DEN-Induced Rat Model Reproduces Key Features of Human Hepatocellular Carcinoma

Abstract

Hepatocellular carcinoma (HCC) is the most common type of liver cancer. The majority of HCC cases are associated with liver fibrosis or cirrhosis developing from chronic liver injuries. The immune system of the liver contributes to the severity of tissue damage, the establishment of fibrosis and the disease’s progression towards HCC. Herein, we provide a detailed characterization of the DEN-induced HCC rat model during fibrosis progression and HCC development with a special focus on the liver’s inflammatory microenvironment. Fischer 344 male rats were treated weekly for 14 weeks with intra-peritoneal injections of 50 mg/kg DEN. The rats were sacrificed before starting DEN-injections at 0 weeks, after 8 weeks, 14 weeks and 20 weeks after the start of DEN-injections. We performed histopathological, immunohistochemical, RT-qPCR, RNA-seq and flow cytometry analysis. Data were compared between tumor and non-tumor samples from the DEN-treated versus untreated rats, as well as versus human HCCs. Chronic DEN injections lead to liver damage, hepatocytes proliferation, liver fibrosis and cirrhosis, disorganized vasculature, and a modulated immune microenvironment that mimics the usual events observed during human HCC development. The RNA-seq results showed that DEN-induced liver tumors in the rat model shared remarkable molecular characteristics with human HCC, especially with HCC associated with high proliferation. In conclusion, our study provides detailed insight into hepatocarcinogenesis in a commonly used model of HCC, facilitating the future use of this model for preclinical testing.

Chronic Intermittent Hypoxia Increases Cell Proliferation in Hepatocellular Carcinoma

“…This induced progressive liver damage and liver carcinogenesis, leading to nodule development in 100% of the animals after 14 weeks and to fully developed HCC at 14 + 6 weeks, as illustrated in Figure 1b. Confirming previous results [15], a DEN treatment strongly promoted hepatocyte proliferation, as assessed by Cyclin D1 staining (Figure 1c, left panels). Similarly, chronic DEN injections led to the development of fibrosis/cirrhosis, as revealed by Sirius red staining (Figure 1c, middle panels).…”

“…Seven-week-old Fischer 344 male rats were treated once a week with intra-peritoneal injections of 50 mg/kg DEN (Sigma-Aldrich, Steinheim am Albuch, Germany) diluted in pure olive oil (Sigma-Aldrich, Steinheim am Albuch, Germany) in order to obtain HCC on a fibrotic/cirrhotic liver after 14 weeks, as published previously [15]. We analyzed three different time points of DEN treatment: (i) the 8-week group was sacrificed after 8 weeks of DEN injection, (ii) the 14-week group was sacrificed after 14 weeks of DEN injection, and (iii) the 14 + 6-week group was sacrificed after 14 weeks of DEN injection followed by 6 weeks without any DEN injections.…”

“…The diethylnitrosamine (DEN) chronically induced rat model reproduces the pathogenesis of HCC, as seen in humans. This includes liver damage, chronic inflammation, hepatocytes proliferation, liver fibrosis and cirrhosis, disorganized vasculature, and modulations of the liver's immune microenvironment [15][16][17]. In general, the animal model of HCC mimics particularly well the "high proliferation" subtype of human HCC, without additional metabolic liver disease.…”

Geraniol ve vitamin C’nin dietilnitrozamin kaynaklı deneysel hepatoselüler karsinogenez üzerindeki etkisi

“…Approximately one out of every 100 cancer cases encountered has been diagnosed with liver cancer. Liver tumors do not show any clinical findings other than fever, abdominal pain, jaundice, and hepatomegaly 2,3,5 . Diethylnitrosamine (DENA), which is a hepatocarcinogen; is known to disrupt nuclear enzymes involved in DNA repair and replication and is used as a carcinogen to induce liver cancer 6,7 .…”

“…Regeneration in the liver plays an important role in the development of hepatocellular carcinogenesis (HCC). HCC, known as malignant neoplasia of the liver, originates from hepatocytes 2,5 . HCC, a primary malignant tumor, is the most common and deadliest type of liver cancer 1 .…”