Involvement of nuclear factor kappa B in the regulation of rat luteal function: potential roles as survival factor and inhibitor of 20alpha-hydroxysteroid dehydrogenase

Abstract

Nuclear factor kappa B (NF B) is an important intracellular conveyor of extracellular signals and modulates a number of gene responses. Due to the potential significance of NF B in regulating ovarian gene expression, we examined in the rat: (i) whether NF B is activated and developmentally regulated in the corpus luteum (CL) throughout pregnancy; (ii) the proteins forming the NF B complex in luteal cells; and (iii) the role of this transcription factor in luteal function. Western analysis and immunohistochemistry revealed that p65 and p50 were highly expressed throughout pregnancy and were located in both the nucleus and cytoplasm of luteal cells. In addition, because NF B is maintained in the cytoplasm bound to I B, whose phosphorylation allows NF B translocation to the nucleus, we studied the developmental expression of phosphorylated and nonphosphorylated forms of I B . Western analysis revealed that I B was present and phosphorylated throughout pregnancy in the CL whereas by protein/DNA array and electromobility shift assays we found that luteal nuclear extracts bind to an NF B consensus sequence, and that the binding activity decreased along pregnancy. The specific binding was supershifted only by an anti-p65 antibody and not by antibodies against p50, p52, cRel, or RelB. Using day 4 postpartum ovaries, we found higher NF B binding activity in the newly formed CL than in old CL of pregnancy. Furthermore, NF B DNA binding activity was enhanced by prolactin in luteinized granulosa cells. In our first functional study, blockade of NF B/p65 binding to DNA with the sesquiterpene lactone helenalin in luteinized granulosa cells correlated with induction of cell death in a dose-dependent manner. In a second functional study, overexpression of NF B/p65 in luteal cells resulted in inhibition of 20 -hydroxysteroid dehydrogenase (20 HSD) promoter activity as well as endogenous 20 HSD mRNA expression. In summary, we have shown that: (i) NF B is expressed within the CL, primary luteinized granulosa cells, and a rat luteal cell line; (ii) NF B activation within the CL is developmentally regulated in pregnancy, depends on the age of the gland, and can be upregulated by prolactin; (iii) inhibition of NF B/p65 binding to an NF B DNA consensus sequence correlates with induction of cell death in ovarian luteinized granulosa cells; and (iv) overexpression of NF B in luteal cells inhibits 20 HSD gene expression. The results further support a role for NF B as a survival factor in the CL.

NF-κB p65 repression by the sesquiterpene lactone, Helenalin, contributes to the induction of autophagy cell death

“…In addition, helenalin interacts with RelA to inhibit DNA binding to its cognate response elements and by inhibiting activation of the transcription factor NF-κB [7]. Blockade of NF-κB/p65 binding to DNA with helenalin correlated with induction of cell death in a dose-dependent manner [14]. Numerous reports have demonstrated NF-κB p65 to play a role in autophagy induced cell death [13,15], however the function in which helenalin participated in autophagy was unknown.…”

Activator protein 1‐mediated expression of monocyte chemoattractant protein 1 in cultured rat luteal cells

“…First, why is the transcription of mcp-1 gene not induced before culturing in luteal cells where NF-kB is already active? Other investigators have also reported the presence of active NF-kB in rat luteal cells (Telleria et al, 2004). A likely explanation for this is that NF-kB is necessary but not sufficient for the activation of JNK.…”

Activation of NF‐κB signaling pathway during HCG‐induced VEGF expression in luteal cells

“…Furthermore, NF‐κB is closed with the inflammatory response through interleukin (IL)‐1β production (Wu et al, ; Yang et al, ), including ovarian grannulosa cells (Yang et al, ). Some reports demonstrated NF‐κB was expressed in human granulosa luteal cells and activated by TNF‐α, resulting in degradation of IκBalpha and mobilization of NF‐κB into the nucleus (Gonzalez‐Navarrete et al, ), and NF‐κB was also involved in the regulation of rat luteal function as a survival factor and inhibitor of 20alpha‐hydroxysteroid dehydrogenase (Telleria et al, ). Interestingly, NF‐κB was implicated for angiogenesis of corpus luteum and ovarian hyperstimulation syndrome (Chen et al, ), which was consistent with the role of HIF‐1α signaling (Zhang et al, , b, ).…”