Reproducibility of histopathological findings in experimental pathology of the mouse: a sorry tail

Perspective Focus on Reproducibility Reproducibility of histopathological findings in experimental pathology of the mouse: a sorry tail © 2017 Nature America, Inc., part of Springer Nature. All rights reserved. Jerrold M Ward1, Paul N Schofield2,3 & John P Sundberg3 Reproducibility of in vivo research using the mouse as a model organism depends on many factors, including experimental design, strain or stock, experimental protocols, and methods of data evaluation. Gross and histopathology are often the endpoints of such research and there is increasing concern about the accuracy and reproducibility of diagnoses in the literature. To reproduce histopathological results, the pathology protocol, including necropsy methods and slide preparation, should be followed by interpretation of the slides by a pathologist familiar with reading mouse slides and familiar with the consensus medical nomenclature used in mouse pathology. Likewise, it is important that pathologists are consulted as reviewers of manuscripts where histopathology is a key part of the investigation. The absence of pathology expertise in planning, executing and reviewing in vivo research using mice leads to questionable pathology-based findings and conclusions from studies, even in high-impact journals. We discuss the various aspects of this problem, give some examples from the literature and suggest solutions. Histopathological descriptions of the frequency and nature of lesions and disease entities are very often the endpoints in biomedical research conducted in model organisms such as the mouse. In contrast with clinical pathology, where endpoints are usually assessed using biochemical and molecular assays, histopathological assessment, despite using molecular markers and imaging as adjunct qualitative and quantitative techniques, is highly dependent on the individual expertise of trained expert pathologists. Pathologists must not only recognize lesions, but also have knowledge of the background diseases of the mice and understand the meaning of the pattern of disease in the whole mouse1–4. Reproducibility of histopathological endpoints therefore depends on the implementation of a common standardized vocabulary, competent work-up and an in-depth knowledge of the mouse strains under investigation so that, for example, background lesions are not mistaken for those that are experimentally induced. Such knowledge is critical in the design of experiments, as well as in understanding the effects of husbandry, the microbiome and diet on the interpretation of results2,5. In recent years, funding agencies and scientific communities alike have expressed increasing concern about the lack of reproducibility of experiments in the biomedical domain. Attention was initially drawn to this issue by pharmaceutical companies, which rely on preclinical, precompetitive research for drug development pipelines6–8. Identification of this problem has been followed by an outpouring of concern from funding agencies such as the US National Institutes of Health9–11 and, to an extent, journals and professional bodies12–17. Although much attention has been paid to the reproducibility of molecular assays, in vitro (cell culture) assays and the inappropriate application of statistical methods, only recently have the issues surrounding reproducibility in animal experimentation been discussed in depth13. Many of these discussions have concerned husbandry and the effect of diet and microbiome on experimental outcomes18–20, particularly in neuroscience14. However, recent papers have addressed the problem of what a sound histopathological investigation should look like, how to use knowledge of pathology in experimental design, based on the ARRIVE and related guidelines, and the confounding effect of the environment and the gut and skin microbiomes5. Here we address some of the issues that affect reproducibility of histopathological findings: a lack of pathology expertise, in author lists and in peer review; poor standards of reporting, as illustrated by common errors seen in papers, and inconsistent pathology nomenclature; and availability of primary data, without which it is impossible to assess a paper without attempting a complete experimental replication21–23. Notably, we emphasize that if pathologists are not involved in designing mouse experiments and interpreting lesions, the accuracy of the diagnoses reported and the conclusions drawn may be questionable. The importance of pathologists Pathology is a medical specialty that requires years of training, experience and board certification as a minimum. Although pathology has many sub-disciplines, such as mouse pathology, 1Global VetPathology, Montgomery Village, Maryland, USA. 2Department of Physiology Development and Neuroscience, University of Cambridge, Downing Street, Cambridge, UK. 3The Jackson Laboratory, Bar Harbor, Maine, USA. Correspondence should be addressed to J.M.W. (). 146    Volume 46, No. 4 | APRIL 2017 www.nature.com/laban Focus on Reproducibility Perspective Table 1 | Evidence of questionable pathology interpretation in publications Figure legends do not accurately reflect what is in the figure Figure legends do not describe anything in the figure Lack of complete or appropriate necropsies and histopathology Misidentification of normal organs and tissues as lesions Diagnoses of non-neoplastic lesions as neoplasms Diagnoses of tumors with unconventional terminology © 2017 Nature America, Inc., part of Springer Nature. All rights reserved. Reporting of benign lesions as malignant Reporting of inflammatory lesions as neoplasms Reporting of novel lesions incorrectly Use of incorrect (accepted) terminology/diagnoses a general pathologist is much more of an expert than a nonpathologist, and is often sufficient to provide substantial benefit to an animal research study24. However, investigators often do not have enough funds to pay for research pathology services and/or believe that they can perform histopathology interpretations themselves. A lack of pathology expertise by investigators leads to inaccurate histopathological descriptions of lesions and often missed, or spurious reporting of, pathological findings in publications. Absence of a pathologist may be noticed in the figure legends, which often do not describe the lesions displayed, or, in some rare cases, in images that are replicated in various orientations for different lesions of mice25. In some cases, a pathologist was not involved in late of final edits or did not review the galley proofs of an accepted manuscript26, leading to a substantial error in reporting. In addition to accurate interpretation of data, pathologists are important for ensuring that proper nomenclature is used when reporting on results. The use of generally accepted pathology nomenclature for unexpected and new findings leads to publications that can be interpreted by readers, including other pathologists. Rodent pathology term (...truncated)