Fig. 3

Intracellular collagen fibril formation and growth by hMSCs in different groups at different time points after removal of extracellular collagen fibrils by treatment with bacterial collagenase type VI. a-p Visualization of internalized collagen after collagenase treatment. a-d 2 h. e-h, m, n 4 h. i-l 6 h. o, p 24 h. a, e, i Control. b, f, j Inhibition of intracellular matrix degradation by cysteinase inhibitor (E64D). c, g, k, m Combination of both intracellular (E64D) and extracellular (GM6001) protease inhibitors. d, h, l, n-p Inhibition of extracellular matrix degradation by broad-spectrum protease inhibitor (GM6001). m 1–2, o1-2 Side views of hMSCs showing that the location of the stained fibrils was within the intracellular space. q-t Quantitative measurement of total fluorescence intensity of internalized collagen per cell on images at 63× (mean + standard deviation, n = 2 to 8). q Control. r Inhibition of intracellular matrix degradation by cysteine proteinase inhibitor (E64D). s Combination of both intracellular (E64D) and extracellular (GM6001) protease inhibitors. t Inhibition of extracellular matrix degradation by broad-spectrum protease inhibitor (GM6001). Green: Alexa Fluor 488-labeled collagen type I; orange: lysozyme; red: Plasma membrane; and grey: reflection. Collagen (10 mg/ml), E64D (20 μM), and GM6001 (25 μM) were used. hMSC human mesenchymal stem cell