Discussion
Results highlight the immune-modulatory impact of AAT on the complement system, involving an important potential role for complement activation in disease pathogenesis in AATD. The association between plasma C3d levels and pulmonary disease severity, that decrease in response to AAT augmentation therapy, supports the exploration of C3d as a candidate biomarker of therapeutic efficacy in AATD.
Methods
To isolate AAT linker proteins, plasma samples were separated by size exclusion chromatography, followed by co-immunoprecipitation. AAT binding proteins were identified by mass spectrometry. Complement turnover and activation was determined by ELISA measurement of C3, C3a and C3d levels in plasma of healthy controls (n=15), AATD (n=51), non-AATD patients with obstructive airway disease (n=10) and AATD patients post AAT augmentation therapy (n=5).
Results
Direct binding of complement C3 to AAT was identified in vivo and in vitro. Compared with healthy controls, a breakdown product of C3, C3d, was increased in AATD (0.04 µg/mL vs 1.96 µg/mL, p=0.0002), with a significant correlation between radiographic pulmonary emphysema and plasma levels of C3d (R2=0.37, p=0.001). In vivo, AAT augmentation therapy significantly reduced plasma levels of C3d in comparison to patients not receiving AAT therapy (0.15 µg/mL vs 2.18 µg/mL, respectively, p=0.001).