Journal: Oncotarget

Article Title: TRPC1/TRPC3 channels mediate lysophosphatidylcholine-induced apoptosis in cultured human coronary artery smooth muscles cells

doi: 10.18632/oncotarget.10853

Figure Lengend Snippet: A. Cell viability was determined with MTT assay in human coronary artery SMCs incubated with 10, 30, 60, and 90 μmol/L lysoPC for 72 h. Data are mean ± SEM (* P < 0.05, ** P < 0.01 vs . control, 0 μmol/L lysoPC). B. Cell viability was determined in human coronary artery SMCs incubated with 10, 30, and 60 μmol/L lysoPC for 24-72 h. Data are mean ± SEM ( n = 3 individual experiments, * P < 0.05, ** P < 0.01 vs . 0 day).

Article Snippet: The primary human coronary artery SMCs were purchased from ScienCell Research Laboratories (Carlsbad, CA, USA) and cultured in α-MEM/F12 (Invitrogen, Hong Kong, China).

Techniques: MTT Assay, Incubation, Control

Journal: Oncotarget

Article Title: TRPC1/TRPC3 channels mediate lysophosphatidylcholine-induced apoptosis in cultured human coronary artery smooth muscles cells

doi: 10.18632/oncotarget.10853

Figure Lengend Snippet: A. Cell cycling progression was not affected by 30 or 60 μmol/L lysoPC (24 h) in human coronary artery SMCs. B. Flow cytometry analysis was made in cells treated with 30 or 60 μmol/L (24 h) and co-stained with PI and Annexin V-FITC (V, viability; D, death; LA, late apoptosis; EA, early apoptosis) C. Percentage of cells that show viability, early apoptosis, late apoptosis and death after treatment with 30 or 60 μmol/L lysoPC ( n = 3, * P < 0.05, ** P < 0.01 vs . vehicle control). D. Representative microphotographs of TUNEL staining in human coronary artery SMCs treated with 30 or 60 μmol/L lysoPC (24 h). TUNEL labelling is stained in green and nuclei are labelled by DAPI staining in blue. Scale bar = 50 μm. E. The percentage of TUNEL-positive cells in total cells. The DAPI staining cells were countered from five randomly picked regions (** P < 0.01 vs . vehicle control). F. Detection of the caspase-3 activity in human coronary artery SMCs treated with 30 or 60 μmol/L lysoPC for 24 h. ( n = 3 individual experiments, * P < 0.05, ** P < 0.01 vs . 0, i.e. vehicle control).

Article Snippet: The primary human coronary artery SMCs were purchased from ScienCell Research Laboratories (Carlsbad, CA, USA) and cultured in α-MEM/F12 (Invitrogen, Hong Kong, China).

Techniques: Flow Cytometry, Staining, Control, TUNEL Assay, Activity Assay

Journal: Oncotarget

Article Title: TRPC1/TRPC3 channels mediate lysophosphatidylcholine-induced apoptosis in cultured human coronary artery smooth muscles cells

doi: 10.18632/oncotarget.10853

Figure Lengend Snippet: A. Cytosolic free Ca 2+ was determined in Tyrode's solution containing 5% FBS with a confocal microscopy technique in human coronary artery SMCs preloaded with Fluo-3 AM (data are mean ± SEM). The pseudoratio dF/F was applied to express intracellular free Ca 2+ level, where dF is the measured fluorescence intensity of fluo 3, and F is the initial level of fluorescence intensity. LysoPC 30 μmol/L ( n = 27) or 60 μmol/L ( n = 27, ** P < 0.01 vs . 10 μmol/L lysoPC), but not 10 μmol/L lysoPC ( n = 25), induced a significant sustained Ca 2+ influx. B. LysoPC (30 μmol/L) induced sustained Ca 2+ increase (data are mean ± SEM) in the presence of 1.8 mmol/L Ca 2+ in bath medium ( n = 28), but not in the medium with 0 Ca 2+ and 1 mmol/L EGTA ( n = 26, ** P < 0.01 vs . 1.8 mmol/L Ca 2+ ). C. Cell apoptosis determined in 1.8 and 0.9 mm/L Ca 2+ medium in human coronary artery SMCs by co-staining with PI and Annexin V-FITC (V, viability; D, death; LA, late apoptosis; EA, early apoptosis.) after treatment with 60 μmol/L lysoPC for 24 h. D. Percent values of viability, early apoptosis, late apoptosis and death after treatment with 60 μmol/L lysoPC. ( n = 3, * P < 0.05, ** P < 0.01 vs . vehicle control; # P < 0.05, ## P < 0.01 vs . 1.8 mmol/L Ca 2+ ).

Article Snippet: The primary human coronary artery SMCs were purchased from ScienCell Research Laboratories (Carlsbad, CA, USA) and cultured in α-MEM/F12 (Invitrogen, Hong Kong, China).

Techniques: Confocal Microscopy, Fluorescence, Staining, Control

Journal: Oncotarget

Article Title: TRPC1/TRPC3 channels mediate lysophosphatidylcholine-induced apoptosis in cultured human coronary artery smooth muscles cells

doi: 10.18632/oncotarget.10853

Figure Lengend Snippet: A. RT-PCR (left) and Western blots (right) of TRPC isoforms in human coronary artery SMCs (passages 3, 4, and 6). B. Western blots and ratio of protein levels of TRPC1, TRPC3 or TRPC4 channels in human coronary artery SMCs transfected with 20 or 50 nmol/L siRNAs targeting TRPC1, TRPC3 or TRPC4 channels (relative to b-actin and lipofectamine 2000, n = 3 individual experiments, * P < 0.05, ** P < 0.01 vs . control siRNA or lipofectamine). C. Ca 2+ influx (data are mean ± SEM) induced by 30 μmol/L lysoPC in cells transfected with 50 nmol/L control siRNA ( n = 27), TRPC1 siRNA ( n = 27, ** P < 0.01 vs . control siRNA), TRPC3 siRNA ( n = 28, ** P < 0.01 vs . control siRNA), or TRPC4 siRNA ( n = 28). D. Co-immunoprecipitation showing the interaction between TRPC1 and TRPC3 proteins in cells treated without or with 60 μmol/L lysoPC. IP indicates the antibody used to pull down the interacting proteins. Ig G represents the negative control.

Article Snippet: The primary human coronary artery SMCs were purchased from ScienCell Research Laboratories (Carlsbad, CA, USA) and cultured in α-MEM/F12 (Invitrogen, Hong Kong, China).

Techniques: Reverse Transcription Polymerase Chain Reaction, Western Blot, Transfection, Control, Immunoprecipitation, Negative Control

Journal: Oncotarget

Article Title: TRPC1/TRPC3 channels mediate lysophosphatidylcholine-induced apoptosis in cultured human coronary artery smooth muscles cells

doi: 10.18632/oncotarget.10853

Figure Lengend Snippet: A. Western blots of Bax, Bcl-2, caspase-3, and p-Akt(S473) in human coronary artery SMCs treated with 60 μmol/L lysoPC for 3 h or 6 h incubation with medium containing 1.8 or 0.9 mmol/L Ca 2+ . B. Relative ratio of Bax, Bcl-2, caspase-3, or p-Akt ( n = 3, * P < 0.05, ** P < 0.01 vs . vehicle; # P < 0.05, ## P < 0.01 vs .1.8 mmol/L Ca 2+ ).

Article Snippet: The primary human coronary artery SMCs were purchased from ScienCell Research Laboratories (Carlsbad, CA, USA) and cultured in α-MEM/F12 (Invitrogen, Hong Kong, China).

Techniques: Western Blot, Incubation

Journal: Oncotarget

Article Title: TRPC1/TRPC3 channels mediate lysophosphatidylcholine-induced apoptosis in cultured human coronary artery smooth muscles cells

doi: 10.18632/oncotarget.10853

Figure Lengend Snippet: A. Western blots of Bax, Bcl-2, caspase-3, and p-Akt in human coronary artery SMCs transfected with 50 nM control siRNA, TRPC1 siRNA, TRPC3 siRNA or TRPC4 siRNA for 72 h, and then treated with vehicle (V) or 60 μmol/L lysoPC for 3 and 6 h. B. Relative mean levels of Bax, Bcl-2, caspase-3, and p-Akt in human coronary artery SMCs with the same treatment as in A ( n = 3, * P < 0.05, ** P < 0.01 vs . vehicle, # P < 0.05, ## P < 0.01 vs . control siRNA).

Article Snippet: The primary human coronary artery SMCs were purchased from ScienCell Research Laboratories (Carlsbad, CA, USA) and cultured in α-MEM/F12 (Invitrogen, Hong Kong, China).

Techniques: Western Blot, Transfection, Control

Journal: ACS biomaterials science & engineering

Article Title: TiO 2 -Based Nanotopographical Cues Attenuate the Restenotic Phenotype in Primary Human Vascular Endothelial and Smooth Muscle Cells

doi: 10.1021/acsbiomaterials.9b01475

Figure Lengend Snippet: Representative HIM images of primary human coronary endothelial muscle cells on (A) flat (scale bar = 2 μm) and (B) NT90 surfaces (scale bar = 200 nm). Representative HIM images of primary human coronary SMCs on (C) flat (scale bar = 1 μm) and (D) NT90 surface (scale bar = 2 μm). Cells were fixed after 2 days of culture. Filopodia are indicated by white arrows.

Article Snippet: Cell Culture and Cell Proliferation Assays Primary human coronary artery ECs and primary human coronary artery SMCs were purchased from PromoCell (Heidelberg, Germany).

Techniques:

Journal: ACS biomaterials science & engineering

Article Title: TiO 2 -Based Nanotopographical Cues Attenuate the Restenotic Phenotype in Primary Human Vascular Endothelial and Smooth Muscle Cells

doi: 10.1021/acsbiomaterials.9b01475

Figure Lengend Snippet: EC and SMC cell coverage on flat and NT surfaces. (A) EC and (E) SMC cell numbers on flat and NT surfaces were measured using a CyQUANT assay, which quantifies DNA content in cell lysates. (B) EC and (F) SMC area were quantified using fluorescence microscopy and ImageJ. Representative images of (C,D) ECs and (G,H) SMCs cultured on flat and NT surfaces are shown. Green: phalloidin. Blue: DAPI. Scale bar: 150 μm.

Article Snippet: Cell Culture and Cell Proliferation Assays Primary human coronary artery ECs and primary human coronary artery SMCs were purchased from PromoCell (Heidelberg, Germany).

Techniques: CyQUANT Assay, Fluorescence, Microscopy, Cell Culture

Journal: ACS biomaterials science & engineering

Article Title: TiO 2 -Based Nanotopographical Cues Attenuate the Restenotic Phenotype in Primary Human Vascular Endothelial and Smooth Muscle Cells

doi: 10.1021/acsbiomaterials.9b01475

Figure Lengend Snippet: Total FAK and pFAK were quantified by ELISA using cell lysates from (A,B) ECs and (C,D) SMCs cultured on flat and NT90 surfaces. Total FAK concentration was normalized to the total protein content. The pFAK concentration was normalized to the total FAK concentration. Data are expressed as mean ± SD.

Article Snippet: Cell Culture and Cell Proliferation Assays Primary human coronary artery ECs and primary human coronary artery SMCs were purchased from PromoCell (Heidelberg, Germany).

Techniques: Enzyme-linked Immunosorbent Assay, Cell Culture, Concentration Assay

Journal: ACS biomaterials science & engineering

Article Title: TiO 2 -Based Nanotopographical Cues Attenuate the Restenotic Phenotype in Primary Human Vascular Endothelial and Smooth Muscle Cells

doi: 10.1021/acsbiomaterials.9b01475

Figure Lengend Snippet: Effect of NT topography on inflammatory response. (A) VCAM-1 gene expression in ECs. (B) SMC cell numbers when cultured on flat or NT90 surfaces in control media or under stimulation with 2 ng/mL TNFα, an inflammatory cytokine and known mitogen for SMCs. (C) MCP-1 secretion by SMCs cultured on flat or NT surfaces was measured in conditioned media using ELISA. Data are expressed as mean ± SD.

Article Snippet: Cell Culture and Cell Proliferation Assays Primary human coronary artery ECs and primary human coronary artery SMCs were purchased from PromoCell (Heidelberg, Germany).

Techniques: Expressing, Cell Culture, Enzyme-linked Immunosorbent Assay

Journal: ACS biomaterials science & engineering

Article Title: TiO 2 -Based Nanotopographical Cues Attenuate the Restenotic Phenotype in Primary Human Vascular Endothelial and Smooth Muscle Cells

doi: 10.1021/acsbiomaterials.9b01475

Figure Lengend Snippet: Effect of varying NT diameter on EC and SMC response. Cell growth rates on flat and NT surfaces using (A) ECs and (D) SMCs. Significant values are represented as follows: x NT30; # NT50; *NT90. One symbol: p < 0.05; two symbols: p < 0.01. (B) Relative VCAM-1 mRNA expression in ECs cultured on flat and NT surfaces. Values are normalized to mRNA expression on flat surfaces. (C) SMC cell numbers cultured on flat and NT surfaces in control media and in media containing 2 ng/mL TNFα. (E) MCP-1 secretion by SMCs cultured on flat and NT surfaces. Note: data shown here for flat and NT90 surfaces are identical as those in previous figures. They are shown here as a reference for comparison purposes.

Article Snippet: Cell Culture and Cell Proliferation Assays Primary human coronary artery ECs and primary human coronary artery SMCs were purchased from PromoCell (Heidelberg, Germany).

Techniques: Expressing, Cell Culture