Zhengri Li

Tear production and ocular surface changes in experimental dry eye after elimination of desiccating stress.

PURPOSE To investigate the severity and duration of desiccating stress-induced dry eye disease between mice with and without a genetic predisposition to spontaneous autoimmunity. METHODS Experimental dry eye was induced in 12- to 16-week-old wild-type C57BL/6 and autoimmune NOD.B10.H2(b) mice by subcutaneous injection of scopolamine with exposure to an air draft for 10 days. Tear volume and corneal smoothness were measured at baseline, 5 and 10 days after desiccating stress, and 3, 7, 14, and 28 days after the removal of desiccating stress. Periodic acid-Schiff staining and immunohistochemistry were performed to evaluate the densities of conjunctival goblet cells and CD4(+) T cells in each group. Interleukin (IL)-1β and IL-6 concentrations in conjunctival tissues were measured by multiplex immunobead assay. RESULTS Signs of experimental dry eye were noted at 5 and 10 days after desiccating stress in both strains. After the removal of desiccating stress, in C57BL/6 mice, tear production and corneal smoothness improved at 3 and 7 days, respectively, and conjunctival goblet cells and CD4(+) T-cell densities and cytokine levels returned to baseline levels at 14 days. In contrast, in NOD.B10.H2(b) mice, none of the parameters recovered to baseline levels during a period of 28 days after the removal of desiccating stress. CONCLUSIONS After the removal of desiccating stress in experimental dry eye, tear volume and ocular surface parameters recovered within 2 weeks in C57BL/6 mice, whereas they remained unchanged in NOD mice. In contrast to autoimmune mice, experimental dry eye can be reversed after the elimination of desiccating stress in nonsusceptible mice.

Expression of CCR5 and its ligands CCL3, -4, and -5 in the tear film and ocular surface of patients with dry eye disease.

Purpose: To evaluate the expression of CCR5 and its ligands CCL3, CCL4, and CCL5 in the tear film and ocular surface and their correlation with disease severity in patients with dry eye disease. Materials and Methods: The concentrations of CCL3, CCL4, and CCL5 were measured using enzyme-linked immunosorbent assay in tear samples obtained from forty-three patients with dry eye (17 SS and 26 non-SS patients) and 20 control subjects. The correlation between chemokine levels and tear film and ocular surface parameters was analyzed. Expression of the chemokines and their receptor in the conjunctiva was evaluated using immunohistochemistry. Flow cytometry was performed to detect CCR4+CD4+, CCR5+CD4+, and CCR6+CD4+ cells in the conjunctiva. Results: The concentrations of CCL3, CCL4, and CCL5 were 25.3 ± 24.2, 4.65 ± 3.21, and 93.12 ± 26.31 pg/mL in control subjects, 92.33 ± 13.23, 263.13 ± 116.13, and 253.64 ± 46.29 pg/mL in patients with non-SS, and 215.56 ± 36.1, 697.85 ± 185.65, and 456.12 ± 92.82 pg/mL in patients with SS. The concentrations showed a significant increase in tears of SS patients compared with those of non-SS patients and control subjects (p < 0.05). CCL5 levels showed significant correlation with tear film break-up time, basal tear secretion, tear clearance rate, keratoepitheliopathy score, and goblet cell density (p < 0.01). Staining for the chemokines and their receptor increased in dry eye patients, especially in those with SS patients. Flow cytometry demonstrated increased numbers of CCR5+CD4+, and CCR6+CD4+ cells in dry eye patients in contrast to CCR4+CD4+ cells. Conclusions: Expression of CCR5 and its ligands CCL3, CCL4, and CCL5 increase in the tear film and ocular surface of patients with dry eye syndrome, especially in those with SS. CCL5 levels correlate significantly with various tear film and ocular surface parameters.

Effects of Eye Drops Containing a Mixture of Omega-3 Essential Fatty Acids and Hyaluronic Acid on the Ocular Surface in Desiccating Stress-induced Murine Dry Eye

Abstract Purpose: To investigate the efficacy of the topical application of omega-3 essential fatty acids (EFAs) and hyaluronic acid (HA) mixtures in a mouse model of experimental dry eye (EDE). Methods: Eye drops consisting of 0.1% HA, 0.02%, or 0.2% omega-3 EFAs alone and mixture of 0.02%, or 0.2% omega-3 EFAs and 0.1% HA were applied in desiccating stress-induced murine dry eye. Corneal irregularity scores and fluorescein staining scores were measured 5 and 10 days after treatment. Levels of interleukin (IL)-1β, -17, and interferon gamma-induced protein (IP)-10 were measured in the conjunctiva at 10 days using a multiplex immunobead assay. The concentrations of hexanoyl-lys (HEL) and 4-hydroxynonenal (4-HNE) in conjunctiva tissue were measured with enzyme-linked immunosorbent assays. Results: Mice treated with the mixture containing 0.2% omega-3 EFAs showed a significant improvement in corneal irregularity scores and corneal fluorescein staining scores compared with EDE, HA, 0.02% or 0.2% omega-3 EFAs alone, and 0.02% omega-3 EFAs mixture-treated mice. A significant decrease in the levels of IL-1β, -17, and IP-10 were observed in the 0.2% EFAs mixture-treated group, compared with the other groups. In the mice treated with the mixture containing 0.2% omega-3 EFAs, the concentration of 4-HNE was also lower than the other groups. Although 0.2% omega-3 EFAs alone group also had a significant improvement in corneal irregularity scores and IL-17, IL-10, and 4 HNE levels compared with the other groups, the efficacy was lower than 0.2% omega-3 mixture group. Conclusions: Topically applied eye drops containing a mixture of omega-3 EFAs and HA could improve corneal irregularity and corneal epithelial barrier disruption, and decrease inflammatory cytokines and oxidative stress markers on the ocular surface. Topical omega-3 EFAs and HA mixture may have a greater therapeutic effect on clinical signs and inflammation of dry eye compared with HA artificial tears.

Therapeutic Effect of Topical Adiponectin in a Mouse Model of Desiccating Stress–Induced Dry Eye

PURPOSE To investigate the therapeutic effect of topical adiponectin in a mouse model of experimental dry eye (EDE). METHODS EDE was created by desiccating stress in 6- to 8-week old female C57BL/6 mice. Eye drops consisting of 0.0001%, 0.001%, or 0.01% adiponectin, or balanced salt solution (BSS), were applied in EDE. Tear volume and corneal irregularity score were measured at 5 and 10 days after treatment. Levels of interleukin (IL)-1β, IL-6, tumor necrosis factor (TNF)-α, interferon (IFN)-γ, and monokine induced by interferon-γ (MIG) were measured in the conjunctiva and lacrimal gland using a multiplex immunobead assay at 10 days. Periodic acid-Schiff staining, immunohistochemistry, and flow cytometry were also performed. RESULTS Mice treated with 0.001% or 0.01% adiponectin showed a significant improvement in tear volume and corneal irregularity compared with the EDE control and BSS-treated groups. A significant decrease in the levels of IL-1β, IL-6, TNF-α, IFN-γ, and MIG and staining intensity of TNF-α was observed in the 0.001% and 0.01% adiponectin-treated groups, compared with the other groups, in the conjunctiva and lacrimal gland. In the 0.001% and 0.01% adiponectin-treated groups, the density of conjunctival goblet cells was higher and the number of CD4+CXCR3+ T cells was lower than in the other groups. However, there were no significant differences in all parameters between the 0.0001% adiponectin and control groups. CONCLUSIONS Topical application of adiponectin can markedly improve clinical signs and decrease inflammation in the ocular surface and lacrimal gland of EDE, suggesting that adiponectin eye drops may be used as a therapeutic agent for dry eye disease.

Effectiveness of topical infliximab in a mouse model of experimental dry eye.

Purpose: To investigate the efficacy of a topical anti–tumor necrosis factor-&agr; agent, infliximab, in a mouse model of experimental dry eye (EDE). Methods: EDE was induced in C57BL/6 mice, with or without topical treatment consisting of balanced salt solution or 0.001%, 0.01%, or 0.1% infliximab solutions. Tear volume and corneal smoothness were measured on days 5 and 10 after treatment. Levels of interleukin (IL)-1&bgr;, IL-6, IL-17, and interferon &ggr; (IFN-&ggr;) were measured in the conjunctiva using a multiplex immunobead assay 10 days after treatment. Periodic acid-Schiff staining, immunohistochemistry, and flow cytometry were also performed 10 days after treatment. Results: Mice treated with 0.01% or 0.1% infliximab showed a significant improvement in tear volume and corneal smoothness compared with controls. The 0.01% and 0.1% infliximab-treated groups showed decreased levels of conjunctival IL-1&bgr;, IL-6, IL-17, and interferon &ggr; and a decreased staining intensity of tumor necrosis factor-&agr;. The density of conjunctival goblet cells was higher, whereas the number of CD4+CXCR3+ T cells was lower, in the 0.01% and 0.1% infliximab-treated groups compared with the EDE and balanced salt solution control groups. However, there was no significant difference in all parameters between the 0.001% infliximab-treated group and control group. Conclusions: Topical application of infliximab can improve tear production and ocular surface irregularity, decrease inflammatory cytokines and cells on the ocular surface, and increase conjunctival goblet cell density. These results suggest that topical infliximab eye drops at a concentration of 0.01% and 0.1% may be useful for the treatment of dry eye disease.

Effect of Topical 5-Aminoimidazole-4-carboxamide-1-β-d-Ribofuranoside in a Mouse Model of Experimental Dry Eye.

PURPOSE To investigate the efficacy of topical 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside (AICAR) in a mouse model of experimental dry eye (EDE). METHODS Eye drops consisting of 0.001% or 0.01% AICAR, 0.05% cyclosporine A (CsA), or balanced salt solution (BSS) were applied for the treatment of EDE. Tear volume, tear film break-up time (BUT), and corneal fluorescein staining scores were measured 10 days after treatment. Levels of interleukin (IL)-1β, IL-6, tumor necrosis factor (TNF)-α, interferon (IFN)-γ, interferon gamma-induced protein 10 (IP-10), and monokine induced by interferon-γ (MIG) were measured in the conjunctiva. In addition, Western blot, periodic acid-Schiff staining for evaluating goblet cell density, flow cytometry for counting the number of CD4+CXCR3+ T cells, and immunohistochemistry for detection of 4-hydroxy-2-nonenal (4HNE) were performed. RESULTS Mice treated with 0.01% AICAR showed a significant improvement in all clinical parameters compared with the EDE control, vehicle control, and 0.001% AICAR groups (P < 0.001). A significant decrease in the levels of IL-1β, IL-6, TNF-α, IFN-γ, IP-10, and MIG, the number of CD4+CXCR3+ T cells, and the number of 4HNE-positive cells were also observed in the 0.01% AICAR group (P < 0.001). Although 0.05% CsA also led to an improvement in clinical parameters and inflammatory molecule levels, its therapeutic effects were comparable or inferior to those of 0.01% AICAR. CONCLUSIONS Topical application of 0.01% AICAR can markedly improve clinical signs and decrease inflammation in the ocular surface of EDE, suggesting that AICAR eye drops may be used as a therapeutic agent for dry eye disease.

Effects of Umbilical Cord Serum Eye Drops in a Mouse Model of Ocular Chemical Burn

Purpose: To evaluate the effect of umbilical cord serum (UCS) eye drops on corneal wound healing and haze in a mouse model of ocular chemical burn and compare with that of peripheral blood serum (PBS) eye drops or artificial tears (AT). Methods: Chemical burn of the ocular surface was induced by 1N NaOH in C57BL/6 mice. Injured mice were topically treated with 20% UCS, 20% PBS, or AT four times daily. The changes of corneal epithelial defects and degree of haze were analyzed at 6 h, 1, 2, 3, 5, and 7 days, and histological examination was performed at 3 and 7 days. The concentration of IL-1β in the cornea was measured by enzyme-linked immunosorbent assay at 7 days after treatment. Results: The UCS group showed lower epithelial defect parameters compared with the PBS group at 1 and 2 days (p < 0.05), and with the AT group from 1–5 days (p < 0.05). The haze scores were significantly lower in the UCS group than in the PBS group at 2 and 3 days (p < 0.05), and in the AT group from 2–7 days (p < 0.05). Histological examination showed better epithelial integrity and lower stromal inflammation and edema in the UCS group than the other groups. IL-1β levels were 99.71 ± 85.22 and 230.76 ± 102.67 pg/ml in the UCS and PBS groups, respectively (p = 0.03). Conclusions: UCS eye drops are more effective in improving corneal wound healing and reducing corneal haze compared with PBS eye drops and AT in experimental chemical burns.

Therapeutic Efficacy of Topically Applied Antioxidant Medicinal Plant Extracts in a Mouse Model of Experimental Dry Eye

Purpose. To investigate the therapeutic effects of topical administration of antioxidant medicinal plant extracts in a mouse model of experimental dry eye (EDE). Methods. Eye drops containing balanced salt solution (BSS) or 0.001%, 0.01%, and 0.1% extracts were applied for the treatment of EDE. Tear volume, tear film break-up time (BUT), and corneal fluorescein staining scores were measured 10 days after desiccating stress. In addition, we evaluated the levels of interleukin- (IL-) 1β, tumor necrosis factor- (TNF-) α, IL-6, interferon- (IFN-) γ, and IFN-γ associated chemokines, percentage of CD4+C-X-C chemokine receptor type 3 positive (CXCR3+) T cells, goblet cell density, number of 4-hydroxy-2-nonenal (4-HNE) positive cells, and extracellular reactive oxygen species (ROS) production. Results. Compared to the EDE and BSS control groups, the mice treated with topical application of the 0.1% extract showed significant improvements in all clinical parameters, IL-1β, IL-6, TNF-α, and IFN-γ levels, percentage of CD4+CXCR3+ T cells, goblet cell density, number of 4-HNE-positive cells, and extracellular ROS production (P < 0.05). Conclusions. Topical application of 0.1% medicinal plant extracts improved clinical signs, decreased inflammation, and ameliorated oxidative stress marker and ROS production on the ocular surface of the EDE model mice.

Effect of hyperglycemia on expression of aquaporins in the rat vagina.

OBJECTIVE To investigate the effect of hyperglycemia on the expression of the aquaporin (AQP) isoforms in the diabetic rat vagina. METHODS Female Sprague-Dawley rats (230-240 g, n = 45) were divided into control (n = 10) and experimental (n = 35) groups. Diabetes in the experimental group was induced by intraperitoneal injection of streptozotocin (STZ, 65 mg/kg). STZ-induced diabetic rats were left untreated or given subcutaneous injections of insulin (3 U/d). After 2 and 4 weeks, the blood glucose was measured, and the vaginal blood flow was assessed by Doppler flowmetry. The expression and cellular localization of AQP1 and AQP2 in the rat vagina were determined by Western blot and immunohistochemistry. RESULTS The vaginal blood flow (mL/min/100 g tissue) after pelvic nerve stimulation was significantly lower in the STZ-induced diabetic rats (21.9 ± 6.5 at 2 weeks and 21 ± 2.8 at 4 weeks) compared with the control group (55.5 ± 8.9 at 2 weeks and 52.9 ± 6.5 at 4 weeks; P < .05). In contrast, the vaginal blood flow response was significantly greater in the insulin-treated diabetic groups (47.7 ± 8.7 at 2 weeks and 47.7 ± 8.4 at 4 weeks) and comparable to that of the control group. The protein expression of AQP2 was significantly lower in the STZ-induced diabetic rats and was restored to the control level after insulin treatment. However, no change was seen in AQP1 expression. Thus, hyperglycemia might cause downregulation of AQP2 expression in the diabetic rat vagina. CONCLUSION These results suggest that decreased vaginal lubrication in diabetic women might result from changes in aquaporin expression, in addition to a reduction in the vaginal blood flow response.

Efficacy of the Mineral Oil and Hyaluronic Acid Mixture Eye Drops in Murine Dry Eye

Purpose To investigate the therapeutic effects of mineral oil (MO) and hyaluronic acid (HA) mixture eye drops on the tear film and ocular surface in a mouse model of experimental dry eye (EDE). Methods Eye drops consisting of 0.1% HA alone or mixed with 0.1%, 0.5%, or 5.0% MO were applied to desiccating stress-induced murine dry eyes. Tear volume, corneal irregularity score, tear film break-up time (TBUT), and corneal fluorescein staining scores were measured at 5 and 10 days after treatment. Ten days after treatment, goblet cells in the conjunctiva were counted after Periodic acid-Schiff staining. Results There was no significant difference in the tear volume between desiccating stress-induced groups. The corneal irregularity score was lower in the 0.5% MO group compared with the EDE and HA groups. The 0.5% and 5.0% MO groups showed a significant improvement in TBUT compared with the EDE group. Mice treated with 0.1% and 0.5% MO mixture eye drops showed a significant improvement in fluorescein staining scores compared with the EDE group and the HA group. The conjunctival goblet cell count was higher in the 0.5% MO group compared with the EDE group and HA group. Conclusions The MO and HA mixture eye drops had a beneficial effect on the tear films and ocular surface of murine dry eye. The application of 0.5% MO and 0.1% HA mixture eye drops could improve corneal irregularity, the corneal fluorescein staining score, and conjunctival goblet cell count compared with 0.1% HA eye drops in the treatment of EDE.