新型气体信号分子二氧化硫对自发性高血压大鼠主动脉平滑肌细胞增殖和凋亡的影响

Effects of sulfur dioxide, on the proliferation and apoptosis of aorta smooth muscle cells in hypertension： experiments with rats

目的 探讨新型气体信号分子二氧化硫（SO2）对自发性高血压大鼠（SHR）主动脉平滑肌细胞增殖与凋亡的调节作用。方法 4周龄SHR大鼠随机分为SHR对照组、Na2SO3／NaHSO3组（外源性SO2供体组），每组8只。4周龄正常血压WistarKyoto（WKY）大鼠8只作为正常对照（WKY对照组）。5周后检测大鼠血压及主动脉形态学指标，以高效液相色谱（HPLC）法测定血浆SO2水平；采用原位缺口末端标记方法（TUNEL）检测大鼠主动脉平滑肌细胞凋亡；采用免疫组织化学方法检测主动脉平滑肌细胞增殖细胞核抗原（PCNA）及凋亡相关蛋白Bcl-2、Fas和半胱氨酸蛋白水解酶（caspase）-3的表达，并进行图像分析。结果 5周后SHR对照组血压、血管壁厚与内径比均明显高于WKY对照组[（172±10）nl／nHgV8（112±9）nl／nHg、0．073±0．004V80．057±0．004，均P〈0．01，1mm Hg=0．133kPa]，血浆SO2水平显著低于WKY对照组[（6．4±1．5）μmol／LV8（11．3±1．0）μmol／L]，主动脉平滑肌细胞增殖指数高（0．32±0．06 vs 0．05±0．03），而凋亡指数小（0．16±0．07 vs 0．30±0．19），主动脉平滑肌细胞Bcl-2蛋白表达高（0．209±0．007 vs 0．202±0．006，P〈0．01），Fas、caspase-3蛋白表达弱（0．205±0．006 vs 0．211±0．005、0．229±0．005 vs 0．244±0．010，均P〈0．01）。外源性SO2供体组与SHR对照组比较，血压低[（128±7）mm Hg]，血管壁厚与内径比低（0．066±0．002），血浆SO2含量高[（8．3±1．0）μmol／L]，主动脉平滑肌细胞增殖指数低（0．14±0．03），凋亡指数高（0．40±0．11），主动脉平滑肌细Bcl-2蛋白表达低（0．199±0．006），Fas和caspase-3蛋白呈高表达（分别为0．218±0．003、0．251±0．011，均P〈0．01）。结论 SO2可抑制高血压大鼠主动脉血管平滑肌细胞增殖、促进其细胞凋亡。SO2促进细胞凋亡的途径可能与其对主动脉平滑肌细胞Bcl-2蛋白表达的抑制作用以及对Fas和caspase-3蛋白表达的促进作用有关。

Objective To explore the effects of sulfur dioxide （SO2 ） on the proliferation and apoptosis of aorta smooth muscle cells in hypertension ratsand possible mechanism thereof. Methods Sixteen 4-week-old male spontaneously hypertensive rats （SHRs） were randomly divided into 2 equal groups：control and Na2SO3/NaHSO3 （a SO2 donor ） -treated group. Eight 4-week-old male WKY （Wistar Kyoto） rats were assigned for normal control group. Five weeks later, the pressure was measured. The rat aortas were dyed with Hart＇s method. The morphometfic parameters were calculated by Leica workstation. The plasma level of SO2 was determined by HPLC method. VSMC apoptosis was measured by TUNEL technique. The expression levels of proliferating cell nuclear antigen （ PCNA）, Bcl-2, Fas and caspase-3 were detected by immunohistochemical assay. Results （ 1 ） Compared with those of the WKY rats, the blood pressure, ratio of media to lumen radius, and proliferation index （ PI ） of the SHRswere increased [（172± 10）mm Hgvs （112±9）mm Hg,0.073 ± 0.004vs0.057± 0.004,0.32±0.06vs0.05±O. 03, respectively], but the plasma level of SO2 and the apoptosis index （AI） were decreased in the SHRs [ （6.4 ±1.5） μmol/L vs （11.3±1. 0） μmol/L, O. 16 s O. 07 vs O. 30 s O. 19, respectively]. The expression of Bcl-2 was increased （0.209±0. 007 vs O. 202±O. 006）, and the expression levels of Fas and caspase-3 of SHRs were both lower than those of the WKY rats （0.205±O. 006 vs O. 211±O. 005, O. 229 ±0.005 vs 0.244 ±0.010, respectively）. （2） Compared with the SHR control group, the systolic blood and the ratio of media to lumen radius were decreased [ （ 128±7 ） mm Hg, O. 066 ± O. 002, respectively], but the plasma level of SO2 was increased [ （8.3± 1 ） μmol/L] for the SHR ＋ Na2SOJ NaHSO3 group. PI was lower （0.14 ±0.03） and AI was higher （0.40 ± O. 11） in SHR ＋ Na2SO3/ NaHSO3 group than those in SHR control group. The expression of Bcl-2 of VSMCs was down-regulated （0. 199 ±O. 006）, but the levels of Fas and caspase-3 were up-regulated （0. 218 ±O. 003 and O. 251±O. 011 respectively） in the SHR ＋ Na2SO3/NaHSO3 group. Conclusion SO2 may attenuate the structural remodeling through reducing the proliferation and enhancing the apoptosis of smooth muscle cells in SHRs. SO2 may modulate the process of apoptosis possibly through the downward regulation of the level of Bcl-2 and enhance the expression of Fas and caspase-3.