Glycation products in aged thioglycollate medium enhance the elicitation of peritoneal macrophages

Thioglycollate medium (TGM) is widely used as a stimulatory agent to induce non-infectious peritoneal inflammation for elicitation of macrophages from mice and rats. It has been known for a long time that aged, autoclaved TGM is more efficient than freshly prepared TGM, however, the mechanism responsible for this enhanced activity of aged TGM remains obscure. The aging of TGM apparently favors the non-enzymatic reactions between proteins and reducing sugars in TGM that may lead to the generation of advanced glycation endproducts (AGEs). We have found that aged TGM contains 40-fold more AGEs than fresh TGM. The formation of AGEs in TGM was completely blocked by co-incubation with an AGE inhibitor, aminoguanidine. Intraperitoneal injection of aged TGM into rats elicited approximately 2-fold more macrophages than fresh TGM. However, the addition of AGE-modified proteins to fresh TGM increased the yield of peritoneal cells to a level which was significantly higher than both fresh and aged TGM. The injection of AGE-modified proteins alone did not elicit significantly more macrophages than the level of resident peritoneal cells. These results suggest that the formation of AGEs during aging of TGM is responsible for the enhanced macrophage-eliciting activity in aged TGM. AGEs may act as an enhancing agent to augment the existing inflammatory responses. AGE-supplemented TGM may provide an efficient method for eliciting peritoneal macrophages or establishing an inflammatory animal model.