Department of Gynecology and Obstetrics, Johns Hopkins University School of Medicine, Baltimore, MD.
Department of Obstetrics and Gynecology, Uniformed Services University of the Health Sciences, Bethesda, MD.
Am J Obstet Gynecol. 2020 Nov;223(5):733.e1-733.e14. doi: 10.1016/j.ajog.2020.05.012. Epub 2020 May 15.
Uterine leiomyomas, the most common tumors of the female reproductive system, are characterized by excessive deposition of disordered stiff extracellular matrix and fundamental alteration in the mechanical signaling pathways. Specifically, these alterations affect the normal dynamic state of responsiveness to mechanical cues in the extracellular environment. These mechanical cues are converted through integrins, cell membrane receptors, to biochemical signals including cytoskeletal signaling pathways to maintain mechanical homeostasis. Leiomyoma cells overexpress β1 integrin and other downstream mechanical signaling proteins. We previously reported that simvastatin, an antihyperlipidemic drug, has antileiomyoma effects through cellular, animal model, and epidemiologic studies.
This study aimed to examine the hypothesis that simvastatin might influence altered mechanotransduction in leiomyoma cells.
This is a laboratory-based experimental study. Primary leiomyoma cells were isolated from 5 patients who underwent hysterectomy at the Department of Gynecology and Obstetrics of the Johns Hopkins University Hospital. Primary and immortalized human uterine leiomyoma cells were treated with simvastatin at increasing concentrations (0.001, 0.01, 0.1, and 1 μM, or control) for 48 hours. Protein and mRNA levels of β1 integrin and extracellular matrix components involved in mechanical signaling were quantified by quantitative real-time polymerase chain reaction, western blotting, and immunofluorescence. In addition, we examined the effect of simvastatin on the activity of Ras homolog family member A using pull-down assay and gel contraction.
We found that simvastatin significantly reduced the protein expression of β1 integrin by 44% and type I collagen by 60% compared with untreated leiomyoma cells. Simvastatin-treated cells reduced phosphorylation of focal adhesion kinase down to 26%-60% of control, whereas it increased total focal adhesion kinase protein expression. Using a Ras homolog family member A pull-down activation assay, we observed reduced levels of active Ras homolog family member A in simvastatin-treated cells by 45%-85% compared with control. Consistent with impaired Ras homolog family member A activation, simvastatin treatment reduced tumor gel contraction where gel area was 122%-153% larger than control. Furthermore, simvastatin treatment led to reduced levels of mechanical signaling proteins involved in β1 integrin downstream signaling, such as A-kinase anchor protein 13, Rho-associated protein kinase 1, myosin light-chain kinase, and cyclin D1.
The results of this study suggest a possible therapeutic role of simvastatin in restoring the altered state of mechanotransduction signaling in leiomyoma. Collectively, these findings are aligned with previous epidemiologic studies and other reports and support the need for clinical trials.
子宫肌瘤是女性生殖系统最常见的肿瘤，其特征是细胞外基质过度沉积、排列紊乱和僵硬，以及机械信号通路的基本改变。具体来说，这些改变会影响细胞对外界环境中机械线索的正常动态反应能力。这些机械线索通过整合素、细胞膜受体转化为生化信号，包括细胞骨架信号通路，以维持机械内稳态。子宫肌瘤细胞过度表达β1 整合素和其他下游机械信号蛋白。我们之前的研究报告表明，辛伐他汀作为一种降脂药物，通过细胞、动物模型和流行病学研究具有抗子宫肌瘤作用。
本研究旨在检验辛伐他汀可能影响子宫肌瘤细胞中改变的机械转导的假设。
这是一项基于实验室的实验研究。从在约翰霍普金斯大学医院妇产科接受子宫切除术的 5 名患者中分离出原代子宫肌瘤细胞。用不同浓度（0.001、0.01、0.1 和 1 μM 或对照）的辛伐他汀处理原代和永生化人子宫肌瘤细胞 48 小时。通过实时定量聚合酶链反应、western blot 和免疫荧光定量分析β1 整合素和细胞外基质成分的蛋白质和 mRNA 水平，这些成分参与机械信号转导。此外，我们使用下拉测定和凝胶收缩实验研究了辛伐他汀对 Ras 同源家族成员 A 活性的影响。
与未处理的子宫肌瘤细胞相比，辛伐他汀使β1 整合素的蛋白表达降低了 44%，I 型胶原降低了 60%。辛伐他汀处理的细胞使粘着斑激酶的磷酸化降低至对照的 26%-60%，而总粘着斑激酶蛋白表达增加。使用 Ras 同源家族成员 A 下拉激活测定法，我们观察到辛伐他汀处理的细胞中活性 Ras 同源家族成员 A 的水平降低了 45%-85%，与对照相比。与 Ras 同源家族成员 A 激活受损一致，辛伐他汀处理导致肿瘤凝胶收缩减少，凝胶面积比对照大 122%-153%。此外，辛伐他汀处理导致参与β1 整合素下游信号的机械信号蛋白水平降低，如 A-激酶锚蛋白 13、Rho 相关蛋白激酶 1、肌球蛋白轻链激酶和细胞周期蛋白 D1。
本研究结果提示辛伐他汀在恢复子宫肌瘤中改变的机械转导信号状态方面可能具有治疗作用。总的来说，这些发现与之前的流行病学研究和其他报告一致，并支持进行临床试验的需要。
