The cardiovasculature is the first functional system in the developing embryo and as such, it plays a crucial role in the proper nourishment and formation of all other body regions and organs. A detailed understanding the mechanisms that regulate cardiac morphogenesis is necessary to develop possible strategies for diagnostics and treatment of cardiovascular diseases.One molecule identified as important for the proper formation of the heart is Hyaluronan synthase 2 (Has2), a membrane protein in charge of assembling the glycosaminoglycan hyaluronan (HA). Mouse embryos lacking Has2 do not produce HA, display severe cardiovascular abnormalities and die during early embryogenesis. Thus, Has2 and HA are necessary for the early stages of heart formation, but many questions remain to be answered in regards to their mechanism of action and their role in later events such as the formation of the coronary vessels. Our current study addresses these questions employing two cell lines: NIH-3T3s, as a model of mesenchymal endocardial cushion cells; and epicardial cells, which have an epithelial phenotype.Here we show that HA induces biological activity in embryonic cells in a manner that is dependent on its molecular size. High molecular weight HA (HMW-HA), but not low molecular weight HA (LMW-HA), induces invasion of NIH-3T3 cells while it promotes differentiation and invasion of epicardial cells. We also demonstrate that stimulation of cells with HMW-HA promotes the association of MEKK1 with the HA receptor CD44. This leads to the activation of two distinct pathways, one ERK-dependent and another NFκB-dependent, which are crucial for the induction of cellular responsesFinally, we have demonstrated that the growth factors TGFβ2 and EGF induce Has2 expression and/or phosphorylation. TGFβ2 governs Has2 via MEKK3-dependent mechanisms, while EGF does not require MEKK3. Increased Has2 activity as a result of TGFβ2 and EGF stimulation leads to enhanced HA synthesis. These increased levels of HA are coincident with enhanced cellular differentiation and invasion. Taken together, these findings underscore how EGF, TGFβ2 and HA signals are integrated to form highly complex networks that are crucial for the proper formation of organs and tissues during development.