Roles of TauT and system A in cytoprotection of rat syncytiotrophoblast cell line exposed to hypertonic stress

Hypertonicity-induced cytotoxicity and cytoprotective effect of taurine in TR-TBT 18d-1 cells. Electrophoresis of DNA extracted from TR-TBT 18d-1 cells cultured in the normal (lane 2) or hypertonic medium (lane 3), revealing a hypertonicity-induced apoptotic ladder (A). Lane 1 is molecular weight markers. Nuclei of TR-TBT 18d-1 cells cultured in the normal (B, left) or hypertonic medium (B, right) were stained with DAPI. The cytoprotective effect of taurine on hypertonicity-induced cytotoxicity was evaluated by measuring caspase-3 activity (C). Each column shows the mean ± S.E.M. (n = 3–4). A statistically significant difference is indicated by an asterisk * (p < 0.05). Hypertonicity-induced cytotoxicity and the cytoprotective effect of taurine were visualized by bright-field imaging (D).

Contribution of TauT to taurine uptake by TR-TBT 18d-1 cells. Uptake of [³H]taurine by TR-TBT 18d-1 cells stably transfected with vector (mock) and with siRNA-TauT was measured for 5 min at 37 °C (A). The uptake is presented as cell-to-medium ratio. Saturable uptake was evaluated by the addition of an excess concentration (2 mM) of unlabeled taurine. mRNA expression of TauT in TR-TBT 18d-1 cells stably transfected with vector (mock) and siRNA-TauT was measured by real-time PCR (B). Each column shows the mean ± S.E.M. (n = 3–4). A statistically significant difference is indicated by an asterisk * (p < 0.05).

Effect of osmotic stress on [³H]taurine and [¹⁴C]betaine uptake and the expression levels of osmosensitive genes in TR-TBT 18d-1 cells. Hypertonicity-induced [³H]taurine (10 nM, A) and [¹⁴C]betaine (3 μM, B) uptake by TR-TBT 18d-1 cells cultured in normal (○) and hypertonic (●) medium was measured at 37 °C. The uptake is presented as cell-to-medium ratio. Each point represents the mean ± S.E.M. (n = 3–4). mRNA expression of putative osmolality-sensitive genes was measured by semi-quantitative RT-PCR (C). Each column represents the mean ± S.E.M. (n = 3). A statistically significant difference is indicated by an asterisk * (p < 0.05).

Inhibition study of [³H]taurine and [¹⁴C]betaine uptake by TR-TBT 18d-1 cells cultured in normal and hypertonic media. TR-TBT 18d-1 cells were cultured for 8 h in 300 mOsm/kg and 500 mOsm/kg medium. Cells were incubated with 10 nM [³H]taurine (A) or 3 μM [¹⁴C]betaine (B) in the absence and presence of inhibitors. Data are presented as the mean ± S.E.M. (n = 3–4). A statistically significant difference is indicated by an asterisk * (p < 0.05).

Effect of osmolytes on proliferation of TR-TBT 18d-1 cells. TR-TBT 18d-1 cells were cultured for 48 h in 300 mOsm/kg (normal condition) and 500 mOsm/kg medium (hypertonic condition). Cells were cultured in the absence (control) and presence of 1 mM osmolyte. Data are presented as the mean ± S.E.M. (n = 4). A statistically significant difference from the control is indicated by an asterisk * (p < 0.05).