Renin–angiotensin system inhibitors suppress azoxymethane-induced colonic preneoplastic lesions in C57BL/KsJ-db/db obese mice

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Abstract

Obesity-related metabolic abnormalities, including chronic inflammation and oxidative stress, increase the risk of colorectal cancer. Dysregulation of the renin–angiotensin system (RAS) also plays a critical role in obesity-related metabolic disorders and in several types of carcinogenesis. In the present study, we examined the effects of an angiotensin-converting enzyme (ACE) inhibitor and angiotensin-II type 1 receptor blocker (ARB), both of which inhibit the RAS, on the development of azoxymethane (AOM)-initiated colonic premalignant lesions in C57BL/KsJ-db/db (db/db) obese mice. Male db/db mice were given 4 weekly subcutaneous injections of AOM (15 mg/kg body weight), and then, they received drinking water containing captopril (ACE inhibitor, 5 mg/kg/day) or telmisartan (ARB, 5 mg/kg/day) for 7 weeks. At sacrifice, administration of either captopril or telmisartan significantly reduced the total number of colonic premalignant lesions, i.e., aberrant crypt foci and β-catenin accumulated crypts, compared to that observed in the control group. The expression levels of TNF-α mRNA in the colonic mucosa of AOM-treated db/db mice were decreased by captopril and telmisartan. Captopril lowered the expression levels of TNF-α, IL-1β, IL-6, and PAI-1 mRNAs, while telmisartan lowered the expression levels of COX-2, IL-1β, IL-6, and PAI-1 mRNAs in the white adipose tissues of these mice. In addition, these agents significantly reduced the levels of urinary 8-OHdG, a surrogate marker of oxidative damage to DNA, in the experimental mice. These findings suggested that both ACE inhibitor and ARB suppress chemically-induced colon carcinogenesis by attenuating chronic inflammation and reducing oxidative stress in obese mice. Therefore, targeting dysregulation of the RAS might be an effective strategy for chemoprevention of colorectal carcinogenesis in obese individuals.

Highlights

► Renin–angiotensin system (RAS) inhibitors (ACE inhibitor and ARB) suppress AOM-induced colorectal carcinogenesis in C57BL/KsJ-db/db obese mice. ► RAS inhibitors decreased the expression levels of inflammatory molecules, including TNF-α, in the experimental mice. ► RAS inhibitors also reduced 8-OHdG, a surrogate marker of oxidative damage to DNA, in the experimental mice. ► These findings suggested that RAS inhibitors suppress chemically-induced colon carcinogenesis by attenuating chronic inflammation and reducing oxidative stress in obese mice.

Introduction

Mounting evidence indicates that obesity, a result of a positive energy balance, and its related metabolic abnormalities raise the risk of colorectal cancer (CRC) [1], [2]. Obesity is regarded as a state of chronic inflammation, which is closely associated with colorectal carcinogenesis [3]. Increased levels of adipose tissue lead to the expression of a variety of pro-inflammatory cytokines, including tumor necrosis factor-α (TNF-α) [4], which stimulates tumor promotion and progression of carcinogenesis [5]. Oxidative stress, which is induced by increased energy availability [6], has also been suggested to play an important role in the development of CRC [1], [2]. Thus, these findings suggest that targeting inflammation and oxidative stress may be an effective strategy for preventing the development of CRC, especially in overweight individuals. For instance, a recent study shows that administration of pitavastatin, a hypolipidemic drug, prevents obesity-related colorectal tumorigenesis by attenuating chronic inflammation [7].

Hyperactivity of the renin–angiotensin system (RAS), an endocrine system with critical roles in cardiovascular function, has been implicated in the etiology of high blood pressure, obesity, and metabolic syndrome [8]. In addition, there is strong evidence that the RAS is frequently dysregulated in human malignancies, which correlates with poor patient outcomes. Abnormalities in the RAS influences cancer cell migration, invasion, and metastasis, all of which are closely associated with chronic inflammation and angiogenesis [9], [10]. In cancer tissues, the RAS is upregulated through systemic oxidative stress and hypoxia mechanisms, which triggers chronic inflammatory processes to remodel the surrounding environment [11].

Drugs that reduce the synthesis (angiotensin-converting enzyme [ACE] inhibitors) or action (angiotensin-II type-1 receptor blockers [ARBs]) of angiotensin-II, the active product of RAS, are widely used for the treatment of hypertension. These agents have also been expected to exert beneficial effects that improve the symptoms of metabolic disorders [12], [13]. In addition, retrospective studies have shown that patients taking ACE inhibitors or ARBs had decreased risk of developing some types of cancers, including CRC [14], [15], [16]. The expression levels of ACE are higher in colorectal adenomas and CRC epithelial cells than in the corresponding non-neoplastic crypt and surface epithelia [17]. In a mouse model of CRC liver metastasis, administration of an ACE inhibitor and ARB significantly reduced tumor volume by blocking the RAS activity [18]. These reports suggest that the RAS might be a critical target for the treatment and/or prevention of certain types of human malignancies, including CRC. However, the possibility of CRC chemoprevention by targeting the RAS is yet to be considered.

The C57BL/KsJ-db/db (db/db) mouse is one of the most widely used models of type 2 diabetes. The development of diabetes in db/db mice results in the activation of RAS and induction of oxidative stress, which promotes progressive inflammation [19]. In the present study, we used male db/db mice injected with azoxymethane (AOM) to examine the effects of captopril (ACE inhibitor) and telmisartan (ARB) on the development of aberrant crypt foci (ACF) and β-catenin accumulated crypts (BCAC), both of which are putative precursor lesions for colonic adenocarcinoma [20], [21], by focusing on the attenuation of inflammation and reduction of oxidative stress. This preclinical animal model is useful for investigating specific agents for their ability to prevent inflammation-related colorectal carcinogenesis caused by obesity [7].

Section snippets

Animals, chemicals, and diet

Male homozygous db/db mice aged 4 weeks (Japan SLC, Inc., Shizuoka, Japan) were maintained at the Gifu University Life Science Research Center in accordance with the Institutional Animal Care Guidelines. AOM, captopril, and telmisartan were purchased from Sigma Chemical Co. (St. Louis, MO, USA).

Experimental procedure

The animal experiment, as described previously [7], [22], [23], was approved by the Committee of the Institutional Animal Experiments of Gifu University. A total of 45 male db/db mice were divided into

General observations

As listed in Table 1, the average body weight and relative liver weight of the AOM-injected groups (Groups 4–6) at the end of the experiment were significantly (p < 0.01) lower than those of the saline-injected group (Group 1). This might be caused by the toxicity of AOM, as observed in previous experiments [7], [22], [23]. No significant differences were observed in the mean relative weight of the kidney among the groups. No histopathological findings suggesting toxicity of captopril or

Discussion

There is accumulating evidence to indicate that abnormalities in the RAS play a critical role in several types of carcinogenesis; therefore, agents targeting the RAS might augment cancer therapies [9], [10]. The results of the present study clearly indicated that the RAS inhibitors captopril and telmisartan effectively suppress the development of colonic preneoplastic lesions, ACF and BCAC, in male db/db obese mice. This is the first report that shows the preventive effect of an ACE inhibitor

Conflicts of interest

The authors declare that no conflicts of interest exist.

Acknowledgments

This work was supported in part by Grants-in-Aid from the Ministry of Education, Science, Sports and Culture of Japan (No. 22790638 to M.S. and No. 21590838 to H.M.) and by a Grant-in-Aid for the 3rd Term Comprehensive 10-Year Strategy for Cancer Control from the Ministry of Health, Labor and Welfare of Japan.

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