Spontaneous primary pleural mesotheliomas in Fischer 344 (F344) or other rat strains have rarely been reported. The objectives of this retrospective study were to develop historical incidence data and better characterize the light-microscopic morphology of these naturally occurring neoplasms in a large cohort of rats of several strains. A retrospective review was performed of National Toxicology Program (NTP) studies in rats conducted between 1980 and 2019 and comprising a total of 104,029 rats (51,326 males, 52,703 females), predominantly (90%) of the F344 strain. Of the 94,062 F344 rats surveyed, there were 30 cases of primary pleural mesotheliomas (22 males, 8 females). Of the 2998 Wistar Han rats surveyed, primary pleural mesotheliomas were present in 2 male rats. No primary pleural mesotheliomas were noted in male and female rats of other strains (6669 Sprague Dawley; 300 Osborne-Mendel). All primary pleural mesotheliomas in control and treated F344 and Wistar Han rats were considered spontaneous and unrelated to treatment. Based on light-microscopic evaluation of paraffin-embedded hematoxylin and eosin stained sections, only epithelioid and biphasic histologic subtypes were observed: 18 and 12 in F344 rats, respectively, and one each in Wistar Han rats. No sarcomatoid subtype cases were noted in any strain of rat.

Receptor-interacting protein 1 (RIP1) has emerged as a key protein for transducing signals induced by several immune receptors, including tumour necrosis factor receptor 1 (TNFR 1).When these receptors are engaged, RIP1 kinase activity can drive cell death (apoptosis and necroptosis) and proinflammatory cytokine production which has been implicated in the pathogenesis of multiple inflammatory diseases 1,2 .To date, although RIP1 kinase has been implicated in driving inflammation in multiple tissues, the data points to a key role of RIP1 in the intestines.Inflammatory bowel disease (IBD) is a multifactorial disorder characterised by chronic intestinal inflammation and comprises two distinct diseases: ulcerative colitis (UC) and Crohn's disease (CD).Human mutations in the RIP1 pathway, as shown in patients carrying a deficiency in NF-κB essential modulator (NEMO) or the linear ubiquitin chain assembly complex (LUBAC), result in TNF-dependent intestinal inflammation 3,4 .Similar to the human data, NEMO IEC-KO mice display Paneth cell apoptosis and microbiota-driven chronic inflammation in the colon 5 .Additionally, it has been shown that SHARPIN-deficient cpdm mice, that have a defect in LUBAC, develop TNFdependent multiorgan inflammation 6 .Moreover, RIP1 might be a key driver of Paneth cell death as suggested by increased necroptosis in human Paneth cells in the terminal ileum of patients with CD 7 .It has also recently been shown that necroptosis is strongly associated with intestinal inflammation in children with IBD and that it contributes to

Abstract Background Inflammatory bowel disease (IBD) is a multifactorial disorder characterised by chronic and relapsing intestinal inflammation. Receptor-interacting protein 1 (RIP1) kinase activity is emerging as a driver of pro-inflammatory cytokine production and cell death and has been implicated in multiple inflammatory diseases including IBD. To this end, recent work has shown that RIP1 kinase inhibition reduces the spontaneous production of cytokines from human ulcerative colitis (UC) and Crohn’s disease explants. Methods The aim of this study was to investigate the anti-inflammatory effect of a highly selective inhibitor of RIP1 kinase activity (GSK547A) in the mouse T-cell transfer model of colitis; a model that shares many features with human IBD. All animal studies were ethically reviewed and carried out in accordance with Animals (Scientific Procedures) Act 1986 and the GSK Policy on the Care, Welfare and Treatment of Animals. Chronic colitis was achieved by transferring CD4+CD45RBhigh T cells into immunodeficient female SCID mice (6/8 weeks old). After confirming the development of pathology using endoscopy and MRI, animals were treated therapeutically with the RIP1 inhibitor GSK547A (50 mg/kg twice a day PO) or vehicle (0.5% hydroxypropyl methylcellulose in water). The severity of colitis was measured 5 weeks after cell transfer both macro- and microscopically. Mucosal damage by endoscopy was also assessed. RT-PCR and MSD analysis were performed to detect colon cytokine/chemokine and calprotectin levels. SAA in the plasma was measured using ELISA. Results We found that treatment with GSK547 significantly ameliorated experimental T-cell-dependent colitis in mice. GSK547A-treated mice displayed decreased weight loss, colon density (ratio weight/length), macroscopic disease activity index, colon thickness compared with vehicle-treated animals. Mucosal damage, assessed by endoscopy and histopathology, was also reduced following treatment with RIP1 kinase inhibitor. In addition, GSK547A reduced the expression of cytokines in the colon (TNF-α, IL-17A, INF-γ, IL-6 and MCP-1), both at a protein and RNA level. Relevant translational biomarkers such as SAA in plasma and RNA calprotectin in the colon were also decreased in the GSK547A treated group when compared with vehicle. Conclusion Our results suggest that RIP1K inhibition is a strong protective factor with anti-inflammatory potential in the progression of chronic colitis, when applied to the translationally relevant T-cell transfer model of colitis. These findings suggest the potential application in the management of inflammatory bowel disease and support the ongoing clinical program evaluating the effect of RIP1 kinase inhibition in UC patients.

The INHAND Project (International Harmonization of Nomenclature and Diagnostic Criteria for Lesions in Rats and Mice) is a joint initiative of the Societies of Toxicologic Pathology from Europe (ESTP), Great Britain (BSTP), Japan (JSTP), and North America (STP) to develop an internationally accepted nomenclature for proliferative and nonproliferative changes in rats and mice. The purpose of this publication is to provide a standardized nomenclature for classifying changes observed in the hematolymphoid organs, including the bone marrow, thymus, spleen, lymph nodes, mucosa-associated lymphoid tissues, and other lymphoid tissues (serosa-associated lymphoid clusters and tertiary lymphoid structures) with color photomicrographs illustrating examples of the lesions. Sources of material included histopathology databases from government, academia, and industrial laboratories throughout the world. Content includes spontaneous lesions as well as lesions induced by exposure to test materials. The nomenclature for these organs is divided into 3 terminologies: descriptive, conventional, and enhanced. Three terms are listed for each diagnosis. The rationale for this approach and guidance for its application to toxicologic pathology are described in detail below.

To test the diagnostic approach described in part 1 of this article, 2 exercises were completed by pathologists from multiple companies/agencies. Pathologist's examination of whole slide image (WSI) heart sections from rats using personal diagnostic approaches (exercise #1) corroborated conclusions from study #1. Using the diagnostic approach described in part 1, these pathologists examined the same WSI heart sections (exercise #2) to determine whether that approach increased consistency of diagnosis of rodent progressive cardiomyopathy (PCM) lesions. In exercise #2, there was improved consistency of categorization of small borderline morphologies and mild lesions, but a decrement in consistency of categorizing minimal lesions. Exercises 1 and 2 suggest the described diagnostic approach is representative of that in use by the majority of toxicologic pathologists across companies/agencies and that application by all may improve diagnostic consistency of PCM/like lesions. Additionally, a criterion of approximately 5% heart section involvement is suggested for separating mild from moderate or greater severity. While evidence is not absolute, until further investigation shows otherwise, microscopic changes resembling PCM, but located in the epicardial and subepicardial region of the right ventricle, may be considered as part of the spectrum of PCM.

Session 3 of the Toxicologic Pathology and the Immune System Symposium, presented as part of the 30th Annual Symposium of the Society of Toxicologic Pathology in 2011, focused on the biological advances in control of selected cellular and secretory components of acquired immunity. Acquired immunity goes beyond innate immunity to provide controlled recognition and memory for specific antigenic challenges. Predominately involving activation of T and B lymphocytes, the resulting cellular- and secretory-mediated activity provides immediate and long-term host defenses to antigenic challenge. This session highlighted the biological advances in function and dysfunction of acquired immunity through regulatory T cells, the pathophysiology of effector cells and secretory molecules in immunosuppression, allergic inflammatory disease, and dysregulation that leads to loss of tolerance and autoimmune disease. A brief overview of major concepts in acquired immunity and summaries of the above themes are covered herein, and discussions of these themes are covered in greater detail in this issue of Toxicologic Pathology.

Unexpected deaths occurred in studies involving a nude mouse model of mammary cancer that required subcutaneous implantation of 0.5 mg twenty-one-day release estrogen pellets for growth of the estrogen-dependent mammary tumor xenograft BT474c. Early deaths occurred in female nude mice and were associated with urinary retention, frequently with cystitis. Drug treatment had no effect on the incidence or severity of cystitis. Histological findings did not alter significantly over various time points following pellet implantation. Changes were not seen in males or in females receiving lower doses of estradiol even when the duration of administration was prolonged, suggesting that a threshold level was required for the onset of urinary retention. Because of the influence of estrogen on micturition, immunohistochemistry for estrogen receptor alpha (ERalpha) in the urinary bladder was carried out, which did not demonstrate any differences between females implanted with 0.5 mg twenty-one-day release estrogen pellets and nonimplanted females. Although previous publications have concentrated on possible mechanisms of action, this paper describes the histopathological changes seen in the urinary bladder of female nude mice resulting from exposure to high levels of estradiol.

Immunotoxicology has developed into an integral regulatory requirement of the toxicological assessment of xenobiotics. Enhanced pathological assessment of tissues can provide genuine insight into perturbations of lymphoid cell populations. To facilitate this we have endeavoured to develop a panel of immunohistochemical techniques to demonstrate T-cells and T-cell subsets in formalin-fixed paraffin-embedded rat lymphoid tissues. We were successful in developing methods for CD3 and CD8 but failed to arrive at a satisfactory technique for the direct demonstration of CD4 in these tissues. Taking the assumption that the majority of mature T-cells are either CD4+ or CD8+ we have combined our methods for CD3 and CD8 in a novel dual-labelling IHC method to simultaneously demonstrate CD3, CD8 and, by implication, CD4.

In 2005, the International conference on harmonization (ICH) recommended that all new human pharmaceuticals be tested for unintended immunomodulatory potential via a tiered approach. Included in this approach is a semiquantitative description of changes in the separate compartments of lymphoid tissue (also called enhanced histopathology). Chlorambucil was administered to Hanover Wistar rats at regular time points, followed by a treatment-free (recovery) period. Groups of treated and control animals were sacrificed regularly during both the treatment and recovery periods. Selected tissues were removed, weighed fresh and fixed in formalin, processed, and stained with hematoxylin and eosin. Blood samples and bone marrow smears were also obtained. With the use of enhanced histopathology, a description of the changes in lymphoid tissues and bone marrow was used as a means of assessing the susceptibility, and recovery, of the different lymphoid cell populations over time. A correlation with organ weights, flow cytometry data, and bone marrow cytology was achieved. The administration of chlorambucil in the Hanover Wistar rat provided a useful tool to examine the rate and sequence of changes in the lymphoid organs and bone marrow during treatment with, and the recovery from the effects of, a potent immunosuppressive agent.

Immunotoxicology has developed into an integral regulatory requirement of the toxicological assessment of xenobiotics. Histopathological assessment of lymphoid tissues can provide genuine insight into perturbations of lymphoid cell populations. To facilitate retrospective examination of lymphoid organs should concerns over immunotoxicity be raised, we have endeavored to develop a panel of immunohistochemical techniques to demonstrate T-cells and T-cell subsets in formalin-fixed, paraffin-embedded rat lymphoid tissues. We were successful in developing methods for CD3 and CD8 but failed to arrive at a satisfactory technique for the direct demonstration of CD4 in these tissues. Taking the assumption that the majority of mature T-cells are either CD4+ or CD8+, we have combined our methods for CD3 and CD8 in a novel dual-labeling IHC method to simultaneously demonstrate CD3, CD8, and, by implication, CD4 in rat spleen, thymus, lymph node, and Peyer's patch.

The thymus is a primary lymphoid organ that manifests dynamic physiological changes as animals age in addition to being exquisitely sensitive to stress and toxic insult. It is typically the first lymphoid tissue to respond to immunotoxic xenobiotics, with the first change being loss of cortical lymphocytes by apoptosis. This is followed by removal of the apoptotic cellular debris and, in the absence of recovery, may lead to loss of the cortico-medullary demarcation and organ atrophy. Nonneoplastic proliferative changes include focal lymphoid hyperplasia and proliferation of medullary epithelial cells, often with formation of ribbons, cords, or tubules. Thymomas are relatively rare tumors that exhibit a wide spectrum of morphologic types but do not metastasize. Thymic lymphomas are common in some mouse strains and can become leukemic with hematogenous spread throughout the body.

The National Toxicology Program (NTP) developed the chronic 2-year bioassay as a mechanism for predicting the carcinogenic potential of chemicals in humans. The cost and duration of these studies has limited their use to small numbers of selected chemicals. Many different short-term methods aimed at increasing predictive accuracy and the number of chemicals evaluated have been developed in attempts to successfully correlate their results with evidence of carcinogenicity (or lack of carcinogenicity). Using NTP studies, the effectiveness of correlating prechronic liver lesions with liver cancer encompassing multiple studies using mice (83 compounds) and rats (87 compounds) was assessed. These lesions include hepatocellular necrosis, hepatocellular hypertrophy, hepatocellular cytomegaly, bile duct hyperplasia, and hepatocellular degeneration, along with increased liver weight. Our results indicate that pooling 3 of these prechronic data points (hepatocellular necrosis, hepatocellular hypertrophy, and hepatocellular cytomegaly) can be very predictive of carcinogenicity in the 2-year study ( p < 0 .05). The inclusion of increased liver weight as an endpoint in the pool of data points increases the number of rodent liver carcinogens that are successfully predicted ( p < 0 .05), but also results in the prediction of increased numbers of noncarcinogenic chemicals as carcinogens. The use of multiple prechronic study endpoints provides supplementary information that enhances the predictivity of identifying chemicals with carcinogenic potential.