Figure 4: Cellular proliferation promoted by the dimeric macrocycles in normal human cells.

(a–c) Dose-dependent titration of proliferation induction in normal human epidermal keratinocytes (NHEK) against concentration of hHGF (red) as a positive control, (a) aML5-PEG3 (blue), (b) aMD4-PEG11 (orange) and aMsD4-C6 (grey) as a negative control, or (c) aMD5-PEG11 (green). NHEK were cultured for 6 days with or without hHGF (0.4 pM–30 nM) or dimeric macrocycles (40 pM–3,000 nM). Cells were stained by calcein-AM and quantified by fluorescence intensity. s.d. was calculated from the results of experiments in triplicate. (d) Time course of proliferation induction in NHEK. NHEK were cultured with or without 1.3 nM hHGF or 50 nM dimeric macrocycles. After 2, 4 and 6 days, cell numbers were counted by means of an automated cell counter (mean±s.d., n=3). *P<0.05, **P<0.01 (unpaired Student’s t-test) compared with Mock. (e) Proliferation of various normal human cells stimulated by hHGF and aMD5-PEG11. HDMEC, normal human dermal microvascular endothelial cells; HUVEC, normal human umbilical vein endothelial cells; RPTEC, normal human renal proximal tubule epithelial cells. The respective cells were cultured with or without 1.3 nM hHGF or 50 nM aMD5-PEG11. After 5 days, cell numbers were counted by means of an automated cell counter (mean±s.d., n≥4). **P<0.01, ***P<0.001 (unpaired Student’s t-test) compared with Mock.