Table 1.
Sequence and Origin of Endothelial Extracellular Matrix-Derived Peptides.
Table 2.
Sequence and Origin of Fibroblastic Extracellular Matrix-Derived Peptides.
Fig 1.
Endothelial and fibroblastic ECM-derived peptides stimulate human dermal microvascular endothelial cell proliferation in vitro.
Capillary endothelial cells grown in 48-well plates were treated with peptides at 100 nM every other day for a period of 5 days, and cell counts were performed on Day 5 as described in “Methods.” Relative proliferation compared with 1% BCS (control) is shown as fold of control, calculated by normalizing the mean cell count for each treatment to the mean cell count obtained for negative control, and error bars represent standard deviation. * p < 0.05, ** p < 0.01, *** p < 0.005, **** p < 0.001 compared with negative control (unpaired Student’s T-test).
Fig 2.
Bioactive endothelial and fibroblastic matrix-derived peptides increase human keratinocyte proliferation in vitro.
Adult normal human epidermal keratinocytes (NHEK) grown in 48-well plates were treated with peptides at 10 nM every other day for a period of 5 days, and cell counts were performed on Day 5 as described in “Methods.” Relative proliferation compared with 50% basal NHEK media (control) is shown as fold of control, calculated by normalizing the mean cell count for each treatment to the mean cell count obtained for negative control, and error bars represent standard deviation. * p < 0.05, ** p < 0.01, *** p < 0.005, **** p < 0.001 compared with negative control (unpaired Student’s T-test).
Fig 3.
Endothelial and fibroblastic ECM-derived peptides enhance human dermal fibroblast proliferation in vitro.
Dermal fibroblasts isolated from healthy adult human donors grown in 48-well plates were treated with peptides at 100 nM every other day for a period of 5 days, and cell counts were performed on Day 5 as described in “Methods.” Relative proliferation compared with 1% FBS (control) is shown as fold of control, calculated by normalizing the mean cell count for each treatment to the mean cell count obtained for negative control, and error bars represent standard deviation. * p < 0.05, ** p < 0.01, *** p < 0.005, **** p < 0.001 compared with negative control (unpaired Student’s T-test).
Fig 4.
Quantitative analysis of peptide-induced dermal microvascular remodeling in vitro.
(A-D) Representative photomicrographs of human dermal capillary endothelial cell morphogenesis following treatment with 1% BCS (A), 100 nM TSN1 (B), 100 nM TSN 8 (C), and 100 nM TSN 10 (D). Scale bar represents 100 μm in all panels. (E) The lengths of human dermal capillary endothelial sprouts cultured on growth factor-reduced Matrigel in the presence or absence of 100 nM ECM-derived peptides were measured as described in “Methods” at 5 hours post-plating, with physiologic doses of FGF2 (0.6 nM) serving as positive controls. Data are displayed as fold increase in tube length, relative to 1% BCS-treated negative controls, and error bars represent standard deviation. * p < 0.05, ** p < 0.01, *** p < 0.001, compared with negative control (unpaired Student’s T-test).
Fig 5.
Quantitative evaluation of peptide-dependent wound re-epithelialization.
Percent wound closure attributable to re-epithelialization was measured from merged photomicrographs of hematoxylin and eosin or trichrome-stained tissue sections using NIH ImageJ as described in “Methods.” (A-F). Representative photomicrographs of H&E-stained sections of wounds treated with CMC (A), 0.1 mg/mL TSN6 (B), 1.0 mg/mL TSN6 (C), 0.1 mg/mL TSN18 (D), 1.0 mg/mL TSN18 (E), and Santyl® Collagenase (F) are shown. Scale bar represents 1.0 mm in all images. (G). Wounds analyzed for each treatment are plotted as a function of the percent re-epithelialization. Mean percent wound closure is displayed, and error bars represent standard deviation. * p < 0.05, ** p < 0.01, compared with CMC-treated controls.
Table 3.
Summary of Endothelial ECM-derived peptide bioactivity in vitro.
(Numbers in each column represent mean fold increase relative to controls).
Table 4.
Summary of Endothelial ECM-derived peptide bioactivity in vitro.
(Numbers in each column represent mean fold increase relative to controls).