Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Zebrafish xenograft model for studying mechanism and treatment of non-small cell lung cancer brain metastasis

doi: 10.1186/s13046-021-02173-5

Figure Lengend Snippet: Establishment of zebrafish NSCLC BM xenograft models. ( A ) Schematic diagram of imaging and collecting zebrafish brain. The zebrafish brain was imaged from vertical view (blue arrow) and excised (red dotted box) to extract RNA. ( B ) The schematic diagram showed zebrafish embryos at 2 dpf. The red dots indicated cancer cells injected into the perivitelline space (PVs). ( C ) Images of brain at different developmental stages of Tg ( fli-1 : EGFP) zebrafish (2-6 dpf). The white short dashed line indicated the midbrain of the zebrafish, and the white long dashed line indicated the zebrafish’s eyes and hindbrain. ( D ) The expression of claudin-5 in the zebrafish brain at different developmental stages (2-7 dpf). ( E ) The expression of mfsd2aa and mfsd2ab in the zebrafish brain at different developmental stages (2-6 dpf). ( F ) Survival curves of 2 dpf zebrafish with different tumor-bearing amount in PVs. About 50-400 H1975 cells (red fluorescence) were injected into the PVs of 2 dpf zebrafish, and the number of deaths was counted till 8 dpi. ( G - I ) Three human NSCLC cell lines were involved: H1975, A549 and H1299. About 100 cells were injected into the PVs of zebrafish at different developmental stages, and the brain of zebrafish was imaged at 1 dpi. The white arrows indicated cancer cells in blood vessels of zebrafish brain. ( J - K ) Quantification of the BM cells number in zebrafish at 1 dpi. Significance was considered when P values were lower than 0.05. (ns) indicated statistical insignificance, (*) indicated statistical significance P < 0.05, (**) P < 0.01 and (***) P < 0.001. dpf: days post fertilization, dpi: days post injection

Article Snippet: Human NSCLC lung cancer cell lines, A549, H1975, H1299, originally from American Type Culture Collection, were cultured in accordance with standard requirements.

Techniques: Imaging, Injection, Expressing, Fluorescence

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Zebrafish xenograft model for studying mechanism and treatment of non-small cell lung cancer brain metastasis

doi: 10.1186/s13046-021-02173-5

Figure Lengend Snippet: NSCLC cells could not proliferate in the midbrain of zebrafish at 4 dpi. ( A ) The schematic diagram showed zebrafish embryos at 2 dpf. The red dots indicated cancer cells injected into the midbrain of zebrafish. ( B ) Survival curves of zebrafish with different tumor-bearing amount at midbrain. About 50-400 H1975 cells (red fluorescence) were injected into the midbrain of 2 dpf Tg ( fli-1 : EGFP) zebrafish, and the number of deaths was counted till 8 dpi. ( C - G ) Five human cancer cell lines were involved: human breast cancer cell lines MDA-MB-231 (positive control) and MCF-7 (negative control), human NSCLC cancer lines H1975, A549 and H1299. About 100 cells were injected into the midbrain of zebrafish at 2 dpf, and the midbrain of zebrafish was imaged at 1 dpi and 4 dpi. (H-M) Quantification of cell proliferation in the midbrain of zebrafish at 1 dpi and 4 dpi. The fold change of cells in the midbrain was determined by dividing the measured values at 1 dpi and 4 dpi by the average measured values at 1 dpi. (ns) indicated statistical insignificance, (*) indicated statistical significance P < 0.05, (**) P < 0.01 and (***) P < 0.001

Article Snippet: Human NSCLC lung cancer cell lines, A549, H1975, H1299, originally from American Type Culture Collection, were cultured in accordance with standard requirements.

Techniques: Injection, Fluorescence, Positive Control, Negative Control

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Zebrafish xenograft model for studying mechanism and treatment of non-small cell lung cancer brain metastasis

doi: 10.1186/s13046-021-02173-5

Figure Lengend Snippet: Zebrafish NSCLC BM xenograft models discriminated the BM potentials of different cell lines. ( A ) Transwell culture system was used to assess cell invasion ability in vitro , and five cancer cell lines were involved: MDA-MB-231, MCF-7, H1975, A549 and H1299. Invaded cells were stained with crystal violet (0.5 %) and imaged after 24 h incubation. ( B ) Quantification and comparison of invaded cells in vitro . Colonies were quantified using Image Pro Plus. ( C ) Five cancer cell lines were involved: MDA-MB-231, MCF-7, H1975, A549 and H1299. About 100 cells (red fluorescence) were injected into the PVs of Tg ( fli-1 : EGFP) zebrafish at 2 dpf, and the brain of zebrafish was imaged at 4 dpi. The white arrows indicated cancer cells in blood vessels of zebrafish brain. ( D ) Quantification of BM rate at 4 dpi. The number of BM cells in the same zebrafish at 4 dpi was divided by the number of BM cells at 1 dpi. The ratio of these two was named as BM potential. If the BM potential was greater than 1, it was considered that the zebrafish had brain metastasis. ( E - I ) Quantification of BM cells at 1 dpi and 4 dpi. The two dots connected by a straight line represented the number of BM cells of the same zebrafish at 1 dpi and 4 dpi. ( J ) Quantification and comparison of cell BM potential. The BM potential was determined by dividing the number of BM cells in the same zebrafish at 4 dpi by the number of BM cells at 1 dpi. (ns) indicated statistical insignificance, (*) indicated statistical significance P < 0.05, (**) P < 0.01 and (***) P < 0.001

Article Snippet: Human NSCLC lung cancer cell lines, A549, H1975, H1299, originally from American Type Culture Collection, were cultured in accordance with standard requirements.

Techniques: In Vitro, Staining, Incubation, Comparison, Fluorescence, Injection

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Zebrafish xenograft model for studying mechanism and treatment of non-small cell lung cancer brain metastasis

doi: 10.1186/s13046-021-02173-5

Figure Lengend Snippet: Zebrafish NSCLC BM xenograft models simultaneously discriminated the BM potentials of different cell lines. ( A ) The schematic diagram showed zebrafish embryos at 2 dpf. The red dots and green dots indicated cancer cells labeled with two different dyes were co-injected into the PVs of zebrafish. ( B ) About 50 H1975 cells (red fluorescence) and 50 H1975 cells (green fluorescence) were co-injected into the PVs of wild zebrafish at 2 dpf, and the brain of zebrafish was imaged at 4 dpi. The white long dashed line indicated the brain of zebrafish. ( C ) About 50 H1975 cells (red fluorescence) and 50 A549 cells (green fluorescence) were co-injected into the PVs of wild zebrafish at 2 dpf, and the brain of zebrafish was imaged at 4 dpi. ( D - G ) Quantification of BM cells in co-injection at 1 dpi and 4 dpi. The two dots connected by a straight line represented the number of BM cells of the same zebrafish at 1 dpi and 4 dpi. ( H and I ) The percentage of cells with two different dyes in the brain of zebrafish. The percentage of the cell line (red/green fluorescence) at 1 dpi was determined by dividing the number of BM cells (red/green fluorescence) by the total number of BM cells at 1 dpi. (ns) indicated statistical insignificance, (*) indicated statistical significance P < 0.05, (**) P < 0.01 and (***) P < 0.001

Article Snippet: Human NSCLC lung cancer cell lines, A549, H1975, H1299, originally from American Type Culture Collection, were cultured in accordance with standard requirements.

Techniques: Labeling, Injection, Fluorescence

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Zebrafish xenograft model for studying mechanism and treatment of non-small cell lung cancer brain metastasis

doi: 10.1186/s13046-021-02173-5

Figure Lengend Snippet: Expression of microRNA-330-3p (miR-330-3p) affected the NSCLC BM potential in zebrafish xenograft models. ( A and B ) Quantification of the miR-330-3p expression in vitro . The appropriate transfection concentration was determined between 1 nM and 50 nM, according to the instructions. ( C and D ) The expression of miR-330-3p in H1975 cell line was knocked down by anti-miR-330-3p inhibitor and the expression of miR-330-3p in A549 cell line was overexpressed by over-miR-330-3p mimics. Transwell culture system was used to evaluate the effectiveness of transfection in vitro . Invaded cells were stained with crystal violet (0.5 %) and imaged after 24 h incubation. ( E and F ) Quantification of invaded cells in vitro . Colonies were quantified using Image Pro Plus. ( G and H ) H1975, H1975 with under-expressed miR-330-3p, A549 and A549 with over-expressed miR-330-3p were involved. About 100 cells (red fluorescence) were injected into the PVs of Tg ( fli-1 : EGFP) zebrafish at 2 dpf to evaluate the effectiveness of transfection in vivo , and the brain of zebrafish was imaged at 4 dpi. The white arrows indicated cancer cells in blood vessels of zebrafish brain. ( I - L ) Quantification of BM cells at 1 dpi and 4 dpi. The two dots connected by a straight line represented the number of BM cells of the same zebrafish at 1 dpi and 4 dpi. ( M and N ) Quantification and comparison of the BM potentials of the transfected cell lines and the original cell lines. (ns) indicated statistical insignificance, (*) indicated statistical significance P < 0.05, (**) P < 0.01 and (***) P < 0.001

Article Snippet: Human NSCLC lung cancer cell lines, A549, H1975, H1299, originally from American Type Culture Collection, were cultured in accordance with standard requirements.

Techniques: Expressing, In Vitro, Transfection, Concentration Assay, Staining, Incubation, Fluorescence, Injection, In Vivo, Comparison

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Zebrafish xenograft model for studying mechanism and treatment of non-small cell lung cancer brain metastasis

doi: 10.1186/s13046-021-02173-5

Figure Lengend Snippet: Zebrafish NSCLC BM xenograft models discriminated different chemosensitivities within 4 days. ( A and B ) NSCLC cell lines H1975 and A549 were involved. Transwell culture system was used to evaluate the effectiveness of osimertinib (1 µM) and gefitinib (2 µM) against cell invasion in vitro . Invaded cells were stained with crystal violet (0.5 %) and imaged after 24 h incubation. ( C and D ) Quantification of invaded cells in vitro. Colonies were quantified using Image Pro Plus. ( E and F ) NSCLC cell lines H1975 and A549 were involved. Tg ( fli-1 : EGFP) zebrafish embryos were injected about 100 cells (red fluorescence) into the PVs at 2 dpf and administered with osimertinib (1 µM) and gefitinib (13 µM) by intracardiac injection. ( G and H ) Quantification of cell brain metastasis at 4 dpi. Fold change of BM cell number was determined by dividing the number of BM cells in the same zebrafish at 4 dpi by the number of BM cells at 1 dpi. (ns) indicated statistical insignificance, (*) indicated statistical significance P < 0.05, (**) P < 0.01 and (***) P < 0.001

Article Snippet: Human NSCLC lung cancer cell lines, A549, H1975, H1299, originally from American Type Culture Collection, were cultured in accordance with standard requirements.

Techniques: In Vitro, Staining, Incubation, Injection, Fluorescence

Journal: Cancer Research

Article Title: Regression of Melanoma in a Murine Model by RLIP76 Depletion

doi: 10.1158/0008-5472.can-05-3534

Figure Lengend Snippet: Figure 1. Comparison of RLIP76 levels in cultured malignant cells versus nonmalignant cells. Aliquots of crude detergent extracts of the membrane fractions of malignant cells (H1618, H358, OVCAR-3, PC-3, MCF-7, B16, HepG2, and DG-1) and nonmalignant cells (HAVSM, HUVEC, HLMVEC, and HLBEC), containing 200 Ag protein were used for SDS-PAGE and Western blotting against anti-RLIP76 IgG as primary antibody and horseradish peroxidase–conjugated goat anti-rabbit IgG as secondary antibody and developed with 4-chloro-1-napthol as chromogenic substrate. Results were quantified by scanning densitometry of the full-length RLIP76 protein band near 109 kDa. h-Actin was used as an internal control.

Article Snippet: Human small cell lung cancer (SCLC) lines H1618 and non–small cell lung cancer (NSCLC) lines H358 (bronchioalveolar), PC-3 (human prostate), and B16 mouse skin melanoma were purchased from American Type Culture Collection (Manassas, VA).

Techniques: Comparison, Cell Culture, Membrane, SDS Page, Western Blot, Control

Journal: Journal of medicinal chemistry

Article Title: A Potent and Highly Efficacious Bcl-2/Bcl-xL Inhibitor

doi: 10.1021/jm4001105

Figure Lengend Snippet: Antitumor activity of compounds 31 and 32 in the H146 small-cell lung cancer xenograft model in SCID mice. Tumors were grown to an average size of 126 mm3 and compound 31 or 32 was administered at 15 mg/kg intravenously, daily, 5 days a week for 2 weeks. (a). Tumor growth. (b). Animal body weight.

Article Snippet: 7 Human small cell lung cancer cell lines H146, H1963, H187, and H1417 were purchased from American Type Culture Collection (ATCC) and were maintained in RPMI-1640 medium containing 10% FBS.

Techniques: Activity Assay