Dataset Information

Corneal Wound Healing Requires IKB kinase ? Signaling in Keratocytes.

ABSTRACT: IkB kinase ? (IKK?) is a key signaling kinase for inflammatory responses, but it also plays diverse cell type-specific roles that are not yet fully understood. Here we investigated the role of IKK? in the cornea using Ikk?(?CS) mice in which the Ikk? gene was specifically deleted in the corneal stromal keratocytes. The Ikk?(?CS) corneas had normal morphology, transparency and thickness; however, they did not heal well from mild alkali burn injury. In contrast to the Ikk?(F/F) corneas that restored transparency in 2 weeks after injury, over 50% of the Ikk?(?CS) corneas failed to fully recover. They instead developed recurrent haze with increased stromal thickness, severe inflammation and apoptosis. This pathogenesis correlated with sustained myofibroblast transformation with increased ? smooth muscle actin (?-SMA) expression, higher levels of senescence ?-Gal activity and scar tissue formation at the late stage of wound healing. In addition, the Ikk?(?CS) corneas displayed elevated expression of hemo-oxygenase-1 (HO-1), a marker of oxidative stress, and activation of stress signaling pathways with increased JNK, c-Jun and SMAD2/3 phosphorylation. These data suggest that IKK? in keratocytes is required to repress oxidative stress and attenuate fibrogenesis and senescence in corneal wound healing.

SUBMITTER: Chen L

PROVIDER: S-EPMC4795706 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Corneal Wound Healing Requires IKB kinase β Signaling in Keratocytes.

Chen Liang L Mongan Maureen M Meng Qinghang Q Wang Qin Q Kao Winston W Xia Ying Y

PloS one 20160317 3

IkB kinase β (IKKβ) is a key signaling kinase for inflammatory responses, but it also plays diverse cell type-specific roles that are not yet fully understood. Here we investigated the role of IKKβ in the cornea using Ikkβ(ΔCS) mice in which the Ikkβ gene was specifically deleted in the corneal stromal keratocytes. The Ikkβ(ΔCS) corneas had normal morphology, transparency and thickness; however, they did not heal well from mild alkali burn injury. In contrast to the Ikkβ(F/F) corneas that restor ...[more]

PMID: 26987064

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Project description:Acellular cornea derived hydrogels provide significant advantages in preserving native corneal stromal keratocyte cells and endothelial cells. However, for clinical application, hydrogel physical properties need to be improved, and their role in corneal epithelial wound healing requires further investigation. In this study, an acellular porcine corneal stroma (APCS) hydrogel (APCS-gel) was successfully prepared from 20 mg/ml APCS, demonstrated optimal light transmittance and gelation kinetic properties and retained critical corneal ECM of collagens and growth factors. Compared with fibrin gel, the APCS-gel had a higher porosity ratio and faster nutrition diffusion with an accompanying improvement in the proliferation of primary rabbit corneal epithelial cells (RCECs) and stromal cells (RCSCs). These corneal cell types also displayed improved viability and cellular infiltration. Furthermore, the APCS-gel provides significant advantages in the preservation of RCECs stemness and enhancement of corneal wound healing in vitro and in vivo. After 7 days of culture, 3-4 layers of RCECs were formed on the APCS-gel in vitro, while only 1-2 layers were found on the fibrin gel. More corneal stem/progenitor cell phenotypes (K12-, p63+, ABCG2+) and proliferation phenotypes (Ki67+) were detected on the APCS-gel than fibrin gel. Using a corneal epithelial wound healing model, we also found faster reepithelization in corneas that received APCS-gel compared to fibrin gel. Additionally, our APCS-gel demonstrated better physical and biological properties when compared to Tisseel, a clinically used type of fibrin gel. In conclusion, our APCS-gel provided better corneal epithelial and stromal cell biocompatibility to fibrin gels and due to its transparency and faster gelation time could potentially be superior for clinical purposes. STATEMENT OF SIGNIFICANCE: Extracellular matrix (ECM) can be used to provide tissue specific physical microstructure and biochemical cues for tissue regeneration. Here, we produced an ECM hydrogel derived from acellular porcine cornea stroma (APCS-gel) that retained critical biological characteristics of the native tissue and provided significant transparency and fast gelation time. Our data demonstrated that the APCS-gel was superior to clinically used fibrin gel, as the APCS-gel showed high porosity and permeability, better corneal stromal keratocytes infiltration, increased cellular proliferation and retention of corneal epithelial cells stemness. The APCS-gel improved corneal wound healing in vitro and in vivo. This APCS-gel may have clinical utility for corneal diseases, and the more general approach used to make this hydrogel might be used in other tissues.

Project description:BackgroundHevin is a matricellular protein involved in tissue repair and remodeling via interaction with the surrounding extracellular matrix (ECM) proteins. In this study, we examined the functional role of hevin using a corneal stromal wound healing model achieved by an excimer laser-induced irregular phototherapeutic keratectomy (IrrPTK) in hevin-null (hevin(-/-)) mice. We also investigated the effects of exogenous supplementation of recombinant human hevin (rhHevin) to rescue the stromal cellular components damaged by the excimer laser.Methodology/principal findingsWild type (WT) and hevin (-/-) mice were divided into three groups at 4 time points- 1, 2, 3 and 4 weeks. Group I served as naïve without any treatment. Group II received epithelial debridement and underwent IrrPTK using excimer laser. Group III received topical application of rhHevin after IrrPTK surgery for 3 days. Eyes were analyzed for corneal haze and matrix remodeling components using slit lamp biomicroscopy, in vivo confocal microscopy, light microscopy (LM), transmission electron microscopy (TEM), immunohistochemistry (IHC) and western blotting (WB). IHC showed upregulation of hevin in IrrPTK-injured WT mice. Hevin (-/-) mice developed corneal haze as early as 1-2 weeks post IrrPTK-treatment compared to the WT group, which peaked at 3-4 weeks. They also exhibited accumulation of inflammatory cells, fibrotic components of ECM proteins and vascularized corneas as seen by IHC and WB. LM and TEM showed activated keratocytes (myofibroblasts), inflammatory debris and vascular tissues in the stroma. Exogenous application of rhHevin for 3 days reinstated inflammatory index of the corneal stroma similar to WT mice.Conclusions/significanceHevin is transiently expressed in the IrrPTK-injured corneas and loss of hevin predisposes them to aberrant wound healing. Hevin (-/-) mice develop early corneal haze characterized by severe chronic inflammation and stromal fibrosis that can be rescued with exogenous administration of rhHevin. Thus, hevin plays a pivotal role in the corneal wound healing.

Dataset Information

Corneal Wound Healing Requires IKB kinase ? Signaling in Keratocytes.

Publications

Corneal Wound Healing Requires IKB kinase β Signaling in Keratocytes.

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