Histopathologic Change in the Anterior Part of the Posterior Fixation Suture After a Faden Operation in an Animal Model.

PURPOSE: To observe postoperative histological changes in the anterior part of the posterior fixation suture after a Faden operation in an animal model. METHODS: A posterior fixation suture was placed at two points 6 mm posterior to the insertion of the extraocular muscle on the superior rectus muscle of the right eye in eight rabbits. The superior rectus muscle of the left eye was used as a control. The eyes were enucleated and the anterior portion of the posterior fixation suture, including the myoscleral junction, was extracted 4 weeks after surgery. Postoperative adhesion was graded from 0 to 4 based on histologic findings (hematoxylin-eosin and Masson's trichrome staining). RESULTS: Histological evaluation revealed diffuse fibrosis at the myoscleral junction and the anterior part of the posterior fixation suture after the Faden operation. The graded scores for fibrosis, acute inflammation, chronic inflammation, and foreign body reactions in the Faden operation group were significantly higher than those in the control group (P < .05). Postoperative diffuse fibrosis of the myoscleral junction and anterior-to-posterior fixation sutures were observed in an animal model. CONCLUSIONS: Histologic changes may affect ocular alignment and motility, making reoperation unpredictable after the Faden procedure. [J Pediatr Ophthalmol Strabismus. 20XX;X(X):XX-XX.].

Effects of particulate matter exposure on the expression of the SARS-CoV-2 ACE2 receptor in ocular surface tissues and cells.

Particulate matter (PM) has been reported to be one of the risk factor for COVID-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, although the ocular surface is deeply affected by both PM exposure and SARS-COV-2 infection, no studies have investigated the effects of PM exposure on the ocular route of SARS-COV-2 infection. To this end, we explored the effects of PM on the expression of SARS-COV-2-associated receptors and proteins in ocular surface. Herein, short- and long-term PM-exposed rat models were established by topically administering PM for 3 and 10 days, respectively. Immortalized human corneal epithelial cells (HCECs) and human conjunctival epithelial cells (HCjECs) were exposed to PM. ACE2, TMPRSS2, CD147, and ADAM17 expression levels were measured by western blot analysis. Our results show that short-term PM exposure had little effect on the expressions of ACE2, TMPRSS2, and CD147 in ocular surface tissues. However, long-term PM exposure decreased the ACE2 expression in conjunctival tissues and increased the CD147 expression in corneal or conjunctival tissues. PM exposure reduced the ACE2 expression by increasing the ADAM17 expression and ACE2 shedding level in HCECs and HCjECs. Our findings suggest that long-term PM exposure down-regulate the expression of the SARS-CoV-2 receptor ACE2 in conjunctival tissues through ADAM17-dependent ACE2 shedding. However, long-term PM exposure up-regulates the expression of another SARS-CoV-2 receptor CD147 in ocular surface tissues, accompanied by ocular surface damage and cytotoxicity. This study provides a new insight into uncovering potential risk factors for infection with SARS-CoV-2 via the ocular route.

Differences in vulnerability to desiccating stress between corneal and conjunctival epithelium in rabbit models of short-term ocular surface exposure.

We evaluate the difference in vulnerability to desiccating stress (DS) between the corneal and conjunctival epithelia to understand different ocular surface staining patterns in dry eye patients. We generated a rabbit model of short-term exposure keratopathy. To induce DS in the ocular surface, rabbit right eyelids were opened for 30 min, with blinking once/minute. Corneal staining scores increased from 3-min post-DS exposure, while conjunctival staining increased from 20-min post-DS. At 20 min, the tear MUC5AC level doubled as compared to pre-DS (p = 0.007). In Western blot analysis, conjunctival AQP5, MUC5AC, and CFTR expression increased significantly in response to DS, compared to control (p = 0.039, 0.002, 0.039, respectively). Immunohistochemistry for CD31 and LYVE-1 were performed. CD31-positive cells and lymphatic space surrounded by LYVE-1-positive cells increased significantly in conjunctival tissue post-DS, compared to control (p = 0.0006, p < 0.0001, respectively). Surface damage was worse in the corneal than in the conjunctival epithelium after DS, by scanning electron microscopy. This study showed that the cornea and conjunctival epithelium show differences in vulnerability to DS. Increased blood vessels and dilated lymphatics, accompanied by increased conjunctival epithelial AQP5, MUC5AC, and CFTR expression, underlie the protective mechanism of the conjunctiva to desiccating stress.

SIRT1 Protects Against Particulate Matter-Induced Oxidative Stress in Human Corneal and Conjunctival Epithelial Cells.

Purpose: Sirtuin1 (SIRT1) as a hot therapeutic target for oxidative stress-associated diseases that has been extensively studied. This study aimed to determine the changes in SIRT1 expression in particulate matter (PM)-induced corneal and conjunctival epithelial cell damage and explore potential drugs to reduce PM-associated ocular surface injury. Methods: Immortalized human corneal epithelial cells (HCECs) and human conjunctival epithelial cells (HCjECs) were exposed to an ambient PM sample. Cytotoxicity was evaluated by water-soluble tetrazolium salt-8 assay. SIRT1 expression was measured by Western blot analysis. Reactive oxygen species (ROS) production, cell apoptosis, mitochondrial function, and cell senescence were assessed by using 2',7'-dichlorofluorescein diacetate assay, annexin V apoptosis assay, tetramethylrhodamine ethyl ester assay, and senescence ß-galactosidase staining, respectively. Results: PM-induced cytotoxicity of HCECs and HCjECs occurred in a dose-dependent manner. Increased ROS production, as well as decreased SIRT1 expression, were observed in HCECs and HCjECs after 200 µg/mL PM exposure. In addition, PM induced oxidative stress-mediated cellular damage, including cell apoptosis, mitochondrial damage, and cell senescence. Interestingly, SRT1720, a SIRT1 activator, increased SIRT1 expression and decreased ROS production and attenuated PM-induced cell damage in HCECs and HCjECs. Conclusions: This study determined that SIRT1 was involved in PM-induced oxidative stress in HCECs and HCjECs and found that ROS overproduction may a key factor in PM-induced SIRT1 downregulation. The SIRT1 activator, SRT1720, can effectively upregulate SIRT1 expression and inhibit ROS production, thereby reversing PM-induced cell damage. This study provides a new potential target for clinical treatment of PM-associated ocular surface diseases.

Increased angiogenic factors in the aqueous and vitreous humors after disinsertion of extraocular muscle and the effects of triamcinolone acetate injection.

The four extraocular rectus muscles in the rabbits were disinserted for induction of anterior segment ischemia (ASI) and the changes in the concentrations of prostaglandin E2 (PGE2), hypoxia-inducible factor-1 (HIF-1α), and vascular endothelial growth factor (VEGF) in the aqueous and vitreous humor were evaluated. Disinsertion of four rectus muscles in rabbits was performed in the right eyes of rabbits (ASI group). The concentrations of PGE2, HIF-1α, and VEGF in the aqueous and vitreous humor were measured at 1, 3, 6, 12, and 24 h by ELISA. The concentrations were compared with those of the fellow eyes (contralateral group) and normal healthy eyes (control group). Subconjunctival injection of triamcinolone acetonide (TA) was administered and three cytokine concentrations in the aqueous humor and vitreous humor were measured at 12 h after TA injection. A total of 48 eyes from 28 rabbits were included. The concentrations of PGE2, HIF-1α, and VEGF in the aqueous humor in the ASI and contralateral groups were significantly higher than those in the control group (p < 0.05, all). The aqueous and vitreous humor concentrations of VEGF in eyes with simultaneous TA injection were significantly lower than were those in the ASI group (p = 0.02, all). The concentration of PGE2, HIF-1α, and VEGF in the aqueous humor was increased after induction of ASI and TA injection seems to be effective in inhibiting VEGF elevation in ASI.

Comparison of cytotoxicity effects induced by four different types of nanoparticles in human corneal and conjunctival epithelial cells.

The impact of particulate matter (PM) on ocular surface health has attracted increased attention in recent years. Previous studies have reported that differences in the chemical composition of PM can affect the toxicological response. However, available information on the toxic effects of chemical components of PM on the ocular surface is insufficient. In this paper, we aimed to investigate the toxicity effects of chemical components of PM on the ocular surface, focusing on the effects of four different types of nanoparticles (NPs) in human corneal epithelial cells (HCECs) and human conjunctival epithelial cells (HCjECs), which include titanium dioxide (TiO2), carbon black (CB), zinc dioxide (ZnO), and silicon dioxide (SiO2). We found that the in vitro cytotoxic effects of CB, ZnO, and SiO2 NPs are dependent on particle properties and cell type as well as the exposure concentration and time. Here, the order of increasing toxicity was SiO2 → CB → ZnO, while TiO2 demonstrated no toxicity. Moreover, toxic effects appearing more severe in HCECs than HCjECs. Reactive oxygen species (ROS)-mediated oxidative stress plays a key role in the toxicity of these three NPs in HCECs and HCjECs, leading to apoptosis and mitochondrial damage, which are also important contributors to aging. Sirtuin1 (SIRT1) as an NAD+-dependent protein deacetylase that seems to play a potential protective role in this process. These findings implied that ROS and/or SIRT1 may become a potential target of clinical treatment of PM- or NP-related ocular surface diseases.

Change in rat meibomian gland structure with external pressure on the eyelid.

PURPOSE: This study investigated structural changes in rat meibomian glands following repeated and sustained application of external pressure on the eyelids using a magnet and then subsequent removal of the external pressure. METHODS: Twenty-eight Sprague-Dawley rats were used. The upper eyelid was externally compressed using a pair of magnets. One magnet was placed inside the upper eyelid, another was placed outside the eyelid, and varying periods of pressure were investigated. Untreated eyes were used as controls. Meibography was performed, and the transverse eyelid tissue sections were stained with hematoxylin and eosin and anti-cytokeratin 5 antibody at one hour, two and four weeks after removing the magnets. RESULTS: Meibography showed increased meibomian gland loss (30.0 ± 5.0%), and tissue sections showed decreased area of secretory acini (0.04 ± 0.08 mm2) at one hour after applying external pressure using magnets versus in the control eyes (5.0 ± 5.0% and 0.08 ± 0.08 mm2, respectively). On the other hand, there was no meibomian gland loss or reduction of the area of secretory acini at two and four weeks after removing the magnets in comparison with the control eyes. CONCLUSIONS: Repeated and sustained application of external pressure on the eyelid could induce meibomian gland loss; however, this meibomian gland loss can be restored when the external pressure is removed. Therefore, the repeated application of external pressure on the eyelid is a safe treatment method for obstructive MGD.

Self-sealing hyaluronic acid-coated 30-gauge intravitreal injection needles for preventing vitreous and drug reflux through needle passage.

Self-sealing hyaluronic acid (HA)-coated self-sealing 30-gauge needles exhibiting instant leakage prevention of intravitreal humor and injected drug were developed in this study. Ninety New Zealand rabbits were used in this study. We assessed dye regurgitation in intravitreal ICG dye injections using HA-coated needles (HA needle group) and conventional needles (control group). Vitreous humor levels of anti-vascular endothelial growth factor (VEGF) were compared between groups one, three, and seven days after intravitreal bevacizumab (0.016 mL) injections. Expression levels of inflammatory cytokines in the aqueous humor and vitreous humor, including prostaglandin E2 (PGE2), interferon-γ, tumor necrosis factor-α, interleukin (IL)-1ß, IL-4, IL-6, IL-17, and IL-8, were compared between HA needle, control, and normal (in which intravitreal injection was not performed) groups following 12 intravitreal injections over a period of one week. In the HA needle group, HA remained at the injection site and blocked the hole after intravitreal injection. Dye regurgitation occurred significantly less frequently in the HA needle group (16.7%) than the control group (55.6%) after intravitreal ICG dye injection. Meanwhile, vitreous anti-VEGF levels were markedly higher in the HA needle group than the control group one and three days after intravitreal bevacizumab injections. After 12 intravitreal injections, expression levels of aqueous and vitreous IL-8 significantly increased in the control group compared to the HA needle and normal groups. Conversely, there were no significant differences in the expression of the other seven cytokines among the three groups. Intravitreal injections using HA-coated self-sealing 30-gauge needles can block the outflow of vitreous humor and drugs through the needle passage.

The Protective Effect of an Eye Wash Solution on the Ocular Surface Damage Induced by Airborne Carbon Black Exposure.

PURPOSE: To investigate the effects of an eye wash solution on the ocular surface damage induced by airborne carbon black (CB) exposure. METHODS: Sprague-Dawley rats were exposed to ambient CB for 5 days. During the 5 days, a commercial eye wash solution (Eyebon-W) was used for irrigation twice daily on CB-exposed rat eyes; normal saline was used as the vehicle control. Lactic dehydrogenase (LDH) activity and matrix metallopeptidase (MMP)-9, histamine, and lactoferrin levels were measured in tears. The expression of inflammatory cytokines in the anterior segment of the eyeball was measured by Western blot analysis. RESULTS: The ocular surface staining scores, tear LDH activity, tear MMP-9, histamine, and lactoferrin concentrations, and the expression of interleukin-4 and interferon-γ in the eye were significantly increased in the CB group versus the normal control group. When compared with CB group, the Eyebon-W eye wash treatment significantly reversed these elevations induced by CB, including ocular staining scores, tear LDH activity, histamine and MMP-9 concentrations in the tear fluid, and the expression of interleukin-4 in the eye. On the other hand, saline irrigation only reduced the concentrations of histamine and MMP-9 in tear fluid and the expression of interferon-γ in the eye. CONCLUSIONS: Both Eyebon-W eye wash treatment and saline irrigation reversed CB-induced ocular surface injury, but the efficacy of Eyebon-W was more significant than that of the saline solution when compared with CB group. The use of an eye wash solution seems to play a protective role for the ocular surface when exposed to airborne particulate matter.

The Protective Effect of a Topical Mucin Secretagogue on Ocular Surface Damage Induced by Airborne Carbon Black Exposure.

Purpose: Exposure to airborne particulate matter can induce ocular surface damage and inflammation. We evaluated the effects of a topical mucin secretagogue on the mitigation of ocular surface damage induced by exposure to airborne carbon black (CB). Methods: Sprague-Dawley rats were exposed to ambient CB for 2 hours twice daily for 5 days. Corneal staining score and tear lactic dehydrogenase (LDH) activity were measured to evaluate ocular surface damage. Serum immunoglobulin (Ig) G and IgE levels and the sizes of cervical lymph nodes were also measured. The expressions of interleukin (IL)-4, IL-17, and interferon (IFN)-γ were measured by Western blot analysis. Diquafosol tetrasodium was instilled six times a day for 5 days, and the extent of ocular surface damage was evaluated. Results: After exposure to airborne CB, the median corneal staining score and LDH activity were significantly increased. Serum IgG and IgE levels and the sizes of cervical lymph nodes were also significantly increased. Additionally, the expression of IL-4 and IFN-γ was elevated in the anterior segment of the eyeball. Furthermore, the expression of IL-4, IL-17, and IFN-γ was elevated in the cervical lymph nodes. When exposed to airborne black carbon, topical diquafosol tetrasodium significantly increased tear MUC5AC concentration and decreased tear LDH activity. Conclusions: Exposure to airborne CB induced ocular surface damage and increased proinflammatory cytokines in the eyes and cervical lymph nodes. Topical mucin secretagogues seem to have a protective effect on the ocular surface against exposure to airborne particulate matters.

Effects of Topical Mucolytic Agents on the Tears and Ocular Surface: A Plausible Animal Model of Mucin-Deficient Dry Eye.

Purpose: A topical mucolytic agent, N-acetylcysteine (NAC), has been used to create an animal model without the intestinal mucus layer. In this study, we investigated the effects of topical NAC on the tears and ocular surface. Methods: NAC-treated models were established by topically administering 10% NAC four times daily for 5 days in male Sprague-Dawley rats. Clinical parameters and the expression of mucin proteins and genes were evaluated. Alterations in the conjunctival epithelium and goblet cells were observed. Results: The NAC group showed significant decreases in tear secretion, corneal wetting ability, tear MUC5AC concentration, and conjunctival goblet cell numbers as compared with the control group (all P < 0.01). In addition, significant increases in corneal fluorescein score and rose bengal scores were observed in the NAC group versus in the control group (P < 0.05 and P < 0.01, respectively). Hematoxylin and eosin (H&E) staining and scanning electron microscopy clearly showed damage in the epithelial cell layer and microvilli of the NAC group. Although there was no significant difference in MUC16 gene expression, the MUC16 concentration of the tear film and ocular surface tissue was significantly increased in the NAC group versus in the control group (P < 0.01 and P < 0.05, respectively). Five-day treatment with 3% diquafosol had minimal therapeutic effect in NAC-treated rat eyes. Conclusions: Topical administration of 10% NAC induced ocular surface damage and tear film instability by prompting MUC16 disruption and release from the ocular surface. This animal model could be used to study dry eye disease, especially the mucin-deficiency subtype.

Effects of Menthol-Containing Artificial Tears on Tear Stimulation and Ocular Surface Integrity in Normal and Dry Eye Rat Models.

PURPOSE: To evaluate the effects of menthol-containing artificial tears on tear stimulation and ocular surface integrity in normal and dry eye rat models. METHODS: A total of 54 male Lewis rats were used. The levels of tear secretion and tear MUC5AC concentrations were compared between the menthol-containing artificial tear-treated group (menthol group) and the vehicle-treated group (vehicle group). The groups were compared after a single instillation to evaluate the immediate effects, and after repeated instillation (five times a day for 5 days) to evaluate the longer-term effects. Tear lactate dehydrogenase (LDH) activity was measured to evaluate eye drop instillation-induced ocular surface damage. The effects of menthol-containing artificial tears were also evaluated in a dry eye rat model. RESULTS: After a single instillation of menthol-containing artificial tears, tear secretion increased from 4.37 (±0.75) mm at baseline to 7.37 (±1.60) mm. However, after repeated instillations, the effects of tear stimulation decreased. The tear MUC5AC concentration was significantly lower in the menthol group than in the vehicle group after a single instillation, but not after repeated instillation. However, the tear LDH concentration was significantly increased in the menthol group after repeated instillation. In the dry eye rat model, the extent of menthol-induced tear stimulation was reduced. CONCLUSIONS: Menthol-containing artificial tears increased tear secretion, but lowered the tear MUC5AC concentration. Menthol-induced tear stimulation was reduced after repeated instillation for 5 days and in the dry eye rat model. Conversely, repeated instillation of menthol-induced ocular surface damage, resulting in increased tear LDH activity.

Meibomian Glands and Ocular Surface Changes After Closure of Meibomian Gland Orifices in Rabbits.

PURPOSE: To investigate changes in the corneal surface, tear film, and meibomian glands after meibomian gland orifice closure. METHODS: Seventy-two eyes of 36 rabbits were used. In the study group (n = 36), the meibomian gland orifices of both upper and lower eyelids in the right eyes were electrosurgically coagulated. The 36 untreated left eyes were used as controls. Corneal wetting properties were measured 1, 3, 7, and 14 days after coagulation. The eyelid sections were stained with anti-cytokeratin (CK)1, CK5, and CK6 antibody 8 weeks after coagulation. The area of the secretory acini around one meibomian gland duct was measured, and meibography of rabbits was performed 8 weeks after meibomian gland orifice closure. RESULTS: Three days after coagulation, the corneal wetting property was decreased compared with controls. The meibomian gland ducts gradually dilated in the study group over time. The epithelium of the central ducts in both groups was stained with CK5 and CK6, but not CK1. Although the mean area of the secretory acini in the study group (0.10 ± 0.04 mm) was significantly smaller than that of the control group (0.18 ± 0.04; P = 0.004), meibography showed normal morphology in both study and control groups. CONCLUSIONS: Meibomian gland orifice closure reduced corneal wetting property and induced meibomian gland duct dilation accompanied by shrinkage of secretory acini. Meibography could not detect early changes in the meibomian gland after closure of the orifice. Therefore, when the orifice is obstructed, more active treatments are needed before structural changes occur.

NUDT15 p.R139C variant is common and strongly associated with azathioprine-induced early leukopenia and severe alopecia in Korean patients with various neurological diseases.

Azathioprine (AZA)-induced leukopenia is a relatively common complication in Korean patients. In addition to variation in TPMT (thiopurine S-methyltransferase), the NUDT15 p.R139C variant was recently identified to have a strong association with AZA-induced leukopenia. We investigated these associations in Korean patients undergoing AZA treatment with various neurological diseases. Among 84 enrolled patients, 20 (23.8%; 7 early, 13 late) exhibited leukopenia. The NUDT15 p.R139C variant was associated with leukopenia (OR: 11.844, 95% CI 3.984-36.024, p=1.327 × 10-5). The allelic frequency of NUDT15 p.R139C was as high as 10.7% and the frequency of the C/C, C/T, and T/T genotypes was 84.5, 10.7, and 5.9%, respectively. All T/T homozygous patients (5/5) developed early severe-grade leukopenia (white blood cells <1000mm-3) and severe alopecia. NUDT15 p.R139C was strongly associated with early leukopenia and severe alopecia (OR for early leukopenia: 107.624, 95% CI 18.857-614.250, p=1.403 × 10-7, OR for severe alopecia: 77.152, 95% CI 17.378-342.526, p=1.101 × 10-8). The sensitivity and specificity for predicting AZA-induced early leukopenia were 85.7% and 92.2%, respectively. Therefore, the NUDT15 p.R139C variant is common and strongly associated with AZA-induced early leukopenia and severe alopecia in Korean patients with various neurological diseases.

Comparing the Effects of Particulate Matter on the Ocular Surfaces of Normal Eyes and a Dry Eye Rat Model.

PURPOSE: To compare the effect of exposure to particulate matter on the ocular surface of normal and experimental dry eye (EDE) rat models. METHODS: Titanium dioxide (TiO2) nanoparticles were used as the particulate matter. Rats were divided into 4 groups: normal control group, TiO2 challenge group of the normal model, EDE control group, and TiO2 challenge group of the EDE model. After 24 hours, corneal clarity was compared and tear samples were collected for quantification of lactate dehydrogenase, MUC5AC, and tumor necrosis factor-α concentrations. The periorbital tissues were used to evaluate the inflammatory cell infiltration and detect apoptotic cells. RESULTS: The corneal clarity score was greater in the EDE model than in the normal model. The score increased after TiO2 challenge in each group compared with each control group (normal control vs. TiO2 challenge group, 0.0 ± 0.0 vs. 0.8 ± 0.6, P = 0.024; EDE control vs. TiO2 challenge group, 2.2 ± 0.6 vs. 3.8 ± 0.4, P = 0.026). The tear lactate dehydrogenase level and inflammatory cell infiltration on the ocular surface were higher in the EDE model than in the normal model. These measurements increased significantly in both normal and EDE models after TiO2 challenge. The tumor necrosis factor-α levels and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive cells were also higher in the EDE model than in the normal model. CONCLUSIONS: TiO2 nanoparticle exposure on the ocular surface had a more prominent effect in the EDE model than it did in the normal model. The ocular surface of dry eyes seems to be more vulnerable to fine dust of air pollution than that of normal eyes.

Immediate Effect of 3% Diquafosol Ophthalmic Solution on Tear MUC5AC Concentration and Corneal Wetting Ability in Normal and Experimental Keratoconjunctivitis Sicca Rat Models.

PURPOSE: To evaluate the immediate effect of 3% diquafosol ophthalmic solution on tear MUC5AC concentration, periodic acid-Schiff (PAS)-positive goblet cells, and tear film stability in normal and keratoconjunctivitis sicca (KCS) rat models. METHODS: Rats were divided into normal and KCS groups. 3% of diquafosol solution was instilled into the right eye and normal saline into the left eye in both groups. To determine the peak time of tear MUC5AC concentration, tears were collected after 3% diquafosol instillation every 5 min up to 20 min. The tear film stability and the numbers of PAS-positive goblet cells were compared in both models. RESULTS: After diquafosol instillation, tear MUC5AC concentration increased steadily for 15 min, at which point the MUC5AC concentration reached its peak. In both normal and KCS groups, the MUC5AC concentration at 15 min was higher after instillation of 3% diquafosol solution (17.77 ± 2.09 ng/ml in the normal group, 9.65 ± 3.51 ng/ml in the KCS group) than that after saline instillation (13.74 ± 2.87 ng/ml in the normal group, 8.19 ± 3.99 ng/ml in the KCS group) (p = 0.018 for both). The corneal wetting ability was significantly longer after instillation of 3% diquafosol solution compared with that after instillation of normal saline in the normal group (p = 0.018). The percentage of PAS-positive goblet cells after the instillation of 3% diquafosol solution was significantly lower than that after instillation of normal saline in both models (p = 0.018 for both). CONCLUSIONS: Diquafosol ophthalmic solution was effective in stimulating mucin secretion in both normal and KCS rat models, and the peak time of tear MUC5AC concentration was 15 min after diquafosol instillation. The increased tear MUC5AC concentration was accompanied by improved tear film stability and a decreased percentage of PAS-positive goblet cells.

The Effect of Ambient Titanium Dioxide Microparticle Exposure to the Ocular Surface on the Expression of Inflammatory Cytokines in the Eye and Cervical Lymph Nodes.

Purpose: To investigate the ocular immune response following exposure to airborne titanium dioxide (TiO2) microparticles. Methods: Rats in the TiO2-exposed group (n = 10) were exposed to TiO2 particles for 2 hours twice daily for 5 days, while the controls (n = 10) were not. Corneal staining score and tear lactic dehydrogenase (LDH) activity were measured to evaluate ocular surface damage, serum immunoglobulin (Ig) G and E were assayed by using enzyme-linked immunosorbent assay, and the size of cervical lymph nodes was measured. In addition, the expression of interleukin (IL)-4, IL-17, and interferon (IFN)-γ in the anterior segment of the eyeball and cervical lymph nodes was measured by immunohistochemistry, real-time reverse transcription-polymerase chain reaction (RT-PCR), and Western blot analysis. Results: Median corneal staining score (3.0), tear LDH activity (0.24 optical density [OD]), and cervical lymph node size (36.9 mm2) were significantly higher in the TiO2-exposed group than in the control group (1.0, 0.13 OD, and 26.7 mm2, respectively). Serum IgG and IgE levels were found to be significantly elevated in the TiO2-exposed group (P = 0.021 and P = 0.021, respectively). Interleukin 4 expression was increased in the anterior segment of the eyeball and lymph nodes following TiO2 exposure, as measured by immunostaining, real-time RT-PCR, and Western blot. In addition, IL-17 and IFN-γ levels were also increased following TiO2 exposure compared to controls as measured by immunostaining. Conclusions: Exposure to airborne TiO2 induced ocular surface damage. The Type 2 helper T-cell pathway seems to play a dominant role in the ocular immune response following airborne TiO2 exposure.

NUDT15 variant is the most common variant associated with thiopurine-induced early leukopenia and alopecia in Korean pediatric patients with Crohn's disease.

PURPOSE: Thiopurine-induced leukopenia is a relatively common adverse event related to thiopurine medication in Korean pediatric Crohn's disease. In addition to the mutations of TPMT gene, the NUDT15 c.415C>T variant was recently identified to have a strong association with thiopurine-induced early leukopenia. We conducted this study to define the incidence of azathioprine (AZA)-related leukopenia and to determine the incidence and characteristics of their genetic variants in Korean pediatric Crohn's disease patients. PATIENTS AND METHODS: Patients diagnosed with pediatric Crohn's disease who had used AZA for more than 3 months were recruited. The dose and duration of medication and data regarding adverse events including leukopenia were collected. TPMT and NUDT15 gene sequencing was performed for patients who had experienced AZA-induced leukopenia. RESULTS: A total of 81 patients had used AZA as a maintenance therapy of Crohn's disease. The mean dose of AZA was 1.88±0.39 mg/kg/day. Nine patients (11.1%) experienced AZA-induced leukopenia, and eight patients (9.9%) experienced AZA-induced early leukopenia. Among the eight early leukopenia patients, six patients (75.0%) harbored the NUDT15 c.415C>T variant and one patient (12.5%) had the TPMT c.719A>G (TPMT*3C) variant. All the three patients with NUDT15 c.415C>T homozygous variant suffered from alopecia totalis, and two of them experienced severe systemic infection. Three patients with the NUDT15 heterozygous variant are currently treated with AZA at a dose of 0.76 mg/kg/day. CONCLUSION: Mutations of the NUDT15 and TPMT gene accounted for ∼88% of cases with thiopurine-induced early leukopenia. Extensive hair loss was a recognizable early symptom in patients with the homozygous NUDT15 c.415C>T variant. Sequencing of the NUDT15 genes can guide the clinicians before thiopurine medication. An alternative immunosuppressive medication is recommended for patients with homozygous NUDT15 c.415C>T variant. For those with the heterozygous variant, half the usual dose of AZA can achieve efficacy comparable to that for wild-type patients.

Effect of Titanium Dioxide Nanoparticle Exposure on the Ocular Surface: An Animal Study.

PURPOSE: To evaluate the effect of titanium dioxide (TiO2) nanoparticle exposure on the ocular surface. METHODS: Eighty eyes of 40 rabbits were used. The TiO2-1D group (n = 20) received a single instillation of TiO2 in the right eye. The TiO2-4D group (n = 20) received a TiO2 instillation in the right eye once a day for four days. The 40 untreated left eyes were used as controls. Ocular surface staining (n = 5 for each group) was performed with rose bengal dye, tear secretion (n = 5) was measured using the phenol red thread test, lactic dehydrogenase (LDH) activity (n = 5) and MUC5AC levels (n = 5) were measured in tears, and the area of the conjunctival goblet cells (n = 5) was measured through impression cytology and scanning electron microscopy 24 hours after the last TiO2 instillation. RESULTS: Ocular surface staining was increased but the tear secretion was not changed after TiO2 exposure. The TiO2-1D (1.39 OD) and TiO2-4D groups (0.58 OD) had higher median tear LDH activity than the control groups (0.57 OD and 0.29 OD, respectively). Although the median tear MUC5AC level in the TiO2-1D group (92.7 ng/ml) was higher than that of control 1 group (37.4 ng/ml), there was no significant difference in MUC5AC levels between the TiO2-4D and control 2 groups. Conjunctival goblet cell area decreased after TiO2 exposure. CONCLUSIONS: Exposure to TiO2 nanoparticles induced ocular surface damage. Although the tear MUC5AC level increased after a single exposure, it decreased to normal levels after repeated exposures. The area of conjunctival goblet cells decreased after TiO2 exposure.

NEB-related core-rod myopathy with distinct clinical and pathological features.

INTRODUCTION: Mutations in the gene encoding nebulin (NEB) are known to cause several types of congenital myopathy including recessive nemaline myopathy and distal nebulin myopathy. Core-rod myopathy has recently been reported to be another type of NEB-related myopathy, and is pathologically characterized by the coexistence of cores and nemaline rods within muscle fibers. METHODS: We describe 2 patients with core-rod myopathy who were analyzed genetically by whole exome sequencing and evaluated clinically and pathologically. Findings were compared with those of patients with the disease of other genetic causes. RESULTS: Three NEB mutations were identified, 2 of which were novel. Mild clinical features, unusual patterns of muscle involvement, and atypical pathological findings were observed. CONCLUSIONS: We propose that the clinical and pathological spectrum of core-rod myopathy should be widened. A significant amount of residual nebulin expression is believed to contribute to the much milder phenotype exhibited by the patients we describe here.