Characterization of Subpopulations of Chicken Mononuclear Phagocytes That Express TIM4 and CSF1R

Abstract

The phosphatidylserine receptor, TIM4, encoded by TIMD4, mediates the phagocytic uptake of apoptotic cells. We applied anti-chicken TIM4 monoclonal antibodies, in combination with CSF1R reporter transgenes to dissect the function of TIM4 in the chick (Gallus gallus). During development in ovo, TIM4 was present on the large majority of macrophages but expression became more heterogeneous post-hatch. Blood monocytes expressed KUL01, class II MHC and CSF1R-mApple uniformly. Around 50% of monocytes were positive for surface TIM4. They also expressed many other monocyte-specific transcripts at a higher level than TIM4- monocytes. In liver, highly-phagocytic TIM4hi cells shared many transcripts with mammalian Kupffer cells and were associated with uptake of apoptotic cells. Although they expressed CSF1R mRNA, Kupffer cells did not express the CSF1R-mApple transgene, suggesting that additional CSF1R transcriptional regulatory elements are required by these cells. By contrast, CSF1R-mApple was detected in liver TIM4lo and TIM4- cells which were not phagocytic and were more abundant than Kupffer cells. These cells expressed CSF1R, alongside high levels of FLT3, MHCII, XCR1 and other markers associated with conventional dendritic cells (cDC) in mice. In bursa, TIM4 was present on the cell surface of two populations. Like Kupffer cells, bursal TIM4hi phagocytes co-expressed many receptors involved in apoptotic cell recognition. TIM4lo cells appear to be a sub-population of bursal B cells. In overview, TIM4 is associated with phagocytes that eliminate apoptotic cells in the chick. In the liver, TIM4 and CSF1R reporters distinguished Kupffer cells from an abundant population of DC-like cells.

Characterization of splenic MRC1hiMHCIIlo and MRC1loMHCIIhi cells from the monocyte/macrophage lineage of White Leghorn chickens

“…Several receptors, such as T cell membrane protein 4 (TIM4) [48], receptor tyrosine kinase Mer (MerTK) [49], and mannose receptor [50], are known for their important role in the recognition and phagocytosis of apoptotic cells and pathogens in mice. Similarly, the role of TIM4 in phagocytosing apoptotic cells has been reported in chickens [18]. Therefore, the higher phagocytic activity of MRC1 hi MHCII lo cells compared to that of MRC1 lo MHCII hi cells is likely linked to the expression level of MRC1, a chicken mannose receptor [14].…”

“…Chicken monocyte/macrophage lineage cells expressing MRC1 have been found to exhibit features similar to those in mammals, including morphological features [15], plasticity [16], and capability for phagocytosis [17,18]. Furthermore, they express several TLRs, and the activation of TLRs by their ligands induces bacterial lysis [19] and pro-inflammatory cytokine secretion [20], suggesting that they induce a similar response during inflammation compared to that of mammals.…”

A transient increase in MHC-IIlow monocytes after experimental infection with Avibacterium paragallinarum (serovar B-1) in SPF chickens

“…Interestingly, MHC-II low monocyte population is increased under inflammatory conditions induced by intraperitoneal LPS injection [ 42 ]. On the other hand, Hu and collaborators also demonstrated the monocytes heterogeneity and separated blood monocytes into two subpopulations based on the expression of TIM4 (a receptor that binds to phosphatidylserine) [ 19 ]. It would be of great interest to determine whether MHC-II downregulation occurs in both subpopulations or if it is restricted to one population.…”

“…Moreover, studies in chickens (also demonstrated in humans and mice) have shown that in response to a biological stimulus, macrophages and other cells of the immune system secrete pro- and anti-inflammatory cytokines that are released into the circulatory system to elicit a systemic immune response [ 13 – 16 ]. One of the target populations of these cytokines is monocytes, described as a heterogeneous population in the avian system, which have chemotactic and phagocytic activities and are capable of generating a respiratory burst [ 17 – 19 ]. During infectious or severe inflammatory insult in humans and mice, this population has an important role in compensatory anti-inflammatory response syndrome (CARS), a biological process that is necessary to prevent overwhelming inflammation and avoid organ failure [ 20 , 21 ].…”

Angiogenesis-Inflammation Cross Talk in Diabetic Retinopathy: Novel Insights From the Chick Embryo Chorioallantoic Membrane/Human Vitreous Platform

“…In chickens, T-cell membrane protein 4 (TIM4) is a receptor expressed primarily by macrophages, binds to phosphatidylserine, and most likely participates in the recognition and clearance of apoptotic cells ( 89 ). Hu and colleagues applied anti-chicken TIM4 monoclonal antibodies in combination with colony stimulating factor 1 receptor reporter transgenes to dissect the function of TIM4 in the chick ( 90 ). They demonstrated that TIM4 was present on the large majority of macrophages during development in ovo and to be expressed also by other cells with phagocytic activity, such as dendritic cells, after hatching ( 90 ).…”