Role of hypoxia-inducible factor-1α and CD146 in epidermal growth factor receptor-mediated angiogenesis in salivary gland adenoid cystic carcinoma

Abstract

Adenoid cystic carcinoma (AdCC) of the salivary gland in the head and neck is characterized by indolent yet persistent growth, multiple local recurrences and early hematogenous metastasis. Considering the possible association between the epidermal growth factor receptor (EGFR) signaling pathway and angiogenesis in various types of cancer and the overexpression of EGFR in AdCC, it is reasonable to examine the correlation between angiogenesis and the EGFR signaling pathway in this carcinoma. In the present study, the expression of EGFR, CD31, CD146 and hypoxia‑inducible factor‑1α (HIF‑1α) were evaluated by immunohistochemical staining with tissue microarray containing normal salivary gland (NSG), pleomorphic adenoma (PMA) and AdCC tissues. Pearson's correlation coefficient was conducted to demonstrate the correlation between EGFR, CD31, CD146 and HIF‑1α. To determine their similarity and intimacy, hierarchical analysis was performed with Cluster 3.0 and then visualized using TreeView software. Immunohistochemical results of tissue microarrays were quantified, revealing that the expression of EGFR, CD146 and HIF‑1α increased in AdCC compared with in PMA and NSG tissues. The association between the expression of EGFR and CD31 was significant and positive. The expression of CD146 and HIF‑1α was positively correlated with EGFR and CD31, respectively. These findings suggest that the EGFR signaling pathway has a vital role in AdCC progression and may be associated with HIF‑1α‑mediated angiogenesis. These results may enhance our understanding of the mechanism underlying AdCC progression and provide potential clinical therapeutic strategies based on the inhibition of EGFR.

Figure 1

Association between the expression of EGFR and CD31 in NSG, PMA and AdCC tissues. Representative immunohistochemical staining of (A) EGFR membranous expression and (B) CD31 membranous expression in human NSG, PMA and cribriform, tubular or solid type AdCC tissues. Scale bar=50 µm. Quantification of EGFR expression levels in (C) human NSG, PMA and AdCC tissues and (D) subtypes of AdCC using an AperioScanscope scanner and software. Data were analyzed by Graph Pad Prism 5 software. Data are presented as the mean ± standard error of the mean. ^*P<0.05, AdCC vs. PMA tissues; ^**P<0.01, AdCC vs. NSG tissues. (E) Correlation between EGFR and CD31 expression levels in human NSG, PMA and AdCC tissues (P<0.01, r=0.2618, n=104) using two-tailed Pearson's test. EGFR, epidermal growth factor receptor; NSG, normal salivary gland; PMA, polymorphism adenoma; AdCC, adenoid cystic carcinoma.

Figure 2

Expression of HIF-1α and CD146 in NSG, PMA and AdCC tissues. Representative immunohistochemical staining of (A) CD146 membranous expression and (B) HIF-lα cytoplasmic and nuclear expression in human NSG, PMA and cribriform, tubular or solid type AdCC tissues. Scale bar=50 µm. (C) Quantification of HIF-1α and CD146 expression levels in human NSG, PMA and AdCC tissues and subtypes of AdCC using an AperioScanscope scanner and software. Data were analyzed using Graph Pad Prism 5 software. Data are expressed as the mean ± standard error of the mean. ^*P<0.05, AdCC vs. PMA tissues in CD146, AdCC vs. NSG tissues in HIF1α, solid vs. cribriform subtype tissues in CD146; ^***P<0.001, AdCC vs. NSG tissues in CD146, AdCC vs. PMA tissues in HIF1α. EGFR, epidermal growth factor receptor; NSG, normal salivary gland; PMA, polymorphism adenoma; AdCC, adenoid cystic carcinoma; HIF-1α, hypoxia-inducible factor-1α.

Figure 3

HIF-1α and CD146 may be involved in angiogenesis in AdCC. Correlation and regression of HIF-1α and CD146 in human NSG, PMA and AdCC tissues. (A) Correlation between HIF-1α with CD31 expression levels in human NSG, PMA and AdCC tissues (P<0.001, r=0.5236, n=104). (B) Correlation between CD146 with CD31 expression levels in human NSG, PMA and AdCC tissues (P<0.001, r=0.3346, n=104) using two-tailed Pearson's test. HIF-1α, hypoxia-inducible factor-1α; NSG, normal salivary gland; PMA, polymorphism adenoma; AdCC, adenoid cystic carcinoma.

Figure 4

Overexpression of EGFR is correlated with HIF-1α and CD146 in human AdCC tissue. (A) Expression of EGFR was positively correlated with HIF1-α and CD146 (P<0.05, r=0.2154, n=104; P<0.001, r=0.4701, n=104, respectively) and the (B) expression of HIF-1α was significantly correlated with CD146 (P<0.001, r=0.3491, n=104) in human NSG, PMA and AdCC tissues by analyzing the tissue microarray immunohistochemical staining. (C) Hierarchical clustering of immunohistochemical results of human AdCC with EGFR, HIF-1α and CD146 (statistics including AdCC tissue only n=74). EGFR, epidermal growth factor receptor; NSG, normal salivary gland; PMA, polymorphism adenoma; AdCC, adenoid cystic carcinoma; HIF-1α, hypoxia-inducible factor-1α.