PTEN在主动脉夹层病变组织中的表达变化及与血管平滑肌细胞收缩表型蛋白的关系

Expression changes of PTEN in aortic dissection diseased tissues and its relationship with contractile phenotypic protein of VSMCs

目的 观察磷酸酶和张力蛋白同源物(PTEN)在主动脉夹层(AD)患者及AD小鼠病变组织中的表达变化,并探讨其表达与血管平滑肌细胞收缩表型蛋白的关系。方法 收集6例Stanford A型患者术中切除的病变主动脉(AD组)和6例供体心脏相应正常节段(对照组),采用HE染色观察主动脉结构,EVG染色观察主动脉弹力纤维,Western blotting法检测主动脉组织PTEN及血管收缩表型蛋白MYH11、α-SMA。另取3~4周龄C57BL/6J雄性小鼠12只,随机分为对照组和AD组各6只;取血管平滑肌细胞特异性敲除PTEN雄性小鼠6只作为PTENCKO组。AD组和PTENCKO组给予0.25%的β-氨基丙腈饲料喂养用于诱导AD,4周后给予血管紧张素Ⅱ1 000 ng/(kg·min)皮下泵注,连续2 d,构建AD模型;对照组给予正常饲料喂养,4周后给予等量生理盐水皮下泵注。处死小鼠,取主动脉组织,采用Western blotting法检测PTEN、MYH11、α-SMA及自噬相关蛋白LC3Ⅱ/Ⅰ、Beclin-1、p62。结果 对照组血管壁组织层次清晰,细胞排列整齐,血管壁弹性纤维排列整齐;AD组血管壁结构层次不清,部分细胞丢失、排列不齐,血管壁弹性纤维崩解断裂,胶原沉积增多。与对照组相比,AD组主动脉组织PTEN蛋白表达升高,MYH11、α-SMA蛋白表达降低(P均<0.05)。与对照组相比,AD组小鼠主动脉组织PTEN、LC3Ⅱ/Ⅰ、Beclin-1蛋白表达升高,MYH11、α-SMA、p62蛋白表达降低(P均<0.05);与AD组相比,PTENCKO组小鼠主动脉组织PTEN、LC3Ⅱ/Ⅰ、Beclin-1蛋白表达降低,MYH11、α-SMA、p62蛋白表达升高(P均<0.05)。结论 PTEN在AD患者及AD小鼠病变组织中高表达;抑制PTEN可减轻细胞自噬,恢复血管收缩表型蛋白表达,可能有助于缓解主动脉中层退行性变。

Objective To observe the expression changes of phosphatase and tension homolog(PTEN)in diseased tissues of patients with aortic dissection(AD)and AD mice,and to explore the relationship between its expression and the contractile phenotypic protein of vascular smooth muscle cells(VSMCs).Methods Six cases of diseased aortas(AD group)were collected from patients undergoing Stanford A-type surgery,along with 6 corresponding normal segments from donor hearts(control group).We utilized HE staining to observe aortic structure,EVG staining to detect elastic fiber dis⁃ruption,and Western blotting to examine the expression of PTEN,myosin heavy chain 11(MYH11)andα-smooth muscle actin(α-SMA).Additionally,12 male C57BL/6J mice aged 3-4 weeks were randomly divided into the control and AD groups,with 6 mice in each group.Six male mice with vascular smooth muscle-specific knockout of PTEN were selected as the PTENCKO group.To establish the AD model,mice in the AD group and the PTENCKO group were fed with a diet contain⁃ing 0.25%β-aminopropionitrile for 4 weeks,followed by subcutaneous infusion of AngⅡat a dose of 1000 ng/(kg·min)for 2 consecutive days.Mice in the control group were fed with a normal diet,and after 4 weeks,an equal volume of nor⁃mal saline was subcutaneously injected.After euthanasia,aortic tissues were collected,and Western blotting was per⁃formed to detect PTEN,MYH11,α-SMA,and autophagy-related proteins LC3Ⅱ/Ⅰ,Beclin-1,and p62.Results In the control group, the vascular wall tissue structure was clear, with orderly cell arrangement and neatly arranged elastic fi⁃ bers in the vascular wall. However, in the AD group, the structure of the vascular wall was unclear, with partial loss and disordered arrangement of cells, as well as disintegration and fracture of elastic fibers in the vascular wall, accompanied by increased collagen deposition. Compared with the control group, the expression of PTEN protein in the aortic tissues in⁃ creased in the AD group, while the expression levels of MYH11 and α-SMA proteins decreased (all P<0. 05). Compared with the control group, the expression of PTEN, LC3Ⅱ/Ⅰ, and Beclin-1 protein in the aortic tissues increased in mice of the AD group, while the expression levels of MYH11, α-SMA, and p62 protein decreased (all P<0. 05). Conversely, compared with the AD group, mice in the PTENCKO group showed decreased expression levels of PTEN, LC3Ⅱ/Ⅰ, and Beclin-1 protein in the aortic tissues, while the expression levels of MYH11, α-SMA, and p62 protein increased (all P<0. 05). Conclusions PTEN is highly expressed in diseased tissues of AD patients and AD mice. Inhibiting PTEN can effectively alleviate autophagy, restore the expression of vascular contraction phenotype protein, and help to attenuate aortic medial degeneration.