Special Focus on Regulated Cell Death: Emerging Mechanisms and Current Perspectives in Biology and Pathology

In the past few years, there has been considerable advancement in our understanding of cell death, including the discovery of a series of highly regulated molecular pathways implicated in determining whether a cell will survive or succumb. ¹² These newly recognized modalities of regulated cell death (RCD) may manifest as part of physiologic programs, such as during embryonic or postnatal development, or for the maintenance of homeostasis in adult tissues. ^1,4 However, RCD mechanisms can also be activated when there is failure of adaptive responses to noxious perturbations of the cellular microenvironment. ⁶

While the field of RCD has been historically dominated by the recognition of predominant morphologies, the Nomenclature Committee on Cell Death (NCCD) has more recently proposed a classification based on the identification of distinct molecular pathways and quantifiable biochemical parameters. ⁶ Starting from 2005, the NCCD has provided regular updates on the increasingly intricate nature of cell death focusing on the definition of the mechanisms that orchestrate the newly emerging forms of cell demise. ⁸ Of note, novel insights were obtained into the mechanisms of caspase-independent regulated cell death manifesting with necrotic features. These included, among others, autophagy-dependent cell death; necroptosis, a mechanism dependent on MLKL and RIPK3 (kinase function); ferroptosis, initiated by severe lipid peroxidation; pyroptosis, triggered by pathogen invasion; and NETosis, where neutrophil extracellular traps (NETs) are present. ^5,7

As veterinary pathologists, we are very familiar with morphological definition of cell death, and terms such as necrosis and apoptosis are frequently included in our diagnoses. ³ However, the old dichotomy of necrosis versus apoptosis is not enough, not anymore, as new players are joining the game. In this context, we must also acknowledge the fact that the diverse forms of RCD have considerably overlapping molecular and morphological features. ⁷ What we would previously consider morphologically consistent with apoptosis might in fact result from another type of RCD. In light of this notion, the long-standing debate over the use of diagnostic terms such as “single cell necrosis” and “single cell apoptosis” (which has fueled controversies and created confusion especially in toxicological pathology settings) now appears pointless and obsolete. The existing recommendations on the use of merely descriptive labels are largely inadequate to address the complexity and diversification of the biological events leading to cell death.

This Veterinary Pathology special focus on RCD opens with a broad overview on the current paradigms of nonaccidental cell death, covering the major aspects of some widely recognized and more recently characterized mechanisms of cell death, highlighting the recent advancements in the understanding and classification of these manifestations, and summarizing their investigation using relevant animal models. ⁹

Santagostino and collaborators discuss the potential limitations of Fluoro-Jade C, a histochemical stain used in the neuropathology field. ¹⁰ While this stain has been used to demonstrate neuronal degeneration and death upon exposure to various insults, ¹¹ the actual mechanism by which Fluoro-Jade C labels degenerating neurons is unknown, thus creating some controversy to the actual pathophysiological status of the labeled cells. It appears that different stages of neuronal degeneration and underlying RCD mechanisms may influence the labeling ability of Fluoro-Jade C, supporting the claim that the investigation of neuronal death must derive from the combined interpretation of the microscopic features and any available staining markers in order to make a comprehensive and informed assessment.

Delaney and collaborators focus on the relationship between the host NLRP3 inflammasomes and Coxiella burnetii, the agent responsible for Q fever. ² Through the use of in vitro and in vivo models, the authors confirmed that C. burnetii primes but avoids cytosolic detection by NLRP3 inflammasomes and does not induce caspase-1 activation or IL-1 beta secretion following infection of macrophages. Furthermore, in vivo studies revealed similarities in the development and resolution of C. burnetii–related lesions in wild-type C57BL/6 mice and mice deficient in Casp1/11 and Nlrp3.

The open question remains, how will veterinary pathologists reconcile this emerging molecular understanding of cell death with the morphologic and diagnostic aspects of our discipline? Given that the different mechanisms of RCD have been discovered to play a role in diseases of domestic and experimental animals, the veterinary pathology community would benefit from new knowledge on different forms of RCD in relevant veterinary diseases. Given the growing interest in developing novel therapeutics to selectively interfere with specific RCD pathways and prevent or treat human diseases, information related to the interplay between distinct RCD signaling pathways and the identification of specific molecular effectors for each RCD type are needed. ¹³ Thus, available in vitro and in vivo biomarkers and functional tests will also be discussed, including genetic and pharmacological inhibition studies to assess, accurately distinguish, and quantify the putative mechanisms involved in RCD. ⁹

Overall, we hope that this special focus of Veterinary Pathology will provide information of interest to its readers, including those professionals working in toxicologic pathology, diagnostic pathology, and experimental pathology settings.