Surfactant protein A (SP-A) and transforming growth factor beta (TGF-) 1 have been shown to modulate the functions of immune cells and specifically to inhibit T lymphocyte proliferation. The aim of the present study was to elucidate if the Smad signaling pathway, activated by TGF-1, also plays a role in SP-A mediated inhibition of CD4⁺ T lymphocytes activation. Recombinant CHO derived rSP-A1m (mammalian), but not recombinant Baculovirus derived rSP-A1^hyp (hydroxyproline-deficient), suppressed T lymphocyte proliferation and IL-2 mRNA expression. In order to test whether SP-A induced Smad signaling, a Smad3/4 specific reporter gene was transfected in CD4⁺ T lymphocytes. Only rSP-A1m, but not rSP-A1^hyp induced Smad specific reporter genes, Smad 2 phosphorylation and Smad7 mRNA expression. The effect of rSP-A1m was mediated through the TGF-RII and could be antagonized by anti-TGF-1 neutralizing antibodies and sTGF-RII. Western blot and ELISA analysis revealed that rSP-A1m, but not rSP-A1^hyp contained TGF-1. TGF-1 was responsible for the differences in inhibition of T lymphocyte proliferation and activation of the Smad signaling pathway between rSP-A1m and rSP-A1^hyp. After acidification, nSP-A, obtained from patients with alveolar proteinosis, also induced Smad signaling in T lymphocytes leading to an increased inhibition of CD4⁺ T lymphocyte proliferation, thus indicating the presence of inactive, latent TGF-1 in nSP-A samples. Association between SP-A and latent TGF-1 provides a possible novel mechanism to regulate TGF-1 mediated inflammation and fibrosis reactions in the lung, but also leads to possible misinterpretation of immune-modulator functions of SP-A. Monitoring of SP-A preparations for possible TGF-1 is essential.