子宮頸がんHeLa細胞の増殖、浸潤、移行、上皮間膜へのmiR-503-5pの効果 2 3

[The Effect of miR-503-5p on the Proliferation, Invasion, Migration and Epithelial Interstitium of Cervical Cancer HeLa Cells via Targeting 2 3].

• Wei Ran
• Yu-Hua Zeng
• Xiao-Jie Ma
• Ping Liao
• Xing-Lan Liu

抄録

目的:

子宮頸がんHeLa細胞の増殖、浸潤、遊走、上皮化に及ぼすmiR-503-5pの影響を調べる。

Objective: To investigate the effect of miR-503-5p on the proliferation, invasion, migration and epithelialization of cervical cancer HeLa cells via targeting 2 3.

方法:

対照群、mimic-NC群、miR-503-5p mimic群、2 3群、miR-503-5p mimic+ 2 3群（mimic+ 2 3群）の4つの子宮頸がんHeLa細胞群を設定した。各群の子宮頸がんHela細胞にプラスミドを個別にまたは共同でリポフェクタミン2000でトランスインフェクションし、トランスインフェクション後、遺伝子予測ソフトで標的遺伝子を予測し、フルオレセイン実験でターゲティング関係を確認した。miR-503-5pと2 3の発現をRT-PCRで検出、細胞増殖をMTTアッセイで検出、Ki67、増殖細胞核抗原（PCNA）、E-カドヘリン、N-カドヘリンの発現をウェスタンブロットで検出、細胞浸潤をTranswellで検出、細胞遊走をスクラッチテストで検出した。ヌードマウスを対照群とmiR-503-5p mimic群に分け、mimic-NCまたはmiR-503-5p mimicをトランスフェクトした子宮頸がんHeLa細胞懸濁液0.2mLをヌードマウスの右後肢の腹側に皮下注射した。注入から３０日後、ヌードマウスを頸椎脱臼により犠牲にした。腫瘍重量を電子天秤で秤量し、腫瘍組織中のKI67およびVimentinの発現を免疫組織化学で検出した。

Methods: Four ccervical cancer HeLa cells groups were set up including control group, mimic-NC group, miR-503-5p mimic group, 2 3 group, miR-503-5p mimic+ 2 3 group (mimic+ 2 3 group). The plasmids were separately or jointly transinfected into cervical cancer Hela cells of each group by Lipofectamine 2000, After transinfection, the target gene was predicted by gene prediction software, the targeting relationship was verified by fluorescein experiment, the expression of miR-503-5p and 2 3 was detected by RT-PCR, cell proliferation was detected by MTT assay, expression of Ki67, proliferating cell nuclear antigen (PCNA), E-cadherin and N-cadherin were detected by Western blot, cell invasion was detected by Transwell, and cell migration was detected by scratch test. Nude mice were divided into control group and miR-503-5p mimic group, and 0.2 mL of cervical cancer HeLa cell suspension transfected with mimic-NC or miR-503-5p mimic was injected subcutaneously into the ventral side of the right hind limb of nude mice. Thirty days post injection, the nude mice were sacrificed by cervical dislocation. The tumor weight was weighed by an electronic balance, and the expression of KI67 and Vimentin in the tumor tissue was detected by immunohistochemistry.

結果:

子宮頸がんHeLa細胞におけるmiR-503-5pの発現レベルは低下しており、miR-503-5pは3'UTR領域の結合部位で2 3と結合して2 3を直接標的としていた。miR-503-5pを過剰発現させると、2 3の発現が抑制され、細胞増殖率、Ki67とPCNAの発現量が有意に減少し、浸潤細胞数が減少し、傷口が広がり、治癒率が低下し、E-カドヘリンの発現が上昇し、N-カドヘリンの発現も低下した（<0.01）。miR-503-5pを過剰発現させると、移植された腫瘍の体積と重さが有意に減少し、Ki67とVimentinの陽性の割合が減少した( <0.01)。

Results: The expression level of miR-503-5p in cervical cancer HeLa cells was down-regulated, miR-503-5p directly targeted 2 3 by binding with 2 3 at binding sites in the 3'UTR region. Over-expressing of miR-503-5p inhibited the expression of 2 3, significantly decreased cell growth rate and the expression level of Ki67 and PCNA, decreased the number of invasive cells, widened the scratches, reduced the healing rate, up-regulated the expression of E-cadherin and also down-regulated the expression of N-cadherin ( <0.01). Over-expressing of miR-503-5p significantly reduced the volume and weight of transplanted tumors, and decreased the proportion of positive Ki67 and Vimentin ( <0.01).

結論:

miR-503-5pは、子宮頸がんHeLa細胞の増殖、浸潤、遊走、上皮化を阻害する。

Conclusion: miR-503-5p inhibits the proliferation, invasion, migration and epithelialization of cervical cancer HeLa cells by targeting 2 3.

Copyright© by Editorial Board of Journal of Sichuan University (Medical Science Edition).