Abstract
BackgroundHuman head and neck squamous carcinoma is the 6th most prevalent carcinoma worldwide. Although many novel therapies have been developed, the clinical treatment for patients remains non-ideal. Chloride intracellular channel 4 (CLIC4), one of the seven members of the CLIC family, is a newly found Cl− channel that participates in various biological processes, including cellular apoptosis and differentiation. Accumulating evidence has revealed the significant role of CLIC4 in regulating the apoptosis of different cancer cells. Here, we investigated the functional role of CLIC4 in the apoptosis of HN4 cells, a human head and neck squamous carcinoma cell line.ResultsIn the present study, we used immunohistochemical staining to demonstrate that the expression level of CLIC4 is elevated in the tissue of human oral squamous carcinoma compared with healthy human gingival tissue. Specific CLIC4 small interfering RNA was used to knockdown the expression of CLIC4. The results showed that knockdown of CLIC4 with or without 100 μM adenosine triphosphate (ATP) treatment significantly increased the expression of Bax, active caspase 3, active caspase 4 and CHOP but suppressed Bcl-2 expression in HN4 cells. Moreover, the results from the TdT-mediated dUTP nick end labeling assay indicated that CLIC4 knockdown induced a higher apoptotic rate in HN4 cells under the induction of ATP. In addition, knockdown of CLIC4 dramatically enhanced ATP-induced mitochondrial membrane depolarization in HN4 cells. Moreover, intracellular Ca2+ measurement revealed that Ca2+ release induced by ATP and thapsigargin, a Ca2+-ATPase inhibitor of the endoplasmic reticulum, was significantly enhanced by the suppression of CLIC4 in HN4 cells.ConclusionsKnockdown of CLIC4 enhanced ATP-induced apoptosis in HN4 cells. Both the pathways of mitochondria and endoplasmic reticulum stress were involved in CLIC4-mediated cell apoptosis. Based on our finding, CLIC4 may be a potential and valuable target for the clinical treatment of head and neck squamous carcinoma.Electronic supplementary materialThe online version of this article (doi:10.1186/s13578-016-0070-1) contains supplementary material, which is available to authorized users.
Highlights
Human head and neck squamous carcinoma is the 6th most prevalent carcinoma worldwide
Knockdown of Chloride intracellular channel 4 (CLIC4) enhances Adenosine triphosphate (ATP)‐induced HN4 cell apoptosis via the mitochondrial pathway Recent studies have reported that CLIC4 was a regulator for cancer cell apoptosis [8]
Our data showed that the percentage of apoptotic cells was significantly increased in CLIC4-specific small interfering RNA (siRNA)-transfected cells compared with scrambled siRNA-transfected ones (Fig. 2a, b)
Summary
Human head and neck squamous carcinoma is the 6th most prevalent carcinoma worldwide. Chloride intracellular channel 4 (CLIC4), one of the seven members of the CLIC family, is a newly found Cl− channel that participates in various biological processes, including cellular apoptosis and differentiation. We investigated the functional role of CLIC4 in the apoptosis of HN4 cells, a human head and neck squamous carcinoma cell line. Multiple studies have investigated the expression profile and regulatory effect on the apoptosis of CLIC4 in cancer cells [8, 10,11,12,13]. The expression profile and regulatory effect on apoptosis of CLIC4 in each specific cancer cell may be different. Because CLIC4 is indicated as an integral part of the cellular stress-response pathway and is a significant regulator of cell apoptosis and growth in live cells, we hypothesize that the regulation of CLIC4 protein expression may affect ATP-induced apoptosis
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