**1. Introduction**

High dietary cholesterol intake may lead to increased risk of diseases such as cardiovascular disease and diabetes [1, 2]. Although the recommendation to restrict daily dietary cholesterol

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

intake (300 mg) was removed from the 2015–2020 Dietary Guidelines for Americans [3], it is still recommended that individuals minimize cholesterol consumption. Animal foods such as egg yolk, meats, dairy products, fish, and poultry are major sources of dietary cholesterol. Meanwhile, dietary cholesterol is not found in plant foods. Instead, many plants contain phytosterols, which are chemically similar to cholesterol, and can therefore compete with it and decrease its absorption in the intestinal tract [4]. The effect of dietary cholesterol on plasma cholesterol levels remains undetermined, since the body may suppress endogenous cholesterol synthesis in response to additional cholesterol ingestion [1]. Some studies have suggested that dietary cholesterol increases serum total cholesterol (TC), low-density lipoprotein (LDL) cholesterol, as well as the ratio of LDL to high-density lipoprotein cholesterol [5–8], which are considered to be associated with risk of vascular diseases.

cholesterol and 60% fat) induced steatohepatitis, cellular ballooning, and fibrosis in the livers of male C57Bl/6J mice [21]. We previously established an HFC diet-induced NASH model

The Role of Cholesterol in the Pathogenesis of Hypertension-Associated Nonalcoholic…

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SHRSP5/Dmcr rats are the fifth substrain of the stroke-prone spontaneously hypertensive (SHRSP) rat [20, 22], which is derived from the SHR strain [23]. To establish this strain, SHRSP rats were fed an HFC diet for 1 week, then those with high serum cholesterol levels (600–900 mg/dL in females and 300–600 mg/dL in males) were selected for brother–sister inbreeding. Selective inbreeding was repeated and offspring with increased hypercholesterolemic responses were obtained. Although the SHRSP5/Dmcr rats, formally known as arteriolipidosis-prone rats, were developed as an animal model of arteriosclerosis, marked enlargement and an abnormal whitish color of the liver were noted in the 47th generation. These findings prompted our studies on HFC diet-induced liver damage in this strain.

In order to determine whether the HFC diet-fed SHRSP5/Dmcr strain was a suitable model of NASH, we investigated hepatic histopathological changes following HFC feeding [20]. Male SHRSP5/Dmcr rats at 10 weeks of age were fed either an HFC (35.3% crude lipid and 5% cholesterol) or control diet (4.8% crude lipid and no additional cholesterol) for 2, 8, and 14 weeks. We found that the HFC diet induced microvesicular steatosis and lymphocyte infiltration at 2 weeks. Macrovesicular steatosis, ballooned hepatocytes with eosinophilic Mallory-Denk bodies, and multilobular necrosis were observed in the livers of rats fed an HFC diet at 8 weeks. The severity of steatosis and hepatocyte ballooning was further increased at 14 weeks. Meanwhile, a progressive deterioration of hepatic fibrosis occurred during HFC feeding. Slight pericellular and perivenular fibrotic changes, bridging fibrosis, and end-stage honeycomb fibrosis were observed at 2, 8, and 14 weeks, respectively. In addition, the HFC diet induced a progressive increase in indicators of liver damage, including serum levels of alanine transaminase (AST), aspartate transaminase (ALT), and γ-glutamyltranspeptidase (γ-GTP). Matteoni et al. classified human NAFLD into four types according to histological analysis of liver biopsy specimens: type 1, fatty liver alone; type 2, fat accumulation and lobular inflammation; type 3, fat accumulation and ballooning degeneration; and type 4, fat accumulation, ballooning degeneration, and hepatic fibrosis [24]. The histological characteristics observed in the liver of the SHRSP5/Dmcr strain at 2, 8, and 14 weeks of HFC feedings were very similar to those in type 2, type 3 or 4, and type 4 human NAFLD, respectively. Therefore, all pathological stages of NAFLD can be observed in the SHRSP5/Dmcr strain during HFC feeding. In addition, obesity, insulin resistance, and diabetes were not observed in this model. Therefore, it represents an excellent model of NAFLD/NASH without obesity and

diabetes, and is useful for studying the pathogenesis and therapeutics of this disease.

We further investigated the molecular mechanisms underlying the progression of HFCinduced NASH in the SHRSP5/Dmcr strain [25]. Rats were fed either an HFC or control diet for 2, 8, and 16 weeks, and expression of genes involved in inflammation and hepatic fibrosis was evaluated. Tumor necrosis factor α (TNF-α), a proinflammatory cytokine, was reported to be upregulated in the livers of NASH patients [26]. We showed that the HFC diet increased the hepatic expression of TNF-α in SHRSP5/Dmcr rats at all time points. Nuclear factor κB (NF-κB; p50/p65) and inhibitor of κBα, the proteins involved in NF-κB signaling, which is regulated by TNF-α and plays an important role in inflammatory response, were also upregulated by the HFC diet. Hepatocyte injury and inflammation led to hepatic fibrosis via hepatic stellate

using hypertensive SHRSP5/Dmcr rats [20].

Dietary cholesterol is also linked to the pathogenesis of nonalcoholic fatty liver disease (NAFLD)/nonalcoholic steatohepatitis (NASH) [9, 10]. NAFLD is one of the most common chronic liver diseases worldwide and comprises a spectrum of liver damage, from simple steatosis (a benign non-progressive condition) to NASH, the advanced form that may progress to hepatic cirrhosis or hepatocellular carcinoma [11]. The pathological characteristics of NASH include steatosis, hepatocellular ballooning, lobular inflammation, and hepatic fibrosis. Cholesterol may contribute to NASH development by being catabolized in the liver into bile acids (BAs), which are hepatotoxic and cause liver damage [12]. Li et al. demonstrated that dietary cholesterol exacerbates liver damage and hepatic inflammation in mice fed a high-fat diet [13]. Subramanian et al. reported that an LDL receptor-deficient mouse fed a high-fat, high-carbohydrate diet was a good animal model of NAFLD/NASH, and showed that dietary cholesterol worsened hepatic steatosis and inflammation in this model [9].

In addition, NAFLD/NASH was described as a hepatic manifestation of metabolic syndrome, and its development was associated with hypertension, obesity, diabetes, and hyperlipidemia [14, 15]. Some studies have shown an increased prevalence of NAFLD/NASH among hypertensive patients [16–18]. Using spontaneously hypertensive (SHR) rats fed a choline-deficient diet as a hypertensive animal model of NASH, and its normotensive control, the Wistar Kyoto (WKY) rat [19], Ikuta et al. revealed that hypertension enhances the progression of NASH. We previously developed a novel animal model of hypertension-associated NASH by feeding stroke-prone spontaneously hypertensive5/Dmcr (SHRSP5/Dmcr) rats a high fat and cholesterol (HFC) diet [20]. Further studies from our group suggested that dietary cholesterol may have a potential effect on the development of hypertension-associated NASH (unpublished).

In this chapter, we will discuss the crucial role of dietary cholesterol in the progression of hypertension-associated NASH.
