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Mechanical forces stimulate HGF release from HSC. Isolated HSC were exposed to either shear stress or stretching to investigate changes in gene expression and HGF release in comparison to mechanically unstimulated cells. (a) Expression analyses by qPCR of ECM‐associated genes, (b) Itga5 and Itgb1 , and (c) Hgf and Il6 using RNA samples obtained from HSC exposed to fluid shear stress for 1 hr. The mRNA expression of HSC under static condition (0 dyn/cm 2 ) was set to 100% for each independent experiment ( n = 3; * p < .05 for shear stress vs. static condition). (d) Hgf expression analyses by qPCR in HSC cultured for 7 days under serum‐free conditions on uncoated or laminin‐coated <t>(LN‐211,</t> <t>LN‐521)</t> culture surfaces. Polystyrene and nanostructured surfaces with a spacing of 150 nm between gold dots were used. The mRNA expression of HSC cultured for one week on uncoated polystyrene dishes was set to 100% (broken line) for each experiment ( n = 5; * p < .05). HGF release by HSC exposed to (e–h) fluid shear stress or (i, j) mechanical stretching as analyzed by ELISA and immunofluorescence. HGF concentration of culture supernatants from static condition (0 dyn/cm 2 ) and unstretched HSC were set to 100% for each independent experiment ( n = 4; * p < .05). Immunofluorescence analysis of HGF (red) in HSC showed a high fluorescence signal intensity under (g) static condition, (h) which was lost under fluid shear stress for 1 hr. Cell nuclei are stained by DAPI (blue; scale bars: 20 µm). (a–d, f, j) Data are presented as means ± SEM
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Mechanical forces stimulate HGF release from HSC. Isolated HSC were exposed to either shear stress or stretching to investigate changes in gene expression and HGF release in comparison to mechanically unstimulated cells. (a) Expression analyses by qPCR of ECM‐associated genes, (b) Itga5 and Itgb1 , and (c) Hgf and Il6 using RNA samples obtained from HSC exposed to fluid shear stress for 1 hr. The mRNA expression of HSC under static condition (0 dyn/cm 2 ) was set to 100% for each independent experiment ( n = 3; * p < .05 for shear stress vs. static condition). (d) Hgf expression analyses by qPCR in HSC cultured for 7 days under serum‐free conditions on uncoated or laminin‐coated (LN‐211, LN‐521) culture surfaces. Polystyrene and nanostructured surfaces with a spacing of 150 nm between gold dots were used. The mRNA expression of HSC cultured for one week on uncoated polystyrene dishes was set to 100% (broken line) for each experiment ( n = 5; * p < .05). HGF release by HSC exposed to (e–h) fluid shear stress or (i, j) mechanical stretching as analyzed by ELISA and immunofluorescence. HGF concentration of culture supernatants from static condition (0 dyn/cm 2 ) and unstretched HSC were set to 100% for each independent experiment ( n = 4; * p < .05). Immunofluorescence analysis of HGF (red) in HSC showed a high fluorescence signal intensity under (g) static condition, (h) which was lost under fluid shear stress for 1 hr. Cell nuclei are stained by DAPI (blue; scale bars: 20 µm). (a–d, f, j) Data are presented as means ± SEM

Journal: Aging Cell

Article Title: Impaired integrin α 5 /β 1 ‐mediated hepatocyte growth factor release by stellate cells of the aged liver

doi: 10.1111/acel.13131

Figure Lengend Snippet: Mechanical forces stimulate HGF release from HSC. Isolated HSC were exposed to either shear stress or stretching to investigate changes in gene expression and HGF release in comparison to mechanically unstimulated cells. (a) Expression analyses by qPCR of ECM‐associated genes, (b) Itga5 and Itgb1 , and (c) Hgf and Il6 using RNA samples obtained from HSC exposed to fluid shear stress for 1 hr. The mRNA expression of HSC under static condition (0 dyn/cm 2 ) was set to 100% for each independent experiment ( n = 3; * p < .05 for shear stress vs. static condition). (d) Hgf expression analyses by qPCR in HSC cultured for 7 days under serum‐free conditions on uncoated or laminin‐coated (LN‐211, LN‐521) culture surfaces. Polystyrene and nanostructured surfaces with a spacing of 150 nm between gold dots were used. The mRNA expression of HSC cultured for one week on uncoated polystyrene dishes was set to 100% (broken line) for each experiment ( n = 5; * p < .05). HGF release by HSC exposed to (e–h) fluid shear stress or (i, j) mechanical stretching as analyzed by ELISA and immunofluorescence. HGF concentration of culture supernatants from static condition (0 dyn/cm 2 ) and unstretched HSC were set to 100% for each independent experiment ( n = 4; * p < .05). Immunofluorescence analysis of HGF (red) in HSC showed a high fluorescence signal intensity under (g) static condition, (h) which was lost under fluid shear stress for 1 hr. Cell nuclei are stained by DAPI (blue; scale bars: 20 µm). (a–d, f, j) Data are presented as means ± SEM

Article Snippet: HSC were seeded (50,000 cells/cm 2 ) either on uncoated polystyrene culture dishes or on dishes coated (1.0 µg/cm 2 ) with fibronectin (FN, Sigma‐Aldrich), COL4 (Sigma‐Aldrich), laminin‐211 (LN‐211), or laminin‐521 (LN‐521) obtained from BioLamina.

Techniques: Isolation, Expressing, Cell Culture, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Concentration Assay, Fluorescence, Staining