TY - JOUR
T1 - Dissociation of transforming growth factors beta 1 and beta 2 from surfactant protein A (SP-A) by deglycosylation or deoxycholate treatment
AU - Willems, Coen H. M. P.
AU - Kloosterboer, Nico
AU - Kunzmann, Steffen
AU - Kramer, Boris W.
AU - Zimmermann, Luc J. I.
AU - van Iwaarden, J. Freek
PY - 2012/1/31
Y1 - 2012/1/31
N2 - We were able to demonstrate the presence of transforming growth factor beta 1 and transforming growth factor beta 2 (TGF-beta 1,2) in human as well as porcine pulmonary surfactants and SP-A purified from these surfactants. Human SP-A contained 480 +/- 74 pg TGF-beta 1 and 61 +/- 16 pg TGF-beta 2 per mg SP-A and human pulmonary surfactant contained 140 +/- 28 pg TGF-beta 1 and 67 +/- 13 TGF-beta 2 per mg protein. Porcine SP-A contained 306 +/- 46 pg TGF-beta 1 and 43 +/- 12 pg TGF-beta 2 per mg SP-A and porcine pulmonary surfactant contained 75 +/- 18 pg TGF-beta 1 and 22 +/- 13 TGF-beta 2 per mg protein. Size-exclusion chromatography indicated binding of TGF-beta 1,2 to SP-A Deglycosylation of SP-A released TGF-beta 1,2 from SP-A indicating a role for the carbohydrate moieties of SP-A in binding of TGF-beta 1,2. TGF-beta-free SP-A was obtained by incubating SP-A with 5 mM 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-beta 1,2 separately. In addition, we demonstrated that after incubation of SP-A with TGF-beta 1,2, only a part of the added TGF-beta 1,2 can be measured, whereas after acid treatment almost all added TGF-beta 1,2 was determined, suggesting that complex formation between SP-A and TGF-beta 1,2 influences the measurements of TGF-beta 1,2 in biological samples.
AB - We were able to demonstrate the presence of transforming growth factor beta 1 and transforming growth factor beta 2 (TGF-beta 1,2) in human as well as porcine pulmonary surfactants and SP-A purified from these surfactants. Human SP-A contained 480 +/- 74 pg TGF-beta 1 and 61 +/- 16 pg TGF-beta 2 per mg SP-A and human pulmonary surfactant contained 140 +/- 28 pg TGF-beta 1 and 67 +/- 13 TGF-beta 2 per mg protein. Porcine SP-A contained 306 +/- 46 pg TGF-beta 1 and 43 +/- 12 pg TGF-beta 2 per mg SP-A and porcine pulmonary surfactant contained 75 +/- 18 pg TGF-beta 1 and 22 +/- 13 TGF-beta 2 per mg protein. Size-exclusion chromatography indicated binding of TGF-beta 1,2 to SP-A Deglycosylation of SP-A released TGF-beta 1,2 from SP-A indicating a role for the carbohydrate moieties of SP-A in binding of TGF-beta 1,2. TGF-beta-free SP-A was obtained by incubating SP-A with 5 mM 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-beta 1,2 separately. In addition, we demonstrated that after incubation of SP-A with TGF-beta 1,2, only a part of the added TGF-beta 1,2 can be measured, whereas after acid treatment almost all added TGF-beta 1,2 was determined, suggesting that complex formation between SP-A and TGF-beta 1,2 influences the measurements of TGF-beta 1,2 in biological samples.
KW - Collectin
KW - Lung
KW - Innate immunity
U2 - 10.1016/j.jim.2011.09.014
DO - 10.1016/j.jim.2011.09.014
M3 - Article
C2 - 21989137
SN - 0022-1759
VL - 375
SP - 111
EP - 117
JO - Journal of Immunological Methods
JF - Journal of Immunological Methods
IS - 1-2
ER -