Table 1.
Sequences of the primers for real time PCR.
Fig 1.
Senescence-associated-β-galactosidase (SA-β-gal) staining in the cerebrum 1 to 14 days after injury.
(A) SA-β-gal-positive cells could be observed in the ipsilateral cerebrum at 1, 4, 7, and 14 days after injury in injury groups. Scale = 100 μm. (B) Stained levels of SA-β-gal-positive cells, digitized for analysis by ImageJ software, in the ipsilateral side of the brain injury in the injury group at 4, 7, and 14 days after injury as compared to that in the corresponding area of the contralateral hemicerebrum of the injury group and the bilateral hemicerebrum of the sham and control groups (*p < 0.05, n = 5 for control group, n = 5 for the sham group, n = 5 for the injury group).
Fig 2.
Cyclin D1 immunohistochemistry and mRNA expression in the cerebrum 1 to 14 days after injury.
(A) Cyclin D1 immunostained cells could be observed in the ipsilateral hemicerebrum at 1, 4, 7, and 14 days after injury in injury groups. Long scale = 50 μm, short scale = 12.5 μm. (B-D) Double immunohistochemistry of the ipsilateral hemicerebrum at 4 days after injury for co-localization of (B) cyclin D1, NeuN, DAPI (4',6-diamidino-2-phenylindole), and merge; (C) cyclin D1, ionized calcium binding adaptor molecule 1 (Iba 1), DAPI, and merge; and (D) cyclin D1, glial fibrillary acidic protein (GFAP), DAPI, and merge. Scale = 30 μm. (E) Graph of the numbers of cyclin D1 immunostained cells from (A). (F-H) Cyclin D1 mRNA expression in the ipsilateral hemicerebrum 1 to 14 days after injury. Expressions were normalized to that of glyceraldehyde-3-phosphate dehydrogenase (Gapdh) (F), hypoxanthine guanine phosphoribosyl transferase (Hprt) (G), and peptidylprolyl isomerase A (Ppia) (H) (*p < 0.05, n = 5 for control group, n = 5 for the sham group, n = 5 for the injury group).
Fig 3.
Proliferating cell nuclear antigen (PCNA) immunohistochemistry and mRNA expression in the cerebrum 1 to 14 days after injury.
(A) PCNA immunostained cells could be observed in the ipsilateral hemicerebrum at 1, 4, 7, and 14 days after injury in injury groups. Long scale = 50 μm, short scale = 12.5 μm. (B-D) Double immunohistochemistry of the ipsilateral hemicerebrum at 4 days after injury for co-localization of (B) PCNA, NeuN, DAPI (4',6-diamidino-2-phenylindole), and merge; (C) PCNA, ionized calcium binding adaptor molecule 1 (Iba 1), DAPI, and merge; and (D) PCNA, glial fibrillary acidic protein (GFAP), DAPI, and merge. Scale = 30 μm. (E) Graph of the numbers of PCNA immunostained cells in (A). (F-H) Pcna mRNA expression in the ipsilateral hemicerebrum 1 to 14 days after injury. Pcna mRNA expressions were normalized to that of glyceraldehyde-3-phosphate dehydrogenase (Gapdh) (F), hypoxanthine guanine phosphoribosyl transferase (Hprt) (G), and peptidylprolyl isomerase A (Ppia) (H) (*p < 0.05, n = 5 for control group, n = 5 for the sham group, n = 5 for the injury group).
Fig 4.
Cyclin-dependent kinase inhibitor 2A (p16) immunohistochemistry and mRNA expression in the cerebrum 1 to 14 days after injury.
(A) p16 immunostained cells could be observed in the ipsilateral hemicerebrum at 4, 7, and 14 days after injury in injury groups. Long scale = 50 μm, short scale = 12.5 μm. (B) Double immunohistochemistry of the ipsilateral hemicerebrum at 7 days after injury for co-localization of p16, glial fibrillary acidic protein (GFAP), DAPI (4',6-diamidino-2-phenylindole), and merge. Scale = 30 μm. (C) Graph of the numbers of p16 immunostained cells in the ipsilateral hemicerebrum in (A). (D-F) p16 mRNA expression in the ipsilateral hemicerebrum 1 to 14 days after injury. p16 mRNA expressions were normalized to that of glyceraldehyde-3-phosphate dehydrogenase (Gapdh) (D), hypoxanthine guanine phosphoribosyl transferase (Hprt) (E), and peptidylprolyl isomerase A (Ppia) (F) (*p < 0.05, n = 5 for control group, n = 5 for the sham group, n = 5 for the injury group).
Fig 5.
Cyclin-dependent kinase inhibitor 1A (p21) immunohistochemistry and mRNA expression in the cerebrum 1 to 14 days after injury.
(A) p21 immunostained cells could be observed in the ipsilateral hemicerebrum at 1, 4, 7, and 14 days after injury in injury groups. Long scale = 50 μm, short scale = 12.5 μm. (B, C) Double immunohistochemistry of the ipsilateral hemicerebrum at 7 days after injury for co-localization of (B) p21, NeuN, DAPI (4',6-diamidino-2-phenylindole), and merge; and (C) p21, ionized calcium binding adaptor molecule 1 (Iba 1), DAPI, and merge. Scale = 30 μm. (D) Graph of the numbers of p21 immunostained cells in the ipsilateral hemicerebrum in the injury group at 1 to 14 days after injury in (A). (E-G) p21 mRNA expression in the ipsilateral hemicerebrum 1 to 14 days after injury. p21 mRNA expressions were normalized to that of glyceraldehyde-3-phosphate dehydrogenase (Gapdh) (E), hypoxanthine guanine phosphoribosyl transferase (Hprt) (F), and peptidylprolyl isomerase A (Ppia) (G) (*p < 0.05, n = 5 for control group, n = 5 for the sham group, n = 5 for the injury group).
Fig 6.
mRNA expression of the transformation related protein 53 (p53) in the ipsilateral hemicerebrum 1 to 14 days after injury.
p53 mRNA expressions were normalized to that of glyceraldehyde-3-phosphate dehydrogenase (Gapdh) (A), hypoxanthine guanine phosphoribosyl transferase (Hprt) (B) and peptidylprolyl isomerase A (Ppia) (C) (*p < 0.05, n = 5 for control group, n = 5 for the sham group, n = 5 for the injury group).