DEVELPOMEN TO NEW TREATMENT AND DETAILS OF THE GENETIC CHANGES UNDERLYING THE NEOPLASTIC TRANSFORMATION OF UTERINE SMOOTH MUSCLE

新疗法的发展和子宫平滑肌肿瘤转化背后的基因变化细节

• 批准号: 13470350
• 负责人: NIKAIDO Toshio
• 金额: $ 9.15万
• 依托单位: Shinshu University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-13470350/
• 关键词: Leiomyosarcoma; uterine; malignancies; tumorigenicity; Calponin h1; therapeutic; smooth muscle cells; electroporation; tumor suppressor; 転写調節; 制御因子

Leiomyosarcoma constitutes only about 1.3% of uterine malignancies and one-quarter of uterine sarcomas, but it is important because it is an extremely malignant neoplasm with high rates of local recurrence and distant metastasis. The introduction of a new therapy for leiomyosarcoma of the uterus is urgently needed since neither radiotherapy nor chemotherapy is effective. However, the details of the genetic changes underlying the neoplastic transformation of uterine smooth muscle cells have not been fully characterized. Calponin hl, an actin-binding protein capable of inhibiting smooth muscle contraction, is a constitutive element of smooth muscle cells (SMCs). Among smooth muscle neoplasms of the uterus, however, leiomyosarcomas exhibit a reduced expression of calponin h1, as we reported previously. Therefore, the effect of calponin hl was assessed by transfecting its gene into leiomyosarcoma cells. A human calponin h1 cDNA construct was transfected into the human leiomyosarcoma cell l … More ines SKN and SK-LMS-lby electroporation. To confirm the gene transfer and expression of calponin h1 protein in the neomycin-resistant colonies, transfected cells were investigated by Southern blotting, RT-PCR, Western blotting and immunohistochemistry. We observed the morphology of calponin hl-transfected cells and evaluated their proliferative activity and the tumorigenicity by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl-2H-tetrazolium bromide (MTT) assay, anchorage-independent growth assay, and nude mice tumorigenicity assay. The cellular arrangement of calponin hl-transfected cells came to resemble that seen in cultured normal myometrial smooth muscle cells. The proliferation of human calponin h1-transfected leiomyosarcoma cells was inhibited to 69% of control in SK-LMS-1, and to 70% in SKN. In assays of anchorage-independent growth and nude mice tumorigenicity, growth and tumorigenicity were both significantly reduced in calponin h1-transfected leiomyosarcoma cells. Calponin h1 may function as a tumor suppressor in leiomyosarcoma. Clinically, induction of the normal differentiated phenotype by transferring the calponin h1 gene into poorly differentiated leiomyosarcoma cells may be of potential therapeutic value. Less

平滑肌肉瘤仅占子宫恶性肿瘤的1.3%和子宫肉瘤的四分之一，但它是一种非常恶性的肿瘤，局部复发率和远处转移率都很高，所以它很重要。由于放疗和化疗都不有效，迫切需要引进一种新的治疗子宫平滑肌肉瘤的方法。然而，子宫平滑肌细胞肿瘤性转化的遗传变化的细节还没有完全确定。Calponin H1是一种肌动蛋白结合蛋白，能抑制平滑肌收缩，是平滑肌细胞(SMC)的组成成分。然而，在子宫平滑肌肿瘤中，正如我们之前所报道的那样，平滑肌肉瘤显示出钙蛋白H1的表达减少。因此，通过将其基因导入平滑肌肉瘤细胞来评价其对细胞的作用。将人钙蛋白H1cDNAc DNA导入人平滑肌肉瘤细胞L…更多的INS SKN和SK-LMS-1通过电穿孔。通过Southern blotting、RT-PCR、Western blotting和免疫组织化学等方法检测细胞对新霉素耐药细胞的基因转移和表达情况。通过3-(4，5-二甲基噻唑-2-基)-2，5-二苯基-2H-四唑溴化法、锚定非依赖性生长实验和裸鼠成瘤实验观察细胞形态，评价细胞的增殖活性和致瘤性。转钙蛋白h1细胞的细胞排列与培养的正常子宫肌层平滑肌细胞相似。在SK-LMS-1和SKN中，转染人Calponin H1的人平滑肌肉瘤细胞的增殖抑制分别为对照组的69%和70%。在锚定非依赖性生长和裸鼠成瘤性实验中，转导钙粘蛋白H1的平滑肌肉瘤细胞的生长和成瘤性均显著降低。在平滑肌肉瘤中，Calponin H1可能作为肿瘤抑制因子发挥作用。在临床上，将钙蛋白H1基因导入低分化的平滑肌肉瘤细胞，诱导其正常分化表型可能具有潜在的治疗价值。较少

项目成果

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Shih HC：“β-连环蛋白的核定位与子宫内膜癌中细胞周期蛋白 D1 的表达相关。”Anticancer Res. 23. 3749-3754 (2003)

Ise H, Ferdous A, Sugihara N, Nikaido T, Negishi N, Akaike T.: "Separation of mouse hepatocytes of distinct biological phenotypes based on their asialoglycoprotein receptor-mediated adhesion to an artificial ligand."J Artif Organs. 4. 342-347 (2001)

Ise H、Ferdous A、Sugihara N、Nikaido T、Negishi N、Akaike T.：“基于脱唾液酸糖蛋白受体介导的与人工配体的粘附，分离具有不同生物表型的小鼠肝细胞。”J Artif Organs。

Takagi Y, Nikaido T, Toki T, Kita N, Kanai M, Ashida T, Ohira S, Konishi I.: "Levels of Oxidative Stress and Redox-related Molecules in the Placenta in Preeclampsia and Fetal Growth Restriction."Virchows Archiv. 444. 49-55 (2004)

Takagi Y、Nikaido T、Toki T、Kita N、Kanai M、Ashida T、Ohira S、Konishi I.：“先兆子痫和胎儿生长受限胎盘中氧化应激和氧化还原相关分子的水平。”Virchows Archive。

Kakegawa T: "Soluble asialoglycoprotein receptors reflect the apoptosis of hepatocytes"Cell Transplant. 11. 407-415 (2002)

Kakekawa T：“可溶性脱唾液酸糖蛋白受体反映肝细胞的凋亡”细胞移植。

Orii A, Masutani H, Nikaido T, Zhai Y-L, Kato K, Kariya M, Konishi I, Yodoi J, Fujii S.: "Altered post-translational modification of redox factor 1 (Ref-1) protein in human uterine smooth muscle tumors."J Clin End Metab.. 87. 3754-3759 (2002)

Orii A、Masutani H、Nikaido T、Zhai Y-L、Kato K、Kariya M、Konishi I、Yodoi J、Fujii S.：“人子宫平滑肌肿瘤中氧化还原因子 1 (Ref-1) 蛋白翻译后修饰的改变