Conjunctival Chalasis Should Not be Overlooked: Investigation Into Postoperative Ocular Surface Changes in Involutional Blepharoptosis.

BACKGROUND: Involutional blepharoptosis is common among elderly people. The tightening of eyelids postptosis surgery could potentially increase friction between the eyelid and the ocular surface, but this hypothesis has not yet been substantiated by research. The authors explored the relationship between involutional blepharoptosis surgery and friction-related diseases, namely conjunctivochalasis, lid wiper epitheliopathy, and superior limbic keratoconjunctivitis. METHODS: We conducted a prospective study involving 31 patients who underwent levator advancement for involutional blepharoptosis. Both preoperatively and 6 weeks postoperatively, the authors assessed a range of outcome measures, including margin reflex distance-1, 2, tear film break-up time, superficial punctate keratopathy, inferior conjunctivochalasis, upper lid wiper epitheliopathy, and superior limbic keratoconjunctivitis. RESULTS: Conjunctivochalasis was detected in 18 eyes preoperatively and 20 eyes postoperatively. Lid wiper epitheliopathy was detected in 2 eyes preoperatively and in no eyes postoperatively. Superior limbic keratoconjunctivitis was detected in 2 eyes preoperatively and 1 eye postoperatively. From preoperative to postoperative assessments, conjunctivochalasis worsened in 11 eyes (17.2%), and there were no eyes with worsening lid wiper epitheliopathy or superior limbic keratoconjunctivitis. There was a significant worsening of superficial punctate keratopathy in the group with exacerbated conjunctivochalasis compared with the unchanged group (0.72 vs. 0.12, P =0.0222). The superficial petechial keratopathy in the 6 cases in which there was worsening of both conjunctivochalasis and superficial petechial keratopathy were all located inferiorly in the cornea. CONCLUSIONS: Conjunctivochalasis can worsen following ptosis surgery, potentially leading to an increase in inferior superficial punctate keratopathy. When performing involutional blepharoptosis surgery, surgeons should be mindful of the potential implications of friction-related diseases, particularly conjunctivochalasis.

Loss of TRPV4 Cation Channel Inhibition of Macrophage Infiltration and Neovascularization in a Mouse Cornea.

Corneal injury-associated inflammation could induce inward-growing neovascularization from the periphery of the tissue. Such neovascularization could cause stromal opacification and curvature disturbance, and both potentially impair visual function. In this study, we determined the effects of the loss of transient receptor potential vanilloid 4 (TRPV4) expression on the development of neovascularization in the corneal stroma in mice by producing a cauterization injury in the central area of the cornea. New vessels were immunohistochemically labeled with anti-TRPV4 antibodies. TRPV4 gene knockout suppressed the growth of such CD31-labeled neovascularization in association with the suppression of infiltration of macrophages and tissue messenger RNA expression of the vascular endothelial cell growth factor A level. Treatment of cultured vascular endothelial cells with supplementation of HC-067047 (0.1 µM, 1 µM, or 10 µM), a TRPV4 antagonist, attenuated the formation of a tube-like structure with sulforaphane (15 µM, for positive control) that modeled the new vessel formation. Therefore, the TRPV4 signal is involved in injury-induced macrophagic inflammation and neovascularization activity by vascular endothelial cells in a mouse corneal stroma. TRPV4 could be a therapeutic target to prevent unfavorable postinjury neovascularization in the cornea.

The Roles of Transient Receptor Potential Vanilloid 1 and 4 in Olfactory Regeneration.

Olfactory disorders, which are closely related to cognitive deterioration, can be caused by several factors, including infections, such as COVID-19; aging; and environmental chemicals. Injured olfactory receptor neurons (ORNs) regenerate after birth, but it is unclear which receptors and sensors are involved in ORN regeneration. Recently, there has been great focus on the involvement of transient receptor potential vanilloid (TRPV) channels, which are nociceptors expressed on sensory nerves during the healing of damaged tissues. The localization of TRPV in the olfactory nervous system has been reported in the past, but its function there are unclear. Here, we investigated how TRPV1 and TRPV4 channels are involved in ORN regeneration. TRPV1 knockout (KO), TRPV4 KO, and wild-type (WT) mice were used to model methimazole-induced olfactory dysfunction. The regeneration of ORNs was evaluated using olfactory behavior, histologic examination, and measurement of growth factors. Both TRPV1 and TRPV4 were found to be expressed in the olfactory epithelium (OE). TRPV1, in particular, existed near ORN axons. TRPV4 was marginally expressed in the basal layer of the OE. The proliferation of ORN progenitor cells was reduced in TRPV1 KO mice, which delayed ORN regeneration and the improvement of olfactory behavior. Postinjury OE thickness improved faster in TRPV4 KO mice than WT mice but without acceleration of ORN maturation. The nerve growth factor and transforming growth factor ß levels in TRPV1 KO mice were similar to those in WT mice, and the transforming growth factor ß level was higher than TRPV4 KO mice. TRPV1 was involved in stimulating the proliferation of progenitor cells. TRPV4 modulated their proliferation and maturation. ORN regeneration was regulated by the interaction between TRPV1 and TRPV4. However, in this study, TRPV4 involvement was limited compared with TRPV1. To our knowledge, this is the first study to demonstrate the involvement of TRPV1 and TRPV4 in OE regeneration.

Engineered Knockout of TRPA1 Inhibits Laser-Induced Choroidal Neovascularization Along With Associated TGFß1 Expression and Neutrophil Infiltration.

We examined the effects of gene ablation and chemical inhibition of transient receptor potential ankyrin 1 (TRPA1) on the growth of experimental argon laser-induced choroidal neovascularization (CNV) in mice. CNV was induced in the eyes of 6- to 8-week-old TRPA1-null (knockout [KO]) and wild-type (WT) mice by argon laser irradiation. Gene expression analysis was performed in laser-injured tissues at days 1 and 3. CNV growth was evaluated at day 14. Reciprocal bone marrow transplantation was performed between each genotype to identify the components responsible for either recipient tissue or bone marrow-derived inflammatory cells. Our results show that laser irradiation successfully induced CNV growth at the site of laser injury. The size of induced CNV was significantly smaller in KO mice than in WT mice at day 14, as determined by angiography with fluorescein isothiocyanate-dextran. Invasion of neutrophils, but not macrophages, was suppressed in association with suppression of the expression of transforming growth factor ß1 and interleukin 6 in laser-irradiated KO tissue. Bone marrow transplantation indicated that the genotype of the recipient mouse, but not of inflammatory cells, is attributable to the KO phenotype. Systemic administration of a TRPA1 antagonist also reduced the CNV in a WT mouse. In conclusion, TRPA1 signaling in local cells is involved in growth of laser-induced CNV. The phenotype was not attributable to vascular endothelial cells and inflammatory cells. Blocking TRPA1 signal may therefore be a potential treatment strategy for CNV-related ocular diseases.

Delayed regression of laser-induced choroidal neovascularization in TNFα-null mice.

We investigated the effects of lacking TNFα on the development and regression of Argon-laser-induced choroidal neovascularization (CNV) in mice. We lasered ocular fundus for induction of CNV in both wild-type (WT) and TNFα-null (KO) mice. Fluorescence angiography was performed to examine the size of CNV lesions. Gene expression pattern of wound healing-related components was examined. The effects of exogenous TNFα on apoptosis of human retinal microvascular endothelial cells (HRMECs) and on the tube-like structure of the cells were investigated in vitro. The results showed that Argon-laser irradiation-induced CNV was significantly larger in KO mice than WT mice on Day 21, but not at other timepoints. Lacking TNFα increased neutrophil population in the lesion. The distribution of cleaved caspase3-labelled apoptotic cells was more frequently observed in the laser-irradiated tissue in a WT mouse as compared with a KO mouse. Exogenous TNFα induced apoptosis of HRMECs and accelerated regression of tube-like structure of HRMECs in cell culture. Taken together, TNFα gene knockout delays the regression of laser-induced CNV in mice. The mechanism underlying the phenotype might include the augmentation of neutrophil population in the treated tissue and attenuation of vascular endothelial cell apoptosis.

Loss of sphingosine 1-phosphate receptor 3 gene function impairs injury-induced stromal angiogenesis in mouse cornea.

Sphingosine 1-phosphate (S1P) is a bioactive sphingolipid generated through sphingosine kinase1 (SPK1)-mediated phosphorylation of sphingosine. We show here that injury-induced S1P upregulation increases corneal neovascularization through stimulating S1PR3, a cognate receptor. since this response was suppressed in S1PR3-knockout mice. Furthermore, Cayman10444, a selective S1PR3 inhibitor, reduced this response in WT mice. Such reductions in neovascularization were associated with reduced vascular endothelial growth factor A (VEGF-A) mRNA expression levels in WT TKE2 corneal epithelial cells and macrophages treated with CAY10444 as well as macrophages isolated from S1PR3 KO mice. S1P increased tube-like vessel formation in human vascular endothelial cells (HUVEC) and human retinal microvascular endothelial cells (HRMECs) cells expressing S1PR3. In S1PR3 KO mice, TGFß1-induced increases in αSMA gene expression levels were suppressed relative to those in the WT counterparts. In S1PR3 deficient macrophages, VEGF-A expression levels were lower than in WT macrophages. Transforming growth factor ß1(TGFß1) upregulated SPK1 expression levels in ocular fibroblasts and TKE2 corneal epithelial cells. CAY10444 blocked S1P-induced increases in VEGF-A mRNA expression levels in TKE2 corneal epithelial cells. Endogenous S1P signaling upregulated VEGF-A and VE-cadherin mRNA expression levels in HUVEC. Unlike in TKE2 cells, SIS3 failed to block TGFß1-induced VEGF-A upregulation in ocular fibroblasts. Taken together, these results indicate that injury-induced TGFß1 upregulation increases S1P generation through increases in SPK1 activity. The rise in S1P formation stimulates the S1PR3-linked signaling pathway, which in turn increases VEGF-A expression levels and angiogenesis in mouse corneas.

Impairment of corneal epithelial wound healing is association with increased neutrophil infiltration and reactive oxygen species activation in tenascin X-deficient mice.

The purpose of the study was to uncover the role of tenascin X in modulation of healing in mouse corneas subjected to epithelium debridement. Healing in corneas with an epithelial defect was evaluated at the levels of gene and protein expression. Wound healing-related mediators and inflammatory cell infiltration were detected by histology, immunohistochemistry and real-time RT-PCR. Tenascin X protein was upregulated in the wounded wild-type (WT) corneal epithelium. The lack of tenascin X impaired closure of an epithelial defect and accelerated infiltration of neutrophils into the wound periphery as compared to the response in WT tissue. Expression of wound healing-related proinflammatory and reparative components, i.e., interleukin-6, transforming growth factor ß, matrix metalloproteinases, were unaffected by the loss of tenascin X expression. Marked accumulation of malondialdehyde (a lipid peroxidation-derived product) was observed in KO healing epithelia as compared with its WT counterpart. Neutropenia induced by systemic administration of a specific antibody rescued the impairment of epithelial healing in KO corneas, with reduction of malondialdehyde levels in the epithelial cells. Finally, we showed that a chemical scavenging reactive oxygen species reversed the impairment of attenuation of epithelial repair with a reduction of tissue levels of malondialdehyde. In conclusion, loss of tenascin X prolonged corneal epithelial wound healing and increased neutrophilic inflammatory response to debridement in mice. Tenascin X contributes to the control of neutrophil infiltration needed to support the regenerative response to injury and prevent the oxidative stress mediators from rising to cytotoxic levels.

Shp2-mediated MAPK pathway regulates ΔNp63 in epithelium to promote corneal innervation and homeostasis.

Corneal nerve fibers serving sensory, reflex, and neurotrophic functions sustain corneal homeostasis and transparency to promote normal visual function. It is not known whether corneal epithelium is also important for the corneal innervation. Herein, we generated a compound transgenic mouse strain, K14rtTA;tetO-Cre (TC);Shp2flox/flox, in which Shp2 was conditionally knocked out from K14-positive cells including corneal epithelium (Shp2K14ce-cko) upon doxycycline (dox) administration. Our data reveal that Shp2K14ce-cko caused corneal denervation. More specifically, corneal epithelium thickness and corneal sensitivity reduced dramatically in Shp2K14ce-cko mice. In addition, corneal epithelial wound healing after debridement was delayed substantially in the mutant mice. These defects manifested in Shp2K14ce-cko mice resemble the symptoms of human neurotrophic keratopathy. Our in vitro study shows that neurite outgrowth of the mouse primary trigeminal ganglion cells (TGCs) was inhibited when as cocultured with mouse corneal epithelial cells (TKE2) transfected by Shp2-, Mek1/2-, or ∆Np63-targeted siRNA but not by Akt1/2-targeted siRNA. Furthermore, ∆Np63 RNA interference downregulated Ngf expression in TKE2 cells. Cotransfection experiments reveal that Shp2 tightly monitored ΔNp63 protein levels in HEK293 and TKE2 cells. Taken together, our data suggest that the Shp2-mediated MAPK pathway regulated ΔNp63, which in turn positively regulated Ngf in epithelium to promote corneal innervation and epithelial homeostasis.

Topical hyaluronan alone promotes corneal epithelial cell migration whereas combination with benzalkonium chloride impairs epithelial wound healing.

Purpose: To evaluate the effects of topical hyaluronan (HA) on corneal epithelial wound healing when administered with or without benzalkonium chloride (BAC).Methods: A cultured human corneal epithelial cell line (HCE-T) was subjected to in vitro scratch assays and in situ epithelial migration was evaluated in organ-cultured rabbit corneas. The corneal epithelium of C57BL/6J mice was also evaluated to determine in vivo wound healing. An in vivo imaging system was also used to evaluate the effects of HA on eye drop retention on the ocular surface.Results: The findings revealed the promotion of HCE-T migration, in situ rabbit corneal epithelial migration, and in vivo wound healing in mouse corneal epithelium by HA. Pre-treatment with HA also protected against delayed epithelial wound healing in BAC in vitro. However, pre-treatment with 3 mg/mL HA did not show a protective effect against BAC in vivo, but instead delayed epithelial wound healing and increased detection of cleaved caspase-3. This suggested that HA promotes the retention of BAC on the ocular surface. The instilled HA was retained after 15 min, at a significantly higher rate than for phosphate-buffered saline.Conclusions: The combination of HA and BAC impaired wound healing in the corneal epithelium.

Effects of a prostaglandin F2alpha derivative glaucoma drug on EGF expression and E-cadherin expression in a corneal epithelial cell line.

Purpose: We examined the effects of travoprost on cell proliferation-related signals and E-cadherin expression in vitro and in situ in order to obtain evidence to support the hypothesis that topical travoprost impairs the integrity of the corneal epithelium.Methods: A human corneal epithelial cell culture was treated with travoprost (0.4 mg/ml) and/or PD168393 (an EGF receptor inhibitor, 10 µM). The culture was then processed for cell proliferation, an mRNA expression analysis of epidermal growth factor (EGF) and E-cadherin, and protein expression analysis of E-cadherin by immunocytochemistry and Western blotting. The eyes of C57/BL6 mice were incubated in serum-free medium plus travoprost (0.4 mg/ml) and/or PD168393 (10 µM). After being cultured for 24 h, the expression patterns of phospho-EGFR, phospho-ERK, E-cadherin, and Ki67 were immunohistochemically examined in paraffin sections.Results: The addition of travoprost up-regulated EGF mRNA expression and cell proliferation in the corneal epithelial cell culture, and this was cancelled by the addition of PD168393. This FP agonist also decreased E-cadherin expression levels in the cell-cell contact zone, and this was cancelled by the addition of PD168393. In the organ culture, the addition of travoprost to the medium up-regulated the expression of phospho-EGFR and phospho-ERK as well as cell proliferation, and down-regulated the expression of E-cadherin in the corneal epithelium, particularly in basal cells, whereas PD168393 reversed these effects.Conclusions: Travoprost activates epithelial cell proliferation by up-regulating an EGF-related signal in association with the suppression of E-cadherin localization in the cell-cell contact zone. Modulation of the EGF signal may be a strategy to minimize the negative impact of this mitogen on reformation of corneal barrier function during epithelial renewal.

Sensory nerve supports epithelial stem cell function in healing of corneal epithelium in mice: the role of trigeminal nerve transient receptor potential vanilloid 4.

In order to understand the pathobiology of neurotrophic keratopathy, we established a mouse model by coagulating the first branch of the trigeminal nerve (V1 nerve). In our model, the sensory nerve in the central cornea disappeared and remaining fibers were sparse in the peripheral limbal region. Impaired corneal epithelial healing in the mouse model was associated with suppression of both cell proliferation and expression of stem cell markers in peripheral/limbal epithelium as well as a reduction of transient receptor potential vanilloid 4 (TRPV4) expression in tissue. TRPV4 gene knockout also suppressed epithelial repair in mouse cornea, although it did not seem to directly modulate migration of epithelium. In a co-culture experiment, TRPV4-introduced KO trigeminal ganglion upregulated nerve growth factor (NGF) in cultured corneal epithelial cells, but ganglion with a control vector did not. TRPV4 gene introduction into a damaged V1 nerve rescues the impairment of epithelial healing in association with partial recovery of the stem/progenitor cell markers and upregulation of cell proliferation and of NGF expression in the peripheral/limbal epithelium. Gene transfer of TRPV4 did not accelerate the regeneration of nerve fibers. Sensory nerve TRPV4 is critical to maintain stemness of peripheral/limbal basal cells, and is one of the major mechanisms of homeostasis maintenance of corneal epithelium.

Suppression of neovascularization in corneal stroma in a TRPA1-null mouse.

Corneal neovascularization and inflammatory fibrosis induced by severe injury or infection leads to tissue opacification and even blindness. Transient receptor potential (TRP) channel subtypes contribute to mediating these maladaptive responses through their interactions with other receptors. TRPV1 is one of the contributing channel isoforms inducing neovascularization in an alkali burn mouse wound healing model. VEGF-A upregulation contributes to neovascularization through interaction with its cognate receptors (VEGFR). Since the TRP isoform in this tissue, TRPA1, is also involved, we determined here if one of the pathways mediating neovascularization and immune cell infiltration involve an interaction between VEGFR and TRPA1 in a cauterization corneal mouse wound healing model. Localization of TRPA1 and endothelial cell (EC) CD31 immunostaining pattern intensity determined if TRPA1 expression was EC delimited during cauterization induced angiogenesis. Quantitative RT-PCR evaluated the effects of the absence of TRPA1 function on VEGF-A and TGF-ß1 mRNA expression during this process. Macrophage infiltration increased based on rises in F4/80 antigen immunoreactivity. TRPA1 immunostaining was absent on CD31-immunostained EC cells undergoing neovascularization, but it was present on other cell type(s) adhering to EC in vivo. Absence of TRPA1 expression suppressed both stromal neovascularization and inhibited macrophage infiltration. Similarly, the increases occurring in both VEGF-A and TGF-ß1 mRNA expression levels in WT tissue were blunted in the TRPA1-/- counterpart. On the other hand, in the macrophages their levels were invariant and their infiltration was inhibited. To determine if promotion by TRPA1 of angiogenesis was dependent on its expression on other unidentified cell types, the effects were compared of pharmacological manipulation of TRPA1 activity on EC proliferation tube formation and migration. In the presence and absence of a fibroblast containing feeder layer. Neither VEGF-induced increases in human vascular endothelial cell (HUVEC) proliferation nor migration were changed by a TRPA1 antagonist HC-030031 in the absence of a feeder layer. However, on a fibroblast feeder layer this antagonist suppressed HUVEC tube formation. In conclusion, during corneal wound healing transactivation by VEGFR of TRPA1 contributes to mediating neovascularization and macrophage infiltration. Such crosstalk is possible because of close proximity between VEGFR delimited expression on EC and TRPA1 expression restricted to cell types adhering to EC.

The murine lens: A model to investigate in vivo epithelial-mesenchymal transition.

Epithelial-mesenchymal transition (EMT) produces myofibroblasts that contribute to the formation of fibrotic tissue with an impairment of tissue homeostasis and functionality. The crystalline lens of the eye is a unique transparent and isolated tissue. The lens vesicle becomes isolated from the surface ectoderm, its cells are all contained as they line the inner surface of the lens capsule. Clinically the formation of fibrotic tissue by the lens epithelial cells causes a type of cataract or opacification and contraction of the lens capsule postcataract surgery. Production of EMT in the intact animal lens by using specific gene transfer to the lens or experimental lens injury has been shown to be a powerful tool to investigate EMT processes. It is not easy to uncover whether the origin of the myofibroblast is epithelial cell-derived or from other cell lineages in fibrotic tissues. However, myofibroblasts that appear in the crystalline lens pathology are totally derived from the lens epithelial cells for the reasons mentioned above. Here, we report on different animal models of lens EMT, using either transgenic approaches or injury to study the biological aspects of EMT. Developmental Dynamics 247:340-345, 2018. © 2017 The Authors Developmental Dynamics published by Wiley Periodicals, Inc. on behalf of American Association of Anatomists.

Loss of tenascin X gene function impairs injury-induced stromal angiogenesis in mouse corneas.

To determine the contribution by tenascin X (Tnx) gene expression to corneal stromal angiogenesis, the effects were determined of its loss on this response in TNX knockout (KO) mice. In parallel, the effects of such a loss were evaluated on vascular endothelial growth factor (VEGF) and transforming growth factor ß1 (TGFß1) gene and protein expression in fibroblasts and macrophages in cell culture. Histological, immunohistochemical and quantitative RT-PCR changes determined if Tnx gene ablation on angiogenic gene expression, inflammatory cell infiltration and neovascularization induced by central corneal stromal cauterization. The role was determined of Tnx function in controlling VEGF-A or TGFß1 gene expression by comparing their expression levels in ocular fibroblasts and macrophages obtained from wild-type (WT) and body-wide Tnx KO mice. Tnx was up-regulated in cauterized cornea. In Tnx KO, macrophage invasion was attenuated, VEGF-A and its cognate receptor mRNA expression along with neovascularization were lessened in Tnx KOs relative to the changes occurring in their WT counterpart. Loss of Tnx instead up-regulated in vivo mRNA expression of anti-angiogenic VEGF-B but not VEGF-A. On the other hand, TGFß1 mRNA expression declined in Tnx KO cultured ocular fibroblasts. Loss of Tnx gene expression caused VEGF-A expression to decline in macrophages. Tnx gene expression contributes to promoting TGFß1 mRNA expression in ocular fibroblasts and VEGF-A in macrophages, macrophage invasion, up-regulation of VEGF-A expression and neovascularization in an injured corneal stroma. On the other hand, it suppresses anti-angiogenic VEGF-B mRNA expression in vivo.

Lack of plakoglobin impairs integrity and wound healing in corneal epithelium in mice.

We generated cornea-specific plakoglobin (Jup; junctional plakoglobin) knockout mice in order to investigate the function of plakoglobin on the maintenance of the homeostasis of corneal epithelium in mice. Cornea epithelium-specific conditional knockouts (JupCEΔ/CEΔ) (cKO) were obtained by breeding keratin12-Cre (Krt12-Cre) mice to Jup-floxed (Jupf/f) mice. Light and transmission electron microscopic and immunohistochemical analyses were carried out to determine consequence of the loss of plakoglobin on maintaining corneal epithelium integrity under mechanical stress, e.g., brushing and wound healing. Immunohistochemistry analysis demonstrated that, although Jup ablation did not affect BrdU incorporation, basal cell-like cells labeled for keratin 14 were ectopically present in the supra-basal layer in mutant corneal epithelium, suggestive of altered cell differentiation. Plakoglobin-deficient epithelium exhibits increased fragility against mechanical intervention when compared to wild-type controls under identical treatment. Closure of an epithelial defect was significantly delayed in JupCEΔ/CEΔ epithelium. Our findings indicate that the lack of plakoglobin significantly affects corneal epithelium differentiation, as well as its structural integrity. Plakoglobin is essential to the maintenance of the structure of the corneal epithelium and its wound healing.

Impaired healing of cornea incision injury in a TRPV1-deficient mouse.

The present study attempts to elucidate the role of TRPV1 cation channel receptor on primary repair in an incision-wounded mouse cornea in vivo. Previous study revealed that blocking TRPV1 suppressed myofibroblast formation and expression of transforming growth factor ß1 (TGFß1) in cultured keratocytes or ocular fibroblasts. Male C57BL/6 (wild-type; WT) mice and male C57BL/6 Trpv1-null (KO) mice incurred a full-thickness incision injury (1.8 mm in length, limbus to limbus) in the central cornea of one eye with a surgical blade under general and topical anesthesia. The injury was not sutured. On days 0, 5, and 10, the eyes were enucleated, processed for histology, immunohistochemistry, and real-time RT-PCR gene expression analysis to evaluate the effects of the loss of TRPV1 on primary healing. Electron microscopy observation was also performed to know the effect of the loss of TRPV1 on ultrastructure of keratocytes. The results showed that the loss of Trpv1 gene delayed closure of corneal stromal incision with hindered myofibroblast transdifferentiation along with declines in the expression of collagen Ia1 and TGFß1. Inflammatory cell infiltration was not affected by the loss of TRPV1. Ultrastructurally endoplasmic reticulum of TRPV1-null keratocytes was more extensively dilated as compared with WT keratocytes, suggesting an impairment of protein secretion by TRPV1-gene knockout. These results indicate that injury-related TRPV1 signal is involved in healing of stromal incision injury in a mouse cornea by selectively stimulating TGFß-induced granulation tissue formation.

Ocular surface inflammation impairs structure and function of meibomian gland.

Dysfunction of the meibomian glands alters secreted meibum quantitatively and qualitatively that can lead to damage to the ocular surface epithelium. In response to an unstable tear film cause by meibomian gland dysfunction, ocular surface epithelium is damaged and expresses inflammatory cytokines leading to secondary ocular inflammation. In turn, inflammatory disorders of the palpebral conjunctiva and lid margin may affect the structure and function of meibomian gland. The disorders include allergic conjunctivitis, long-term usage of contact lenses, dermatological diseases that affect conjunctival homeostasis, Stevens-Johnson's syndrome or chemical burning of the ocular surface and lid margin.

Bacteriology of the conjunctiva in pre-cataract surgery patients with occluded nasolacrimal ducts and the operation outcomes in Japanese patients.

BACKGROUND: Contamination of the conjunctiva in association with nasolacrimal duct obstruction is by all accounts a risk factor for infectious endophthalmitis post-cataract surgery. METHODS: All patients who underwent cataract day surgery routinely received nasolacrimal duct syringing with normal saline at the Wakayama Medical University Hospital, Japan, from 2011 to 2013. The microorganisms isolated from conjunctival swab samples of patients with occluded nasolacrimal ducts and their susceptibility to antibiotics, as well as the operation outcomes in all the patients were retrospectively investigated. RESULTS: Nasolacrimal duct obstruction was observed in 125 eyes of 90 patients (3.3%; 42 eyes of 30 male individuals, and 83 eyes of 60 female individuals) from a total of 3754 eyes of 2384 patients by using irrigation samples of nasolacrimal ducts. The mean age of the subjects with duct obstruction was 79 ± 8.5 years.. In bacterial cultures of swabs from these 125 individuals, microbial growth was detected in 56 samples (i.e. 44.8%). Coagulase-negative Staphylococcus was detected in 28 eyes, and Corynebacterium species was detected in 17 eyes. Staphylococcus aureus, excluding methicillin-resistant S. aureus was detected in seven eyes with nasolacrimal duct obstruction. Methicillin-resistant S. aureus was isolated in two eyes with nasolacrimal duct obstruction. Each case was treated with topical antibiotics based on the results of antibiotic sensitivity tests. After culturing of cotton swab samples from the conjunctiva, and using direct micrography of bacteria every 2 or 3 days after starting treatment, and once the results were negative (consecutively tested three times), the patients received cataract surgery. In the current case series, bacteria were not detected in conjunctival swabs obtained consecutively three times for 3 weeks after starting topical antibiotics in 118 eyes from 125 eyes (94.4%), and later in the remaining patients. No patient required dacryocystorhinostomy to eliminate bacterial contamination in the conjunctiva following topical antibiotic therapy. No patient developed infectious endophthalmitis at least 1-year post-cataract surgery. CONCLUSIONS: All the patients receiving cataract day surgery underwent the operation after the elimination of conjunctival microorganism contamination in association with nasolacrimal duct obstruction by using appropriate topical antibiotics.

Inhibition of development of laser-induced choroidal neovascularization with suppression of infiltration of macrophages in Smad3-null mice.

We evaluated the effects of the loss of Smad3 on the development of experimental argon laser-induced choroidal neovascularization (CNV) in mice. An in vitro angiogenesis model was also used to examine the role of transforming growth factor-ß1 (TGFß1)/Smad3 signaling in vessel-like tube formation by human umbilical vein endothelial cells (HUVECs). CNV was induced in eyes of 8-12-week-old B6.129-background Smad3-deficient (KO) mice (n=47) and wild-type (WT) mice (n=47) by argon laser irradiation. Results showed that the size of the CNV induced was significantly smaller in KO mice as compared with WT mice at day 14 as revealed by high-resolution angiography with fluorescein isothiocyanate-dextran. Immunohistochemistry and real-time reverse transcription-polymerase chain reaction of RNA extracted from laser-irradiated choroidal tissues were conducted on specimens at specific timepoints. Invasion of macrophages (F4/80+), but not neutrophils (myeloperoxidase+), and appearance of myofibroblasts (α-smooth muscle actin+) were suppressed in laser-irradiated KO tissues. mRNA expression of inflammation-related factors, that is, vascular endothelial growth factor (VEGF), macrophage-chemoattractant protein-1 (MCP-1), interleukin-6 (IL-6) and TGFß1 in choroidal tissues was suppressed by the loss of Smad3. We then examined the effects of adding a Smad3 inhibitor, SIS3, or an ALK5 inhibitor, SB431542, on tube formation promoted by TGFß1 or VEGF in HUVECs cocultured with fibroblast feeder. Further addition of SIS3 or SB431542 augmented vessel-like tube formation by HUVECs in the presence of TGFß1 or VEGF. In conclusion, lack of Smad3 attenuated the growth of laser-induced CNV with suppression of inflammation by macrophages in mice. Because blocking TGFß1/Smad3 signal stimulated the activity of angiogenesis of HUVECs in vitro, the reduction of CNV in vivo in KO mice is attributed to a decrease in growth factor levels in the tissue by the loss of Smad3.