Silencing SPARC inhibits high fluoride-induced apoptosis of thyrocytes / 西安交通大学学报(医学版)

ABSTRACT

Objective: To investigate the effects and mechanisms of secreted protein, acidic and rich in cysteine (SPARC) on high fluoride-induced apoptosis of thyrocytes. Methods: Human thyroid cells (Nthy-ori 3-1) were cultured and treated with various concentrations (0.1, 1 and 10 mmol/L) of NaF for 24 h, and the expression of SPARC was evaluated using Real-time PCR and Western blot, respectively. The cells were divided into four groups control group, NaF group, si-SPARC group (cells were transfected with SPARC siRNA for 48 h and then exposed to NaF for 24 h), and si-NC group (cells were transfected with negative control siRNA for 48 h and then exposed to NaF for 24 h). Cytotoxicity was assayed using CCK-8 and LDH; cell apoptosis rate was detected with ELISA. The expressions of cleaved caspase3 (c-caspase3) and IGF-1R were measured by Western blot. In addition, si-SPARC and si-IGF-1R were co-transfected into thyrocytes to further explore mechanisms of SAPRC by evaluating apoptosis. Results: The mRNA and protein levels of SPARC were augmented with the increase of NaF (P<0.05). Cell viability was significantly higher in si-NC group than that in si-SPARC group [(84.02±9.51)% vs. (58.31±6.86)%, P<0.05], and the release rate of LDH was lower [(134.25±18.98)% vs. (195.18±23.50)%, P<0.05]. Cell apoptosis rate was lower in si-SPARC group than that in si-NC group [(124.67±19.44)% vs. (175.24±16.46)%, P<0.05]. In addition, silencing SPARC upregulated the expression of IGF-1R (1.95±0.24 vs. 0.93±0.08, P<0.05), and inhibition of IGF-1R reversed the effect of SPARC on apoptosis. Conclusion: Inhibition of SPARC reduces high fluoride-induced cytotoxicity and blocks cell apoptosis. The possible mechanism is through the negative regulation of IGF-1R.