Mice Deficient in TAZ (Wwtr1) Demonstrate Clinical Features of Late-Onset Fuchs' Endothelial Corneal Dystrophy.

Purpose: We sought to define the role of Wwtr1 in murine ocular structure and function and determine the role of mechanotransduction in Fuchs' endothelial corneal dystrophy (FECD), with emphasis on interactions between corneal endothelial cells (CEnCs) and Descemet's membrane (DM). Methods: A Wwtr1 deficient mouse colony was established, and advanced ocular imaging, atomic force microscope (AFM), and histology/immunofluorescence were performed. Corneal endothelial wound healing was assessed using cryoinjury and phototherapeutic keratectomy in Wwtr1 deficient mice. Expression of WWTR1/TAZ was determined in the corneal endothelium from normal and FECD-affected patients; WWTR1 was screened for coding sequence variants in this FECD cohort. Results: Mice deficient in Wwtr1 had reduced CEnC density, abnormal CEnC morphology, softer DM, and thinner corneas versus wildtype controls by 2 months of age. Additionally, CEnCs had altered expression and localization of Na/K-ATPase and ZO-1. Further, Wwtr1 deficient mice had impaired CEnC wound healing. The WWTR1 transcript was highly expressed in healthy human CEnCs comparable to other genes implicated in FECD pathogenesis. Although WWTR1 mRNA expression was comparable between healthy and FECD-affected patients, WWTR1/TAZ protein concentrations were higher and localized to the nucleus surrounding guttae. No genetic associations were found in WWTR1 and FECD in a patient cohort compared to controls. Conclusions: There are common phenotypic abnormalities seen between Wwtr1 deficient and FECD-affected patients, suggesting that Wwtr1 deficient mice could function as a murine model of late-onset FECD. Despite the lack of a genetic association between FECD and WWTR1, aberrant WWTR1/TAZ protein subcellular localization and degradation may play critical roles in the pathogenesis of FECD.

Andrographolide Inhibits Corneal Fibroblast to Myofibroblast Differentiation In Vitro.

Corneal opacification due to fibrosis is a leading cause of blindness worldwide. Fibrosis occurs from many causes including trauma, photorefractive surgery, microbial keratitis (infection of the cornea), and chemical burns, yet there is a paucity of therapeutics to prevent or treat corneal fibrosis. This study aimed to determine if andrographolide, a labdane diterpenoid found in Andrographis paniculate, has anti-fibrotic properties. Furthermore, we evaluated if andrographolide could prevent the differentiation of fibroblasts to myofibroblasts in vitro, given that the transforming growth factor beta-1(TGF-ß1) stimulated persistence of myofibroblasts in the cornea is a primary component of fibrosis. We demonstrated that andrographolide inhibited the upregulation of alpha smooth muscle actin (αSMA) mRNA and protein in rabbit corneal fibroblasts (RCFs), thus, demonstrating a reduction in the transdifferentiation of myofibroblasts. Immunofluorescent staining of TGF-ß1-stimulated RCFs confirmed a dose-dependent decrease in αSMA expression when treated with andrographolide. Additionally, andrographolide was well tolerated in vivo and had no impact on corneal epithelialization in a rat debridement model. These data support future studies investigating the use of andrographolide as an anti-fibrotic in corneal wound healing.

Antimicrobial Peptide Expression at the Ocular Surface and Their Therapeutic Use in the Treatment of Microbial Keratitis.

Microbial keratitis is a common cause of ocular pain and visual impairment worldwide. The ocular surface has a relatively paucicellular microbial community, mostly found in the conjunctiva, while the cornea would be considered relatively sterile. However, in patients with microbial keratitis, the cornea can be infected with multiple pathogens including Staphylococcus aureus, Pseudomonas aeruginosa, and Fusarium sp. Treatment with topical antimicrobials serves as the standard of care for microbial keratitis, however, due to high rates of pathogen resistance to current antimicrobial medications, alternative therapeutic strategies must be developed. Multiple studies have characterized the expression and activity of antimicrobial peptides (AMPs), endogenous peptides with key antimicrobial and wound healing properties, on the ocular surface. Recent studies and clinical trials provide promise for the use of AMPs as therapeutic agents. This article reviews the repertoire of AMPs expressed at the ocular surface, how expression of these AMPs can be modulated, and the potential for harnessing the AMPs as potential therapeutics for patients with microbial keratitis.

Non-specific protection from respiratory tract infections in cattle generated by intranasal administration of an innate immune stimulant.

Alternatives to antibiotics for prevention of respiratory tract infections in cattle are urgently needed given the increasing public and regulatory pressure to reduce overall antibiotic usage. Activation of local innate immune defenses in the upper respiratory tract is one strategy to induce non-specific protection against infection with the diverse array of viral and bacterial pathogens associated with bovine respiratory disease complex (BRDC), while avoiding the use of antibiotics. Our prior studies in rodent models demonstrated that intranasal administration of liposome-TLR complexes (LTC) as a non-specific immune stimulant generated high levels of protection against lethal bacterial and viral pathogens. Therefore, we conducted studies to assess LTC induction of local immune responses and protective immunity to BRDC in cattle. In vitro, LTC were shown to activate peripheral blood mononuclear cells in cattle, which was associated with secretion of INFγ and IL-6. Macrophage activation with LTC triggered intracellular killing of Mannheimia hemolytica and several other bacterial pathogens. In studies in cattle, intranasal administration of LTC demonstrated dose-dependent activation of local innate immune responses in the nasopharynx, including recruitment of monocytes and prolonged upregulation (at least 2 weeks) of innate immune cytokine gene expression by nasopharyngeal mucosal cells. In a BRDC challenge study, intranasal administration of LTC prior to pathogen exposure resulted in significant reduction in both clinical signs of infection and disease-associated euthanasia rates. These findings indicate that intranasal administration of a non-specific innate immune stimulant can be an effective method of rapidly generating generalized protection from mixed viral and bacterial respiratory tract infections in cattle.