Abalone is often called as 'The emperor of the seashells' in Korea and mostly consumed as stamina food for sick and weak individuals because of its high content of proteins and vitamins [
5]. Abalone mainly live on sea weeds that contain rich and concentrated nutritional components [
2]. Apart from its known nutritional importance, there are not many studies on the effects of the abalone visceral extract. One of the known effects of abalone visceral extract is its antioxidant activity as demonstrated [
4,
5]. However, there are still few
in vivo evidence and no detailed action mechanisms for its anti-tumor effects [
7]. In the present study, we have demonstrated the potent anti-tumor efficacy of abalone visceral extract and have elucidated its underlying mechanism using a mouse breast cancer model that have high malignancy in tumor growth and metastasis [
13,
14]. Oral administration of abalone visceral extract significantly lowered tumor progression and metastasis by down-regulating the tumor-associated growth factors such as Cox-2, EGF, VEGF and FGF, while increasing the proliferation and cytolytic activity of CD8
+ T cells.
Cyclooxygenase-2 (Cox-2) is an enzyme that catalyzes arachidonic acid to prostaglandins. Cox-2 is predominantly expressed in synoviocytes, fibroblasts, osteoblasts, activated endothelial cells and tumor cells [
34,
35]. Cox-2 expression is induced by pro-inflammatory and mitogenic stimuli such as growth factors (EGF, FGF and VEGF) [
36] and cytokines (TNF-α and IL1-β). Enhanced expression of Cox-2 is linked with tumor progression by inducing immune suppression as well as angiogenic and metastatic progression [
34,
37,
38]. Elevated Cox-2 expression is associated with increased tumor size during breast cancer progression [
22,
39]. Modulation of Cox-2 expression by specific inhibitors is regarded as good chemopreventive approach for cancer treatment. However, Cox-2 inhibitors affect multiple cellular pathways and show some side effects [
34,
40]. Therefore, use of nutritive supplementation substances might be regarded as potential cancer preventive approach [
11,
12]. We have found that the oral administration of abalone visceral extract exerted anti-tumor growth effects by inhibiting tumor volume (up to 30%) compared with control feeding group (Figure.
1). The mouse breast tumor induced by 4T1 tumor cells mimics human breast cancer in the aspect of spontaneous metastasis to lung, lymph nodes, liver and bone [
41]. Cox-2 expression has been identified as the marker for selective lung metastasis [
27] in breast cancer model [
20]. In this study, we used 4T1 mammary adenocarcinoma cells for tumor implantation. Oral administration of abalone visceral extract significantly inhibited tumor metastasis by modulating Cox-2 expression (Figure.
4). In accordance with our result, a previous study also demonstrated that treatment of either non-selective Cox inhibitor or selective Cox-2 inhibitor significantly reduced primary tumor growth and metastasis of 4T1 breast cancer cells[
42]. Even though reduction of Cox-2 expression does not exactly match with inhibition of Cox-2 activity by known inhibitors, specific knockdown of Cox-2 directly could reduce level of PGE2 synthesis in 4T1 cells [
23]. In line with aforementioned reports, we investigated whether abalone visceral extract changes Cox-2 expression upon treatment. Oral administration of abalone visceral extract reduced the metastatic splenomegaly (Figure.
2A and
2B) [
17,
18] and lymphomegaly (data not shown) [
43]. Metastatic breast cancer has a strong tendency to propagate into lung and bone [
19‐
21,
26]. Treatment of abalone visceral extract significantly inhibited lung metastasis (Figure.
2C and
2D) by decreasing Cox-2 expression level (Figure.
4A and
4B). Numerous evidences show that decreased level of Cox-2 is well correlated with metastatic inhibition from variety kinds of cancers [
44]. Furthermore, previous data suggested that Cox-2 expression is associated with angiogenesis, lymph node metastasis, and apoptosis in human breast cancer [
43] along with enhanced MMP-13 expression [
28]. Interestingly, the expression levels of VEGF, EGF and MMP-13 are all decreased upon abalone visceral extract treatment (Figure.
4C). Collectively, oral administration of abalone visceral extract reduced metastatic progression by lowering Cox-2 expression and other target molecules including angiogenic factors and metalloproteinases in the metastatic tissues.
The tumor microenvironment induces active immune tolerance and escapes immune surveillance. Boosting the immune response can be one of the indirect ways to eliminate or suppress tumor growth via regulating immune homeostasis [
30]. CD8
+ T cells are known to have anti-tumor activity by killing the tumor antigens in an antigen specific or antigen non-specific way [
44,
45]. Tumor specific CD8
+ T cells possess increased proliferation, cytolytic activity and induce expression of death related proteins and cytokines [
31]. However, CD8
+ T cells at tumor sites or tumor draining lymph nodes frequently exhibit functional defects such as defective antigen specific cytolytic activity [
46], lack of perforin expression [
47], defective cytokine production and abnormal proliferation [
48,
49]. Enhanced CD8
+ T cell activity is therefore critical to eradicate tumor cells, especially in tumor regions. In this study, oral administration of abalone visceral extract significantly inhibited tumor growth compared with the control (PBS-treated) group (Figure.
1 and Figure.
2). Administration of abalone visceral extract enhanced the cytolytic activity of CD8
+ T cells by increasing the expression of effector molecules such as cytokines (IFN-γ and TNF-α) and cytolytic molecules (FasL, Gzm B and Gzm C) (Figure.
5B and
5C). Even though inflammatory cytokine signaling is the known stimulation for Cox-2 expression [
24], increased expression of the cytokine in CD8+ T cells upon abalone visceral extract treatment can be explained by other mechanisms apart from Cox-2 regulation by abalone visceral extract in tumor cells. In addition, abalone visceral extract significantly increased the specific lysis rate in the JAM test (Figure.
5D). Therefore, the enhanced effector function of CD8
+ T cells by administration of abalone visceral extract may enhance anti-tumor immunity, which leads to suppression of tumor growth and metastasis to different organs.