Urea Biosensor Based on Conducting Polymer Transducers

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Role of Transforming Growth Factor Beta in Corneal Function, Biology and Pathology

Transforming growth factor-beta (TGFβ) is a pleiotropic multifunctional cytokine that regulates several essential cellular processes in many parts of the body including the cornea. Three isoforms of TGFβ are known in mammals and the human cornea expresses all of them. TGFβ1 has been shown to play a central role in scar formation in adult corneas whereas TGFβ2 and TGFβ3 have been implicated to play a critical role in corneal development and scarless wound healing during embryogenesis. The biological effects of TGFβ in the cornea have been shown to follow SMAD dependent as well as SMAD-independent signaling pathways depending upon cellular responses and microenvironment. Corneal TGFβ expression is necessary for maintaining corneal integrity and corneal wound healing. On the other hand, TGFβ is perhaps the most important cytokine in the pathogenesis of fibrotic disease in the cornea. Although the transformation of keratocytes to myofibroblasts induced by TGFβ is largely believed to cause corneal fibrosis or scarring, the precise molecular mechanism(s) involved in this process is still unknown. Currently no drugs are available to treat corneal scarring effectively without causing significant side effects. Many approaches to treat TGFβ-mediated corneal scarring are under investigation. These include blocking of TGFβ, TGFβ receptor, TGFβ function and/or TGFβ maturation. Other strategies such as modulating keratocyte proliferation, apoptosis, transcription and DNA condensation are also being investigated. The potential of gene therapy to neutralize the pathologic effects of TGFβ has also been demonstrated recently

Can human xylosyltransferase-1 serve as a biomarker and therapeutic target for corneal fibrosis?

Tested was the hypothesis that XYLT1 plays an important role in corneal wound healing and scarring and may allow development of newer strategies for curing corneal fibrosis. The specific aims were: 1) to characterize XYLT1 expression in normal and wounded human and rabbit corneas, 2) investigate its role in corneal wound healing, and 2) determine whether XYLT1 can serve as a biomarker for corneal fibrosis

Effect of Prophylactic and Therapeutic Mitomycin C on Corneal Apoptosis, Cellular Proliferation, Haze, and Long-Term Keratocyte Density in Rabbits

PURPOSE—To determine the mechanism through which topical mitomycin C prevents and treats corneal haze after photorefractive keratectomy (PRK) and to examine the effects of dosage and duration of exposure. METHODS—In 224 New Zealand rabbits, −9.0 diopter PRK with mitomycin C or balanced salt solution was performed. Haze level was graded at the slit-lamp. Rabbits were sacrificed at 4 hours, 24 hours, 4 weeks, or 6 months after surgery and immunohistochemistry was performed with TUNEL assay, Ki67 and α-SMA. RESULTS—TUNEL-positive apoptotic cells marginally increased in all mitomycin C groups whereas Ki67-positive mitotic cells decreased significantly following mitomycin C application. A greater decrease in myofibroblasts was noted with prophylactic mitomycin C treatment than therapeutic mitomycin C treatment. There was, however, an anterior stromal acellular zone (approximately 20% of the total stroma) in eyes treated with mitomycin C, which persisted to the maximum follow-up of 6 months. CONCLUSIONS—Mitomycin C treatment induces apoptosis of keratocytes and myofibroblasts, but the predominate effect in inhibiting or treating haze appears to be at the level of blocked replication of keratocytes or other progenitor cells of myofibroblasts. Treatment with 0.002% mitomycin C for 12 seconds to 1 minute appears to be just as effective as higher concentrations for longer duration in the rabbit model. However, a persistent decrease in keratocyte density in the anterior stroma could be a warning sign for future complications and treatment should be reserved for patients with significant risk of developing haze after PRK

Efficacy and Safety Comparison Between Suberoylanilide Hydroxamic Acid and Mitomycin C in Reducing the Risk of Corneal Haze After PRK Treatment In Vivo

PURPOSE: This study compared the efficacy and safety of suberoylanilide hydroxamic acid (SAHA) and mitomycin C (MMC) up to 4 months in the prevention of corneal haze induced by photorefractive keratectomy (PRK) in rabbits in vivo. METHODS: Corneal haze in rabbits was produced with −9.00 diopter PRK. A single application of SAHA (25 μM) or MMC (0.02%) was applied topically immediately after PRK. Effects of the two drugs were analyzed by slit-lamp microscope, specular microscope, TUNEL assay, and immunofluorescence. RESULTS: Single topical adjunct use of SAHA (25 μM) or MMC (0.02%) after PRK attenuated more than 95% corneal haze and myofibroblast formation (P \u3c .001). SAHA did not reduce keratocyte density, cause keratocyte apoptosis, or increase immune cell infiltration compared to MMC (P \u3c .01 or .001). Furthermore, SAHA dosing did not compromise corneal endothelial phenotype, density, or function in rabbit eyes, whereas MMC application did (P \u3c .01 or .001). CONCLUSIONS: SAHA and MMC significantly decreased corneal haze after PRK in rabbits in vivo. SAHA exhibited significantly reduced short- and long-term damage to the corneal endothelium compared to MMC in rabbits. SAHA is an effective and potentially safer alternative to MMC for the prevention of corneal haze after PRK. Clinical trials are warranted

Targeted AAV5-Smad7 Gene Therapy Inhibits Corneal Scarring \u3cem\u3ein vivo\u3c/em\u3e

Corneal scarring is due to aberrant activity of the transforming growth factor β (TGFβ) signaling pathway following traumatic, mechanical, infectious, or surgical injury. Altered TGFβ signaling cascade leads to downstream Smad (Suppressor of mothers against decapentaplegic) protein-mediated signaling events that regulate expression of extracellular matrix and myogenic proteins. These events lead to transdifferentiation of keratocytes into myofibroblasts through fibroblasts and often results in permanent corneal scarring. Hence, therapeutic targets that reduce transdifferentiation of fibroblasts into myofibroblasts may provide a clinically relevant approach to treat corneal fibrosis and improve long-term visual outcomes. Smad7 protein regulates the functional effects of TGFβ signaling during corneal wound healing. We tested that targeted delivery of Smad7 using recombinant adeno-associated virus serotype 5 (AAV5-Smad7) delivered to the corneal stroma can inhibit corneal haze post photorefractive keratectomy (PRK) in vivo in a rabbit corneal injury model. We demonstrate that a single topical application of AAV5-Smad7 in rabbit cornea post-PRK led to a significant decrease in corneal haze and corneal fibrosis. Further, histopathology revealed lack of immune cell infiltration following AAV5-Smad7 gene transfer into the corneal stroma. Our data demonstrates that AAV5-Smad7 gene therapy is relatively safe with significant potential for the treatment of corneal disease currently resulting in fibrosis and impaired vision

Epigenetic Modification Prevents Excessive Wound Healing and Scar Formation After Glaucoma Filtration Surgery

PURPOSE. The purpose of this study was to determine the efficacy of suberoylanilide hydroxamic acid (SAHA), a histone deacetylase inhibitor (HDACi), in prevention of excessive wound healing and scar formation in a rabbit model of glaucoma filtration surgery (GFS). METHODS. A rabbit model of GFS was used. Rabbits that underwent GFS received balanced salt solution, or SAHA (50 lM), or mitomycin C (0.02%). Clinical scores of IOP, bleb vascularity, and slit-lamp examination were performed. On postoperative day 14, rabbits were killed and the bleb tissues were collected for evaluation of tissue fibrosis with hematoxylin and eosin, Masson trichrome, a-smooth muscle actin (aSMA), and F-actin staining. Furthermore, SAHA-mediated acetylation of histones in corneal fibroblasts and conjunctiva were determined by Western blot analysis. RESULTS. Suberoylanilide hydroxamic acid treatment after GFS showed no signs of edema, corneal opacity, endophthalmitis, or cataract formation. Morphometric analysis of SAHA-treated eyes showed higher bleb length (P \u3c 0.001), bleb area (P \u3c 0.05), lower IOP (P \u3c 0.01), and decreased vascularity compared to control. Furthermore, SAHA treatment showed significantly reduced levels of aSMA (P \u3c 0.001), F-actin (P \u3c 0.01), and collagen deposition (P \u3c 0.05) at the sclerotomy site. In addition, SAHA treatment increased the acetylation status of H3 and H4 histones in corneal fibroblasts and conjunctiva. CONCLUSIONS. This study demonstrates that HDAC inhibition is an attractive pharmacologic target to modulate GFS wound healing, and SAHA, an HDACi, can be a useful adjunct to improve the GFS outcome

Typhoidal Salmonella and Emerging Resistance in Outbreak Proportions

Introduction: Typhoidal Salmonella causes an invasive infection resulting in 200 000 deaths among 20 million patients annually. Typhoid remains a public health problem in Southeast Asia, the Indian subcontinent, Africa, and South America. Traveler’s diarrhea caused by Salmonella is common in Asia. Outbreaks of typhoidal Salmonella resistant to ampicillin, chloramphenicol, and trimethoprim-sulfamethoxazole in the 1990s pushed therapy to ciprofloxacin which was replaced by ceftriaxone due to fluoroquinolone resistance. Methods: This prospective study characterizes demographical, etiological, and resistance patterns in typhoidal Salmonella at a 1000-bed teaching hospital in New Delhi, India. Two hundred inpatients in pediatrics, obstetrics-gynecology, medicine, intensive care, and OPD in whom Salmonella bacteremia was detected were characterized by routine and automated microbiology techniques. Results: The mean age of patients in this study was 21.4 years. Overall, 71% of patients suffered from Salmonella Typhi followed by 26% from Salmonella Paratyphi A. Four cases of Salmonella resistance to ampicillin, trimethoprim-sulfamethoxazole, and chloramphenicol were encountered. A high degree of partial and complete resistance to fluoroquinolones was seen among Salmonella Typhi, Salmonella Paratyphi A, and Salmonella Paratyphi B cases. Resistance to ciprofloxacin was 48% among Salmonella Typhi and 100% among Salmonella Paratyphi A cases. Only 18% of Salmonella Typhi cases were completely resistant to quinolones, while 79% were partially resistant. A total of 92% of Salmonella Paratyphi A cases were partially resistant to quinolones. Four Salmonella cases were resistant to ceftriaxone. Conclusion: Salmonella Typhi remains the predominant serotype, followed by Salmonella Paratyphi A. The high prevalence of quinolone resistance in Salmonella Typhi and Salmonella Paratyphi A is a serious problem limiting empirical therapy to non-quinolone-based therapy such as ceftriaxone. Multidrug-resistant Salmonella is an emerging problem requiring active surveillance among residents and travelers presenting with tropical fever

Targeted AAV5-Smad7 gene therapy inhibits corneal scarring in vivo.

Corneal scarring is due to aberrant activity of the transforming growth factor β (TGFβ) signaling pathway following traumatic, mechanical, infectious, or surgical injury. Altered TGFβ signaling cascade leads to downstream Smad (Suppressor of mothers against decapentaplegic) protein-mediated signaling events that regulate expression of extracellular matrix and myogenic proteins. These events lead to transdifferentiation of keratocytes into myofibroblasts through fibroblasts and often results in permanent corneal scarring. Hence, therapeutic targets that reduce transdifferentiation of fibroblasts into myofibroblasts may provide a clinically relevant approach to treat corneal fibrosis and improve long-term visual outcomes. Smad7 protein regulates the functional effects of TGFβ signaling during corneal wound healing. We tested that targeted delivery of Smad7 using recombinant adeno-associated virus serotype 5 (AAV5-Smad7) delivered to the corneal stroma can inhibit corneal haze post photorefractive keratectomy (PRK) in vivo in a rabbit corneal injury model. We demonstrate that a single topical application of AAV5-Smad7 in rabbit cornea post-PRK led to a significant decrease in corneal haze and corneal fibrosis. Further, histopathology revealed lack of immune cell infiltration following AAV5-Smad7 gene transfer into the corneal stroma. Our data demonstrates that AAV5-Smad7 gene therapy is relatively safe with significant potential for the treatment of corneal disease currently resulting in fibrosis and impaired vision

Corneal Injury Is Associated With Stromal and Vascular Alterations Within Cranial Dura Mater

The cornea and cranial dura mater share sensory innervation. This link raises the possibility that pathological impulses mediated by corneal injury may be transmitted to the cranial dura, trigger dural perivascular/connective tissue nociceptor responses, and induce vascular and stromal alterations affecting dura mater blood and lymphatic vessel functionality. In this study, using a mouse model, we demonstrate for the first time that two weeks after the initial insult, alkaline injury to the cornea leads to remote pathological changes within the coronal suture area of the dura mater. Specifically, we detected significant pro-fibrotic changes in the dural stroma, as well as vascular remodeling characterized by alterations in vascular smooth muscle cell (VSMC) morphology, reduced blood vessel VSMC coverage, endothelial cell expression of the fibroblast specific protein 1, and significant increase in the number of podoplanin-positive lymphatic sprouts. Intriguingly, the deficiency of a major extracellular matrix component, small leucine-rich proteoglycan decorin, modifies both the direction and the extent of these changes. As the dura mater is the most important route for the brain metabolic clearance, these results are of clinical relevance and provide a much-needed link explaining the association between ophthalmic conditions and the development of neurodegenerative diseases