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Juvenile hormone (JH) plays a pivotal role in almost every aspect of insect development and reproduction. The chemical structure of the JH in heteropteran species has long remained elusive until methyl (2R,3S,10R)-2,3;10,11-bisepoxyfarnesoate, commonly named as juvenile hormone III skipped bisepoxide (JHSB3), was isolated from Plautia stali (Hemiptera: Heteroptera: Pentatomidae). Recently, several groups reported the presence of JHSB3 in other heteropteran species. However, most of the studies paid no attention to the determination of the relative and absolute structure of the JH. In this study, we investigated the JH of the cabbage bug Eurydema rugosa (Hemiptera: Heteroptera: Pentatomidae), known as a pest for wild and cultivated crucifers. JHSB3 was detected in the hexane extract from the corpus allatum (CA) product using a chiral ultraperformance liquid chromatography-tandem mass spectrometer (UPLC-MS/MS) which can inform the absolute stereochemistry of the JH. Its stereoisomers were not detected. Topical application of the synthetic JHSB3 to the last instar nymphs inhibited their metamorphosis and induced nymphal-type colouration of the dorsal abdomen in a dose-dependent manner. Additionally, the topical application of JHSB3 effectively terminated summer and winter diapauses in females. These results indicate that the JH of E. rugosa is JHSB3. Although individuals in summer and winter diapauses are physiologically distinct in E. rugosa, the results suggest that the physiological differences between these diapauses are based, not on the responsiveness to JH, but on the processes governing activation of the CA or on its upstream cascades.

Corneal fibroblasts are embedded within an extracellular matrix composed largely of collagen type 1, proteoglycans, and other proteins in the corneal stroma, and their morphology and function are subject to continuous regulation by collagen. During wound healing and in various pathological conditions, corneal fibroblasts differentiate into myofibroblasts characterized by the expression of α-smooth muscle actin (α-SMA). Endo180, also known as urokinase-type plasminogen activator (uPA) receptor-associated protein (uPARAP), is a collagen receptor. Here we investigated whether targeting of Endo180 and the uPA receptor (uPAR) by uPA might play a role in the regulation of α-SMA expression by culturing corneal fibroblasts derived from uPA-deficient ( uPA−/−) or wild-type ( uPA+/+) mice in a collagen gel or on plastic. The expression of α-SMA was upregulated, the amounts of full-length Endo180 and uPAR were increased, and the levels of both transforming growth factor-β (TGF-β) expression and Smad3 phosphorylation were higher in uPA−/− corneal fibroblasts compared with uPA+/+ cells under the collagen gel culture condition. Antibodies to Endo180 inhibited these effects of uPA deficiency on α-SMA and TGF-β expression, whereas a TGF-β signaling inhibitor blocked the effects on Smad3 phosphorylation and α-SMA expression. Our results suggest that uPA deficiency might promote the interaction between collagen and Endo180 and thereby increase α-SMA expression in a manner dependent on TGF-β signaling. Expression of α-SMA is thus negatively regulated by uPA through targeting of Endo180 and uPAR.

The introduction of fetal or neonatal signatures such as cell cycle promoting genes into damaged adult hearts has been vigorously pursued as a promising strategy for stimulating proliferation and regeneration of adult cardiomyocytes, which normally cannot divide. However, cell division of cardiomyocytes requires preceding dedifferentiation with sarcomere disassembly and calcium dysregulation, which, in principle, compromises contractile function. To overcome this intrinsic dilemma, we explored the feasibility of optimizing the induction protocol of the cell cycle promoter in mice. As a model of this approach, we used Fam64a, a fetal-specific cardiomyocyte cell cycle promoter that we have recently identified. We first analyzed transgenic mice maintaining long-term cardiomyocyte-specific expression of Fam64a after birth, when endogenous expression was abolished. Despite having an enhanced proliferation of postnatal cardiomyocytes, these mice showed age-related cardiac dysfunction characterized by sustained cardiomyocyte dedifferentiation, which was reminiscent of the dilemma. Mechanistically, Fam64a inhibited glucocorticoid receptor-mediated transcriptional activation of Klf15, a key regulator that drives cardiomyocyte differentiation, thereby directing cardiomyocytes toward immature undifferentiated states. In contrast, transient induction of Fam64a in cryoinjured wildtype adult mice hearts improved functional recovery with augmented cell cycle activation of cardiomyocytes. These data indicate that optimizing the intensity and duration of the stimulant to avoid excessive cardiomyocyte dedifferentiation could pave the way toward developing efficient strategy for successful heart regeneration.

Purpose. To report a case of corneal perforation, in a patient with a history of herpetic keratitis, during combination chemotherapy including cetuximab. Case. We report the case of a 71-year-old man who was diagnosed with a hypopharyngeal carcinoma and received radiation therapy combined with cetuximab, the epidermal growth factor receptor (EGFR) inhibitor monoclonal antibody. He was referred to us because of ocular hyperemia and corneal perforation in his left eye. In spite of conservative therapy, his corneal perforation was exacerbated, with iris incarceration into the wound site and exposure to the surface of the cornea. He therefore discontinued treatment with the combination chemotherapy and underwent lamellar keratoplasty using a preserved donor cornea. After treatment with cetuximab resumed, there was no recurrence of the corneal perforation. Conclusion. We have presented the first case of cetuximab-related corneal perforation in a patient who had a history of recurrent herpetic keratitis. EGFR inhibitors, such as cetuximab, can induce corneal perforation in cases with a history of herpetic stromal keratitis.

Purpose: To report a case of neurotrophic keratopathy associated with nasopharyngeal carcinoma. Case Report: A 59-year-old man who had been diagnosed with a nasopharyngeal carcinoma was referred to the authors because of visual disturbance and pain in his right eye. Slit-lamp examination revealed a corneal epithelial defect and corneal stromal edema surrounding the epithelial defect area in his right eye. Magnetic resonance imaging showed a mass in his cavernous sinus, which was identified as nasopharyngeal carcinoma (NPC). We diagnosed neurotrophic keratopathy associated with NPC and initiated treatment with preservative-free artificial tears, antibiotic eye drops, fibronectin, a therapeutic contact lens, and amniotic membrane transplantation. However, the persistent corneal epithelial defect was unresponsive to these treatments. Conclusion: Neurotrophic keratopathy secondary to NPC is thought to be rare. We presented a case of neurotrophic keratopathy associated with cavernous sinus metastasis of an NPC. The development of new and more effective treatments for this refractory disease is anticipated.

Connectin, also called titin, is the largest protein with a critical function as a molecular spring during contraction and relaxation of striated muscle; its mutation leads to severe myopathy and cardiomyopathy. To uncover the cause of this pathogenesis, zebrafish have recently been used as disease models because they are easier to genetically modify than mice. Although the gene structures and putative primary structures of zebrafish connectin have been determined, the actual primary structures of zebrafish connectin in heart and skeletal muscles remain unclear because of its large size and the PCR amplification-associated difficulties. In this research, using RT-PCR amplification from zebrafish heart and skeletal muscles, we determined the complete primary structures of zebrafish connectin in the I-band region at which mechanical property is modulated by alternative splicing. Our results showed that the domain structures of zebrafish connectins were largely similar to those of human connectins; however, the splicing pathways in the middle-Ig segment and the PEVK segment were highly diverse in every isoform. We also found that a set of 10 Ig domains in the middle-Ig segment of zebrafish connectin had been triplicated in human connectin. Because these triplicate regions are expressed in human leg and diaphragm, our findings may provide insight into the establishment of walking with limbs and lung respiration during tetrapod evolution.

Keratocytes, corneal resident cells in the corneal stroma, exist between collagen lamellae and maintain the corneal stromal structure. When the corneal stroma is damaged, keratocytes are transformed to myofibroblasts to aid corneal wound healing by phagocytizing debris. Keratocytes and extracellular collagen influence each other because keratocytes cultured in a 3D collagen gel undergo morphological changes and keratocytes produce metalloproteases that degrade extracellular collagen. IL-1 and plasminogen are critical mediators for collagen degradation. The plasminogen system contributes to tissue repair by activating matrix metalloproteinases (MMPs), releasing growth factors from the extracellular matrix and extracellular matrix degradation. Urokinase-type plasminogen activator (uPA) is thought to be involved in corneal disorders and regulates corneal wound healing. uPA is a serine protease synthesized by various cells such as corneal epithelial cells, corneal fibroblasts, vascular endothelial cells, smooth muscle cells, monocytes, macrophages, and malignant tumor cells of different origins. Here, we review the role of uPA in corneal stromal wound healing. uPA is expressed in leukocytes and corneal fibroblasts in the corneas of patients with corneal ulcerations suggesting it is a key regulator of corneal stromal wound healing. uPA is directly involved in plasmin-mediated collagen degradation induced by IL-1. Moreover, uPA is critically involved in promoting leukocyte infiltration in corneal inflammation by activating MMP-9. This activation is presumably directly and indirectly mediated by the plasminogen/plasmin cascade. Moreover, uPA mediates the release of inflammatory cytokines from corneal fibroblasts to promote leukocyte infiltration.

The interaction of keratocytes with extracellular matrix components plays an important role in the maintenance of corneal transparency and shape as well as in the healing of corneal wounds. In particular, the interaction of these cells with collagen and cell-mediated collagen contraction contribute to wound closure. Endo180 is a receptor for collagen that mediates its cellular internalization. We have now examined the role of Endo180 in collagen contraction mediated by corneal fibroblasts (activated keratocytes). Antibodies to Endo180 inhibited the contractile activity of mouse corneal fibroblasts embedded in a three-dimensional collagen gel and cultured in the presence of serum, with this effect being both concentration and time dependent and essentially complete at an antibody concentration of 0.2 μg/ml. Whereas corneal fibroblasts cultured in a collagen gel manifested a flattened morphology with prominent stress fibers under control conditions, they showed a spindlelike shape with few stress fibers in the presence of antibodies to Endo180. Antibodies to Endo180 had no effect on the expression of α–smooth muscle actin or the extent of collagen degradation in collagen gel cultures of corneal fibroblasts. Immunohistofluorescence analysis did not detect the expression of Endo180 in the unwounded mouse cornea. However, Endo180 expression was detected in keratocytes migrating into the wound area at 3 days after a corneal incisional injury. Together, our results suggest that Endo180 is required for the contraction of collagen matrix mediated by corneal fibroblasts and that its expression in these cells may contribute to the healing of corneal stromal wounds.

Loss of cardiomyocyte proliferative capacity after birth is a major obstacle for therapeutic heart regeneration in adult mammals. We and others have recently shown the importance of hypoxic in utero environments for active foetal cardiomyocyte proliferation. Here, we report the unexpected expression of novex-3, the short splice variant of the giant sarcomeric protein connectin (titin), in the cardiomyocyte nucleus specifically during the hypoxic foetal stage in mice. This nuclear localisation appeared to be regulated by the N-terminal region of novex-3, which contains the nuclear localisation signal. Importantly, the nuclear expression of novex-3 in hypoxic foetal cardiomyocytes was repressed at the postnatal stage following the onset of breathing and the resulting elevation of oxygen tension, whereas the sarcomeric expression remained unchanged. Novex-3 knockdown in foetal cardiomyocytes repressed cell cycle-promoting genes and proliferation, whereas novex-3 overexpression enhanced proliferation. Mechanical analysis by atomic force microscopy and microneedle-based tensile tests demonstrated that novex-3 expression in hypoxic foetal cardiomyocytes contributes to the elasticity/compliance of the nucleus at interphase and facilitates proliferation, by promoting phosphorylation-induced disassembly of multimer structures of nuclear lamins. We propose that novex-3 has a previously unrecognised role in promoting cardiomyocyte proliferation specifically at the hypoxic foetal stage.

Purpose. To report a case of persistent corneal epithelial defect that had occurred after a trigeminal nerve block. Case Presentation. A 75-year-old female had suffered from postherpetic neuralgia for 8 years. She underwent Gasserian ganglion block surgery and noticed declining visual acuity in the right eye on the following day. She presented with severe hyperemia and corneal epithelial defects in the right eye and experienced remarkable reduction of sensitivity in the right cornea. She was diagnosed with neurotrophic keratopathy. Ofloxacin eye ointment and rebamipide ophthalmic suspension ameliorated the corneal epithelial defects but superficial punctate keratopathy, corneal superficial neovascularization, and Descemet’s fold persisted. Although the epithelial defects occasionally recurred, the corneal sensation and epithelial defects, Descemet’s fold, and corneal superficial neovascularization all improved around 5 months after trigeminal nerve block. The HRT II Rostock Cornea Module (RCM) could not detect any corneal subbasal nerve fibers at postoperative 4 months; however, it could detect them at postoperative 6 months. Conclusions. As the nerve block effect wore off, the corneal subbasal nerve fibers slowly regenerated. As the corneal sensation improved, the corneal epithelial defects and superficial neovascularization also improved. The HRT II RCM appeared useful for observing loss and regeneration of the corneal subbasal nerve fibers.

Background Graft detachment is a complication of non-Descemet stripping automated endothelial keratoplasty (nDSAEK). We report a case of spontaneous reattachment of an extensively dislocated graft after nDSAEK. Case presentation A 54-year-old male underwent penetrating keratoplasty (PKP) for keratoconus in his left eye in 2001. Following graft opacity due to rejection, a second PKP was implemented in May 2014. The graft was kept in good condition after the reoperation and yet, visual acuity (VA) declined due to cataract. PEA+IOL was then performed in May 2015. Because edema appeared in the graft 6 months after the PEA+IOL, nDSAEK was carried out in May 2016. Although the donor graft well attached immediately after the nDSAEK, the graft was almost completely dislocated 3 h later except a temporal part. Air was reinjected into the anterior chamber on the following day and the detachment was resolved. Despite of the treatment, about 1/5 of the graft remained detached and the detachment deteriorated to 3/4 of the graft 9 days later. Because the patient could not decide whether to undergo another operation immediately, we decided to follow him up first and found that the partially detached graft reattached spontaneously 1 month later during the follow-up. Although the cornea had a mild edema remaining in the superior temporal area, his BCVA improved to 1.0. Three months later, the graft remained in position and the cornea kept its transparency. Conclusions Spontaneous reattachment was observed during the follow-up in a case that had shown a comparatively extensive graft dislocation after nDSAEK.

Fetal cardiomyocytes actively proliferate to form the primitive heart in utero in mammals, but they stop dividing shortly after birth. The identification of essential molecules maintaining this active cardiomyocyte proliferation is indispensable for potential adult heart regeneration. A recent study has shown that this proliferation depends on a low fetal oxygen condition before the onset of breathing at birth. We have established an isolation protocol for mouse fetal cardiomyocytes, performed under strict low oxygen conditions to mimic the intrauterine environment, that gives the highest proliferative activities thus far reported. Oxygen exposure during isolation/culture markedly inhibited cell division and repressed cell cycle-promoting genes, and subsequent genome-wide analysis identified Fam64a as a novel regulatory molecule. Fam64a was abundantly expressed in hypoxic fetal cardiomyocyte nuclei, but this expression was drastically repressed by oxygen exposure, and in postnatal cardiomyocytes following the onset of breathing and the resulting elevation of oxygen tension. Fam64a knockdown inhibited and its overexpression enhanced cardiomyocyte proliferation. Expression of a non-degradable Fam64a mutant suggested that optimum Fam64a expression and subsequent degradation by anaphase-promoting complex/cyclosome (APC/C) during the metaphase-to-anaphase transition are required for fetal cardiomyocyte division. We propose that Fam64a is a novel cell cycle promoter of hypoxic fetal cardiomyocytes in mice.

Purpose Plasminogen has been detected in the corneal stroma after tissue injury and interacts with corneal fibroblasts during wound healing. We examined the effect of plasminogen on phagocytic activity of corneal fibroblasts. Methods Cultured human corneal fibroblasts were exposed to plasminogen and then incubated with fluorescent microparticles before measurement of phagocytic activity by confocal microscopy or flow cytometry. The binding of corneal fibroblasts to immobilized plasminogen was quantitated with a real-time biomolecular interaction assay. The production of urokinase-type plasminogen activator (uPA), matrix metalloproteinases (MMPs), and IL-1β by corneal fibroblasts was measured by fibrin zymography, by immunoblot analysis or gelatin zymography, or with an enzyme-linked immunosorbent assay, respectively. Results Plasminogen increased phagocytic activity of corneal fibroblasts in a concentration- and time-dependent manner, with the maximal effect apparent at 30 μg/mL and 24 hours. Corneal fibroblasts bound to immobilized plasminogen in a manner dependent on time and cell number, and the stimulatory effect of plasminogen on phagocytic activity was blocked in the presence of epsilon-aminocaproic acid, an inhibitor of plasminogen binding to cell surface receptors. Plasminogen-induced phagocytic activity was not associated with changes in the production of uPA, MMPs, or IL-1β by corneal fibroblasts. Conclusions Plasminogen induced phagocytic activity in corneal fibroblasts in a manner dependent on its binding to the cell surface. This effect was not associated with increased production of proteases or IL-1β. Thus, plasminogen may promote the clearance of foreign particles or damaged tissue components by corneal fibroblasts early after tissue injury.

Transforming growth factor-beta (TGF-β) is one of the main epithelial-mesenchymal transition (EMT)-inducing factors. In general, TGF-β-induced EMT promotes cell migration and invasion. TGF-β also acts as a potent regulator of pericellular proteolysis by regulating the expression and secretion of plasminogen activators. Urokinase-type plasminogen activator (uPA) is a serine protease that binds to its cell surface receptor (uPAR) with high affinity. uPA binding to uPAR stimulates uPAR's interaction with transmembrane proteins, such as integrins, to regulate cytoskeletal reorganization and cell migration, differentiation and proliferation. However, the influence of TGF-β and the uPA/uPAR system on EMT in retinal pigment epithelial (RPE) cells is still unclear. The purpose of this study was to determine the effect of TGF-β2, which is the predominant isoform in the retina, and the uPA/uPAR system on RPE cells. In this study, we first examined the effect of TGF-β2 and/or the inhibitor of uPA (u-PA-STOP(®)) on the proliferation of a human retinal pigment epithelial cell line (ARPE-19 cells). Treatment with TGF-β2 or u-PA-STOP(®) suppressed cell proliferation. Combination treatment of TGF-β2 and u-PA-STOP(®) enhanced cell growth suppression. Furthermore, western blot analysis, fibrin zymography and real-time reverse transcription PCR showed that that TGF-β2 induced EMT in ARPE-19 cells and that the expression of uPA and uPAR expression was up-regulated during EMT. The TGF-β inhibitor SB431542 suppressed TGF-β2-stimulated uPA expression and secretion but did not suppress uPAR expression. Furthermore, we seeded ARPE-19 cells onto Transwell chambers and allowed them to invade the collagen matrix in the presence of TGF-β2 alone or with TGF-β2 and u-PA-STOP(®). TGF-β2 treatment induced ARPE-19 cell invasion into the collagen gel. Treatment with a combination of TGF-β2 and the uPA inhibitor strongly inhibited ARPE-19 cell invasion compared with treatment with TGF-β2 alone. Furthermore, the interaction between uPA and ARPE-19 cells was analyzed using a surface plasmon biosensor system. The binding of uPA to ARPE-19 cells was observed. In addition, TGF-β2 significantly promoted the binding activity of uPA to ARPE-19 cells in a time-dependent or cell-number-dependent fashion. These results indicate that TGF-β-induced EMT-associated phenotype changes in ARPE-19 cells and the invasiveness of ARPE-19 cells into a collagen gel matrix are mediated, at least in part, by uPA.

Previous reports showed that cosmetic cleansing oil for removing makeup, which contains mineral oil and surfactant, can deform some silicone hydrogel contact lenses (SHCLs) when applied directly to the lenses, although plasma-coated SHCLs (lotrafilcon A and B) were not affected. In the present study, we investigated hydrogel lenses and SHCLs in both wet and dry conditions.Several brands of hydrogel and SHCLs were immersed in a cleansing oil solution containing Sudan Black B for 5 min under wet and dry conditions. The lenses under the wet condition were simply picked up from the saline, whereas those under the dry condition were blotted with paper wipes. After immersing, the excess solution remaining on the lenses was removed by finger rubbing with a multipurpose solution. The lenses were then examined using a stereomicroscope, and their mean brightness was measured and compared.The cosmetic cleansing oil was not absorbed by the hydrogel lenses under wet or dry conditions. However, four of seven brands of SHCLs absorbed the cosmetic cleansing oil under both conditions (dry and wet), whereas asmofilcon A absorbed it only under the dry condition. Lotrafilcon B and delefilcon A did not absorb cleansing oil even under the dry condition.Hydrogel lenses resist cosmetic cleansing oil. However, SHCLs have different degrees of resistance depending on the lens material. Some SHCLs absorbed cosmetic cleansing oil more under dry conditions than under wet conditions.

Purpose Staphylococcus aureus is a common cause of corneal ulceration, and staphylokinase (SAK) produced by this bacterium is a plasminogen activator. To investigate the pathogenesis of corneal ulceration induced by S. aureus, we examined the effects of bacterial culture broth and SAK on collagen degradation in a culture model in which human corneal fibroblasts are embedded in a collagen gel. Methods Corneal fibroblasts embedded in collagen were exposed to S. aureus culture broth or SAK. Collagen degradation was assessed by measurement of hydroxyproline in acid hydrolysates of culture supernatants. Expression of pro-matrix metalloproteinase-1 (pro-MMP-1) was detected by immunoblot analysis as well as reverse transcription and real-time polymerase chain reaction analysis. Results Both S. aureus culture broth and SAK markedly increased collagen degradation in the presence of corneal fibroblasts and plasminogen. This effect of the culture broth was dependent on cell number to a greater extent than was that of SAK. Whereas the culture broth also increased the expression of pro-MMP-1 in corneal fibroblasts at both mRNA and protein levels, SAK did not. Conclusions Our results suggest that S. aureus may promote collagen degradation both by upregulating pro-MMP1 expression in corneal fibroblasts, with pro-MMP-1 then being converted to active MMP-1 by plasmin, and by directing plasmin activity toward collagen in a SAK-dependent manner.

Corneal wound healing is a complex process involving the integrated actions of various growth factors, cytokines and extracellular matrix produced by corneal cells and inflammatory cells. Connective tissue growth factor (CTGF) has been linked to wound healing, and fibronectin (FN) is a major component of the extracellular matrix. However, the functions of CTGF and FN in corneal epithelial cells are not well understood. We therefore investigated the coordinated function of CTGF and FN in the attachment and migration of corneal epithelial cells. Treatment of human corneal epithelial cells (HCECs) with transforming growth factor (TGF) beta1 up-regulated the expression of CTGF, but did not noticeably affect FN expression, as judged by immunoblot analysis of cell lysates. In contrast, the amount of FN accumulated in the cultured media was increased in a time-dependent manner, but CTGF was undetectable in the cultured media. The expression level of FN was decreased by the knockdown of CTGF expression with a specific short hairpin RNA, indicating that CTGF acts as an upstream mediator of FN expression. CTGF augmented the FN-mediated increase in the attachment of HCEC by about twofold, although CTGF alone did not influence the attachment. Moreover, the migration assay with rabbit corneal blocks revealed that CTGF (390 nM) alone or in combination of FN (10 microg/mL) promoted corneal epithelial migration; the mean migration distances of control, CTGF, and CTGF + FN were 272, 325, and 626, microm, respectively. In conclusion, CTGF cooperates with FN in enhancing the attachment and migration of corneal epithelial cells.

Purpose Keratocytes maintain homeostasis of the corneal stroma through synthesis, secretion, and degradation of collagen fibrils of the extracellular matrix. Given that these cells are essentially embedded in a collagen matrix, keratocyte-collagen interactions may play a key role in regulation of the expression or activation of enzymes responsible for matrix degradation including urokinase-type plasminogen activator (uPA), plasmin, and matrix metalloproteinases (MMPs). We examined the effect of extracellular collagen on the production of uPA by corneal fibroblasts (activated keratocytes) stimulated with the proinflammatory cytokine interleukin-1β (IL-1β). Methods Human corneal fibroblasts were cultured either on plastic or in a three-dimensional gel of type I collagen. Plasminogen activators were detected by fibrin zymography, whereas the IL-1 receptor (IL-1R) and MMPs were detected by immunoblot analysis. Collagen degradation by corneal fibroblasts was assessed by measurement of hydroxyproline in acid hydrolysates of culture supernatants. Results Collagen and IL-1β synergistically increased the synthesis and secretion of uPA in corneal fibroblasts. Collagen also upregulated IL-1R expression in the cells in a concentration-dependent manner. The conversion of extracellular plasminogen to plasmin, as well as the plasminogen-dependent activation of MMP-1 and MMP-3 and degradation of collagen apparent in three-dimensional cultures of corneal fibroblasts exposed to IL-1β, were all abolished by a selective uPA inhibitor. Conclusions Collagen and IL-1β cooperate to upregulate uPA production by corneal fibroblasts. Furthermore, IL-1β-induced collagen degradation by these cells appears to be strictly dependent on uPA expression and mediated by a uPA-plasmin-MMP pathway.

Purpose Urokinase-type plasminogen activator (u-PA) plays an important role in corneal wound healing, yet its role in corneal inflammation remains poorly understood. We investigated the role of u-PA in a murine model of lipopolysaccharide (LPS)-induced corneal inflammation. Methods The corneal epithelium was scraped and LPS was applied to u-PA wild-type (u-PA(+/+)) and u-PA-deficient (u-PA(-/-)) mice. Corneal re-epithelialization and opacity were measured by stereomicroscopy. Fibrin zymography was performed to detect plasminogen activators in corneas from u-PA(+/+) and u-PA(-/-) mice. Neutrophil, macrophage, and u-PA receptor (u-PAR) expression were determined by immunohistochemistry. Gene expression of corneal macrophage chemoattractant protein (MCP)-1 and macrophage inflammatory protein (MIP)-2 was assessed with reverse transcription-polymerase chain reaction. The in vitro effects of endogenous u-PA on MCP-1, MIP-2, matrix metalloproteinase (MMP)-2, and MMP-9 expression, and macrophage migration activity in mouse ocular fibroblasts stimulated by LPS, were examined. Results The u-PA(+/+) mice showed enhanced corneal inflammation as compared with u-PA(-/-) mice. The u-PA expression was increased by LPS stimulation. Immunohistochemical analyses indicated that more neutrophils and macrophages were present in corneas from u-PA(+/+) mice than u-PA(-/-) mice. The u-PAR expression was detected in inflammatory cells and in the leading edges of the epithelial migrating cells. Enhanced mRNA expression of MCP-1 and MIP-2 was observed in corneas from u-PA(+/+) mice compared to u-PA(-/-) mice. Macrophage chemoattractant protein-1, MIP-2, and MMP-9, but not MMP-2, significantly increased in corneal fibroblasts from u-PA(+/+) mice compared with u-PA(-/-) mice. Conclusions These data indicate that u-PA promotes LPS-induced leukocyte infiltration in cornea and that u-PA is an important component in LPS-induced corneal inflammatory responses.

Neovascular glaucoma (NVG) secondary to internal carotid artery (ICA) occlusion is usually resistant to treatment. We report a case of NVG with ICA occlusion improved by intravitreal bevacizumab (IVB) injection and carotid artery stent replacement (CAS), even though we did not perform panretinal photocoagulation. A 67-year-old male with NVG noted visual loss in his left eye. Magnetic resonance angiography showed left ICA occlusion. He was diagnosed with NVG secondary to ICA occlusion. The next day, we carried out IVB injection in his left eye, following which the iris and angle neovascularization regressed, and the intraocular pressure decreased to normal within a day after the injection. CAS was performed on his left ICA at a month post injection. Two months later, we reinjected bevacizumab in his left eye. His condition remained stable with no recurrence over two years. This case indicates that IVB injection and CAS are useful for early-stage NVG secondary to ICA occlusion.

Background The exact pathogenetic mechanisms of Coats’ disease remain unknown. In this report, we show two cases of Coats’ disease that achieved a favorable prognosis with the combined treatment of intravitreal bevacizumab (IVB) injection prior to photocoagulation, although both initially resisted photocoagulation therapy. Case presentations Case 1 was a 15-year-old boy with initial visual acuity of 0.4 OD. At the temporal retina, aneurysms and abnormal telangiectatic vessels were observed. Hard exudates and an exudative retinal detachment extended to the fovea. He was diagnosed as having Coats’ disease at stage 3A and we performed laser photocoagulation as an initial approach to treat peripheral aneurysms and telangiectatic vessels. After the treatment, the exudative retinal detachment was eased and visual acuity improved to 1.0; however, recurrence occurred after 5 months. The exudative change was resistant against laser photocoagulation therapy and we therefore added IVB as an adjuvant before photocoagulation. Fourteen days after IVB injection phased laser photocoagulation was given to cover the abnormal capillaries, aneurysms and the leakage area spotted in FA. A good prognosis was obtained with decreased exudation and improved visual acuity. Case 2 was an 11-year-old boy with decreased visual acuity of 0.15 OS at the initial visit. Hard exudates, retinal edema and serous retinal detachment were seen at the macula and peripheral retina. Fluorescein angiography revealed telangiectatic capillaries at the temporal retina. Our diagnosis was Coats’ disease at stage 3A. Extensive photocoagulation was performed as an initial treatment to the lesion. However, the exudative change was severe and resistant against the photocoagulation treatment. Therefore, we added IVB as an adjuvant before photocoagulation. Exudative change in the retina seemed to be eased 7 days after IVB injection, therefore, phased laser phototherapy was added to cover the abnormal capillaries. After the combination therapy, exudative change was remarkably ameliorated and better visual acuity was achieved. Conclusion Bevacizumab is considered an effective adjuvant for Coats’ disease with exudative change resistant to retinal photocoagulation therapy.

Background Corneal ulceration leading to perforation is associated with infectious and non-infectious destructive conditions in the cornea. The fibrinolytic (plasminogen/plasmin) system is considered to contribute to tissue remodeling in the wound healing process and it is believed to play an important role in proteolysis and fibrosis. To determine the localization of urokinase-type plasminogen activator (u-PA), u-PA receptor (u-PAR) and α2-antiplasmin (α2AP) in the tissue of a corneal perforation, we investigated immunohistochemical expressions of u-PA, u-PAR, α2AP, CD68, and α-smooth muscle actin (α-SMA) in a patient with corneal perforation that developed from an ulcer of no clear cause. Case presentation The patient was a 77-year-old woman who presented with a perforated corneal ulcer in her right eye. The cause of her corneal ulcer was unknown. Double immunohistochemistry was performed for the combinations of u-PA with u-PAR, CD68 or α-SMA and α2AP with CD68 or α-SMA to detect the localization of u-PA and α2AP. u-PA and u-PAR co-localization was seen in the corneal ulceration area. u-PA was mainly observed in CD68-positive cells and in some α-SMA positive cells. On the other hand, α2AP was not expressed in CD68-positive cells, but was expressed in α-SMA positive cells. Conclusion We identified expression of the u-PA/u-PAR complex and α2AP in a patient with a corneal ulcer. These two molecules are believed to play a crucial role in inflammatory cell recruitment, ECM synthesis and degradation during corneal wound healing.