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Background. Nonadherence to immunosuppression in liver transplant recipients (LTRs) leads to deterioration in health outcomes. Once-dailyextended-release tacrolimus (TAC-ER) may improve adherence when compared to twice-dailyimmediate-release tacrolimus (TAC-IR). Methods. We conducted a randomized controlled study to evaluate medication adherence, clinical efficacy, and safety of TAC-ER in stable LTR. All patients >18 years who underwent liver transplantation before 6 months were eligible. Patients were randomized 1 : 1 to continued TAC-IR or conversion to TAC-ER. The primary outcome was change in medication adherence from baseline to 9 months, assessed using BAASIS. Secondary outcomes were tacrolimus trough levels, safety, and quality of life. Results. Thirty-one patients were consented and randomized to either of the two groups: conversion to TAC-ER (n = 15) or continued TAC-IR (n = 16). Six patients in the TAC-ER group withdrew after randomization due to apprehension about switching medication (n = 2), unwillingness to travel (n = 2), and increased liver tests after conversion (n = 2, both were acute rejections despite therapeutic tacrolimus levels and were considered unrelated to TAC-ER). We compared the results of nine patients in the TAC-ER group that completed the study with those of sixteen in the TAC-IR group. At baseline, there was no difference in tacrolimus trough levels between groups. Improved adherence was observed in the TAC-ER group as 100% of patients reported at least one period of full adherence during the study period (100% vs. 62.6%, p=0.035). Tacrolimus trough levels and liver tests were comparable between groups throughout the study. There were no differences in eGFR, HbA1c, or QoL between the groups. Conclusion. TAC-ER improved medication adherence while maintaining comparable trough levels, liver function, and QoL as TAC-IR in LTR.

Objectives: Asians represent the fastest growing ethnic group in the United States. Despite significant diversity within the group, many transplant studies treat Asians as a homogeneous entity. We compared patient and graft survival among major Asian eth- nicities to determine whether any subgroup has superior out- comes. Methods: We conducted a retrospective analysis of kidney trans- plants on Asian and White patients between 2001 and 2012. Co- variates included gender, age, comorbidities, and donor category. Primary outcomes included one-year patient and graft survival. Secondary outcomes included delayed graft function (DGF) and rejection as cause of graft loss and death. Results: Ninety-one Asian patients were identified. Due to the large proportion of Chinese patients (n=37), we grouped other Asians into one entity (n=54) for statistical comparison among Chinese, other Asians, and Whites (n=346). Chinese subjects had significantly lower body mass index (BMI) (p=0.001) and had the lowest proportion of living donors (p>0.001). Patient survival was highest in our Chinese cohort (p>0.001) Discussion: Our study confirms outcome differences among Asian subgroups in kidney transplantation. Chinese demonstrate better patient survival at one year than Whites and non-Chinese Asians despite fewer live donors. Lower BMI scores may partly explain this. Larger, long-term studies are needed to elucidate outcome disparities among Asian subgroups

Central venous catheters play a pivotal role in the perioperative support of critically ill patients. They are used for administration of fluids, vasopressors, blood products, and various medications; however, their use may be associated with serious complications, such as catheter fracture and embolization. While most data on catheter fracture embolization consist of isolated case reports, only a few studies have examined patients with central venous catheter embolism. We report a traumatic inadvertent transection of central venous catheter that migrated through a donor transplanted liver and was found to be lodged in the recipient’s right ventricle. The catheter was retrieved under fluoroscopy using a trilobed snare device.

Drug-induced kidney injury (DIKI) is of significant concern, both during drug development and in clinical practice. We report a patient-centric approach for clinical implementation of the FDA-qualified kidney safety biomarker panel, highlighting Phase 1 and 2 trials for candidate therapeutics in Pfizer's portfolio (PFE-1 and PFE-2, respectively) that induced kidney tubular injury in rat toxicity studies. Clusterin (CLU), cystatin-C (CysC), kidney injury molecule-1 (KIM-1), N-acetyl-beta-d-glucosaminidase (NAG), neutrophil gelatinase-associated lipocalin (NGAL), and osteopontin (OPN) were measured in urine samples from (i) Phase 1 healthy volunteers (HVs; n = 12) dosed with PFE-1, (ii) Phase 2 rheumatoid arthritis (RA) patients (n = 266) dosed with PFE-2, (iii) lupus patients on standard-of-care therapies (n = 121), and (iv) healthy volunteers (n = 60). The FDA-defined composite measure (CM), calculated as the geometric mean response across the 6 biomarkers, was increased ∼30% in HVs administered 100 mg PFE-1 relative to placebo, providing evidence of DIKI. In contrast, the CM for RA patients dosed with PFE-2 was comparable to placebo controls, helping to de-risk the concern for DIKI at clinically relevant doses. Comparing individual biomarker concentrations across disease states revealed that CLU, KIM-1, NAG, NGAL, and OPN are elevated in the urine of RA and lupus patients (those without severe active proliferative lupus nephritis) relative to HVs. Overall, these case studies demonstrate the value of using the FDA-qualified kidney biomarker panel to guide risk assessment, dose selection, and clinical decision making for novel therapeutics, both in HVs and patient populations., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Abrocitinib is a Janus kinase (JAK) 1 selective inhibitor approved for the treatment of atopic dermatitis. Female reproductive tissues were unaffected in general toxicity studies, but an initial female rat fertility study resulted in adverse effects at all doses evaluated. A second rat fertility study was conducted to evaluate lower doses and potential for recovery., Methods: This second study had 4 groups of 20 females each administered abrocitinib (0, 3, 10, or 70 mg/kg/day) 2 weeks prior to cohabitation through gestation day (GD) 7. In addition, 2 groups of 20 rats (0 or 70 mg/kg/day) were dosed for 3 weeks followed by a 4-week recovery period before mating. All mated females were evaluated on GD 14., Results: No effects were observed at ≤10 mg/kg/day. At 70 mg/kg/day (29x human exposure), decreased pregnancy rate, implantation sites, and viable embryos were observed. All these effects reversed 4 weeks after the last dose., Conclusions: Based on these data and literature on the potential role of JAK signaling in implantation, we hypothesize that these effects may be related to JAK1 inhibition and, generally, that peri-implantation effects such as these, in the absence of cycling or microscopic changes in nonpregnant female reproductive tissues, are anticipated to be reversible., (© 2024 Wiley Periodicals LLC.)

A high incidence of thymic lymphoma has been noted in mice deficient of retinoid-related orphan receptor γ2 (RORγ2), which is required for differentiation of naïve CD4 + T cells into T H 17 cells. Using a RORγ homozygous knockout (KO) mouse model of thymic lymphoma, we characterized this tumor progression and investigated the utility of 5-hydroxymethylcytosine (5hmC) signatures as a non-invasive circulating biomarker for early prediction of malignancy. No evidence for malignancy was noted in the wild-type mice, while primary thymic lymphoma with multi-organ metastasis was observed microscopically in 97% of the homozygous RORγ KO mice. The severity of thymic lymphoma was not age-dependent in the KO mice of 2 to 4 months old. Differential enrichment of 5hmC in thymic DNA and plasma cell-free DNA (cfDNA) was compared across different stages of tumor progression. Random forest modeling of plasma cfDNA achieved good predictivity (AUC = 0.74) in distinguishing early non-metastatic thymic lymphoma compared to cancer-free controls, while perfect predictivity was achieved with advanced multi-organ metastatic disease (AUC = 1.00). Lymphoid-specific genes involved in thymocyte selection during T cell development (Themis, Tox) were differentially enriched in both plasma and thymic tissue. This could help in differentiating thymic lymphoma from other tumors commonly detected in rodent carcinogenicity studies used in pharmaceutical drug development to inform human malignancy risk. Overall, these results provide a proof-of-concept for using circulating cfDNA profiles in rodent carcinogenicity studies for early risk assessment of novel pharmaceutical targets., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Kelly A. Fader, Mark M. Gosink, Shuhua Xia, Thomas A. Lanz, Charles Halsey, Vishal S. Vaidya, Zaher A. Radi report a relationship with Pfizer Inc. that includes: employment. K.A.F., M.M.G., S. X., T.A.L., C.H., V.S.V., and Z.A.R. are/were employees of Pfizer Inc. at the time of their contribution to the manuscript., (Copyright © 2023. Published by Elsevier Inc.)

Inflammatory bowel disease (IBD), comprising Crohn's disease and ulcerative colitis, is a lifelong disease that manifests with chronic intestinal inflammation, sequential fibrosis, and an increased risk of colitis-associated colon cancer (CAC). The combined effects of genetic, immunological, environmental, and microbial factors render it difficult to determine the specific mechanism underlying the induction and perpetuation of IBD. Various animal models of IBD have contributed enormously to the understanding of IBD pathogenesis in terms of genomics, transcriptomics, proteomics, microbiome, and drug development of novel therapeutics. Although comprehensive research on IBD has been enabled by advanced technologies, such as genetically engineered models, there is a great need to develop relevant in vivo models of colitis and fibrosis. Here, we review 4 categories of animal models of acute and chronic intestinal inflammation, fibrosis, and CAC: chemically induced, genetically engineered, T cell transfer, and spontaneous gene mutation models.

Thymoma, a tumor of thymic lymphocytes or thymic epithelial cells (TECs), is a common spontaneous tumor in Wistar Han rats, especially in females with up to 18% incidence in controls. In addition to sex, there are rat strain differences in background incidence of thymomas such as Sprague Dawley versus Wistar Han rats. Human thymomas are very rare and without clear differences in incidence between males and females. Immunomodulatory and anti-inflammatory pharmaceutical drug classes, including Janus kinase inhibitors, increase the incidence of benign thymoma in two-year rat carcinogenicity studies. Potential non-genotoxic mechanisms that might contribute to the pathogenesis of thymoma development in one sex (female) Wistar Han rats include: (1) hormonal differences, (2) high proliferation rate of TECs, (3) delayed physiologic thymic involution, and/or (4) significant level of immunosuppression at high doses of a pharmaceutical drug. Factors to consider in the human cancer risk assessment of pharmaceutical-induced thymoma are: the genotoxicity of the test article, sex and strain of rats, exposure safety margins, and pathophysiologic differences and similarities of thymoma between rats and humans. Totality of weight of evidence approach and available data suggest thymomas observed in carcinogenicity studies of pharmaceutical drugs are not relevant for human risk at clinically relevant therapeutic doses., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests. Zaher Radi and Nasir Khan reports a relationship with Pfizer Global Research and Development that includes: employment and equity or stocks. The authors are employed by Pfizer Inc. and declare that there are no conflict of interest., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Influx and efflux kidney tubular transporters are major determinants of the disposition of xenobiotics, including pharmaceutical drugs. On the basolateral membrane of proximal tubular cells, there are influx transporters, such as organic cation transporters. On the apical membrane of proximal tubular cells, there are efflux transporters, such as multidrug and toxin extrusion proteins. The secretion process across the apical membrane into the lumen occurs via efflux transporters which plays an important role in serum creatinine (sCr) elimination in urine. The interference of a pharmaceutical drug with transporters can lead to changes in sCr with no alterations in biomarkers or light microscopic evidence indicative of renal injury. Identification of transporters that influence drug disposition, toxicity, and overall nonclinical safety assessment is important in drug discovery and development programs. This mini review describes some key aspects of kidney tubular transporters and drug-induced renal toxicities in safety risk assessment and drug development.

Immune tolerance is defined by an active state of immune system unresponsiveness to foreign and self-antigens. Loss of immune tolerance to self-antigens and the resulting overexpression of autoantibodies can lead to tissue injury and development of various autoimmune diseases. In drug development, the goal of newly emerging immune tolerance therapies is to treat autoimmune disorders by restoring the immunoregulatory capacity of the immune system. Development of immune tolerance targets is initiated with the establishment of pharmacological efficacy in relevant disease animal models, followed by their stepwise translation to humans. This review discusses the major challenges to developing tolerance inducing pharmaceutical drugs, including the selection of appropriate disease models to establish efficacy, adequate, and acceptable in vitro and in vivo safety assessments, relevant biomarkers of human safety and efficacy, and finally, some regulatory guidelines to successfully develop immune tolerance therapeutics. [Box: see text].

Anatomically, the kidneys are paired, bean-shaped (in most mammals), excretory organs that lie in the retroperitoneum. High blood flow to the kidneys, together with high oxygen consumption, makes them more vulnerable to exposure, via the circulation, and subsequent injury related to high concentrations of xenobiotics and chemicals. In preclinical drug development and safety assessment of new investigational drugs, changes in kidney structure and/or function following drug administration in experimental laboratory animals need to be put in context with interspecies differences in kidney functional anatomy, physiology, spontaneous pathologies, and toxicopathological responses to injury. In addition, translation to human relevance to avoid premature drug termination from development is vital. Thus, detection and characterization of kidney toxicity in preclinical species and human relevance will depend on the preclinical safety testing strategy and collective weight-of-evidence approach including new investigational drug mechanism of action (MOA), preclinical and clinical interspecies differences, and MOA relevance to humans. This review describes kidney macroscopic and microscopic functional anatomy, physiology, pathophysiology, toxicology, and drug-induced kidney toxicities in safety risk assessment and drug development.

Non-steroidal anti-inflammatory drugs (NSAIDs) are among the most widely used therapeutic class in clinical medicine. These are sub-divided based on their selectivity for inhibition of cyclooxygenase (COX) isoforms (COX-1 and COX-2) into: (1) non-selective (ns-NSAIDs), and (2) selective NSAIDs (s-NSAIDs) with preferential inhibition of COX-2 isozyme. The safety and pathophysiology of NSAIDs on the renal and cardiovascular systems have continued to evolve over the years following short- and long-term treatment in both preclinical models and humans. This review summarizes major learnings on cardiac and renal complications associated with pharmaceutical inhibition of COX-1 and COX-2 with focus on preclinical to clinical translatability of cardio-renal data.

Pathophysiologically, the classification of acute kidney injury (AKI) can be divided into three categories: (1) prerenal, (2) intrinsic, and (3) postrenal. Emerging evidence supports the involvement of renal tubular epithelial cells and the innate and adaptive arms of the immune system in the pathogenesis of intrinsic AKI. Pro-inflammatory damage-associated molecular patterns, pathogen-associated molecular patterns, hypoxia inducible factors, toll-like receptors, complement system, oxidative stress, adhesion molecules, cell death, resident renal dendritic cells, neutrophils, T and B lymphocytes, macrophages, natural killer T cells, cytokines, and secreted chemokines contribute to the immunopathogenesis of AKI. However, other immune cells and pathways such as M2 macrophages, regulatory T cells, progranulin, and autophagy exhibit anti-inflammatory properties and facilitate kidney tissue repair after AKI. Thus, therapies for AKI include agents such as anti-inflammatory (e.g., recombinant alkaline phosphatase), antioxidants (iron chelators), and apoptosis inhibitors. In preclinical toxicity studies, drug-induced kidney injury can be seen after exposure to a nephrotoxicant test article due to immune mechanisms and dysregulation of innate, and/or adaptive cellular immunity. The focus of this review will be on intrinsic AKI, as it relates to the immune and renal systems cross talks focusing on the cellular and pathophysiologic mechanisms of AKI.

Cellular development and homeostasis are regulated via programmed cell death (PCD; apoptosis), which is a genetically regulated cellular process. Accidental cell death (ACD; necrosis) can be triggered by chemical, physical, or mechanical stress. Necrosis is the presence of dead tissues or cells in a living organism regardless of the initiating process and can be observed in infectious and non-infectious diseases and toxicities. This article describes tissue-based immunohistotechnical protocols used for assessing PCD and necrosis in formalin-fixed tissues obtained from preclinical species used in investigative and toxicologic pathology. Two commonly employed protocols for the identification of PCD and necrosis are described in this article: immunohistochemistry (IHC) for cleaved caspase 3, and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL). TUNEL has been used to detect DNA fragmentation by labeling the terminal ends of nucleic acids in necrotic and apoptotic cells. © 2018 by John Wiley & Sons, Inc., (Copyright © 2018 John Wiley & Sons, Inc.)

Glomerulopathy and body weight gain were noted after chronic oral administration of a novel nonsteroidal dissociated agonist of the glucocorticoid receptor compound, fosdagrocorat, to beagle dogs fed an ad libitum diet. To further investigate the role of diet and treatment with either fosdagrocorat or the glucocorticoid comparator, prednisone, on renal safety, a 13-week investigative study was conducted in beagle dogs. Renal histopathology, clinical chemistry, urinalysis, glomerular filtration rate (GFR), body weight, heart rate, blood pressure (BP), and hematology were investigated in restricted- and ad libitum-fed dogs administered prednisone (2.2 mg/kg/d), fosdagrocorat (5 mg/kg/d), or vehicle for 13 weeks. Glomerulopathy was primarily observed in fosdagrocorat- and prednisone-treated ad libitum but not in feed-restricted or ad libitum vehicle-treated dogs. Kidneys in dogs from the prednisone-treated ad libitum had the greatest incidence and severity of tubular degenerative changes. Increased urine volume and decreased urine-specific gravity were present in prednisone- and fosdagrocorat-treated dogs, regardless of diet. These changes were not associated with consistent changes in GFR. Fosdagrocorat or prednisone treatment ad libitum dogs had the greatest increase in body weight gain. Sporadic changes in systolic and diastolic BP were noted in fosdagrocorat- and prednisone-treated groups. Significant reductions in serum cortisol and absolute eosinophils were noted in both ad libitum- and restriction-fed prednisone- and fosdagrocorat-treated dogs. In conclusion, prednisone-treated dogs fed ad libitum had greater glucocorticoid-induced renal effects than those dosed with fosdagrocorat.

Tofacitinib, an oral Janus kinase (JAK) inhibitor for treatment of rheumatoid arthritis, targets JAK1, JAK3, and to a lesser extent JAK2 and TYK2. JAK1/3 inhibition impairs gamma common chain cytokine receptor signaling, important in lymphocyte development, homeostasis and function. Adult and juvenile cynomolgus monkey and rat studies were conducted and the impact of tofacitinib on immune parameters (lymphoid tissues and lymphocyte subsets) and function (T-dependent antibody response (TDAR), mitogen-induced T cell proliferation) assessed. Tofacitinib administration decreased circulating T cells and NK cells in juvenile and adult animals of both species. B cell decreases were observed only in rats. These changes and decreased lymphoid tissue cellularity are consistent with the expected pharmacology of tofacitinib. No differences were observed between juvenile and adult animals, either in terms of doses at which effects were observed or differential effects on immune endpoints. Lymphomas were observed in three adult monkeys. Tofacitinib impaired the primary TDAR in juvenile monkeys, although a recall response was generated. Complete or partial reversal of the effects on the immune system was observed., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Tofacitinib is an oral JAK inhibitor for the treatment of rheumatoid arthritis. In the 2-year carcinogenicity study with tofacitinib, increased incidence of hibernoma (a neoplasm of brown adipose tissue [BAT]) was noted in female rats at ≥30 mg/kg/day (≥41x human exposure multiples). Thus, signaling pathways within BAT were investigated by measuring BAT: weight, cell proliferation biomarkers, content of basal and prolactin-induced phosphorylated Signal Transducer and Activator of Transcription (STAT), and uncoupling protein 1 (UCP-1). The relationship between cardiovascular hemodynamics and plasma norepinephrine (NE) levels was also investigated. Tofacitinib administered to female rats at doses of 10, 30, or 75 mg/kg/day for 14 days increased BAT weight at 75 mg/kg/day and cell proliferation at ≥30 mg/kg/day. JAK inhibition, observed as lower pSTAT3 and pSTAT5 in BAT, was noted at ≥10 mg/kg/day, while lower activity of BAT was observed as lower UCP-1 protein at ≥30 mg/kg/day. In cultured brown adipocytes, prolactin-induced increase in pSTAT5 and pSTAT3 were inhibited by tofacitinib in a concentration-dependent manner. Tofacitinib lowered blood pressure, increased heart rate, and resulted in dose-dependent increases in circulating NE. Thus, JAK/STAT inhibition in BAT and sympathetic stimulation are two factors which might contribute to the genesis of hibernomas by tofacitinib in rats., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Tofacitinib is an oral Janus kinase (JAK) inhibitor for the treatment of rheumatoid arthritis. Tofacitinib preferentially inhibits receptor signaling through JAK3 and JAK1, relative to JAK2. In the 2-year rat carcinogenicity study, there were tofacitinib, dose-related increases in the incidences of testicular Leydig cell hyperplasia and benign adenomas in male rats, and decreased incidences of mammary tumors and duct dilatation/galactocele in female rats. Such findings in rats are typical of agents, such as dopamine agonists, which decrease prolactin (PRL) activity. Since prolactin signals through the JAK2 pathway, we hypothesized that these findings were off-target effects due to inhibition of PRL signaling via JAK2. The studies reported here were designed to investigate the interruption of PRL signaling pathways in Leydig cells. In isolated primary rat Leydig cells, PRL increased phosphorylated Signal Transducer and Activator of Transcription-5 protein, and mRNA levels for luteinizing hormone receptor. Tofacitinib, at concentrations observed in the rat carcinogenicity study, dose-dependently inhibited these effects. These observations illustrate a novel mechanism, the inhibition of prolactin signaling by which modulation of JAK activity can modulate PRL signaling pathways to induce Leydig cell tumors in rats. Since human Leydig cells lack this PRL dependence for normal function, these rodent tumors do not indicate a health risk to human patients., (© The Author 2016. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Optimal histochemical staining is critical to ensure excellent quality stained sections to enable light microscopic and histomorphometric image analysis.  Verhoeff-van Gieson is the most widely used histochemical stain for the visualization of vascular elastic fibers. However, it is notoriously difficult to differentiate fine elastic fibers of small vasculature to enable histomorphometric image analysis of vasculature size characterization especially in organs such as the lung. A tissue fixation regime of 10% neutral buffered formalin with subsequent fixation in 70% ethanol further compounds the problem of small vessel staining and identification.  Therefore, a modified Verhoeff's elastin stain was developed as a reliable method to optimally highlight the internal and external elastic lamina of small arteries (50-100 µm) and intra-acinar vessels (10-50 µm) in 3 µm thick lung tissue sections from models of pulmonary arterial hypertension.  This modified Verhoeff's elastin stain demonstrated optimal staining of fine elastic fibers of pulmonary blood vessels. As a result, high-quality histomorphometric image analysis evaluation of vessel wall thickness in small arteries and intra-acinar vessels was successfully accomplished.  In conclusion, modification of the standard Verhoeff-van Gieson histochemical stain is needed to visualize small caliber vessels' elastic fibers especially in tissues not fixed in 10% neutral buffered formalin only.

An incidental, asymptomatic, focal inflammatory lesion was detected in brain cerebrum of an approximately 6-year-old, female cynomolgus macaque from a chronic toxicology study. No gross lesions were noted at necropsy. Microscopically, the lesion contained a cross-section of larvae approximately 70-80 μm in diameter, a centrally located intestine flanked on either side by large triangular excretory columns, and prominent single lateral cuticular alae. Mixed inflammatory cells of eosinophils, macrophages, and lymphocytes admixed with abundant connective tissue stroma and necrosis surrounded the larvae. Histochemical stains for trichrome revealed significant amount of fibrous connective tissue. The morphology of the larvae was compatible with Baylisascaris spp. Based on the microscopic and histochemical examination, a diagnosis of neural Baylisascaris spp. larva migrans was made., (Copyright © 2014 Elsevier GmbH. All rights reserved.)

An incidental, asymptomatic, well-circumscribed, solitary, submucosal nodular mass was detected on the mucosal surface of the inner lower lip in a female cynomolgus macaque (age, approximately 2.4 y) during a juvenile chronic toxicology study. Grossly, the nodule was soft with brown to tan discoloration and measured approximately 4 mm in diameter. Microscopically, the nodule was covered by normal stratified squamous epithelium and composed of well-circumscribed irregular lobules containing hyperplastic and normal-appearing mucinous salivary gland acini and ducts, which were separated by thick connective tissue septae. In light of the gross pathology and results of microscopic examination, salivary gland hamartoma was diagnosed. This lesion resembles adenomatoid hyperplasia of mucous salivary glands in humans, which is a rare nonneoplastic swelling. To our knowledge, this case description is the first report of a cynomolgus macaque with the rare entity of lip salivary gland hamartoma, which likely represents adenomatous hyperplasia in humans.

Sphingosine-1-phosphate (S1P) is a major bioactive phospholipid, which binds to and activates a family of five G-protein-coupled receptors designated as S1P 1 (S1P1) through S1P5. The S1P1 receptor subtype, expressed primarily on lymphocytes, is known to play a critical role in the regulation of lymphocyte trafficking. S1P1 inhibitors result in the inhibition of lymphoid cell trafficking and are of interest to treat various inflammatory conditions. In this study, we describe a gastric finding associated with oral gavage administration of a small molecule S1P1 inhibitor to Sprague-Dawley rats. Rats were administered an S1P1 inhibitor once daily for 4 weeks and necropsies were conducted at the end of the dosing phase, and clinical pathology and histopathologic examination were performed. Lymphopenia and changes in lymphoid tissues were noted and were consistent with the pharmacodynamic effects for S1P1 inhibitory action. Histopathologic examination of the stomach revealed atrophy and depletion of gastric parietal cells in the glandular portion of the stomach. There are no literature data to suggest that this gastric effect is related to S1P1 pharmacology. Therefore, the mechanism of the observed gastric lesion is likely chemotype mediated.

Renal tubular inclusion bodies are rarely associated with drug administration. The authors describe the finding of renal cortical tubular intranuclear and intracytoplasmic inclusion bodies associated with the oral administration of a norepinephrine/serotonin reuptake inhibitor (NSRI) test article in Sprague-Dawley (SD) rats. Rats were given an NSRI daily for 4 weeks, and kidney histopathologic, ultrastructural pathology, and immunohistochemical examinations were performed. Round eosinophilic intranuclear inclusion bodies were observed histologically in the tubular epithelial cells of the renal cortex in male and female SD rats given the NSRI compound. No evidence of degeneration or necrosis was noted in the inclusion-containing renal cells. By ultrastructural pathology, inclusion bodies consisted of finely granular, amorphous, and uniformly stained nonmembrane-bound material. By immunohistochemistry, inclusion bodies stained positive for d-amino acid oxidase (DAO) protein. In addition, similar inclusion bodies were noted in the cytoplasmic tubular epithelial compartment by ultrastructural and immunohistochemical examination.  This is the first description of these renal inclusion bodies after an NSRI test article administration in SD rats. Such drug-induced renal inclusion bodies are rat-specific, do not represent an expression of nephrotoxicity, represent altered metabolism of d-amino acids, and are not relevant to human safety risk assessment.

The occurrence of mesenteric lymph node angiomas (benign vascular neoplasms including lymphangioma and hemangioma) in untreated control rats in 2-year carcinogenicity studies can range from rare to common depending on the strain used. This lesion is most common in male rats. Factors and conditions that may contribute to the etiopathogenesis of lymph node angiomas in rats include: (1) genetic drift, (2) congenital/developmental malformation, (3) sinus vascular transformation/venous obstruction of outflow, (4) "inflammatory" pseudo-tumors, and/or (5) defects of endothelial lymphatic vascular secretion/permeability. Lymph node angiomas in humans are extremely rare, not reported in mesenteric lymph nodes, and more common in females than males. The evaluation of increased mesenteric lymph node angiomas in rats for overall human safety risk assessment of novel pharmaceutical therapeutics should consider: genotoxicity of the test article, occurrence of vascular neoplasms in other locations in rats and in mice, occurrence of proliferative vascular lesions in nonclinical toxicology studies in non-rodent species, dose/exposure response, and pathophysiologic/morphologic differences and similarities of lymph node angiomas between rats and humans. Angiomas are independent lesions from angiosarcomas and are not precursors for angiosarcomas in either humans or animals. Mesenteric lymph node angiomas in rats are unlikely to be relevant for human risk assessment of pharmaceutical agents., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Objective: The mechanistic link between Janus kinase (JAK) signaling and structural damage to arthritic joints in rheumatoid arthritis (RA) is poorly understood. This study was undertaken to investigate how selective inhibition of JAK with tofacitinib (CP-690,550) affects osteoclast-mediated bone resorption in a rat adjuvant-induced arthritis (AIA) model, as well as human T lymphocyte RANKL production and human osteoclast differentiation and function., Methods: Hind paw edema, inflammatory cell infiltration, and osteoclast-mediated bone resorption in rat AIA were assessed using plethysmography, histopathologic analysis, and immunohistochemistry; plasma and hind paw tissue levels of cytokines and chemokines (including RANKL) were also assessed. In vitro RANKL production by activated human T lymphocytes was evaluated by immunoassay, while human osteoclast differentiation and function were assessed via quantitative tartrate-resistant acid phosphatase staining and degradation of human bone collagen, respectively., Results: Edema, inflammation, and osteoclast-mediated bone resorption in rats with AIA were dramatically reduced after 7 days of treatment with the JAK inhibitor, which correlated with reduced numbers of CD68/ED-1+, CD3+, and RANKL+ cells in the paws; interleukin-6 (transcript and protein) levels were rapidly reduced in paw tissue within 4 hours of the first dose, whereas it took 4-7 days of therapy for RANKL levels to decrease. Tofacitinib did not impact human osteoclast differentiation or function, but did decrease human T lymphocyte RANKL production in a concentration-dependent manner., Conclusion: These results suggest that the JAK inhibitor tofacitinib suppresses osteoclast-mediated structural damage to arthritic joints, and this effect is secondary to decreased RANKL production., (Copyright © 2012 by the American College of Rheumatology.)

This unit describes in detail the in vitro methods for measuring the interaction of new chemical entities (NCEs) with human renal transporters (hOAT1, hOAT2, and hOCT2) as both a substrate and inhibitor. Renal transporter substrate assays help in the identification of renal secretion mechanisms and assessment of the potential renal drug-drug interactions (DDIs) for NCE as a target, as well as to predict its renal clearance in humans. Human renal transporter (hOAT1, hOAT2, and hOCT2) inhibition assays characterize the inhibition potency of NCE and predict the potential for renal DDIs as a perpetrator with xenobiotics and drugs that are mainly renally cleared. In addition, such inhibition assays enable a better assessment of the potential for renal transporter-mediated nephrotoxicity and pathology. Therefore, renal transporter substrate and inhibition assays are pivotal in drug discovery and development for renally cleared drugs and those that are co-administered with marketed compounds mainly eliminated via the kidney., (© 2012 by John Wiley & Sons, Inc.)

The utility of inflammatory bowel disease (IBD) models in evaluating pharmacologic efficacy of novel drug candidates is reviewed. IBD models are generally classified into six groups based on the etiopathogenesis: chemically- and hapten-induced, spontaneously developed, T-cells, transgenic and immunoregulatory knockout models. The chemically- and hapten-induced models are the most widely utilized for evaluating pharmacologic efficacy of novel drug candidates because they are technically simple and rapid to induce gut pathology. In contrast, the T-cells adoptive transfer model is technically more complex to execute with longer study duration, resulting in the rare utility of this model in pharmacologic efficacy studies. Spontaneously developed, transgenic and immunoregulatory knockout IBD models gradually develop spontaneous colitis or ileitis as they age. In this critical review, the pathological and immunological characteristics of various IBD animal models, and the pharmacologic efficacy of current therapeutic agents and drug candidates for IBD in these animal models are compared. Moreover, perspectives on experimental conditions, and applicability to evaluation of prophylactic and therapeutic pharmacologic efficacy of drug candidates in drug discovery and development are discussed.

Background: 2,4,6-Trinitrobenzenesulfonic acid (TNBS)-induced colitis has been used as an inflammatory bowel disease (IBD), Crohn's disease (CD), preclinical model. However, published data on pharmacologic and therapeutic efficacy testing of this model are limited. FTY720 inhibits lymphoid cell trafficking in inflammatory conditions and is of interest to treat IBD., Aim: We investigated the pharmacologic therapeutic efficacy of sulfasalazine, FTY720, and anti-IL-12/23p40, in a TNBS CD model., Methods: Female, 7-week-old, BALB/c mice were given sulfasalazine orally (PO) and intraperitoneally (IP) at 10 mg/kg, FTY720 at 3 mg/kg PO, and mouse anti-IL-12/23p40 at 25 mg/kg IP. Vehicle groups given PO either phosphate-buffered saline/water or 40% ethanol served as controls. Pharmacologic efficacy was assessed using body weight loss, clinical scores of diarrhea and intestinal gross pathology, and colon weight parameters., Results: Sulfasalazine and FTY720 treatment did not prevent body weight loss or reduce clinical scores of diarrhea or intestinal gross pathology, when compared with vehicle treatment. However, anti-IL-12/23p40 treatment showed significant efficacy by preventing body weight loss, reducing clinical scores of diarrhea, and reducing intestinal gross pathologic lesions, when compared with vehicle-treated animals. Sulfasalazine, anti-IL-12/23p40, and FTY720 were not effective in reducing colon weight., Conclusion: With the exception of anti-IL-12/23p40, sulfasalazine, and FTY720 did not demonstrate full pharmacologic efficacy in our TNBS CD model.

Macrophage colony-stimulating factor (M-CSF) is a hematopoietic growth factor that is responsible for the survival and proliferation of monocytes and the differentiation of monocytes into macrophages, including Kupffer cells (KCs) in the liver. KCs play an important role in the clearance of several serum enzymes, including aspartate aminotransferase and creatine kinase, that are typically elevated as a result of liver or skeletal muscle injury. We used three distinct animal models to investigate the hypothesis that increases in the levels of serum enzymes can be the result of decreases in KCs in the apparent absence of hepatic or skeletal muscle injury. Specifically, neutralizing M-CSF activity via a novel human monoclonal antibody reduced the CD14(+)CD16(+) monocyte population, depleted KCs, and increased aspartate aminotransferase and creatine kinase serum enzyme levels in cynomolgus macaques. In addition, the treatment of rats with clodronate liposomes depleted KCs and led to increased serum enzyme levels, again without evidence of tissue injury. Finally, in the osteopetrotic (Csf1(op)/Csf1(op)) mice lacking functional M-CSF and having reduced levels of KCs, the levels of serum enzymes are higher than in wild-type littermates. Together, these findings support a mechanism for increases in serum enzyme levels through M-CSF regulation of tissue macrophage homeostasis without concomitant histopathological changes in either the hepatic or skeletal system., (Copyright © 2011 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Prostaglandins synthesized by enzymatic reactions such as cyclooxygenases have been implicated in lung pathophysiology. The goal of this study was to delineate the pulmonary ontogeny of cyclooxygenase enzymes (COX-1 and COX-2) immunohistochemical expression and cellular localization in various microanatomic locations of lungs from pre-term, term, and post-natal lambs. Lung tissues were obtained at 115 and 130 days of gestation from pre-term lambs, 145 days (term; complete gestation), and 15 days post-natally. No significant differences were seen in lung COX-1 expression at various microanatomic locations during pre-term, term, or postnatally. Moderate to strong COX-1 expression was present in macrophages, alveolar septa, bronchial smooth muscle cells, bronchiolar smooth muscle cells, vascular endothelial cells, and vascular smooth muscle cells. Minimal COX-1 expression was present in bronchial and bronchiolar epithelial cells. Most microanatomic locations lacked COX-2 expression with the exception of weak expression that was present in bronchial and bronchiolar epithelial cells at 145 days of full gestation and 15 days post-natally. This work suggests that: (a) COX-1 is constitutively expressed in lungs from pre-term, term, and post-natal lambs in various microanatomic pulmonary locations, (b) there is differential expression of COX-1 and COX-2 in the developing lung, and (c) COX-2 does not appear to play a role in lung fetal development, at least in neonatal lambs., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Inflammatory bowel disease (IBD) is a chronic disease that affects not only the young adults, but also the elderly. The elderly are more vulnerable and at higher risk from complications related to IBD. In this review we focus on IBD important features in the elderly and discuss the disease (1) epidemiology, (2) pathophysiology, (3) clinical manifestations and diagnosis, (4) prognosis, (6) therapy and (7) potential future research directions.

Inhibitors of the JAK family of nonreceptor tyrosine kinases have demonstrated clinical efficacy in rheumatoid arthritis and other inflammatory disorders; however, the precise mechanisms by which JAK inhibition improves inflammatory immune responses remain unclear. In this study, we examined the mode of action of tofacitinib (CP-690,550) on JAK/STAT signaling pathways involved in adaptive and innate immune responses. To determine the extent of inhibition of specific JAK/STAT-dependent pathways, we analyzed cytokine stimulation of mouse and human T cells in vitro. We also investigated the consequences of CP-690,550 treatment on Th cell differentiation of naive murine CD4(+) T cells. CP-690,550 inhibited IL-4-dependent Th2 cell differentiation and interestingly also interfered with Th17 cell differentiation. Expression of IL-23 receptor and the Th17 cytokines IL-17A, IL-17F, and IL-22 were blocked when naive Th cells were stimulated with IL-6 and IL-23. In contrast, IL-17A production was enhanced when Th17 cells were differentiated in the presence of TGF-β. Moreover, CP-690,550 also prevented the activation of STAT1, induction of T-bet, and subsequent generation of Th1 cells. In a model of established arthritis, CP-690,550 rapidly improved disease by inhibiting the production of inflammatory mediators and suppressing STAT1-dependent genes in joint tissue. Furthermore, efficacy in this disease model correlated with the inhibition of both JAK1 and JAK3 signaling pathways. CP-690,550 also modulated innate responses to LPS in vivo through a mechanism likely involving the inhibition of STAT1 signaling. Thus, CP-690,550 may improve autoimmune diseases and prevent transplant rejection by suppressing the differentiation of pathogenic Th1 and Th17 cells as well as innate immune cell signaling.

Cyclooxygenase (COX) catalyzes the conversion of arachidonic acid into prostaglandin H(2) (PGH(2)), which is subsequently converted to the prostanoids PGE(2), PGI(2), PGF(2alpha), and thromboxane A(2). COX has 2 distinct membrane-anchored isoenzymes: COX-1 and COX-2. COX-1 is constitutively expressed in most normal tissues; COX-2 is highly induced by proinflammatory mediators in the setting of inflammation, injury, and pain. Inhibitors of COX activity include conventional nonselective nonsteroidal anti-inflammatory drugs and selective nonsteroidal anti-inflammatory drugs, such as COX-2 inhibitors. The adverse effects of COX inhibitors on the cardiovascular system have been addressed in the last few years. In general, COX inhibitors have many effects, but those most important to the cardiovascular system can be direct (through the effects of prostanoids) and indirect (through alterations in fluid dynamics). Despite reports of detrimental human cardiovascular events associated with COX inhibitors, short, long, and lifetime preclinical toxicology studies in rodents and nonrodents have failed to identify these risks. This article focuses on the expression and function of COX enzymes in normal and pathologic conditions of the cardiovascular system and discusses the cardiovascular pathophysiologic complications associated with COX inhibition.

The authors describe a case in which well-circumscribed, expansile, and nonencapsulated nodular masses with missing digits were detected on the right and left feet in a 6-year-old female cynomolgus macaque from a routine toxicology study. Grossly, these masses were composed of variably sized and firm nodules containing white, chalklike material in the subcutaneous tissue on cross section. Microscopically, the nodules were composed of irregular lobules containing amorphous to granular, light to dark basophilic material that was surrounded by macrophages and multinucleated giant cells and separated by fibrous connective tissue. The nodule's contents were von Kossa and Alizarin red S positive. Serum calcium and phosphorus levels of this monkey were within normal ranges. Based on the gross pathology, histopathology, serum chemistry, and histochemistry, a diagnosis of dystrophic calcinosis circumscripta was made. Dystrophic calcinosis circumscripta is an uncommon syndrome of mineralization that occurs following tissue damage, without abnormalities in calcium and phosphorus homeostasis, and it is characterized by deposition of calcium salts in soft tissues. To the best of the authors' knowledge, this is the first report of dystrophic calcinosis circumscripta in a cynomolgus macaque.

Prostaglandins (PGs) play an important role in pulmonary physiology and various pathophysiological processes following infection. The initial step in the biosynthesis of PGs is regulated by two distinct cyclooxygenase enzymes, cyclooxygenase-1 (COX-1) and COX-2. The goal of this study was to investigate the pulmonary cellular localization and distribution of COX-1 and COX-2 in a neonatal lamb model following respiratory syncytial virus (RSV) and parainfluenza virus 3 (PI3) infection, organisms that also cause significant respiratory disease in children. No significant differences were seen in pulmonary COX-1 expression at various microanatomical locations following RSV or PI3 infection compared to controls. In contrast, COX-2 was upregulated following RSV and PI3 infection. Strong expression was restricted to bronchial and bronchiolar epithelial cells and macrophages, while minimal expression was present in the same microanatomical locations in the uninfected lungs. Other microanatomical locations in both the controls and the infected lungs lacked expression. This work suggests that during RSV or PI3 infection: (1) COX-1 cellular expression is not altered, (2) COX-2 cellular expression is upregulated in airway bronchiolar and bronchial epithelial cells and macrophages, (3) respiratory epithelium along with macrophages are important microanatomical compartments regulating the host inflammatory response during viral infection, and (4) COX-2 may be a potential target for RSV and PI3 therapy.

The p38 mitogen-activated protein kinases (MAPKs) are members of discrete signal transduction pathways that have significant regulatory roles in a variety of biological processes, depending on the cell, tissue and organ type. p38 MAPKs are involved in inflammation, cell growth and differentiation and cell cycle. In the female reproductive system, p38 MAPKs are known to regulate various aspects of the reproductive process such as mammalian estrous and menstrual cycles as well as early pregnancy and parturition. p38 MAPKs have also been implicated in alterations and pathologies observed in the female reproductive system. Therefore, pharmacologic modulation of p38 MAPKs, and inter-connected signaling pathways (e.g., estrogen receptor signaling, c-fos, c-jun), may influence reproductive physiology and function. This article provides a critical, comparative review of available data on the roles of p38 MAPKs in the mammalian female reproductive system and in reproductive pathophysiology in humans and preclinical species. We first introduce fundamental differences and similarities of the mammalian female reproductive system that should be considered by toxicologists and toxicologic pathologists when assessing the effects of new pharmacologic agents on the female reproductive system. We then explore in detail the known roles for p38 MAPKs and related molecules in female reproduction. This foundation is then extended to pathological conditions in which p38 MAPKs are thought to play an integral role.

1. Hypertensive mice expressing the human renin (REN) and angiotensinogen (AGT) genes are used as a model for human hypertension. 2. The aim of the present study was to investigate the cellular expression and distribution of inducible nitric oxide synthase (iNOS) using immunohistochemistry in lung, heart and kidney tissues from a model of human hypertension using male and female double-transgenic (h-Ang 204/1h-Ren6) mice and wild-type C57/BI6J mice as controls. 3. In the kidney, the pattern of iNOS expression in various renal microanatomical regions during hypertension was similar to that of age-matched controls, except in the medullary ascending limb (MAL). In hypertension, iNOS expression was downregulated in the MAL. No significant differences in iNOS expression were seen between control or hypertensive mice in various cardiac microanatomical locations. In the lungs of hypertensive mice, iNOS expression was upregulated in bronchial airway epithelium and bronchial and vascular smooth muscle cells, but downregulated in alveolar macrophages, alveolar septa and pulmonary vascular endothelial cells. Expression of iNOS was similar between male and female mice in the kidney, heart and lungs. 4. In conclusion, iNOS regulation in hypertension is complex and depends on the cell type in which it is expressed and the localization of the cell type in the cardiorenal and pulmonary systems.

A well-circumscribed, expansile, and nonencapsulated cardiac rhabdomyoma composed of tightly arranged, large, variably sized, ovoid to irregular, swollen myocytes with deeply eosinophilic cytoplasm and varying degrees of cytoplasmic vacuolation was detected in an eight- to nine-month-old female beagle dog in a routine toxicology study. By histochemistry, the neoplasm was periodic acid-Schiff positive. By immunohistochemistry (IHC), neoplastic cells were positive for desmin and myoglobin and negative for vimentin and smooth muscle actin. Spontaneous lesions in the heart of young beagle dogs are rare in drug safety studies. On the basis of histopathology, histochemistry, and IHC findings, a diagnosis of cardiac rhabdomyoma was made. Cardiac rhabdomyoma is one of the most frequently occurring primary tumors of the heart and, by far, the most common neoplasm in human infants and children. To our knowledge, this is the first report of the canine cardiac rhabdomyoma.

Mice that are homozygous for the recessive osteopetrosis spontaneous mutation (op/op) develop severe osteopetrosis due to a defect in the production of macrophage colony-stimulating factor (M-CSF) and a deficiency in monocyte-derived osteoclasts. Our study describes a novel soft tissue finding in an osteopetrosis (B6C3Fe a/a-Csf1(op)/J) mouse model. Tissues were obtained from B6C3Fe a/a-Csf1(op)/J mice and age-matched wild-type mice, processed for hematoxylin and eosin sections, and comprehensive light microscopic tissue evaluation was performed. Mutant mice had characteristic traits of op/op deficiency including missing incisors and domed skulls. Histologically, the bone marrow cavity was effaced by interweaving thick bony trabeculae consistent with osteopetrosis. An increase in a finely granular, basophilic interstitial extracellular matrix (ECM) was observed in the subcutaneous connective tissue of the op/op mice when compared with controls. Histochemically, the ECM was negative with periodic acid Schiff and stained dark blue with alcian blue at a pH of 2.5, indicating that it is composed primarily of nonsulfated glycosaminoglycans (GAGs). This work suggests an increased ECM that is composed mainly of GAGs located in the subcutaneous tissue in op/op mice. This increase in ECM may be related to altered matrix production or turnover because of changes in M-CSF production., (2009 S. Karger AG, Basel.)

Cyclooxygenase (COX) catalyzes the conversion of arachidonic acid into prostaglandins (PGs), which play a significant role in health and disease in the gastrointestinal tract (GI) and in the renal, skeletal, and ocular systems. COX-1 is constitutively expressed and found in most normal tissues, whereas COX-2 can be expressed at low levels in normal tissues and is highly induced by pro-inflammatory mediators. Inhibitors of COX activity include: (1) conventional nonselective, nonsteroidal anti-inflammatory drugs (ns-NSAIDs) and (2) COX-2 selective nonsteroidal anti-inflammatory drugs (COX-2 s-NSAIDs). Inhibition of COX-1 often elicits GI toxicity in animals and humans. Therefore, COX-2 s-NSAIDs were developed to provide a selective COX-2 agent, while minimizing the attendant COX-1-mediated GI toxicities. Rats and dogs overpredict COX inhibition for renal effects such as renal handling of electrolytes in humans. COX inhibitors are shown to have both beneficial and detrimental effects, such as on healing of ligament or tendon tears, on the skeletal system in animal models. Certain ophthalmic conditions such as glaucoma and keratitis are associated with increased COX-2 expression, suggesting a potential role in their pathophysiology.

Age- and gender-related lymphoid tissue variability in control male and female monkeys of various ages (under three years; three to six years; seven to fifteen years) was characterized. Spleen and thymus organ weights, organ-to-body and organ-to-brain ratios, morphology by light microscopy, and B- and T-cell immunohistochemistry (IHC) were evaluated. Splenic weights and ratios were not significantly different between various age groups or genders, except males and females in the three-to-six-years age group, who exhibited statistically significant changes from the under-three-years age group. No differences in the number of primary follicles, secondary follicles with germinal centers, B-cell follicles, and periarterial lymphoid sheath were seen between age groups or genders, and no trends were noted in the spleen. By IHC, no differences were observed in B- and T-cell splenic densities. Several age- and gender-related changes in weights and ratios were noted in the thymus. The thymus had a trend toward increased interlobular fat infiltration with increasing age in both males and females. Thymic delineation of the cortex and medulla was significantly decreased in the seven-to-fifteen-years age group for males only. The cortex-to-medulla ratio was significantly lower only in males in the seven-to-fifteen-years age group. B- and T-cell cellular density did not change across various ages.

Cyclooxygenase isoenzymes (COX-1 and -2) catalyze the conversion of arachidonic acid to prostaglandins (PGs) and play a significant role in the health and disease of the eye. Experimental animal models of ocular diseases have been used to assess the effects of the selective COX inhibitors (COXIBs), and nonselective, nonsteroidal anti-inflammatory drugs (ns-NSAIDs) are currently employed in the management of various ophthalmic conditions. This paper provides a review of the comparative expression of COX-1 and COX-2 in the eye under normal and pathologic conditions, including the pathophysiologic role of PGs, and the effects of ns-NSAIDs and selective COX-2 inhibitors in the treatment of ophthalmic conditions.

Rats have an average estrous cycle of 4-5 days. There are four phases (proestrus, estrus, metesterus, and diestrus) in the estrous cycle in rodents. Histologic staging of the rodent estrous cycle is challenging and requires expertise. Thus, utilizing additional parameters such as cellular proliferation of the various components of the uterine microanatomy may assist with this process. Having an alternative method by which a pathologist can correctly identify the stages of the rodent estrous cycle would be valuable to the assessment and interpretation of safety studies for new drug candidates. This study was performed to investigate the microanatomic location of the uterine proliferative activity by image analysis and immunohistochemistry using Ki-67, a well-established marker of proliferating cells. Each stage of the rodent estrous cycle exhibited a different pattern of cellular proliferation. During proestrus, the lowest degree of cellular proliferation occurred in the glandular epithelial cells and the highest occurred in the myometrial cells. In estrus, lower levels of cellular proliferation were seen in the luminal and glandular epithelial cells, while a higher rate of proliferation occurred in myometrial cells followed by the stromal cells. At the metestrus stage, the highest cellular proliferation occurred in stromal and myometrial cells, while lesser proliferation was observed in luminal and glandular epithelial cells. This work demonstrates that in the rodent uterus there are cyclic changes in cellular proliferation in specific microanatomic uterine locations which can aid in the staging of the estrous cycle.

Chronic lymphocytic thyroiditis characterized by multifocal follicular lymphoid cell infiltrates with germinal centers, thyroid acinar atrophy and pituitary cell hyperplasia/hypertrophy of the adenohypophysis was detected in a vehicle control, 4-year-old female Cynomolgus macaque in a routine toxicology study. Lymphoid cells of germinal centers were positive for the B-cell marker CD20 by immunohistochemistry (IHC), while remaining lymphocytes were positive for the T-cell marker CD3. Hypertrophied/hyperplastic pituitary cells were positive for thyroid stimulating hormone (TSH) by IHC, consistent with an adaptive response due to removal of hormonal negative feedback from the diseased thyroid gland. Features of this case are similar to chronic lymphocytic thyroiditis in humans, an autoimmune disorder also known as Hashimoto's disease. Chronic lymphocytic thyroiditis with compensatory pituitary changes may occur spontaneously in young, clinically normal cynomolgus macaques and its presence in drug treated animals should be interpreted with caution.

Hypertensive mice that express the human renin and angiotensinogen genes are used as a model for human hypertension because they develop hypertension secondary to increased renin-angiotensin system activity. Our study investigated the cellular localization and distribution of COX-1, COX-2, mPGES-1, and mPGES-2 in organ tissues from a mouse model of human hypertension. Male (n = 15) and female (n = 15) double transgenic mice (h-Ang 204/1 h-Ren 9) were used in the study. Lung, kidney, and heart tissues were obtained from mice at necropsy and fixed in 10% neutral buffered formalin followed by embedding in paraffin wax. Cut sections were stained immunohistochemically with antibodies to COX-1, COX-2, mPGES-1, and mPGES-2 and analyzed by light microscopy. Renal expression of COX-1 was the highest in the distal convoluted tubules, cortical collecting ducts, and medullary collecting ducts; while proximal convoluted tubules lacked COX-1 expression. Bronchial and bronchiolar epithelial cells, alveolar macrophages, and cardiac vascular endothelial cells also had strong COX-1 expression, with other renal, pulmonary, or cardiac microanatomic locations having mild-to-moderate expression. mPGES-2 expression was strong in the bronchial and bronchiolar epithelial cells, mild to moderate in various renal microanatomic locations, and absent in cardiac tissues. COX-2 expression was strong in the proximal and distal convoluted tubules, alveolar macrophages, and bronchial and bronchiolar epithelial cells. Marked mPGES-1 was present only in bronchial and bronchiolar epithelial cells; while mild-to-moderate expression was present in other pulmonary, renal, or cardiac microanatomic locations. Expression of these molecules was similar between males and females. Our work suggests that in hypertensive mice, there are (a) significant microanatomic variations in the pulmonary, renal, and cardiac distribution and cellular localization of COX-1, COX-2, mPGES-1, and mPGES-2, and (b) no differences in expression between genders.

Cyclooxygenase (COX) is a rate-limiting enzyme that catalyzes the conversion of arachidonic acid, an essential fatty acid present in cell membrane phospholipids and liberated by phospholipase, into prostaglandins (PGs) and prostanoids. COX has two distinct membrane-anchored isoenzymes; COX-1 and COX-2. COX-1 is a constitutively expressed and found in most normal body tissues; COX-2 is expressed in normal tissues at low levels and is highly induced by pro-inflammatory mediators in the setting of inflammation, injury, and pain. Inhibitors of COX activity include: (1) conventional non-selective non-steroidal anti-inflammatory drugs (ns-NSAIDs); (2) selective COX-2 inhibitors (COXIBs); and (3) COX-1 inhibitors. Non-selective NSAIDs, at therapeutic doses, inhibit both COX-1 and COX-2. The anti-inflammatory benefits of these drugs are primarily derived from COX-2 inhibition, while inhibition of COX-1 often elicits gastrointestinal (GI) toxicity. Therefore, COXIBs were developed to provide a selective COX-2 agent, i.e., one, that at fully therapeutic doses demonstrated comparable therapeutic benefit to non-selective NSAIDs, without the attendant COX-1-mediated GI toxicities. In this review, we evaluate available literature describing the pathophysiologic role of cyclooxygenases and the effects of their inhibition in GI system in experimental and domestic animal species.

An ulcerated, 1 x 0.5 cm, subcutaneous mass on the craniolateral aspect of the right tibiotarsus of a 4-year-old male cockatiel was removed. Histologically, the neoplasm was non-encapsulated, infiltrative and composed of irregular vascular channels lined by branching and variably sized spindle-shaped cells with large vesicular nuclei, prominent nucleoli and rare mitoses. Surrounding these vascular channels were fibroblasts and mixed inflammatory cells. Neoplastic cells had diffuse immunoreactivity to factor VIII supporting a diagnosis of haemangiosarcoma.

An ear (auricular) neoplasm from a 1-year-old male rat was removed surgically and examined histologically. Macroscopically, the neoplasm was firm, white and measured (0.5 x 0.5 cm). Microscopically, the neoplasm was expansile, non-encapsulated, and composed of large, pleomorphic, polygonal to spindle-shaped cells containing multiple nuclei. Using immunohistochemical and chemical stains, the neoplastic cells were positive for vimentin, myoglobin, phosphotungstic acid haematoxylin and desmin, but had no immunoreactivity for cytokeratin or alpha-smooth muscle actin. On the basis of histopathological, immunohistochemical and histochemical stains, a diagnosis of auricular rhabdomyosarcoma was made. Although reported infrequently in human, this is, to the author's knowledge, the first report that describes the detailed gross, histopathological, histochemical and immunohistochemical findings of auricular rhabdomyosarcoma in a rat.

The uterine cellular expression and distribution of c-fos, ERalpha and p38alpha was compared in humans, nonhuman primates, and rats using immunohistochemistry. ERalpha and c-fos were present in the glandular (GE) and luminal epithelial cells (LE) of humans and nonhuman primates, with differing expression patterns evident between proliferative and secretory cycle phases. In rats, the highest and lowest expression of c-fos was present during proestrus and estrus, respectively, in the LE and GE. The most intense ERalpha staining in rats was observed during proestrus in the GE, while the least intense staining was seen in the LE during proestrus. Strong LE and GE expression of p38alpha was present in rats in all stages of the estrous cycle and during the proliferative phase in both humans and nonhuman primates. No p38alpha expression was observed during the secretory phase in either humans or nonhuman primates. Our work suggests that c-fos, ERalpha and p38alpha (a) are primarily expressed during the proliferative phase, but not the secretory phase and exhibit interspecies expression variability, and (b) rats exhibit cyclic changes in the expression of c-fos and ERalpha.