Updated project metadata. A central and unique aspect of high-grade serous ovarian carcinoma (HGSC) is the extensive transcoelomic spreading of tumor cell via the peritoneal fluid or malignant ascites. We and others identified tumor-associated macrophages (TAM) in the ascites as promoters of metastasis-associated processes like extracellular matrix (ECM) remodeling, tumor cell migration, adhesion and invasion. The precise mechanisms and mediators involved in these functions of TAM are, however, largely unknown. By comparing monocyte-derived macrophages (MDM) differentiated in ascites to TAM-like asc-MDM, by LPS/IFN to inflammatory m1-MDM and by IL-10 to alternatively activated m2c-MDM, we found that conditioned media from both, asc-MDM and m2c-MDM, stimulated migration of patient-derived tumor cells, while m1-MDM failed to do so. Secretome analysis by mass spectrometry identified an overlapping set of 9 proteins secreted by both asc-MDM and m2c-MDM, but not by m1-MDM, in all tested donors. Of these, three proteins, namely transforming growth factor beta induced protein (TGFBI), tenascin C (TNC) and fibronectin (FN1), have been associated with migration-related functions. Intriguingly, increased ascites concentrations of TGFBI, TNC and fibronectin were associated with short progression-free survival. Furthermore, transcriptome and secretome analyses point to TAM as major producers of these proteins, further supporting an essential role for TAM in promoting HGSC progression. Consistent with this hypothesis, we were able to demonstrate that the migration-inducing potential of asc-MDM and m2c-MDM secretomes may be inhibited, at least partially, by neutralizing antibodies against TGFBI and TNC or siRNA-mediated silencing of TGFBI expression. In conclusion, the present study provides the first experimental evidence that TAM-derived TGFBI and TNC in ascites promote HGSC progression.