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. 2016 Sep 12;126(10):3879–3893. doi: 10.1172/JCI84164

Figure 5. Antisenescence effect of JH chemicals on normal aged cells.

Figure 5

(A) Nuclear deformation of aged fibroblasts was ameliorated by JH4. Normal cells obtained from 9-year-old (N9) and 81-year-old (N81) subjects were incubated with JH4 (5 μM) for 48 hours and stained with anti–lamin A/C Ab (green). DAPI was used for DNA staining. Scale bars: 10 μm. (B) JH4 could induce cell proliferation in aged fibroblasts. After seeding 7.5 × 104 cells per well, cells were maintained for a maximum of 6 days with the indicated JH chemicals (5 μM). Cell numbers were calculated every 48 hours. The slow proliferation of cells from N81 compared with that of cells from N9 (Figure 4B) was completely rescued by JH4 treatment. *P = 0.013. The P value was determined by Student’s t test for analysis of statistical significance between 2 groups. (C) JH4 suppressed senescence of N81 fibroblasts. An SA–β-gal assay was performed on cells from N81 after treatment with the indicated chemicals (5 μM) for 48 hours. Original magnification, ×40. (D) JH4 induced H3K9Me3 expression in normal aged cells. Immunofluorescence for H3K9Me3 (green) was performed in N9 and N81 cells under treatment with the indicated chemicals (5 μM) for 48 hours. Scale bars: 10 μm. (E) JH4 suppressed DCR2 and p16INK4A in normal human cells. Reverse transcriptase-PCR (RT-PCR) analysis of the indicated genes was performed in normal cell lines (N9, N29, and N81) after treatment with JH4 for 48 hours.