Botulinum neurotoxins E (BoNT/E) and A (BoNT/A) act by cleaving Synaptosome-Associated Protein 25 (SNAP25) at two different C-terminal sites, but they display very distinct durations of action, BoNT/E being short acting and BoNT/A long acting. We investigated the duration of action, spread and neuronal transport of BoNT/E (6.5 ng/kg) and BoNT/A (125 pg/kg) after single intramuscular administrations of high equivalent efficacious doses, in rats, over a 30- or 75-day periods, respectively. To achieve this, we used (i) digit abduction score assay, (ii) immunohistochemistry for SNAP25 (N-ter part; SNAP25N-ter and C-ter part; SNAP25C-ter) and its cleavage sites (cleaved SNAP25; c-SNAP25E and c-SNAP25A) and (iii) muscular changes in histopathology evaluation. Combined in vivo observation and immunohistochemistry analysis revealed that, compared to BoNT/A, BoNT/E induces minimal muscular changes, possesses a lower duration of action, a reduced ability to spread and a decreased capacity to be transported to the lumbar spinal cord. Interestingly, SNAP25C-ter completely disappeared for both toxins during the peak of efficacy, suggesting that the persistence of toxin effects is driven by the persistence of proteases in tissues. These data unveil some new molecular mechanisms of action of the short-acting BoNT/E and long-acting BoNT/A, and reinforce their overall safety profiles.

Harmonization of diagnostic terminology used during the histopathologic analysis of rodent tissue sections from nonclinical toxicity studies will improve the consistency of data sets produced by laboratories located around the world. The INHAND Project (International Harmonization of Nomenclature and Diagnostic Criteria for Lesions in Rats and Mice) is a cooperative enterprise of 4 major societies of toxicologic pathology to develop a globally accepted standard vocabulary for proliferative and nonproliferative lesions in rodents. A prior manuscript (Toxicol Pathol 2012;40[4 Suppl]:87S-157S) defined multiple diagnostic terms for toxicant-induced lesions, common spontaneous and age-related changes, and principal confounding artifacts in the rat and mouse central nervous system (CNS) and peripheral nervous system (PNS). The current article defines 9 new diagnostic terms and updates 2 previous terms for findings in the rodent CNS and PNS, the need for which has become evident in the years since the publication of the initial INHAND nomenclature for findings in rodent neural tissues. The nomenclature presented in this document is also available electronically on the Internet at the goRENI website (http://www.goreni.org/).

Glucagon-like peptide 1 (GLP1) analogs have been associated with an increased incidence of thyroid C-cell hyperplasia and tumors in rodents. This effect may be due to a GLP1 receptor (GLP1R)-dependent mechanism. As the expression of GLP1R is much lower in primates than in rodents, the described C-cell proliferative lesions may not be relevant to man. Here, we aimed to establish primary thyroid cell cultures of rat and human to evaluate the expression and function of GLP1R in C-cells. In our experiments, GLP1R expression was observed in primary rat C-cells ( in situ hybridization) but was not detected in primary human C-cells (mRNA and protein levels). The functional response of the cultures to the stimulation with GLP1R agonists is an indirect measure of the presence of functional receptor. Liraglutide and taspoglutide elicited a modest increase in calcitonin release and in calcitonin expression in rat primary thyroid cultures. Contrarily, no functional response to GLP1R agonists was observed in human thyroid cultures, despite the presence of few calcitonin-positive C-cells. Thus, the lack of functional response of the human cultures adds to the weight of evidence indicating that healthy human C-cells have very low levels or completely lack GLP1R. In summary, our results support the hypothesis that the GLP1R agonist-induced C-cell responses in rodents may not be relevant to primates. In addition, the established cell culture method represents a useful tool to study the physiological and/or pathological roles of GLP1 and GLP1R agonists on normal, non-transformed primary C-cells from rats and man.

Pathology peer review verifies and improves the accuracy and quality of pathology diagnoses and interpretations. Pathology peer review is recommended when important risk assessment or business decisions are based on nonclinical studies. For pathology peer review conducted before study completion, the peer-review pathologist reviews sufficient slides and pathology data to assist the study pathologist in refining pathology diagnoses and interpretations. Materials to be reviewed are selected by the peer-review pathologist. Consultations with additional experts or a formal (documented) pathology working group may be used to resolve discrepancies. The study pathologist is solely responsible for the content of the final pathology data and report, makes changes resulting from peer-review discussions, initiates the audit trail for microscopic observations after all changes resulting from peer-review have been made, and signs the final pathologist’s report. The peer-review pathologist creates a signed peer-review memo describing the peer-review process and confirming that the study pathologist’s report accurately and appropriately reflects the pathology data. The study pathologist also may sign a statement of consensus. It is not necessary to archive working notes created during the peer-review process.

JENNIFER S. MCKAY, ERIO BARALE-THOMAS, BRAD BOLON, CATHERINE GEORGE, JERRY HARDISTY, SUNAO MANABE, FREDERIC SCHORSCH, MUNEHIRO TERANISHI, AND KLAUS WEBER AstraZeneca, Macclesfield, SK10 4TG, United Kingdom Johnson and Johnson PRD, 2340 Beerse, Belgium GEMpath Inc., Longmont, CO 80503-2339, USA Ipsen, 91966 Les Ulis Cedex, France Experimental Pathology Laboratories, Research Triangle Park NC 22709, USA Daiichi Sankyo Co., Ltd., Japan Bayer CropScience, 69009 Lyon, France Harlan Laboratories, 4452 Itingen, Switzerland

The International Federation of Societies of Toxicologic Pathologists (IFSTP) proposes a common global framework for training future toxicologic pathologists who will support regulatory-type, nonclinical toxicology studies. Optimally, trainees should undertake a scientific curriculum of at least five years at an accredited institution leading to a clinical degree (veterinary medicine or medicine). Trainees should then obtain four or more years of intensive pathology practice during a residency and/or on-the-job "apprenticeship," at least two years of which must be focused on regulatory-type toxicologic pathology topics. Possession of a recognized pathology qualification (i.e., certification) is highly recommended. A nonclinical pathway (e.g., a graduate degree in medical biology or pathology) may be possible if medically trained pathologists are scarce, but this option is not optimal. Regular, lifelong continuing education (peer review of nonclinical studies, professional meetings, reading, short courses) will be necessary to maintain and enhance one's understanding of current toxicologic pathology knowledge, skills, and tools. This framework should provide a rigorous yet flexible way to reliably train future toxicologic pathologists to generate, interpret, integrate, and communicate data in regulatory-type, nonclinical toxicology studies.

Subcutaneous injection of pharmaceutical compounds into the dorsal skin of rats is common in preclinical and nonclinical studies. However, no detailed histologic description of this anatomic location has been published to date. Following the observation of vascular lesions in the dorsum of rats in a thirteen-week toxicity study, a complementary study was performed on untreated Sprague-Dawley rats to evaluate the normal histology of the skin and subcutis, the potential effect of chronic subcutaneous injection on the morphology of the skin and its vasculature, and the spontaneous vascular pathology in the areas used as injection sites in the principal study. This study showed that saline injection did not fundamentally alter the morphology of the injection sites used for the principal study. Skin thickness was greater in males than in females. Although acellular intimal thickening occurred spontaneously in the dorsal skin of untreated males and females, only males had a spontaneous incidence of intimal hyperplasia. No site predilection for intimal lesions was apparent for either sex. Saline injection, or the physical trauma of injection, may induce intimal hyperplasia; males appear more likely to develop the lesion than do females. It is possible that acellular intimal thickening can progress to intimal hyperplasia under appropriate conditions.

The International Federation of Societies of Toxicologic Pathologists (IFSTP) proposes a common global framework for training future toxicologic pathologists who will support regulatory-type, nonclinical toxicology studies. Optimally, trainees should undertake a scientific curriculum of at least five years at an accredited institution leading to a clinical degree (veterinary medicine or medicine). Trainees should then obtain four or more years of intensive pathology practice during a residency and/or on-the-job ‘‘apprenticeship,’’ at least two years of which must be focused on regulatory-type toxicologic pathology topics. Possession of a recognized pathology qualification (i.e., certification) is highly recommended. A nonclinical pathway (e.g., a graduate degree in medical biology or pathology) may be possible if medically trained pathologists are scarce, but this option is not optimal. Regular, lifelong continuing education (peer review of nonclinical studies, professional meetings, reading, short courses) will be necessary to maintain and enhance one’s understanding of current toxicologic pathology knowledge, skills, and tools. This framework should provide a rigorous yet flexible way to reliably train future toxicologic pathologists to generate, interpret, integrate, and communicate data in regulatory-type, nonclinical toxicology studies.

As part of the ICH process of harmonization of testing guidelines for immunotoxicity, the European Society of Toxicologic Pathology (ESTP) has contributed to the scientific discussion on methods and evaluation of immunotoxicity studies with technical and scientific recommendations on toxicologic pathology. The weighing and sampling of immune organs is discussed taking into consideration specifically the value of lymph node weighing and the selection of appropriate lymph nodes for the detection of local and systemic effects. The different techniques of bone marrow preparation are considered for routine and extended investigations. Criteria are given for the gross and histopathological detection of effects in Peyer's patches. For the histopathological evaluation it is strongly recommended that each compartment within the different lymphoid organs is investigated separately and semiquantitatively since this approach has shown to increase the sensitivity and specificity of immunohistopathology.