitative analysis (m). ∗ p ≤ 0.05. Three independent experiments were performed with similar results. Data are shown as the mean ± SEM.

Article Snippet: The human ovarian cancer cell line A2780 (RRID: CVCL_0134) was purchased from Shanghai Pituo Biological Technology Co., Ltd. (China), and the cell line SKOV3 (RRID: CVCL_0532) was purchased from Solarbio Science & Technology Co., Ltd. (China).

Techniques: Inhibition, Activity Assay, Migration, Staining, Flow Cytometry, Expressing, Luciferase, Software, Western Blot, Co-Immunoprecipitation Assay, Immunoprecipitation

Journal: International Journal of Nanomedicine

Article Title: Low-intensity focused ultrasound (LIFU)-induced acoustic droplet vaporization in phase-transition perfluoropentane nanodroplets modified by folate for ultrasound molecular imaging

doi: 10.2147/IJN.S122667

Figure Lengend Snippet: In vitro FA-NDs targeting efficiency. Notes: CLSM images of SKOV3 cells after incubation with FA-NDs or non-NDs. In comparison, SKOV3 cells were first incubated with excess free FA and then FA-ND nanoemulsions under the same conditions (scale bar: 50 μm). Blue, DAPI-stained nuclei; green, DiO-labeled plasmalemma; red, DiI-labeled nanodroplets. Abbreviations: FA-ND, folate-targeted perfluoropentane nanodroplet; CLSM, confocal laser scanning microscopy; DAPI, 4′,6-diamidino-2-phenylindole; DiO, 3,3′-dioctadecyloxacarbocyanine perchlorate; DiI, 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate.

Article Snippet: The SKOV3 human ovarian cancer cell line, which was obtained from Chongqing Key Laboratory of Ultrasound Molecular Imaging (Chongqing, People’s Republic of China), was selected for this research because it overexpresses the FR.

Techniques: In Vitro, Incubation, Comparison, Staining, Labeling, Confocal Laser Scanning Microscopy