For many years the placental cytotrophoblasts obtained by disaggregation of placental tissue have served as the most widely investigated model for the study of placental function in vitro. However, the purity of cytotrophoblasts was not consistent. The IEVT cells used in the study exhibit similar cellular function to normal EVT cells  and they provide a promising model to study the function of EVT cells. Using these cells, we have demonstrated that activin-A stimulates, while inhibin-A inhibits, ActRI mRNA levels.
Our results showed that ActRI mRNA accumulation at 12 hours after treatment was stimulated in a dose-dependent manner by activin-A. The time course of the activin-A effect on ActRI mRNA showed responses at 3 to 12 hours after treatment, with a maximal increase occurring at 6 hours. This response pattern of activin-A on ActRI mRNA suggests that activin-A exerts a positive feedback effect on its own receptor and that the action of activin-A on ActRI mRNA level is transient. The transient effect of activin-A is likely due to the termination of activin signaling. Activin signaling has been shown to be terminated by negative feedback mechanisms involving the activation of Smad7 by activin, and in turn, Smad7 inhibits further activin signaling by blocking the phosphorylation of Smad2/3 by activin type I receptors [28, 29]. In addition, activin signaling can also be terminated by the degradation of Smad proteins through the ubiquitination pathway .
The changes in ActRI mRNA levels as determined by cPCR in this study could have resulted from changes in the transcriptional rate of ActRI gene and/or the stability of the ActRI mRNA. Activin is known to regulate gene expression at the level of transcription via the Smad signaling pathway [5–7] and expression of Smads in trophoblast cells including the HTR8/SVsno have been demonstrated [31, 32], it is therefore possible that activin-A may regulate its receptor expression at the level of gene transcription. Future studies are also needed to confirm that there is a correlated change at the protein level.
Although this is the first study demonstrating that activin-A regulates its own receptor expression in human placenta, several studies in other tissues have also observed modulation of activin signaling by activin itself. In the rat pituitary, activin-A stimulated ActRI and ActRIIB mRNA levels without altering ActRII mRNA expression . On the other hand, activin-A downregulated ActRIB, ActRII and ActRIIB mRNA levels in a cell line derived from a p53(-/-)inhibin-α (-/-) mouse testicular tumor .
Follistatin is widely distributed and produced by many activin-responsive tissues [1, 25] and may, therefore, serve to anatomically and temporally limit the local activities of activin. Follistatin mRNA transcripts and immunoreactivities have been detected in placental trophoblast cells  Similar to many other systems [24, 25], follistatin also neutralizes activin actions in human placenta . In the present study, we also found that the stimulatory effect of activin-A on ActRI mRNA levels could be completely blocked by follistatin. Since it is known that follistatin inhibits activin actions by binding to activin and thus preventing the interaction between activin and its type II receptor  the neutralization of activin-A action on ActRI mRNA levels by follistatin suggests that the action of activin is a receptor-mediated event.
Inhibin and activin possess opposing activities in several biological systems including pituitary FSH secretion, erythroid differentiation, and gonadal sex-steroid production . In the present study, we found that inhibin-A caused a 60 % reduction in basal ActRI mRNA levels and completely inhibited activin-A-induced ActRI mRNA expression. This finding is in agreement with the observation that inhibin counteracts the effects of activin on cultured first trimester trophoblast cells .
In summary, we have demonstrated that in activin-A upregulates mRNA levels of it own receptor ActRI in human extravillous trophoblast cells. Since activin-A has been shown to stimulate the differentiation of extravillous trophoblast cells , the up-regulation of ActRI by activin-A suggests a positive feedback regulatory mechanism by which activin-A modulates its own signaling in these cells.