Three-dimensional(3D)cell culture methods have been validated that can replicate the tumor environment in vivo to a large extent,providing an effective tool for studying tumors.In this study,we demonstrated the use of...Three-dimensional(3D)cell culture methods have been validated that can replicate the tumor environment in vivo to a large extent,providing an effective tool for studying tumors.In this study,we demonstrated the use of standard laboratory pipette tips as micro vessels for generating 3D cell spheroids.No microfabrication or wet-chemistry surface modifications were involved in the procedure.Spheroids consisting of single or multiple cell types were generated within 24 h just by pipetting and incubating a cell suspension in pipette tips.Scanning electron microscope and optical microscope proved that the cells grew together tightly,and suggested that while gravity force might have initiated the sedimentation of cells at the bottom of the tip,the active aggregation of cells to form tight cell-cell interactions drove the formation of spheroids.Using common laboratory micropipettes and pipette tips,the rate of spheroid generation and the generation reproducibility was characterized from five boxes each with 80 tips.The ease of transferring reagents allowed modeling of the growth of microvascular endothelial cells in tumor spheroids.Moreover,the pairing and fusion of tumor spheroids could be manipulated in the pipette tips,suggesting the potential for building and assembling heterogeneous micro-tumor tissues in vitro to mimic solid tumors in vivo.This study demonstrated that spheroids can be readily and cost-effectively generated in standard biological laboratories in a timely manner using pipette tips.展开更多
OBJECTIVE To investigate apoptosis induced by photodynamic therapy with benzoporphyrin derivative monoacid ring A （BPD-MA） and explore its potential mechanism in human bladder cancer cells. METHODS Photosensitizatio...OBJECTIVE To investigate apoptosis induced by photodynamic therapy with benzoporphyrin derivative monoacid ring A （BPD-MA） and explore its potential mechanism in human bladder cancer cells. METHODS Photosensitization of BPD-MA was activated with a red light Laser （632.8nm） delivered at 10 mW/cm^2 to give a total dose of 2.4 J/cm^2. Cellular apoptosis was measured with flow cytometry analysis and an insitu terminal deoxyuridine nick end-labeling （TUNEL） assay. Changes in mitochondrial membrane potential （△φm） were monitored by a flow cy-tometric method with Rhodamine 123 staining and the expression of bcl- 2 in BIU-87 cells was detected with immunocytochemical staining. RESULTS At 8 h following photodynamic treatment, the degree of apoptosis was significantly increased when analyzed with flow cytometry and TUNEL assay. Treatment of the BIU-87 cells by PDT with BPD-MA resulted in the collapse of the △φm and a decrease of bcl-2 expression. CONCLUSION BPD-MA-mediated PDT can effectively induce apoptosis in BIU-87 cells. The mechanism probably is through a mitochondrial-initiated pathway.展开更多
OBJECTIVE To explore the effect of photodynamic therapy with benzoporphyrin derivative monoacid ring A （BPD-MA） on the proliferation and apoptosis of human bladder cancer cells. METHODS Photosensitization of BPD-MA ...OBJECTIVE To explore the effect of photodynamic therapy with benzoporphyrin derivative monoacid ring A （BPD-MA） on the proliferation and apoptosis of human bladder cancer cells. METHODS Photosensitization of BPD-MA was activated with a red light laser （632.8 nm） delivered at 10 mw/cm^2 to give a total dose of 2.4 J/cm^2. Cellular proliferative activity was measured using the 3-（4,5.- dimethylethiazil-2-yl）-2,5-Diph3-eyl tetrazolium bromide （MFi-） assay and 3H-thymidine incorporation. Cell apoptosis was determined with flow cytometry analysis and the terminal deoxyuridine nicked-labeling （TUNEL） assay. RESULTS At 24 h post photodynamic treatment, photodynamic therapy significantly decreased cellular proliferative activity. The rate of apoptosis in BIU-87 cells 8 h after photodynamic treatment significantly increased up to 26.11± 2.59% as analyzed with flow cytometry. In situ labeling of DNA cleavage products with the terminal deoxyuridine nicked-labeling （TUNEL） assay reinforced these observations, BPD-MA-mediated photosensitization increased the number of TUNEL-positive cells compared to the controls. However, laser irradiation alone, BPD-MA alone and sham radiation did not affect cellular proliferative activity or apoptosis of the human bladder cancer BIU-87 cells. CONCLUSION Photodynamic therapy with BPD-MA significantly decreases cellular proliferative activity and enhances apoptosis. Therapy using this method might be a promising approach to treat patients with bladder cancer.展开更多
基金supported by the National Natural Science Foundation of China(No.32171401)the Natural Science Foundation of Chongqing(No.CSTB2022NSCQ-MSX0808)the Specific Research Fund of the Innovation Platform for Academicians of Hainan Province(No.YSPTZX202126),China.
文摘Three-dimensional(3D)cell culture methods have been validated that can replicate the tumor environment in vivo to a large extent,providing an effective tool for studying tumors.In this study,we demonstrated the use of standard laboratory pipette tips as micro vessels for generating 3D cell spheroids.No microfabrication or wet-chemistry surface modifications were involved in the procedure.Spheroids consisting of single or multiple cell types were generated within 24 h just by pipetting and incubating a cell suspension in pipette tips.Scanning electron microscope and optical microscope proved that the cells grew together tightly,and suggested that while gravity force might have initiated the sedimentation of cells at the bottom of the tip,the active aggregation of cells to form tight cell-cell interactions drove the formation of spheroids.Using common laboratory micropipettes and pipette tips,the rate of spheroid generation and the generation reproducibility was characterized from five boxes each with 80 tips.The ease of transferring reagents allowed modeling of the growth of microvascular endothelial cells in tumor spheroids.Moreover,the pairing and fusion of tumor spheroids could be manipulated in the pipette tips,suggesting the potential for building and assembling heterogeneous micro-tumor tissues in vitro to mimic solid tumors in vivo.This study demonstrated that spheroids can be readily and cost-effectively generated in standard biological laboratories in a timely manner using pipette tips.
文摘OBJECTIVE To investigate apoptosis induced by photodynamic therapy with benzoporphyrin derivative monoacid ring A （BPD-MA） and explore its potential mechanism in human bladder cancer cells. METHODS Photosensitization of BPD-MA was activated with a red light Laser （632.8nm） delivered at 10 mW/cm^2 to give a total dose of 2.4 J/cm^2. Cellular apoptosis was measured with flow cytometry analysis and an insitu terminal deoxyuridine nick end-labeling （TUNEL） assay. Changes in mitochondrial membrane potential （△φm） were monitored by a flow cy-tometric method with Rhodamine 123 staining and the expression of bcl- 2 in BIU-87 cells was detected with immunocytochemical staining. RESULTS At 8 h following photodynamic treatment, the degree of apoptosis was significantly increased when analyzed with flow cytometry and TUNEL assay. Treatment of the BIU-87 cells by PDT with BPD-MA resulted in the collapse of the △φm and a decrease of bcl-2 expression. CONCLUSION BPD-MA-mediated PDT can effectively induce apoptosis in BIU-87 cells. The mechanism probably is through a mitochondrial-initiated pathway.
文摘OBJECTIVE To explore the effect of photodynamic therapy with benzoporphyrin derivative monoacid ring A （BPD-MA） on the proliferation and apoptosis of human bladder cancer cells. METHODS Photosensitization of BPD-MA was activated with a red light laser （632.8 nm） delivered at 10 mw/cm^2 to give a total dose of 2.4 J/cm^2. Cellular proliferative activity was measured using the 3-（4,5.- dimethylethiazil-2-yl）-2,5-Diph3-eyl tetrazolium bromide （MFi-） assay and 3H-thymidine incorporation. Cell apoptosis was determined with flow cytometry analysis and the terminal deoxyuridine nicked-labeling （TUNEL） assay. RESULTS At 24 h post photodynamic treatment, photodynamic therapy significantly decreased cellular proliferative activity. The rate of apoptosis in BIU-87 cells 8 h after photodynamic treatment significantly increased up to 26.11± 2.59% as analyzed with flow cytometry. In situ labeling of DNA cleavage products with the terminal deoxyuridine nicked-labeling （TUNEL） assay reinforced these observations, BPD-MA-mediated photosensitization increased the number of TUNEL-positive cells compared to the controls. However, laser irradiation alone, BPD-MA alone and sham radiation did not affect cellular proliferative activity or apoptosis of the human bladder cancer BIU-87 cells. CONCLUSION Photodynamic therapy with BPD-MA significantly decreases cellular proliferative activity and enhances apoptosis. Therapy using this method might be a promising approach to treat patients with bladder cancer.
