FK506 inhibits the mice glomerular mesangial cells proliferation by affecting the transforming growth factor-β and Smads signal pathways

Figures & data

Figure 1. The comparisons of cell proliferation between the FK506 group and the control. Notes: cells were seeded into 96-well plates at a density of 3 ×10⁴ cells/well. After starving in the serum-free medium overnight, the cells were incubated in the medium for 8, 24, and 48 h. Cell proliferation was assessed by using MTT assay. Absorbance was measured by ELISA plate-reader at 570 nm. The values were expressed as a mean ± SD. The presented data are based on at least three independent experiments. *p < 0.05, compared to normal group. The proliferation of FK506 group treated with FK506 10, 30, and 60 ng/mL for 8, 24, and 48 h was assessed by MTT cell proliferation assay. The data shown here (mean± SD) were from one representative experiment performed in triplicate, repeated three times with similar results.

Figure 2. FK506 reduced the expressions of Smad2 protein increased by TGF-β1 in mesangial cells. Notes: Mesangial cells were cultured and treated with TGF-β1 5 ng/mL with or without the addition of FK506 (10, 30, and 60 ng/mL) for 48 h. Total cell protein lysates were prepared and subjected to Western blotting analysis with antibody against Smad2 and GAPDH. Experiments were repeated thrice with similar results. *p < 0.05 as compared with control group; #p < 0.05 as compared with TGF-β1 stimulated group. Lane 1: control; Lane 2: TGF-β1 5 ng/mL treatment; Lane 3: TGF-β1 5 ng/mL + FK506 10 ng/mL treatment; Lane 4: TGF-β1 5 ng/mL + FK506 30 ng/mL treatment; Lane 5: TGF-β1 5 ng/mL + FK506 60 ng/mL treatment.

Table 1. FK506 reduced the expressions of Smad2 mRNA increased by TGF-β1 in mesangial cells.

	8 h	24 h	48 h
The control	0.151 ± 0.002	0.172 ± 0.014	0.194 ± 0.004
FK506 10 ng/mL	0.150 ± 0.014	0.171 ± 0.041	0.191 ± 0.006
FK506 30 ng/mL	0.148 ± 0.020	0.169 ± 0.037	0.190 ± 0.010
FK506 60 ng/mL	0.149 ± 0.031	0.168 ± 0.021	0.187 ± 0.014
TGF-β1 5 ng/mL	1.101 ± 0.020#	1.192 ± 0.047#	1.221 ± 0.010#
TGF-β1 + FK506 10 ng/mL	0.671 ± 0.010*	0.761 ± 0.014*	0.897 ± 0.011*
TGF-β1 + FK506 30 ng/mL	0.473 ± 0.022*	0.583 ± 0.031*	0.578 ± 0.006*
TGF-β1 + FK506 60 ng/mL	0.311 ± 0.012*	0.452 ± 0.016*	0.491 ± 0.015*

Notes: Cells were treated with TGF-β1 5 ng/mL followed by FK506 10, 30, and 60 ng/mL and then incubated at indicated time points for the determination of Smad2 mRNA. The data shown here (mean ± SD) are from one representative experiment performed in triplicate, repeated thrice with similar results.

*p < 0.05, #p < 0.01 as compared with the TGF-β1 stimulated group.