Dissociation of transforming growth factors β1 and β2 from surfactant protein A (SP-A) by deglycosylation or deoxycholate treatment

We were able to demonstrate the presence of transforming growth factor β1 and transforming growth factor β2 (TGF-β1,2) in human as well as porcine pulmonary surfactants and SP-A purified from these surfactants. Human SP-A contained 480±74pg TGF-β1 and 61±16pg TGF-β2 per mg SP-A and human pulmonary surfactant contained 140±28pg TGF-β1 and 67±13 TGF-β2 per mg protein. Porcine SP-A contained 306±46pg TGF-β1 and 43±12pg TGF-β2 per mg SP-A and porcine pulmonary surfactant contained 75±18pg TGF-β1 and 22±13 TGF-β2 per mg protein. Size-exclusion chromatography indicated binding of TGF-β1,2 to SP-A. Deglycosylation of SP-A released TGF-β1,2 from SP-A indicating a role for the carbohydrate moieties of SP-A in binding of TGF-β1,2. TGF-β-free SP-A was obtained by incubating SP-A with 5mM deoxycholate at pH 9.2 followed by size-exclusion chromatography, a protocol which can be used to study the biological activities of SP-A and TGF-β1,2 separately. In addition, we demonstrated that after incubation of SP-A with TGF-β1,2, only a part of the added TGF-β1,2 can be measured, whereas after acid treatment almost all added TGF-β1,2 was determined, suggesting that complex formation between SP-A and TGF-β1,2 influences the measurements of TGF-β1,2 in biological samples. ► The presence of TGF-β1,2 was demonstrated in human and porcine pulmonary surfactant and SP-A. ► Binding of TGF-β1,2 to the carbohydrate moieties of SP-A was shown. ► A protocol is described to isolate TGF-β1,2 free intact SP-A. ► Binding of TGF-β1,2 to SP-A influences the measurement of TGF-β1,2.