Precancerous lesions in the glandular stomach of Wistar rats induced with dimethylamine hydrochloride and sodium nitrite

Chun Qi Li,Wei Wen Liu

Subject headings   stomach neoplasms; precancerous lesions; dimethylamine hydrochloride; sodium nitrite

Li CQ, Liu WW. Precancerous lesions in the glandular stomach of Wistar rats induced with dimethylamine hydrochloride and sodium nitrite. China Nati J New Gastroenterol, 1996;2(4):209-211

Absrtact

AIM To observe the effects of both dimethylamine hydrochloride and sodiumnitrite on the glandular stomach of Wistar rats.

METHODS  Both dimethylamine hydrochloride and sodium nitrite were administered to male Wistar rats at a concentration of 0.06% in the drinking water ad libitum for 56 weeks, and the rats were killed subsequently. The sequential histological changes induced in the glandular stomach of the rats by both dimethylamine hydrochloride and sodium nitrite were also observed.

RESULTS  It was found that the rate of intestinal metaplasia in the glandular stomach of rats was 88.9% (40/45), 55.0% (22/40) of which was the sulphomucin-positive intestinal metaplasia, and that of mild, moderate and severe dysplasia was 44.5% (20/45), 33.3% (15/45), and 22.2% (10/45) respectively. Cystic dilatation of gastric glands was found in 28.9% (13/45). No gastric cancer occurred. The sequential histological changes were also reported.

CONCLUSION   Both dimethylamine hydrochloride and sodium nitrite can induce the precancerous lesions of glandular stomach in Wistar rats and may be one of the pathogenic factors of gastric cancer.




INTRODUCTION
Since the demonstration by Magee and Barnes^£Û1£Ý in 1956 of the carcinogenicity of N-nitrosodimethylamine (NDMA), the carcinogenic potiential of over 200 N-nitrosocompounds (N-NC) has been established in all 40 animal species so far tested with varying degrees of susceptibility in different organs including the liver, stomach, oesophagus, intestines, bladder, brain, lung and kidney^£Û2,3£Ý. There is now convincing experimental epidemiological evidence that N-NC associated with human cancer affects the oral cavity, urinary bladder and oesophagus and its possible role in the pathogenesis of gastric cancer has also been strongly postulated^£Û4£Ý. Recently NDMA and some other volatile N-nitrosamines and the precursors of N-NC were detected in fish-sauce by Chinese researchers in the high-risk areas of gastric cancer^£Û5,6£Ý. Lu Shiª²Xin ª«et al^£Û7£Ý, further found that the concentration of N-NC was relatively low in foods but that in the fasting gastric juice was very high in the highª²risk population  of oesophageal cancer, postulating that the endogenous synthesis ª«in vivoª« was the main source of Nª²NC. Both dimethylamine and sodium nitrite are the precursors of NDMA synthesis, and whether they can induce the gastric cancer is unknown. In present study, we observed the effects of both dimethylamine hydrochloride and sodium nitrite on the glandular stomach of rats.ª¤ª¤

MATERIALS AND METHODS
One hundred and forty male Wistar rats, weighing 120-140g, were randomized into 3 groups. Groups 1 and 2 consisted of 50 rats respectively. They were given a standard diet and received a mixed drinking water containing respectively 0.06% dimethylamine hydrochloride and sodium nitrite ad libitum. The control group consisted of 40 rats, and was fed with a regular diet and water. All the rats were housed in 10 in a plastic cage in a room with controlled temperature and humidity. Animals were maintained under the guidance set forth in the ¡°Guide for the Care and Use of Laboratory Animals¡± by the National Laboratory Animals Administration Council, China. From the beginning of experiments every 2-4 weeks, 3-4 rats in Group 1 were killed during 56 weeks. The rats in group 2 and control group were killed at the 56th week.ª¥
         The stomaches of the rats were quickly excised and opened along the great curvature. Each stomach was washed several times with normal saline for gross examination, and fixed in 10% neutral formalin and cut into 8 longitudinal strips about 2mm in width. The strips were embedded in paraffin and serially sectioned at 4¦Ìm in thickness. Sections were routinely stained with hematoxylin and eosin, and with alcian blue/periodic acidª²Schiff (AB/PAS) and high iron diamine/alcian blue (HID/AB). The livers of the rats were also treated routinely for the histopathological observations.ª¥The pathological diagnosis and the grading of the lesions were identified according to the criteria of the National Antiª²cancer Association of China in 1990.ª¤

Fig. 1 IM of gastric mucosa was observed. Goblet cells and columnar cells secreting abundant acid mucin mixed with neutral mucin were shown. AB(pH2.5)/PAS¡Á192ª¤

Fig. 2 Gastric dysplasia with inflammatory cells infiltration and glandular atrophy. HE¡Á100ª¤

Fig. 3 Cystic dilatation of gastric glands. AB(pH2.5)/PAS¡Á63

RESULTS

Group 1
During the 2nd to 12th week, the gastric mucosal congestion, edema and erosion were grossly observed under light microscope. There was a lot of inflammatory cell infiltration. Intestinal metaplasia (IM) of the glandular stomach was first found in the glandular neck with characteristic goblet cells in the 18th week and after wards extended to the surface or bottom of the mucosa (Fig. 1). Mild dysplasia of the gastric mucosa occurred at the 24th week, moderate dysplasia at the 32th week and severe dysplasia at the 46th week (Fig. 2). Single and multiple cystic dilatation of the gastric glands was noted at the 22nd week. In the forestomaches of rats, hyperplasia or mild dysplasia of squamous epithelia appeared after the 36th week. At the 48th week, under light microscope the obvious degeneration and necrosis of liver cells and inflammatory cells infiltration in the portal area were observed. At the 54th week, greyish white nodule in the livers could be seen at gross examination and under light microscope, the most typical feature was the hyperplasia of liver cells and fibro-tissues and adenoma-like dilatation of some intrahepatic biliary ducts.ª¤ª¤

Group 2
Total intake of dimethylamine hydrochloride or sodium nitrite was 5.88gªª and the mean daily intake was respectively 15mgªª for a rat. Five rats died in the course of experimentation. It was found that the rates of IM in the glandular stomaches of the rats were 88.9% (40/45), 55.0% (22/40) respectively of which was the sulphomucinª²positive IM, and the rates of mild, moderate and severe dysplasia were 44.5 (20/45), 33.3% (15/45) and 22.2% (10/45) respectively. Cystic dilatation of gastric glands was found in 28.9% (13/45). No gastric cancer occurred. Hyperplasia of the squamous epithelia was 26.7% (12/45) and mild dysplasia 17.8% (8/45) in the forestomaches of the rats. Ten cases (22.2%) of liver cirrhosis without liver cancer were diagnosed.ª¤ª¤

Control group
IM was observed in 3 rats (7.5%) in the pyloric gland areas of stomaches.ª¤ª¤

DISCUSSION
N-NC have been identified in various environmental situations, such as food products, water, drugs, cosmetics, tobacco products, agricultural and industrial products and so on. Exposure in human beings may be through ingestion, inhalation, skin contact and ª«in vivoª« formation, the latter mainly in the gastrointestinal tract which probably represent the main sources of human exposure. The site most commonly regarded as risk from endogenous N-NC synthesis ª«in vivoª« is the stomach. N-NC synthesis ª«in vivoª« can be carried out by chemical or biological synthesis^£Û8£Ý. Dimethylamine hydrochloride or sodium nitrite itself is not carcinogenic but they can be changed into NDMA by chemical synthesis in the acidic environment of stomach.ª¥
        It is widely accepted that epithelial dysplasia of the gastric mucosa is a precancerous lesion and IM, especially sulphomucin-positive IM is closely related to gastric cancer^£Û9-11£Ý. We previously found that the sequential lesions of gastric glandular mucosa induced with both N-methyl-N-nitro-N-nitrosoguanidine (MNNG) and ranitidine were from IM, dysplasia to gastric cancer or from dysplasia to gastric cancer^£Û12£Ý. In this study both dimethylamine hydrochloride and sodium nitrite were administered to Wistar rats in the drinking water ad libitum for 56 weeks, the incidences of IM and moderate and severe dysplasia were respectively 88.9% and 55.5%, suggesting that the two precursors of NDMA synthesis can induce the precancerous lesions of the stomach in the rats. This finding enriches the contents of etiology of the gastric cancer and supplies new experimental evidence for the primary prevention of gastric cancer.ª¥
         We previously reported that 0.01% NDMA liquid was administered to the rats for 16 weeks ad libitum and the incidence of liver cancer was 100% at the 18th week^£Û13£Ý. In the present study, both dimethylamine hydrochloride and sodium nitrite, the precursors of NDMA synthesis, were fed to the rats for 56 weeks, 22.2% of the rats developed liver cirrhosis. Epidemic investigations suggested that NDMA could be one of the pathogenic factors of oesophageal cancer. Oesophageal lesions were not observed in the study but only hyperplasia and mild dysplasia of the squamous epithelium was found in forestomaches of the rats. We supposed that this may be related to the low synthesizing quantity of NDMA in the stomaches of the rats. The lesions of the liver and forestomach may be more severe if higher concentration of both dimethylamine hydrochioride and sodium nitrite was administrated to the rats.ª¤ª¤

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Department of Gastroenterology, Air Force General Hospital,Beijing 100036, China.ª¤
Dr Li Chun Qi, Physicianª²inª²Charge, having 50 papers and two books published.
Dr. Liu Weiª²Wen, Professor of Internal Medicine, Southwest Hospital, Third Military
Medical College, Chongqing 630038, China.ª¤
*This study was supported by ¡°8¡¤5¡± Medical Science Foundation of Chinese PLA.ª¤
Correspondence to Dr. Li Chun Qi,ª¤Department of Gastroenterology, Air Force
General Hospital,Beijing 100036, China.
Tel:+86¡¤010¡¤68410099 EXT 8301.
Received 29th July 1996.