INTRODUCTION

The National Center for Toxicological Research (NCTR) and the Carcinogenesis Testing Program of the National Cancer Institute (NCI) have performed numerous toxicity and carcinogenesis studies in mice. Over the last several years these studies have resulted in the development of a large data base of both spontaneous and induced neoplastic and nonneoplastic lesions. Although papers have been published by the staffs of both research programs on the incidence of a variety of spontaneous and induced lesions, no single document describing the morphology of both common and rare spontaneous and induced lesions in mice has appeared in the literature. The purpose of this monograph is to present morphologic descriptions of both nonneoplastic and neoplastic spontaneous and selected induced lesions in mice to interested investigators. The lesions are predominantly from the BALB/c strain and B6C3F1 hybrid mice, but are typical of those seen in all mouse strains and stocks studied.

The BALB/cStCrlfC3H/Nctr inbred mouse subtrain was developed at the National Center for Toxicological Research (NCTR), Jefferson, Arkansas. The strain was Caesarian-derived and foster nursed on gnotobiotic C3H/HeSchGN mothers. Most of the studies at NCTR using the BALB/c mouse have been conducted with specific pathogen free-defined flora (SPF-DF) animals in a barrier-type facility (Littlefield et al., 1979). The current NCTR data base contains microscopic findings on a total of over 60,000 BALB/c mice of which 6,500 are controls. Various BALB/c substrains have been widely used in toxicology and carcinogenesis studies.

The B6C3F1 (C57BL/6N x C3H/HeN)F1 mouse is the F1 hybrid of the C57BL/6N female and the C3H/HeN male. Because of its hardiness, this hybrid has been used for a number of years in bioassays for chemical carcinogens at both NCI and NCTR. The parent strains originated from the Veterinary Resources Branch of the National Institutes of Health, Bethesda, Maryland. The hybrids were usually produced at the National Cancer Institute's Frederick Cancer Research Center or at the Charels River Breeding Laboratories, Wilmington, Massachusetts.

Complete necropsies were performed on most mice, and blocks from approximately 40 organs and tissues were collected for study. All organs were placed directly in Bouin's solution (BALB/c) or 10% neutral buffered foralin (B6C3F1). After18-48 hours in Bouin's solution, the tissues were removed from the fixative, trimmed, and specified groups of tissues were placed in plastic cassettes with perforated bottoms and stainless steel tops. The cassettes were prelabeled with the animal number. The cassettes, each containing multiple tissues from a given animal, were dehydrated in alcohol, cleared in xylene, and infiltrated with paraffin on automatic tissue processors. After processing, the tissues were embedded in paraffin utilizing an embedding center. Sections were routinely cut at 5 μm on rotary microtomes, and sections from 2 paraffin blocks were placed on a single slide when possible. The slides were labeled with the same number as the blocks and routinely stained with hematoxylin and eosin.

The types and incidence of spontaneous lesions in aging mice varies with mouse strain and sex. Several papers and books have documented the incidence of these lesions in aging mice (see General References on Mouse Pathology). Tumor incidence depends, in part, on strain, sex, age (Sheldon and Greenman, 1979; Ward et al., 1979a; Frith et al., 1983a) and necropsy protocol (Frith et al., 1976). Knowledge of naturally occurring lesions provides background information necessary to evaluate the significance of pathologic findings in a rodent safety assessment assay. Since infectious diseases may also affect the background lesions seen in mice, specific pathogen-free (SPF) mice, now commercially available, provide the best choice for use in safety testing.

Classification of nonneoplastic and neoplastic lesions of mice is dependent on available information on the specific lesions; this includes morphologic, histochemical, immunocytochemical, ultrastructural and biologic information. Classification may also vary with one's life experience and personal bias. In this monograph, we use classifications based on information currently available on the biology, morphology, and histochemistry of murine tumors. This atlas illustrates the application of avidin-biotin immunocytochemistry for diagnoses of lesions in mice.

The majority of natural and experimentally induced neoplasms pass through a stage of focal or diffuse hyperplasia in a tissue (Fig. 1), especially for epithelial tissues. Murine neoplasms may arise within areas of hyperplasia or in situ from normal tissue. They often develop first as well differentiated neplasms appearing benign in morphology and behavior. As they grow in size, they often become nodular, well circumscribed or delineated, compress adjacent normal tissue and are thus diagnosed as adenomas. In some organs, such as the liver and lung, focal areas of the benign tumors may suddenly become cytologically malignant. These malignant areas may eventually overgrow the benign tumor. The malignant tumor, a carcinoma if developed from epithelial tissues or a sarcoma if derived from mesenchymal tissues, may invade adjacent tissues and metastasize to regional lymph nodes, lungs and elsewhere. Transplantation of tumors to syngeneic hosts may prove their neoplastic rather than malignant nature, since morphologically and biologically benign tumors can occasionally be transplanted. Many carcinogens induce specific morphologic types of tumors (Ward, 1984a). The causes and pathogenesis of many spontaneous lesions in aging mice are unknown. Viruses are known to cause some sarcomas and leukemias. Genetics also play a role in susceptibility to some tumors. Nonneoplastic lesions often have no apparent cause, and some are related to aging changes in the endocrine system. Infectious diseases often have characteristic lesions which are diagnostic for the etiologic agent and are not seen in SPF mice.

The severity of nonneoplastic age-realted degenerative lesions may be graded as minimal (1 +), mild (2 +), moderate (3 +)or severe or marked (4 +). Occassionally, exposure to the test chemical or agent may increase or decrease the severity of the aging lesions. In mice, age-related lesions are strain dependent. They may occur in any tissue but most commonly are seen in the brain, liver, gonads, reproductive tract and kidney.

Toxic agents or infections may reduce the proportion of mice reaching the tumor age of various neoplasms, thereby reducing the overall incidence of tumors and necessitating comparing lesions in control and experimental animals by comparable age. Toxic agents may impair immunity and increase overall incidence of a lesion so there is no statistical difference in the overall incidence of the lesion between treated mice and controls; however, in treated mice, the lesion appears earlier and is more severe on the average, so that an analysis of the incidence of more severe forms of the lesion at a given age often proves to be significantly greater in treated mice than in controls.