PI3K/AKT pathway mediates induction of IL-1RA by TSH in fibrocytes: modulation by PTEN.
BACKGROUND
TSH provokes expression of inflammatory genes in CD34(+) fibrocytes. These cells appear to infiltrate the orbit in Graves' disease (GD), where they putatively become the CD34(+) orbital fibroblast subset (GD-OF). This may have importance in solving the pathogenesis of thyroid-associated ophthalmopathy. The IL-1 family is targeted by TSH in fibrocytes and OFs by inducing secreted IL-1 receptor antagonist (IL-1RA) and intracellular IL-1RA in a cell-specific pattern. Phosphoinositide 3-kinase (PI3K) mediates several TSH actions in thyroid. This pathway is modulated by phosphatase and tensin homolog deleted on chromosome 10 (PTEN). Vanishingly little is known currently about TSHR signaling to IL-1RA expression in nonthyroidal cells. Furthermore, factors modulating TSH action in these cells are largely unexplored.


OBJECTIVE
To characterize intermediate signaling between TSHR and IL-1RA in fibrocytes and GD-OFs and to begin to identify the proximate regulators of TSHR signaling in nonepithelial, extrathyroidal cells as a strategy for developing therapies for thyroid-associated ophthalmopathy.


METHODS
Fibrocytes and GD-OFs were collected and analyzed from healthy individuals and those with GD in an academic clinical practice.


METHODS
Real-time PCR, Western blot analysis, cell transfections, and chromatin immunoprecipitation analysis.


RESULTS
TSH induces IL-1RA in fibrocytes and GD-OFs by activating the PI3K/AKT pathway. Interrupting either PI3K or AKT with small molecule inhibitors or by knocking down their expression with targeting small interfering RNA attenuates the actions of TSH. OFs exhibit greater basal PTEN activity and lower constitutive AKT phosphorylation than do fibrocytes. Patterns of PTEN induction diverge in the two cell types.


CONCLUSIONS
The current findings identify the PI3K/AKT pathway as critical to the induction by TSH of IL-1RA in fibrocytes and GD-OFs. Furthermore, PTEN modulates the amplitude of the induction. In GD-OFs, relatively high basal PTEN levels prevent secreted IL-1RA expression or release. Knocking down PTEN allows GD-OFs to exhibit a pattern of IL-1RA expression resembling fibrocytes.
Departments of Ophthalmology and Visual Sciences (B.L., T.J.S.) and Internal Medicine (T.J.S.), Division of Metabolism, Endocrinology, and Diabetes, University of Michigan Medical School, Ann Arbor, Michigan 48105.
Bin
B
Li
Terry J
TJ
Smith
D002478:Cells, Cultured; D005347:Fibroblasts / Q000378:metabolism; D015870:Gene Expression / Q000502:physiology; D055785:Gene Knockdown Techniques; D006111:Graves Disease / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D053590:Interleukin 1 Receptor Antagonist Protein / Q000235:genetics / Q000378:metabolism*; D015850:Interleukin-6 / Q000378:metabolism; D009915:Orbit / Q000166:cytology; D051059:PTEN Phosphohydrolase / Q000235:genetics / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation / Q000502:physiology; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology; D013972:Thyrotropin / Q000378:metabolism*
C507763:IL1RN protein, human; C508600:IL6 protein, human; D053590:Interleukin 1 Receptor Antagonist Protein; D015850:Interleukin-6; D013972:Thyrotropin; D019869:Phosphatidylinositol 3-Kinases; C494918:AKT1 protein, human; D051057:Proto-Oncogene Proteins c-akt; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human