氢氯噻嗪和美托拉宗促进成骨样细胞增殖及成骨分化

A Study on the Proliferation and Differentiation of Osteoblast-like Cells Promoted by Hydrochloridethiazide and Metolazone

目的观察钠氯共转运体(NCC)阻断剂氢氯噻嗪和美托拉宗阻断NCC后,对UMR106成骨样细胞增殖及成骨分化的影响。方法将细胞分为对照组、氢氯噻嗪组及美托拉宗组,分别加入不同浓度(1、10、100 M)的氢氯噻嗪及美托拉宗与UMR106细胞共培养,在不同时间点用MTT法检测细胞的增殖,碱性磷酸酶(ALP)试剂盒检测ALP活性,实时荧光定量PCR法(qRT-PCR)检测成骨相关因子Runx2和Osterix的mRNA表达。结果加药48h后,1)氢氯噻嗪组在1 M与10 M浓度的吸光度(OD值)较对照组提高了(38.0±5.6)%和(30.3±3.3)%,差异有统计学意义(P<0.05);美托拉宗组在1、10、100 M浓度的OD值较对照组提高(24.2±1.8)%、(50.8±6.6)%及(26.8±2.1)%,差异有统计学意义(P<0.05);2)氢氯噻嗪组和美托拉宗组在1、10、100 M浓度的ALP活性较对照组分别提高了(169.6±19.8)%、(86.6±8.7)%、(118.2±12.5)%以及(73.4±6.8)%、(145.6±14.5)%、(63.0±7.9)%,差异均有统计学意义(P<0.05);3)加药培养24h后,氢氯噻嗪组和美托拉宗组在1、10、100 M浓度Runx2mRNA的表达增多,分别是对照组的(3.52±0.31)、(2.85±0.15)、(2.50±0.41)倍及(2.05±0.45)、(4.57±0.98)、(2.93±0.34)倍,差异均有统计学意义(P<0.05)。结论氢氯噻嗪和美托拉宗在加药24h可提高Runx2mRNA的表达,48h促进细胞增殖及ALP活性。两药可能通过上调成骨相关因子Runx2mRNA的表达促进UMR106细胞的增殖及成骨分化。

Objective To investigate the effect of the sodium and chloride co-transporter （NCC） blockers Hydrochlorothiazide （HCTZ） and Metolazone on the proliferation and osteoblast differentiation of UMR106 cells. Methods The Cells were divided into the control group, HCTZ group and metolazone group. HCTZ and Metolazone of different concentration （1M, 10M and 100M） were co-cultured with UMR106 cells respectively. The cell proliferation, ALP activity, and expression of Runx2 and Osterix mRNA were detected by the MTT method, ALP kit, and qRT-PCR repectively at different time points. Results Compared with the control group after 48 hours of being co-cultured, the OD values in the HCTZ groups with the concentration of 1M and 10M increased by （38.0±5.6） % and （30.3±3.3）% respectively and those in the Metolazone groups with the concentration of 1M, 10M and 100M increased by （24.2±1.8）%, （50.8±6.6）% and （26.8±2.1）% respectively, and the differences were statistically significant （P〈0.05）. The ALP activities in the HCTZ groups with the concentration of 1M, 10M and 100M increased by （169.6 ± 19.8）%, （86.6±8.7） % and （118.2 ± 12.5）%, and those in the Metolazone groups with the concentration of 1M, 10M and 100M increased by （73.4 ±6.8）%, （145.6± 14.5）% and （63.0±7.9）% respectively, and the differences were statistically significant （P〈 0.05）. Compared with the control group after 48 hours of being co-cultured, the expression levels of Runx2 and Osterix mRNA in the HCTZ groups with the concentration of 1M, 10 M and 100 M increased to （3. 52±0.31）times, （2. 85±0.15）times and （2. 50±0.41）times, and the those in the Metolazone groups with the concentration of 1M, 10M and 100M increased to （2.05 ±0.45） times, （4.57± 0.98） times and （2.93 ± 0.34） times, and the differences were statistically significant （P〈0.05）. Conclusion HCTZ and metolazone significantly increase the expression of Runx2 mRNA after 24 hours of being co-cultured and promote the cell proliferation and ALP activity after 48 hours of being cocultured. Both drugs may promote the proliferation and osteoblast differentiation of UMR106 ceils by up- regulating the expression of osteogenesis-related factor Runx2 mRNA.