Modification in CLIC4 Expression is Associated with P53, TGF-β, TNF-α and Myofibroblasts in Lip Carcinogenesis

Abstract Chloride intracellular channel-4 (CLIC4) is regulated by p53 and tumor necrosis factor-α (TNF-α), it is linked to the increase of transforming growth factor-β (TGF-β), and myofibroblastic differentiation in skin carcinogenesis. This study analyzed the immunoexpression of CLIC4, p53, TGF-β, TNF-α, and α-SMA in 50 actinic cheilitis (AC) and 50 lower lip squamous cell carcinoma (LLSCC). AC and LLSCC immunoexpression were categorized as score 1 (<5% positive cells), 2 (5-50%) or 3 (>50%). For CLIC4, nuclear and cytoplasmic immunostaining of epithelial cells was considered individually. For morphologic analysis, the World Health Organization criteria were used to epithelial dysplasia grade of ACs, and Bryne grading of malignancy system was applied for LLSCC. Higher nuclear CLIC4 (CLIC4n) and TGF-β were observed in ACs with low-risk of transformation, while cytoplasmic CLIC4 (CLIC4c), p53 and TNF-α were higher in the high-risk cases (p<0.05). In LLSCCs, CLIC4c was higher in cases with lymph node metastasis, advanced clinical stages, and histological high-grade malignancy. p53 expression was higher in high-grade LLSCCs, whereas TGF-β decreased as the clinical stage and morphological grade progressed (p<0.05). ACs showed an increased expression of CLIC4n and TGF-β, while CLIC4c and α-SMA were higher in LLSCCs (p<0.0001). Both lesions showed negative correlation between CLIC4n and CLIC4c, while in LLSCCs, negative correlation was also verified between CLIC4c and p53, as well as CLIC4c and TGF-β (p<0.05). Change of CLIC4 from the nucleus to cytoplasm and alterations in p53, TGF-β, TNF-α, and α-SMA expression are involved in lip carcinogenesis.

摘要 细胞内氯离子通道4（Chloride intracellular channel-4, CLIC4）受p53与肿瘤坏死因子-α（tumor necrosis factor-α, TNF-α）调控，与转化生长因子-β（transforming growth factor-β, TGF-β）表达上调及皮肤癌变过程中的肌成纤维细胞分化密切相关。本研究对50例光化性唇炎（actinic cheilitis, AC）及50例下唇鳞状细胞癌（lower lip squamous cell carcinoma, LLSCC）组织中CLIC4、p53、TGF-β、TNF-α及α-平滑肌肌动蛋白（α-smooth muscle actin, α-SMA）的免疫表达进行了分析。AC与LLSCC的免疫表达被分为3个评分等级：评分1（阳性细胞占比<5%）、评分2（5%~50%）及评分3（>50%）。对于CLIC4，需分别评估上皮细胞的细胞核与细胞质免疫染色情况。形态学分析方面，采用世界卫生组织（World Health Organization, WHO）标准对AC的上皮异型增生程度进行分级，同时采用Bryne恶性分级系统对LLSCC进行恶性程度评估。结果显示，在低转化风险的AC组织中，细胞核CLIC4（CLIC4n）与TGF-β的表达水平更高；而高风险AC组织中，细胞质CLIC4（CLIC4c）、p53及TNF-α的表达水平显著升高（p<0.05）。在LLSCC组织中，伴淋巴结转移、临床分期较晚及组织学高级别恶性的病例，其CLIC4c表达水平更高。高级别LLSCC组织中p53表达水平更高，而TGF-β的表达则随临床分期与组织学分级升高而降低（p<0.05）。AC组织中CLIC4n与TGF-β的表达上调，而LLSCC组织中CLIC4c与α-SMA的表达水平更高（p<0.0001）。两种病变组织均显示CLIC4n与CLIC4c呈负相关；在LLSCC组织中，CLIC4c还分别与p53及TGF-β呈负相关（p<0.05）。CLIC4从细胞核向细胞质的表达移位，以及p53、TGF-β、TNF-α及α-SMA的表达异常，均参与了唇部癌变的发生发展过程。