Morphological features of serosa‐associated lymphoid clusters of the rat parietal pleura: Exploring a relatively unexplored system

Serosal membranes contain secondary lymphoid structures that monitor body cavity fluid and can initiate immune responses. While serosa-associated lymphoid clusters (SALCs) of the omental peritoneum (also known as milky spots) are well studied, research on pleural (p)SALCs is limited. This study aimed to examine the distribution, 3D in situ morphology, and cellular composition of SALCs in the parietal pleura in rats. By providing detailed anatomical data, this research may guide future studies on their presence in humans and for potential extrapolation of functional characteristics from rats to humans, if morphological similarities exist. The thorax of fixed Wild type Long Evans female rat cadavers was opened, followed by in toto staining with hematoxylin to improve in situ SALC visibility. Pleural segments, as previously described for rats, were allocated and subsequently stereomicroscopically studied for the presence of pSALCs to gain insight into their topographical distribution and in situ 3D morphological characteristics. To confirm that these structures indeed represent SALCs, tissue sections of each morphological subtype were microscopically studied for the presence of lymphocytes and macrophages, the primary constituents of SALCs. SALCs were observed in decreasing order in the retropericardial pleural folds (RPF), the adipose streak, pulmonary ligaments, and the ventral mediastinal pleura (VMP), whereas no SALCs were observed in pericardial, paravertebral, costal, and diaphragmatic pleura. In situ, pSALCs could be divided into four 3D morphological categories: flat, vascular, adipose, and protruding. Microscopic investigation showed that all pSALC subtypes contained T cells, B cells, and macrophages, confirming the lymphoid nature of these structures. Data provided by this study forms the basis for future morphological comparisons with humans and the potential to extrapolate functional characteristics of pSALCs from previous experimental studies in rats to humans. Our study shows that some SALC containing pleural segments are unique to rats and these interspecies differences should be considered carefully when designing future studies in humans. Furthermore, the phenomenon of distinct 3D SALC morphological subtypes requires further investigation to determine their functional implications, including any relationship between three-dimensional structures and e.g. immune activation.