Abstract:
Objective To investigate the effects of recombinant human thyroid-stimulating hormone (rhTSH) on thyroglobulin (Tg) secretion, sodium-iodide symporter (NIS) expression, and 131I uptake function in differentiated thyroid carcinoma cells.
Methods Real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) was used to screen the KTC-1, TPC-1, and BCPAP human papillary thyroid carcinoma cell lines to identify the cell line with the highest thyroid-stimulating hormone receptor (TSHR) gene expression level for subsequent experiments. By taking this cell line as the research object, concentration–effect experiments were first performed (control group (treated with 0 ng/ml rhTSH, recorded as group N) and experimental groups (treated with 50, 100, 200, 300, and 500 ng/ml rhTSH concentrations and recorded as groups R1, R2, R3, R4, and R5, respectively), with all groups treated for 24 h). Subsequently, on the basis of the determined optimal concentration, time–effect experiments were conducted (experimental group (optimal concentration of rhTSH for 2, 6, 12, 18, and 24 h) and control group (cultured with the same medium without rhTSH for 2, 6, 12, 18, and 24 h)). Western blot analysis was used to detect the relative expression levels of TSHR and NIS proteins; enzyme linked immunosorbent assay was employed to detect the concentration of Tg in the cell supernatant; and a thyroid function analyzer was applied to measure the 24 h 131I uptake rate of cells. Comparisons between two groups of measurement data were performed by using independent samples t-test, and comparisons among multiple groups were conducted through one-way analysis of variance.
Results The relative expression level of the TSHR gene in BCPAP cells was significantly higher than those in TPC-1 and KTC-1 cells, and the differences were statistically significant (2−ΔΔCT: (0.88±0.32) vs. (0.30±0.10) vs. (0.01±0.00), t=0.575, 0.872, P=0.008, P<0.001). Therefore, BCPAP cells were selected for subsequent experiments. (1) In the concentration–effect experiment, the Tg concentrations in groups R3, R4, and R5 were significantly higher than that in group N, and the differences were statistically significant ((6.13±0.16) ng/ml vs. (5.39±0.28) ng/ml, (6.63±0.57) ng/ml vs. (5.39±0.28) ng/ml, (6.42±0.39) ng/ml vs. (5.39±0.28) ng/ml, t=4.015, 3.367, 3.701, P=0.016, 0.028, 0.021). The relative expression level of the NIS protein in group R4 was significantly higher than that in groups N, R1, R2, R3, and R5, and the differences were statistically significant ((1.02±0.22) vs. (0.30±0.01) vs. (0.46±0.02) vs. (0.59±0.01) vs. (0.70±0.01) vs. (0.83±0.01), t=9.769–27.959, all P≤0.001). The 131I uptake counts in group R4 were significantly higher than those in groups N, R1, R2, and R3, and the differences were statistically significant ((1 404.00±8.19) cmp vs. (1 297.67±17.90) cmp vs. (1 301.67±30.09) cmp vs. (1 304.67±24.83) cmp vs. (1 365.33±17.79) cmp, t=3.421–9.358, all P<0.05) ). (2) In the time–effect experiment, 300 ng/ml rhTSH continuously promoted Tg secretion within 24 h ((6.07±0.56) ng/ml vs. (5.66±0.51) ng/ml, t=2.124, P=0.043). At 24 h, NIS protein expression significantly increased ((1.09±0.87) vs. (0.83±0.09), t=3.537, P=0.024). The 131I uptake rates of the cells in the experimental group at 12–24 h were significantly higher than those in the corresponding control group (12 h: (1251.33±52.70) cmp vs. (1078.00±16.00) cmp, 18 h: (1312.00±14.00) cmp vs. (1201.00±24.27) cmp, 24 h: (1404.00±48.66) cmp vs. (1309.67±32.52) cmp, t=5.451, 6.862, 2.792, P=0.006, 0.002, 0.049).
Conclusion rhTSH at 300 ng/ml acting on BCPAP cells within 24 h can effectively promote Tg secretion, NIS protein expression, and 131I uptake function in the cells.
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