PDGF-driven proliferation, migration, and IL8 chemokine secretion in human corneal fibroblasts involve JAK2-STAT3 signaling pathway.
OBJECTIVE
Platelet-derived growth factor (PDGF) is associated with corneal fibroblast migration and proliferation and plays an important role in corneal wound healing. However, the intracellular mechanisms of PDGF-mediated functions in corneal fibroblasts are poorly understood. We tested the hypothesis that PDGF functional activities in the cornea involve the Janus kinase-2/signal transducers and activators of transcription-3 (JAK2-STAT3) signaling pathway and whether PDGF induces the expression of suppressors of cytokine signaling 3 (SOCS3), belonging to the novel family of feedback regulators of cytokine and growth factor activities.


METHODS
Human corneal fibroblast (HSF) cultures were used as an in vitro model for functional analysis. Real-time polymerase chain reactions were performed to quantify gene expression. Immunoprecipitation and immunoblotting techniques were used to measure protein expression. Cell growth, migration, and ELISA assays were used for functional validation.


RESULTS
Low endogenous levels of STAT3 and SOCS3 mRNA and protein expression were noted in HSFs. PDGF treatment of HSF significantly induced SOCS3 mRNA (3.0-4.5 fold) and protein (1.5-2.5 fold) expression in a time-dependent manner. Similarly, PDGF treatment of HSF significantly increased STAT3 protein expression at two tested time points (2.5-2.96 fold). Cultures exposed to vehicle (control) did not show any change in SOCS3 and STAT3 mRNA or protein expression. An addition of AG-490, a selective inhibitor of the JAK2-STAT3 pathway, significantly inhibited PDGF-mediated STAT3 induction and cell growth and migration in HSF. We also observed that PDGF induced interleukin-8 (IL8) chemokine secretion (2 fold) and AG-490 inhibited IL8 secretion.


CONCLUSIONS
Our data showed that PDGF induced STAT3, SOCS3, and IL8 chemokine secretion in human corneal fibroblasts. Further, PDGF-induced cell growth, migration, and IL8 secretion in corneal fibroblast involve the JAK2-STAT3 signaling pathway.
Vij
Neeraj
N
Mason Eye Institute, School of Medicine, University of Missouri-Columbia, Columbia, MO 65212, USA.
Sharma
Ajay
A
Thakkar
Mahesh
M
Sinha
Sunilima
S
Mohan
Rajiv R
RR
D002465:Cell Movement / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D003315:Cornea / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D006801:Humans; D016209:Interleukin-8 / Q000378:metabolism*; D053614:Janus Kinase 2 / Q000378:metabolism*; D008954:Models, Biological; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology*; D050796:STAT3 Transcription Factor / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D000071223:Suppressor of Cytokine Signaling 3 Protein; D050826:Suppressor of Cytokine Signaling Proteins / Q000378:metabolism
D016209:Interleukin-8; D010982:Platelet-Derived Growth Factor; C106644:SOCS3 protein, human; D050796:STAT3 Transcription Factor; D000071223:Suppressor of Cytokine Signaling 3 Protein; D050826:Suppressor of Cytokine Signaling Proteins; D053614:Janus Kinase 2