Objective: To compare the effects of levo-praziquantel (L-PZQ) and dextro-praziquantel (D-PZQ) on the proliferation and activation of the human hepatic stellate cell line LX-2 in vitro.
Methods: LX-2 cells were stimulated with transforming growth factor-β (TGF-β). LX-2 cell proliferation was measured using the CCK-8 assay after 24 h stimulation with 0 to 50 μg/mL concentrations of praziquantel, and the gene and protein expression of type Ⅰ collagen (collagen Ⅰ), type Ⅲ collagen (collagen Ⅲ) and α-smooth muscle actin (α-SMA) was quantified in LX-2 cells using quantitative real-time PCR (qPCR) and Western blotting assays 24 h and 48 h following stimulation with 15 μg/mL praziquantel to detect LX-2 cell activation.
Results: There were significant differences in the survival rate of LX-2 cells between L-PZQ and D-PZQ treatments at all concentrations (F = 6.119 and 79.180, both P values < 0.05). Either L-PZQ or D-PZQ at a concentration of < 30 μg/mL showed no remarkableeffectsonthe LX-2 cell proliferation (both P values > 0.05), and L-PZQ at a concentration of > 50 μg/mL and D-PZQ at a concentration of > 40 μg/mL inhibited the LX-2 cell proliferation (both P values < 0.05), while D-PZQ at concentrations of 40 μg/mL and 50 μg/mL showed greater inhibition on LX-2 cell proliferation than L-PZQ (t = 3.419 and 8.776, both P values < 0.05). There were significant differences in the collagen Ⅰ, collagen Ⅲ and α-SMA expression in LX-2 cells at both transcriptional (F = 21.55, 79.99 and 46.70, all P values < 0.05) and translational levels (F = 20.12, 30.29 and 32.93, all P values < 0.05) among the blank control group, TGF-β stimulation group, L-PZQ treatment group and D-PZQ treatment group. L-PZQ treatment resulted in remarkable inhibition on collagen Ⅲ and α-SMA gene expression in LX-2 cells (both P values < 0.05); however, the treatment showed no remarkable inhibition collagen Ⅰ gene expression or collagen Ⅰ, collagen Ⅲ or α-SMA protein expression in LX-2 cells (all P values > 0.05). In addition, D-PZQ treatment resulted in significant inhibition on collagen Ⅰ, collagen Ⅲ and α-SMA expression in LX-2 cells at both translational and transcriptional levels (all P values < 0.05), and D-PZQ showed higher inhibition on collagen Ⅰ, collagen Ⅲ and α-SMA gene expression in LX-2 cells than L-PZQ (all P values < 0.05).
Conclusions: Both L-PZQ and D-PZQ inhibit the proliferation and activation of LX-2 cells, and D-PZQ shows a higher inhibitory activity than L-PZQ.
［摘要］ 目的 观察左旋吡喹酮 (L-PZQ) 和右旋吡喹酮 (D-PZQ) 体外对人肝星状LX-2细胞系增殖及活化作用差异。方法 利用转化生长因子-β (TGF-β) 刺激激活LX-2细胞; 采用CCK-8法检测以0~50 μg/mL梯度浓度吡喹酮刺激24 h后LX-2细胞增殖情况; 采用实时荧光定量PCR (qPCR) 和免疫印迹试验检测15 mg/mL吡喹酮刺激LX-2细胞24 h 后的Ⅰ型胶原、Ⅲ型胶原和α-平滑肌肌动蛋白 (α-SMA) 基因表达水平和48 h后蛋白表达水平, 以分析LX-2细胞活化情况。结果 L-PZQ和D-PZQ两药物各浓度组LX-2细胞存活率差异均有统计学意义 (F = 6.119、79.180, P 均< 0.05); 药物浓度< 30 μg/mL时, L-PZQ和D-PZQ对激活型LX-2增殖能力均无显著影响 (P 均> 0.05); L-PZQ浓度> 50 μg/mL和D-PZQ浓度> 40 μg/mL时, 均对激活型LX-2增殖能力产生抑制作用 (P 均< 0.05); D-PZQ浓度为40 μg/mL和50 μg/mL时, 对激活型LX-2增殖的抑制作用均强于L-PZQ (t = 3.419、8.776, P 均< 0.05)。空白组、TGF-β诱导组、L-PZQ组和D-PZQ组LX-2细胞中Ⅰ型胶原、Ⅲ型胶原和α-SMA 基因 (F = 21.55、79.99、46.70, P 均< 0.05) 和蛋白表达水平 (F = 20.12、30.29、32.93, P 均< 0.05) 差异均有统计学意义。L-PZQ对激活型LX-2细胞中Ⅲ型胶原和α-SMA 基因表达水平具有显著抑制作用 (P 均< 0.05), 但对Ⅰ型胶原基因表达水平及Ⅰ型胶原、Ⅲ型胶原和α-SMA蛋白表达水平均无显著影响 (P 均> 0.05); D-PZQ对激活型LX-2细胞中Ⅰ型胶原、Ⅲ型胶原和α-SMA 基因和蛋白表达水平均有显著抑制作用 (P 均< 0.05); D-PZQ 对激活型LX-2细胞中Ⅰ型胶原、Ⅲ型胶原和α-SMA 基因表达抑制作用强于L-PZQ (P 均< 0.05)。结论 L-PZQ和D-PZQ对LX-2细胞增殖及活化均有一定抑制作用, 且D-PZQ的抑制作用强于L-PZQ。.
Keywords: Activation; Hepatic stellate cell; Isomer; Praziquantel; Proliferation.