Synergistic Effect of Hyperglycemia and p27^kip1 Suppression on Adult Mouse Islet Beta Cell Replication

Table 1

Effect of p27^Kip1 silencing on the expression of cell cycle proteins in adult mouse islets. Cellular lysates of freshly isolated mouse islets and cultured islets at 54 and 96 h after infection with lentivirus to silence p27^Kip1 were separated on 12% sodium dodecyl sulfate-polyacrylamide gel and electrophoretically transferred onto a nitrocellulose membrane, which was then incubated with antibodies against β-actin, p27^Kip1, cyclin B1, cyclin D1, and FoxM1. Cellular lysates of control islets were collected at 24, 54, and 96 h after infection with virus carrying non-target shRNAs for the determination of p27^Kip1 protein level. Bound antibodies were detected by using a chemiluminescence kit. The levels of cell cycle proteins were expressed as the ratio of the band density of each protein over that of β-actin of the same batch of islet proteins. The protein content of freshly isolated islets was used as the standard for comparison to express the changes in the levels of cell cycle proteins after virus infection. Data were expressed as mean ± standard error. ^aP < 0.05, ^bP < 0.01, ^cP < 0.005 when the indicated mean was analyzed using the single t-test.

Hours after viral infection	p27^Kip1/actin non-targeting control	p27^Kip1/actin targeting	B1/actin targeting	D1/actin targeting	FoxM1/actin targeting
0	1	1	1	1	1
24	0.56±0.11^a, 𝑛=6
54	0.80±0.18, 𝑛=4	0.22±0.06^b, 𝑛=4	9.40±2.42^a, 𝑛=4	0.97±0.11^a, 𝑛=4	1.31±0.10^b, 𝑛=4
96	0.95±0.24, 𝑛=4	0.19±0.03^c, 𝑛=4	3.26±0.85^a, 𝑛=4	0.93±0.12^b, 𝑛=4	1.88±0.46^a, 𝑛=4