![Figure 2. CDM and type I collagen support lobopodia- and lamellipodia-based 3D migration, respectively. (A) HFF-generated CDM has an aligned, fibrillar structure (top left), whereas polymerized 1.7 mg/ml type I collagen forms a random meshwork (bottom left). Both images are maximum projections of 30-µm confocal stacks. Collagen is remodeled by migrating HFFs (GFP, green) along the axis of migration (bottom right), whereas the organization of the CDM is unaffected by migrating HFFs (top right). The CDM was labeled with Alexa Fluor 633, and collagen was visualized by reflection microscopy. (B) Matrix stiffness (Young’s modulus [E]) of the indicated 3D matrices. (C) Strain-stiffening (Ehigh/Emed) behavior of the indicated 3D matrices. Ehigh/Emed > 1 indicates nonlinear elasticity, whereas Ehigh/Emed = 1 indicates linear elasticity (dashed red line). (D) Collagen and 2D CDM support lamellipodia-based migration, whereas 3D CDM triggers lobopodia-based motility. (top) Maximum projections of HFFs expressing GFP migrating inside the 3D CDM, on top of the 2D CDM, or inside type I collagen. LM, lamellipodium; LB, lobopodium. (bottom) The orthogonal views of the corresponding panel above, with the CDM (Alexa Fluor 633) and type I collagen (reflection microscopy) in red. Arrowheads indicate lateral blebs. (E) Cortactin is not enriched at the leading edge during lobopodia-based migration. HFFs migrating in the indicated ECM were fixed and immunostained for cortactin. Arrows indicate the local accumulation of cortactin at the leading edge. Bottom graphs correspond with their respective top images and represent the mean cortactin intensity measured from the leading edge (0 µm) toward the cell center. Each cortactin intensity profile was averaged from 13 cells, with three measurements per cell. Bars: (A and E) 5 µm; (D, top left and middle) 10 µm; (D, top right) 20 µm. All cells are oriented with the leading edge toward the right of the figure. Error bars show means ± SEM. *, P < 0.001 versus the dermal explant. a.u., arbitrary unit.](https://cdn.rupress.org/rup/content_public/journal/jcb/197/3/10.1083_jcb.201201124/6/jcb_201201124_fig2.jpeg?Expires=1773539899&Signature=V83zfWt9sRlz8ry2C6fXMsrqokxNirYvHyTGz4IfZ6q4EDsva1~UiWYTjTnmz6UYEOac8harUdkminTGQwCVefUz0V7ZuyPVyOYO~As~JhAYQExQKG6HHrgZnfez8wHf2stlgaiOjC3YONsJXPkJcDH2NU6eZ5hvz2j3yiMYSUWdTV6zMIoMjI1L2gRqNRKFlDxreuQIqafu1ABk8EgoLcPfrSww3bVbGpOqunpc0kbEpKlcW4i14lcme0SPd14OZnxjseUq~SQVnU8nA-UZ2f~l4KlqtvtwDxLvhUa4LVo2pasHIXVV6R8phn7uJRPkNpNqRbPk3svEZ8bYtqzOuQ__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
CDM and type I collagen support lobopodia- and lamellipodia-based 3D migration, respectively. (A) HFF-generated CDM has an aligned, fibrillar structure (top left), whereas polymerized 1.7 mg/ml type I collagen forms a random meshwork (bottom left). Both images are maximum projections of 30-µm confocal stacks. Collagen is remodeled by migrating HFFs (GFP, green) along the axis of migration (bottom right), whereas the organization of the CDM is unaffected by migrating HFFs (top right). The CDM was labeled with Alexa Fluor 633, and collagen was visualized by reflection microscopy. (B) Matrix stiffness (Young’s modulus [E]) of the indicated 3D matrices. (C) Strain-stiffening (Ehigh/Emed) behavior of the indicated 3D matrices. Ehigh/Emed > 1 indicates nonlinear elasticity, whereas Ehigh/Emed = 1 indicates linear elasticity (dashed red line). (D) Collagen and 2D CDM support lamellipodia-based migration, whereas 3D CDM triggers lobopodia-based motility. (top) Maximum projections of HFFs expressing GFP migrating inside the 3D CDM, on top of the 2D CDM, or inside type I collagen. LM, lamellipodium; LB, lobopodium. (bottom) The orthogonal views of the corresponding panel above, with the CDM (Alexa Fluor 633) and type I collagen (reflection microscopy) in red. Arrowheads indicate lateral blebs. (E) Cortactin is not enriched at the leading edge during lobopodia-based migration. HFFs migrating in the indicated ECM were fixed and immunostained for cortactin. Arrows indicate the local accumulation of cortactin at the leading edge. Bottom graphs correspond with their respective top images and represent the mean cortactin intensity measured from the leading edge (0 µm) toward the cell center. Each cortactin intensity profile was averaged from 13 cells, with three measurements per cell. Bars: (A and E) 5 µm; (D, top left and middle) 10 µm; (D, top right) 20 µm. All cells are oriented with the leading edge toward the right of the figure. Error bars show means ± SEM. *, P < 0.001 versus the dermal explant. a.u., arbitrary unit.
