Activin A is a critical component of the inflammatory response, and its binding protein, follistatin, reduces mortality in endotoxemia

Abstract

Activin A is a member of the transforming growth factor-beta superfamily, which we have identified as having a role in inflammatory responses. We show that circulating levels of activin increase rapidly after LPS-induced challenge through activation of Toll-like receptor 4 and the key adaptor protein, MyD88. Treatment with the activin-binding protein, follistatin, alters the profiles of TNF, IL-1beta, and IL-6 after LPS stimulation, indicating that activin modulates the release of several key proinflammatory cytokines. Further, mice administered one 10-mug dose of follistatin to block activin effects have increased survival after a lethal dose of LPS, and the circulating levels of activin correlate with survival outcome. These findings demonstrate activin A's crucial role in the inflammatory response and show that blocking its actions by the use of follistatin has significant therapeutic potential to reduce the severity of inflammatory diseases.

Fig. 1.

Release of activin A, follistatin, and other cytokines in mice challenged with phenol-purified LPS. (A) Activin A showed a typical rapid but transient increase within 1 h of LPS. (B) Follistatin was significantly elevated 3 h after LPS. (C) TNF concentrations peaked sharply and were significantly elevated 30 min after LPS injection, peaking at 1 h (P < 0.01). (D) IL-6 peaked at 2 h after LPS (P < 0.01) and returned to basal levels between 5 and 8 h. (E) IL-1β slowly increased, peaked at 5 h, and then returned to basal levels. All data are mean ± SE.

Fig. 2.

Release of activin A by LPS is downstream of MyD88. Treatment of the TM4 cell line with LPS for 5 h caused significant release of activin A. Transient transfection of increasing amounts of a dominant-negative MyD88 construct with a nonfunctional TIR domain progressively blocked this activin release, confirming that the LPS stimulation occurs in the TLR4 pathway after binding to MyD88. Data are presented as mean ± SE.

Fig. 3.

Cytokine responses in mice administered 1 μg of follistatin to block the effects of activin after LPS challenge. (A) Activin A after LPS was not affected by follistatin treatment. (B) Follistatin in plasma was significantly suppressed at 5–8 h. (C) Peak plasma concentrations of TNF were significantly suppressed by blockade of activin. (D) The IL-6 response after LPS and follistatin treatment was both elevated and earlier. (E) Peak concentrations of IL-1β were ≈35% of LPS-administered animals, and the peak occurred earlier, shifting from 5 to 2 h. The equivalent response to LPS alone is depicted by a dotted line for comparison. All data are mean ± SE.

Fig. 4.

Activin A induces IL-6 release into the circulation. Injection of 1 μg of human recombinant activin A into mice results in an increase in IL-6 concentrations in the circulation within 30 min. Data are mean ± SE.

Fig. 5.

Survival of mice was increased in mice administered follistatin before a dose of LPS that causes significant shock and death. (A) Mice administered follistatin before LPS (FS, open circles) displayed an increased survival compared with LPS alone (filled circles). (B) Activin A levels in the circulation of mice treated with the lethal dose of LPS, irrespective of follistatin administration, correlate with outcome, with significantly (P < 0.001) higher concentrations in those animals that died compared with those that survived the endotoxemia. The dotted line shows that there were no overlapping values between the two outcomes.