Histochem Cell Biol DOI 10.1007/s00418-008-0553-1 123 ORIGINAL PAPER Metallophilic macrophages are fully developed in the thymus of autoimmune regulator (Aire)-deWcient mice Novica M. MiliTeviT · Livana MiliTeviT · Milon D. MiljkoviT · Milica LabudoviT-BoroviT · Martti Laan · Pärt Peterson · Kai Kisand · Hamish S. Scott · Ning Qu · Jürgen Westermann Accepted: 23 December 2008  Springer-Verlag 2009 Abstract Thymic metallophilic macrophages represent a signiWcant component in the thymus physiology. Recently, we showed their presence to be dependent on functional lymphotoxin- receptor (LTR) signaling pathway. However, it is unknown whether the development of metallophilic macrophages also requires the Autoimmune regulator (Aire) transcription factor, as suggested by some studies for medullary thymic epithelial cells, or perhaps the presence of Aire-expressing thymic epithelial cells them- selves. Therefore, we investigated the presence of metallo- philic macrophages in Aire-deWcient thymus. Our study shows that the metallophilic macrophages are fully devel- oped in the Aire-deWcient thymus; their development is not regulated via Aire transcription factor and does not require the presence of Aire-expressing epithelial cells. On the con- trary, in alymphoplasia (ALY) mice (deWcient in nuclear factor-kappaB-inducing kinase, NIK), which we used as negative control, thymic metallophilic macrophages are completely lacking, similarly as in LTR-deWcient animals. Together, these results show that the development/mainte- nance of thymic metallophilic macrophages is executed via LTR circumventing the Aire transcription factor. Thus, we shed a new light on the molecular requirements for development of these cells and also show that LTR path- way is a common developmental regulator of metallophilic macrophages in diVerent lymphatic organs (i.e., thymus and spleen). Keywords Autoimmune regulator · Thymus · Macrophages · Nuclear factor-kappaB-inducing kinase · Alymphoplastic ALY mice Introduction T lymphocytes selected for export from the thymus into the peripheral lymphoid organs have to be tolerant to self tis- sue-speciWc antigens (TSAs) to avoid the autoimmunity. This is for the most part achieved by the process of negative selection that aVects thymocytes during migration through the thymic medulla (Anderson et al. 2002; Derbinski et al. 2001; Sprent and Kishimoto 2002). In recent years, the knowledge on the process of negative selection has been dramatically advanced. BrieXy, it has been shown that med- ullary thymic epithelial cells (mTEC) are capable of expressing a wide variety of genes and synthesize a large number of TSAs under the control of the transcription regu- lator Aire (Autoimmune regulator) (Anderson et al. 2002; N. M. MiliTeviT (&) · L. MiliTeviT · M. D. MiljkoviT · M. LabudoviT-BoroviT Institute of Histology and Embryology, University of Belgrade Medical School, Belgrade, Serbia e-mail: emilicen@etf.bg.ac.yu M. Laan · P. Peterson · K. Kisand Molecular Pathology, Institute of General and Molecular Pathology, Tartu University, Tartu, Estonia H. S. Scott Division of Molecular Pathology, Institute of Medical and Veterinary Science and The Hanson Institute, and School of Medicine, University of Adelaide, Adelaide, Australia N. Qu Department of Anatomy, Tokyo Medical University, Tokyo, Japan J. Westermann Center for Structural and Cell Biology in Medicine, Institute of Anatomy, University of Lübeck, Lübeck, Germany