Dataset Information

Role of SH2-containing tyrosine phosphatase Shp2 in mouse corneal epithelial stratification.

ABSTRACT:

Purpose

Shp2 protein tyrosine phosphatase mediates a wide variety of receptor tyrosine kinases (RTK) cell signaling. Herein, we investigate the role of Shp2 in corneal morphogenesis and homeostasis.

Methods

Shp2 was conditionally knocked out (Shp2(cko)) in Krt14-rtTA;tet-O-Cre;Shp2(f/f) triple transgenic mice administrated with doxycycline (Dox) from postnatal day 1 (P1) to P10, P15, and P25, respectively. In addition, corneal epithelial debridement was performed in adult (P42) mice treated with or without Dox for 8 days (from P42-P50). Mouse eyes were then subjected to histology and immunohistochemistry.

Results

Shp2(cko) revealed impaired stratification of conjunctival and corneal epithelia during morphogenesis. Likewise, Shp2(cko) failed to restore epithelial stratification after a corneal epithelial wound in adult Shp2(cko). At the cellular level, the ratio of proliferating cell nuclear antigen (PCNA-positive)/total basal cells remained unchanged, but cells in G2/M (survivin-positive) phase was significantly increased in Shp2(cko) as compared with those in the control littermate. Interestingly, deltaN-p63 (?Np63) expression and the asymmetric division of the basal cells were coincidentally dampened in Shp2(cko). Transmission electron microscopic study showed that desmosome and hemidesmosome densities were reduced in the corneal epithelium of Shp2(cko). Immunohistochemistry also demonstrated that expression of E-cadherin/?-catenin junction and laminin-?1 was extensively downregulated in Shp2(cko). On the other hand, corneal epithelium lacking Shp2 remained positive for K14, Pax-6, and keratin 12 (K12), suggesting that Shp2 was dispensable for the corneal epithelial-type differentiation.

Conclusions

These data argued that Shp2 deficiency predominantly impacted p63-dependent cell division and cell adhesive ability, which resulted in the impairment of stratification during corneal epithelial development and wound healing.

SUBMITTER: Ng GY

PROVIDER: S-EPMC3850669 | biostudies-literature | 2013 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:BackgroundWe have previously shown that SHP2 downregulation may predispose fibroblasts to differentiate into myofibroblasts and proposed a role for SHP2 downregulation in the pathogenesis of idiopathic pulmonary fibrosis (IPF). Recent data have shown that SHP2 localizes to the mitochondrial intercristae, and its overexpression enhances mitochondrial metabolism leading to oxidative stress and senescence.ObjectiveTo determine the effect of SHP2 on fibrotic responses.Methods and resultsPrimary mouse lung fibroblasts derived from mice carrying a conditional knock-in mutation (D61G/+), rendering the SHP2 catalytic domain constitutively active, had reduced proliferation (1.6-fold, p < 0.05), migration (2-fold, p < 0.05), as well as reduced responsiveness of TGFB-1 induced fibroblasts-to-myofibroblasts differentiation, compared to wild-type ones. Electron microscope analysis revealed that SHP2 D61G/+ mouse lung fibroblasts were characterized by mitochondrial abnormalities, including swollen mitochondria with disrupted electron-lucent cristae and an increased number of autophagosomes compared to wild-type ones. SHP2 D61G/+ MLFs exhibited increased protein levels of autophagy markers, including LC3B-II and p-62, evidence that was confirmed by immunofluorescence analysis. Mitochondrial function analysis revealed that stable (genotype D61G/+) overexpression of SHP2 led to impaired mitochondrial function, as assessed by decreased mitochondrial membrane potential (1.29-fold, p < 0.05), coupling efficiency (1.82 fold, p < 0.05), oxygen consumption rate (1.9-fold, p < 0.05), and increased reactive oxygen species production both at baseline (1.75-fold, p < 0.05) and following H2O2 stimulation (1.63-fold, p < 0.05) compared to wild-type ones (SHP2+/+). SHP2 D61G/+ mouse lung fibroblasts showed enhanced AMPK activity, as well as decreased activation of the mTORC1 signaling pathway, potentially leading to ineffective mitochondrial metabolism and increased autophagy.ConclusionsSHP2 attenuates fibrotic responses in fibroblast cell lines through negative regulation of mitochondrial metabolism and induction of autophagy. SHP2 activation may represent a promising therapeutic strategy for patients with fibrotic lung diseases.