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OBJECTIVE To investigate the effect of sputum ubiquitin ligase (Cbl-b) gene known-down on the cytotoxicity of H9 T lymphocytes against human laryngeal squamous cancer Hep-2 cells and explore the underlying mechanism. METHODS CD4+ T lymphocytes isolated from 12 patients with laryngeal squamous carcinoma and 12 healthy individuals were examined for Cbl-b mRNA expressions using RT-PCR. H9 T lymphocytes cultured in 96-well plates were transfected with Cbl-b siRNA via liposomes followed by treatment with an anti-IL-2 monoclonal antibody, with H9 T lymphocytes transfected with a scrambled sequence as the negative control. The expressions of Cbl-b mRNA and protein in the cells were detected using real-time fluorescent quantitative PCR and Western blotting, respectively. The killing effect of the treated T lymphocytes against Hep-2 cells was assessed using the cell counting kit (CCK-8). The positive expression rates of CD69 and CD25 on the surface of H9 T lymphocytes were determined using flow cytometry, and the levels of interleukin-2 (IL-2) and interferon-gamma (INF-γ) in the culture supernatants of H9 T lymphocytes were detected with ELISA. RESULTS The CD4+ T lymphocytes from patients with laryngeal squamous carcinoma showed significantly increased Cbl-b mRNA level compared with those from healthy individuals (P < 0.05). Transfection of H9 T lymphocytes with Cbl-b siRNA significantly reduced the expression levels of Cbl-b mRNA and protein (P < 0.05), which were not significantly affected by subsequent treatment of the cells with the anti-IL-2 antibody (P>0.05). At different target-effector ratios, the Cbl-b siRNA-transfected cells showed significantly higher Hep-2 cell killing rates and higher positivity rates of CD69 and CD25 expressions than the blank and negative control cells and the cells with both Cbl-b siRNA transfection and anti-IL-2 treatment (P < 0.05). Cbl-b silencing in H9 T lymphocytes resulted in significantly increased levels of IL-2 and INF-γ in the supernatant as compared with those in the blank and negative control groups (P < 0.05). CONCLUSIONS Cbl-b gene silencing effectively enhances the killing effect of H9 T lymphocytes against Hep-2 cells in vitro probably as the result of enhanced IL-2 secretion and T lymphocyte activation.
Saiming Chen, and
Zhiqun Li, and
Limin Zhou, and
Yunxia Zhang
January 2015,
International journal of clinical and experimental pathology,
Saiming Chen, and
Zhiqun Li, and
Limin Zhou, and
Yunxia Zhang
February 2010,
Oncology reports,
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Zhiqun Li, and
Limin Zhou, and
Yunxia Zhang
August 2010,
Zhonghua er bi yan hou tou jing wai ke za zhi = Chinese journal of otorhinolaryngology head and neck surgery,
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Zhiqun Li, and
Limin Zhou, and
Yunxia Zhang
June 2007,
Lin chuang er bi yan hou tou jing wai ke za zhi = Journal of clinical otorhinolaryngology, head, and neck surgery,
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Zhiqun Li, and
Limin Zhou, and
Yunxia Zhang
June 2014,
Oncology letters,
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Zhiqun Li, and
Limin Zhou, and
Yunxia Zhang
April 2016,
Apoptosis : an international journal on programmed cell death,
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Zhiqun Li, and
Limin Zhou, and
Yunxia Zhang
June 2003,
Lin chuang er bi yan hou ke za zhi = Journal of clinical otorhinolaryngology,
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Zhiqun Li, and
Limin Zhou, and
Yunxia Zhang
February 2008,
Lin chuang er bi yan hou tou jing wai ke za zhi = Journal of clinical otorhinolaryngology, head, and neck surgery,
Saiming Chen, and
Zhiqun Li, and
Limin Zhou, and
Yunxia Zhang
February 2011,
Lin chuang er bi yan hou tou jing wai ke za zhi = Journal of clinical otorhinolaryngology, head, and neck surgery,
Saiming Chen, and
Zhiqun Li, and
Limin Zhou, and
Yunxia Zhang
September 2008,
Human immunology,
