Targeted Suppression of HO-2 Gene Expression Impairs the Innate Anti-inflammatory and Repair Responses of the Cornea to Injury

Overview

Journal Mol Vis

Specialties Cell Biology
Molecular Biology
Ophthalmology

Date 2011 May 10

PMID 21552471

Citations 8

Authors

Lars Bellner

Kiran A Patil

Kirkland Castellano

Adna Halilovic

Michael W Dunn

Michal Laniado Schwartzman

Affiliations

Soon will be listed here.

Abstract

Purpose: Heme oxygenase (HO)-2 is highly expressed in the corneal epithelium and is a component of the heme oxygenase system that represents an intrinsic cytoprotective and anti-inflammatory system based on its ability to modulate leukocyte migration and to inhibit expression of inflammatory cytokines and proteins via its products biliverdin/bilirubin and carbon monoxide (CO). We have shown that in HO-2 null mice epithelial injury leads to unresolved corneal inflammation and chronic inflammatory complications including ulceration, perforation and neovascularization. In this study, we explore whether a localized corneal suppression of HO-2 is sufficient for disrupting the innate anti-inflammatory and repair capability of the cornea.

Methods: Silencing hairpin RNA (shRNA) against HO-2 was administered subconjunctivally (100 ng/eye) as well as topically (100 ng/eye) starting one day before corneal epithelial debridement and once daily, thereafter. The corneal epithelium was removed using an Alger Brush in anesthetized mice. Re-epithelialization was assessed by fluorescein staining using a dissecting microscope and image analysis. Inflammatory response was quantified by myeloperoxidase activity. Levels of mRNA were measured by RT-PCR.

Results: Local injection of HO-2-specific shRNA led to a 50% reduction in corneal HO-2 mRNA. Administration of HO-2-specific shRNA delayed corneal re-epithelialization when compared with the control shRNA-treated group by 14%, 20%, and 12% at days 3, 4, and 7 after injury, respectively (n=18-24). The observed delay in the wound repair process in HO-2 shRNA treated mice was accompanied by a threefold and 3.5 fold increase in the neovascular response at days 4 and 7 after injury. Further, local knockdown of HO-2 lead to an aberrant chronic inflammatory response, as shown by presence of high numbers of inflammatory cells still present in the cornea at day 7 after injury; 1.04±0.45×10(6) in HO-2 knockdown mice versus 0.14±0.03×10(6) inflammatory cells in control mice. Matrix metalloproteinase-2 (MMP-2) but not MMP-9 increased following injury and remained elevated in the injured corneas of the HO-2 shRNA-treated eyes.

Conclusions: Corneal knockdown of HO-2 via local administration of HO-2-specific shRNA leads to delayed re-epithelialization, increased neovascularization and an aberrant inflammatory response similar to what is observed in the HO-2 null mouse. The elevated MMP-2 expression may contribute to the increase in neovascularization in corneas in which HO-2 expression is suppressed.

Citing Articles

The different facets of heme-oxygenase 1 in innate and adaptive immunity.

Silva R, Vasconcelos L, Travassos L Cell Biochem Biophys. 2022; 80(4):609-631.

PMID: 36018440 DOI: 10.1007/s12013-022-01087-z.

Heme oxygenase-2 protects against ischemic acute kidney injury: influence of age and sex.

Nath K, Garovic V, Grande J, Croatt A, Ackerman A, Farrugia G Am J Physiol Renal Physiol. 2019; 317(3):F695-F704.

PMID: 31215802 PMC: 6842883. DOI: 10.1152/ajprenal.00085.2019.

Heme oxygenase-2 deletion impairs macrophage function: implication in wound healing.

Bellner L, Marrazzo G, van Rooijen N, Dunn M, Abraham N, Schwartzman M FASEB J. 2014; 29(1):105-15.

PMID: 25342128 PMC: 4285548. DOI: 10.1096/fj.14-256503.

Pro-healing effects of bilirubin in open excision wound model in rats.

Ahanger A, Leo M, Gopal A, Kant V, Tandan S, Kumar D Int Wound J. 2014; 13(3):398-402.

PMID: 24947136 PMC: 7950029. DOI: 10.1111/iwj.12319.

Heme oxygenase-2 suppress TNF-α and IL6 expression via TLR4/MyD88-dependent signaling pathway in mouse cerebral vascular endothelial cells.

Chen R, Yuan H, Zhang T, Wang Z, Hu A, Wu L Mol Neurobiol. 2014; 50(3):971-8.

PMID: 24788682 DOI: 10.1007/s12035-014-8693-x.

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