Activin inhibits the human Pit-1 gene promoter through the p38 kinase pathway in a Smad-independent manner.

The pituitary transcription factor Pit-1 regulates hormonal production from the anterior pituitary gland. However, the mechanisms by which Pit-1 gene expression is regulated in humans are poorly understood. Activin, a member of the TGFbeta superfamily, acts as a negative regulator of cell growth and prolactin gene expression in lactotrope cells. In this study, we show that activin negatively regulates the human Pit-1 gene promoter. We defined a 117-bp element within the Pit-1 promoter that is sufficient to relay these inhibitory effects. We further investigated the signaling pathways that mediate activin-induced inhibition of Pit-1 gene promoter in pituitary lactotrope cells. We found that the activin effects on Pit-1 gene regulation are Smad independent and require the p38 MAPK pathway. Specifically, blocking p38 kinase activity reverses activin-mediated inhibition of the Pit-1 gene promoter. Together, our results highlight the p38 MAPK pathway as a key regulator of activin function in pituitary lactotrope cells and further emphasizes the critical role played by activin in regulating hormonal production in the pituitary gland.

Age-related decrease in high-density lipoproteins antioxidant activity is due to an alteration in the PON1's free sulfhydryl groups.

The aim of this study was to evaluate the antioxidant activity of HDL with aging and to investigate the implication of PON1 in this process. The study involved 54 healthy subjects distributed in two age groups, young (20-25 years) and elderly (65-85 years). Lipid peroxidation was induced by *OH and O2*- oxygen free radicals produced by gamma-radiolysis of water. LDL oxidation was followed by the measurement of conjugated diene (CD), lipid peroxide (LP) and malondialdehyde (MDA) formation. PON1 was purified separately from young (Y-PON1) and elderly subjects (E-PON1). PON1 activity and structure was followed by measurement of PON1 paraoxonase (p.ase) activity, titration of the SH groups, and electrophoretic mobility by SDS-PAGE. Our results show a significant decrease in the HDL antioxidant activity: percentage of protection against CD formation=27.70% (p<0.01) for E-HDL versus 73.08% (p<0.001) for Y-HDL. Moreover, E-PON1 showed a lower antioxidant activity when compared to Y-PON1 47.08% versus 78.14%, respectively (p<0.0001). Exposition of PON1 to *OH and O2*- oxygen free radicals induced a significant decrease in PON1 p.ase activity as well as a reduction in the number of PON1's free sulfhydryl groups. Moreover, our results show a close association between PON1's free sulfhydryl groups and its capacity to protect LDL against lipid peroxidation. There was a significant decrease in the number of free sulfhydryls between Y-PON1 and E-PON1 with respect to cysteine-284 amino acid residues (p<0.0092).

Activin induces hepatocyte cell growth arrest through induction of the cyclin-dependent kinase inhibitor p15INK4B and Sp1.

In this report, we examined the role of activin in the regulation of cell growth inhibition of human hepatocarcinoma cells. Using RNase protection assay for various cell cycle regulators and Western blotting experiments, we show that activin treatment of HepG2 cells leads to increased gene expression of the cyclin-dependent kinase inhibitor (CDKI) p15INK4B. Furthermore, transient co-transfection studies of the p15INK4B promoter/luciferase construct performed in HepG2 cells demonstrates that activin induction of the p15INK4B promoter is mediated through the Smad pathway. p15INK4B gene promoter mapping analysis revealed a 66-bp region within the proximal domain of the promoter, which contains a consensus site for the transcription factor Sp1, as critical for mediating the activin effect on p15INK4B gene expression. Finally, gel mobility shift experiments, using the Sp1 consensus site, revealed increased DNA binding of Sp1 in response to activin treatment of HepG2 cells, further confirming the involvement of Sp1 in activin-mediated p15INK4B gene promoter activation. Together, our data indicates an important role for the cyclin-dependent kinase inhibitor p15INK4B in activin-induced cell cycle arrest in liver cells.

Activin inhibits pituitary prolactin expression and cell growth through Smads, Pit-1 and menin.

Activin, a member of the TGFbeta superfamily, is a negative regulator of cell growth and prolactin (PRL) production in pituitary lactotrope cells. However, the mechanisms by which this growth factor exerts its growth-inhibitory and -repressive effect on PRL remain unclear. In this study, we show that activin negatively regulates PRL expression at the transcriptional level through the Smad pathway and the multiple endocrine neoplasia type 1 gene product, menin. Our results also demonstrate that the tumor suppressor menin is required for activin-induced growth arrest of somatolactotrope cells. Moreover, we show that activin represses transcription and expression of Pit-1, a pituitary transcription factor that is essential for maintenance and development of lactotrope cells. We defined two Pit-1 DNA-binding sites in the proximal region of the PRL promoter as critical for the activin-mediated inhibition. Together, our results highlight the Smad pathway and the tumor suppressor menin as key regulators of activin effects on PRL and Pit-1 expression, as well as on cell growth inhibition, and emphasize the critical role of activin in the regulation of pituitary function.