Journal: International Journal of Biological Sciences

Article Title: Cancer-associated adipocytes promote the invasion and metastasis in breast cancer through LIF/CXCLs positive feedback loop

doi: 10.7150/ijbs.65227

Figure Lengend Snippet: LIF promotes breast cancer cell migration and invasion by activating Stat3 signaling. ( A ) After co-culturing with breast epithelial cells or breast cancer cells for 24 h, the LIF mRNA expression of adipocytes in each group was detected by q-PCR. ( B ) After co-culturing, the breast epithelial cells or breast cancer cells in the upper chamber were removed, adipocytes were further cultured with serum-free DMEM for another 24 h, and the LIF in the medium was quantified by ELISA. ( C ) Breast cancer MDA-MD-231 cells were co-cultured with adipocytes for 24 h, q-PCR detected the expression of LIFR and gp130 mRNA in breast cancer cells. ( D ) IHC detected the expression of LIF in normal breast tissue and breast cancer tissue, the highly expressed LIF is pointed by the black arrow. The corresponding quantization chart is shown on the right, scale bar: 200 μm. ( E and F ) MDA-MD-231 cells were treated with rhLIF and subjected to migration ( E ) and Transwell Matrigel invasion ( F ) assays, the corresponding quantization chart is shown on the right. ( G and H ) MDA-MD-231 cells were treated with 0, 20, 40, 80, 160 ng/mL rhLIF for 15 min and were treated with rhLIF for different time (0, 5, 15, 30, 60 min), the Stat3 phosphorylation was analyzed by western blot. Typical microscopic fields and blots are shown and quantitative data are presented as mean ± SD from at least three independent experiments. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: MDA-MB-231, MCF-7 and 3T3-L1 cells were maintained in DMEM (#10-013-CV, Corning) containing 10% FBS (#10099141C, Gibco), BT549 cells were maintained in RPMI Medium 1640 (#10-040-CV, Corning) containing 10% FBS, 4T1 were maintained in RPMI 1640 [+] L-glutamine (#99-595-CM, Corning) containing 10% FBS and MCF10A cells were maintained in MCF10A special medium (#CM-0525, Procell).

Techniques: Migration, Expressing, Cell Culture, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: International Journal of Biological Sciences

Article Title: Cancer-associated adipocytes promote the invasion and metastasis in breast cancer through LIF/CXCLs positive feedback loop

doi: 10.7150/ijbs.65227

Figure Lengend Snippet: LIF neutralizing antibody inhibits the migration and invasion capability and the Stat3 phosphorylation induced by CAA-CM on breast cancer cells. (A and B) MDA-MD-231 cells cultured in Adi-CM or CAA-CM or control DMEM were treated with LIF neutralizing antibody/IgG. The cell migration ( A ) and invasion ( B ) situation were monitored under a phase contrast microscope. The corresponding quantization chart is shown on the right. ( C ) MDA-MB-231 cells were treated with different culture media for 15 min, and Stat3 phosphorylation was analyzed by western blot. ( D ) MDA-MD-231 cells cultured in DMEM or Adi-CM or CAA-CM were treated with LIF neutralizing antibody or IgG for 15 min, and Stat3 phosphorylation was analyzed by western blot. Typical microscopic fields and blots are shown and quantitative data are presented as mean ± SD from at least three independent experiments. # p < 0.05, ## p < 0.01, ### p < 0.001, CAA-CM VS. Mock; * p < 0.05, ** p < 0.01, *** p < 0.001, CAA-CM + α-LIF VS. CAA-CM.

Article Snippet: MDA-MB-231, MCF-7 and 3T3-L1 cells were maintained in DMEM (#10-013-CV, Corning) containing 10% FBS (#10099141C, Gibco), BT549 cells were maintained in RPMI Medium 1640 (#10-040-CV, Corning) containing 10% FBS, 4T1 were maintained in RPMI 1640 [+] L-glutamine (#99-595-CM, Corning) containing 10% FBS and MCF10A cells were maintained in MCF10A special medium (#CM-0525, Procell).

Techniques: Migration, Cell Culture, Microscopy, Western Blot

Journal: International Journal of Biological Sciences

Article Title: Cancer-associated adipocytes promote the invasion and metastasis in breast cancer through LIF/CXCLs positive feedback loop

doi: 10.7150/ijbs.65227

Figure Lengend Snippet: Stattic inhibits the migration and invasion of MDA-MB-231 cells and Stat3 phosphorylation induced by rhLIF and CAA-CM. ( A and B ) MDA-MD-231 cells were stimulated with rhLIF or DMEM with Stattic and subjected to migration ( A ) and Transwell Matrigel invasion ( B ) assays. The corresponding quantization chart is shown on the right. ( C ) MDA-MD-231 cells were stimulated by rhLIF combined with Stattic for 15 min, and Stat3 phosphorylation was analyzed by western blot. ( D and E ) MDA-MD-231 cells were stimulated by DMEM, Adi-CM or CAA-CM combined with Stattic and subjected to migration ( D ) and Transwell Matrigel invasion ( E ) assays. The corresponding quantization chart is shown on the right. ( F ) MDA-MD-231 cells were treated with DMEM or different adipocyte culture medium and Stattic for 15 min, and Stat3 phosphorylation was analyzed by western blot. Typical microscopic fields and blots are shown and quantitative data are presented as mean ± SD from at least three independent experiments. # p < 0.05, ## p < 0.01, ### p < 0.001, rhLIF or CAA-CM VS. Con; * p < 0.05, ** p < 0.01, *** p < 0.001, rhLIF + Stattic VS. rhLIF or CAA-CM + Stattic VS. CAA-CM.

Article Snippet: MDA-MB-231, MCF-7 and 3T3-L1 cells were maintained in DMEM (#10-013-CV, Corning) containing 10% FBS (#10099141C, Gibco), BT549 cells were maintained in RPMI Medium 1640 (#10-040-CV, Corning) containing 10% FBS, 4T1 were maintained in RPMI 1640 [+] L-glutamine (#99-595-CM, Corning) containing 10% FBS and MCF10A cells were maintained in MCF10A special medium (#CM-0525, Procell).

Techniques: Migration, Western Blot

Journal: Oncology Research

Article Title: Comparative assessment of antitumor effects between doxorubicin and mitochondria-targeted doxorubicin in combination with radiotherapy

doi: 10.32604/or.2025.058997

Figure Lengend Snippet: TPP-DOX induces cell apoptosis in 4T1 cells while decreasing mitochondrial membrane and increasing ROS levels. (A) Detection of apoptosis level of breast cancer 4T1 cells treated with DOX or TPP-DOX by flow cytometry; (B) ATP secretion (n = 3, * p < 0.05, *** p < 0.001); (C and D) Semi-quantitative analysis of mitochondrial membrane potential (bar = 50 μm) and green fluorescence intensity of JC-1 monomer in 4T1 cells before and after treatment with DOX or TPP-DOX combined with radiotherapy (n = 3, ns, p > 0.05, *** p < 0.001); (E) mtDNA quantity and quality (n = 3, ** p < 0.01); (F and G) Relative production of ROS in 4T1 cells before and after DOX or TPP-DOX combined with radiotherapy (n = 18, ** p < 0.01, *** p < 0.001, **** p < 0.0001); (H) transmission electron microscopy (TEM) of mitochondria (bar = 1 µm). The red arrow indicates the magnified area of the image, where the mitochondrial morphology is prominently displayed.

Article Snippet: Gibco fetal bovine serum (Thermo Fisher Scientific, A5670701, Shanghai, China); RPMI-1640 complete medium (Zhejiang Senrui Biotechnology Co., Ltd., CR-31800, Huzhou, China); 0.25% trypsin & 0.02% EDTA (Zhejiang Senrui Biotechnology Co., Ltd., CR-25200); 4% Paraformaldehyde (Beijing Suolaibao Technology Co., Ltd., P1110, Beijing, China); Penicillin-streptomycin double antibody solution (Tianjin Haoyang Biological Products Technology Co., Ltd., TBD20180091, Tianjin, China); CCK-8 cytotoxicity detection kit (Shanghai Beyotime Biotechnology Co., Ltd., C0037, Shanghai, China); Apoptosis detection Reagent kit (Shanghai Yisheng Biotechnology Co., Ltd., 40311ES20, Shanghai, China); Active oxygen detection kit (Shanghai Beyotime Biotechnology Co., Ltd., S0033S); Hoechst 33342 staining solution (Shanghai Beyotime Biotechnology Co., Ltd., C1022); Hanks buffer (Shanghai Macklin Biochemical Technology Co., Ltd., H917811); Calcein AM/PI double staining kit (Elabscience Biotechnology Co., Ltd, E-CK-A354, Wuhan, China); Mitochondrial membrane potential detection kit (JC-1) (Shanghai Beyotime Biotechnology Co., Ltd., C1071M); Mitochondrial fluorescent probe (Mito-Tracker) (Shanghai Beyotime Biotechnology Co., Ltd., C1032); Caspase kit (Shanghai Beyotime Biotechnology Co., Ltd., C1168M); Ki67 Cell Proliferation Assay Kit (Shanghai Beyotime Biotechnology Co., Ltd., C2312S); Cytochrome C antibody at dilution of 1:200 (Shanghai Beyotime Biotechnology Co., Ltd., AC909); CD31 Rabbit Polyclonal Antibody at dilution of 1:100 (Shanghai Beyotime Biotechnology Co., Ltd., AF6408); Immunol Staining Blocking Buffer (Shanghai Beyotime Biotechnology Co., Ltd., P0102); p63 Rabbit Monoclonal Antibody (Shanghai Beyotime Biotechnology Co., Ltd., AG8597); TUNEL Apoptosis Detection Kit (YEASEN biotechnology (Shanghai) Co., Ltd., 40306ES60, Shanghai, China); Hematoxylin-eosin (H&E) staining (Shanghai Beyotime Biotechnology Co., Ltd., C0105S); Mitochondrial DNA detection kit (Wuhan Servicebio Technology CO., LTD, GM2019, Wuhan, China); ATP Assay Kit (MedChemExpress, HY-K0314, Shanghai, China); MCF-10A cell-specific medium (Wuhan Pricella Biotechnology Co., Ltd., CM-0525, Wuhan, China).

Techniques: Membrane, Flow Cytometry, Fluorescence, Transmission Assay, Electron Microscopy

Journal: Oncology Research

Article Title: Comparative assessment of antitumor effects between doxorubicin and mitochondria-targeted doxorubicin in combination with radiotherapy

doi: 10.32604/or.2025.058997

Figure Lengend Snippet: TPP-DOX induces cell apoptosis in 4T1 cells while decreasing mitochondrial membrane and increasing ROS levels. (A) Detection of apoptosis level of breast cancer 4T1 cells treated with DOX or TPP-DOX by flow cytometry; (B) ATP secretion (n = 3, * p < 0.05, *** p < 0.001); (C and D) Semi-quantitative analysis of mitochondrial membrane potential (bar = 50 μm) and green fluorescence intensity of JC-1 monomer in 4T1 cells before and after treatment with DOX or TPP-DOX combined with radiotherapy (n = 3, ns, p > 0.05, *** p < 0.001); (E) mtDNA quantity and quality (n = 3, ** p < 0.01); (F and G) Relative production of ROS in 4T1 cells before and after DOX or TPP-DOX combined with radiotherapy (n = 18, ** p < 0.01, *** p < 0.001, **** p < 0.0001); (H) transmission electron microscopy (TEM) of mitochondria (bar = 1 µm). The red arrow indicates the magnified area of the image, where the mitochondrial morphology is prominently displayed.

Article Snippet: Gibco fetal bovine serum (Thermo Fisher Scientific, A5670701, Shanghai, China); RPMI-1640 complete medium (Zhejiang Senrui Biotechnology Co., Ltd., CR-31800, Huzhou, China); 0.25% trypsin & 0.02% EDTA (Zhejiang Senrui Biotechnology Co., Ltd., CR-25200); 4% Paraformaldehyde (Beijing Suolaibao Technology Co., Ltd., P1110, Beijing, China); Penicillin-streptomycin double antibody solution (Tianjin Haoyang Biological Products Technology Co., Ltd., TBD20180091, Tianjin, China); CCK-8 cytotoxicity detection kit (Shanghai Beyotime Biotechnology Co., Ltd., C0037, Shanghai, China); Apoptosis detection Reagent kit (Shanghai Yisheng Biotechnology Co., Ltd., 40311ES20, Shanghai, China); Active oxygen detection kit (Shanghai Beyotime Biotechnology Co., Ltd., S0033S); Hoechst 33342 staining solution (Shanghai Beyotime Biotechnology Co., Ltd., C1022); Hanks buffer (Shanghai Macklin Biochemical Technology Co., Ltd., H917811); Calcein AM/PI double staining kit (Elabscience Biotechnology Co., Ltd, E-CK-A354, Wuhan, China); Mitochondrial membrane potential detection kit (JC-1) (Shanghai Beyotime Biotechnology Co., Ltd., C1071M); Mitochondrial fluorescent probe (Mito-Tracker) (Shanghai Beyotime Biotechnology Co., Ltd., C1032); Caspase kit (Shanghai Beyotime Biotechnology Co., Ltd., C1168M); Ki67 Cell Proliferation Assay Kit (Shanghai Beyotime Biotechnology Co., Ltd., C2312S); Cytochrome C antibody at dilution of 1:200 (Shanghai Beyotime Biotechnology Co., Ltd., AC909); CD31 Rabbit Polyclonal Antibody at dilution of 1:100 (Shanghai Beyotime Biotechnology Co., Ltd., AF6408); Immunol Staining Blocking Buffer (Shanghai Beyotime Biotechnology Co., Ltd., P0102); p63 Rabbit Monoclonal Antibody (Shanghai Beyotime Biotechnology Co., Ltd., AG8597); TUNEL Apoptosis Detection Kit (YEASEN biotechnology (Shanghai) Co., Ltd., 40306ES60, Shanghai, China); Hematoxylin-eosin (H&E) staining (Shanghai Beyotime Biotechnology Co., Ltd., C0105S); Mitochondrial DNA detection kit (Wuhan Servicebio Technology CO., LTD, GM2019, Wuhan, China); ATP Assay Kit (MedChemExpress, HY-K0314, Shanghai, China); MCF-10A cell-specific medium (Wuhan Pricella Biotechnology Co., Ltd., CM-0525, Wuhan, China).

Techniques: Membrane, Flow Cytometry, Fluorescence, Transmission Assay, Electron Microscopy

Journal: Oncology Research

Article Title: Comparative assessment of antitumor effects between doxorubicin and mitochondria-targeted doxorubicin in combination with radiotherapy

doi: 10.32604/or.2025.058997

Figure Lengend Snippet: CYT C, Ki67, CD31, TUNEL staining image of mouse tumor tissue after treatment (bar = 50 μm).

Article Snippet: Gibco fetal bovine serum (Thermo Fisher Scientific, A5670701, Shanghai, China); RPMI-1640 complete medium (Zhejiang Senrui Biotechnology Co., Ltd., CR-31800, Huzhou, China); 0.25% trypsin & 0.02% EDTA (Zhejiang Senrui Biotechnology Co., Ltd., CR-25200); 4% Paraformaldehyde (Beijing Suolaibao Technology Co., Ltd., P1110, Beijing, China); Penicillin-streptomycin double antibody solution (Tianjin Haoyang Biological Products Technology Co., Ltd., TBD20180091, Tianjin, China); CCK-8 cytotoxicity detection kit (Shanghai Beyotime Biotechnology Co., Ltd., C0037, Shanghai, China); Apoptosis detection Reagent kit (Shanghai Yisheng Biotechnology Co., Ltd., 40311ES20, Shanghai, China); Active oxygen detection kit (Shanghai Beyotime Biotechnology Co., Ltd., S0033S); Hoechst 33342 staining solution (Shanghai Beyotime Biotechnology Co., Ltd., C1022); Hanks buffer (Shanghai Macklin Biochemical Technology Co., Ltd., H917811); Calcein AM/PI double staining kit (Elabscience Biotechnology Co., Ltd, E-CK-A354, Wuhan, China); Mitochondrial membrane potential detection kit (JC-1) (Shanghai Beyotime Biotechnology Co., Ltd., C1071M); Mitochondrial fluorescent probe (Mito-Tracker) (Shanghai Beyotime Biotechnology Co., Ltd., C1032); Caspase kit (Shanghai Beyotime Biotechnology Co., Ltd., C1168M); Ki67 Cell Proliferation Assay Kit (Shanghai Beyotime Biotechnology Co., Ltd., C2312S); Cytochrome C antibody at dilution of 1:200 (Shanghai Beyotime Biotechnology Co., Ltd., AC909); CD31 Rabbit Polyclonal Antibody at dilution of 1:100 (Shanghai Beyotime Biotechnology Co., Ltd., AF6408); Immunol Staining Blocking Buffer (Shanghai Beyotime Biotechnology Co., Ltd., P0102); p63 Rabbit Monoclonal Antibody (Shanghai Beyotime Biotechnology Co., Ltd., AG8597); TUNEL Apoptosis Detection Kit (YEASEN biotechnology (Shanghai) Co., Ltd., 40306ES60, Shanghai, China); Hematoxylin-eosin (H&E) staining (Shanghai Beyotime Biotechnology Co., Ltd., C0105S); Mitochondrial DNA detection kit (Wuhan Servicebio Technology CO., LTD, GM2019, Wuhan, China); ATP Assay Kit (MedChemExpress, HY-K0314, Shanghai, China); MCF-10A cell-specific medium (Wuhan Pricella Biotechnology Co., Ltd., CM-0525, Wuhan, China).

Techniques: TUNEL Assay, Staining