Xuanwei Liang

A Rabbit Dry Eye Model Induced by Topical Medication of a Preservative Benzalkonium Chloride

PURPOSE To establish a rabbit dry eye model with topical medication of the ocular preparation preservative benzalkonium chloride (BAC). METHODS Sixteen white rabbits were used. One eye of each rabbit was chosen randomly for topical administration of 0.1% BAC twice daily for 14 days. The other untreated eyes served as controls. Schirmer test, fluorescein, and rose bengal staining were performed before and after BAC treatment on days 3, 5, 7, and 14. Conjunctiva impression cytology specimens were collected on days 0, 7, and 14. The rabbits were killed after day 14. Immunofluorescence staining was performed to detect mucin-5 subtype AC (MUC5AC) on conjunctival cryosections. Cornea and conjunctiva structures were evaluated by light and electron microscopy. RESULTS Compared with untreated controls, BAC-treated eyes showed significant decreases in Schirmer scores (P = 0.01) and increases in fluorescein scores (P < 0.001) on days 5, 7, and 14. A significant increase in rose bengal scores was noticed as early as day 3 (P = 0.001). Decreases in goblet cell density occurred on days 7 and 14 (P = 0.001). Decreased MUC5AC and histopathologic and ultrastructural disorders of the cornea and conjunctiva were also observed in the BAC group. CONCLUSIONS These findings demonstrated that an ophthalmic preservative, benzalkonium chloride, induced a dry eye syndrome in rabbits with damage to the cornea and conjunctiva, decreased aqueous tear basal secretion, goblet cell loss, and MUC5AC deficiency. This rabbit model was consistent with human dry eye syndrome in both aqueous tear and mucin deficiency and may be appropriate for studying dry eye syndrome.

Using acellular porcine limbal stroma for rabbit limbal stem cell microenvironment reconstruction.

To investigate the feasibility of using acellular porcine limbal stroma for limbal stem cell microenvironment reconstruction. Limbal reconstruction was performed in rabbit partial limbal defect models. Rabbits were randomly divided into four groups: acellular porcine limbal stroma, de-epithelized rabbit limbal autograft stroma, de-epithelized porcine limbal stroma and acellular porcine corneal stroma transplantation groups. In both the acellular porcine limbal stroma and de-epithelized rabbit limbal autograft stroma groups, cornea transparency and epithelium integrity were sustained and graft rejection was not observed. The basal epithelial cells of the grafts showed the K3+/P63+/Ki67+ phenotype at postoperative month 1, but it returned to K3-/P63+/Ki67+(phenotype characteristic of limbal epithelium) by postoperative months 3 and 6. In the de-epithelized porcine limbal stroma group, acute and serious immune rejection occurred by postoperative days 8-10. The basal epithelial cells of the grafts showed the K3+/P63+/Ki67+ phenotype at postoperative month 1. In the acellular porcine corneal stroma group, there were some new vessel invasion into the peripheral cornea and mild corneal opacity. The basal epithelial cells of the grafts showed the K3+/P63+/Ki67+ phenotype at postoperative months 1, 3, and 6. In conclusion, acellular porcine limbal stroma possessed very low immunogenicity, retained a good original limbal ECM microenvironment, and thus the reconstructed rabbit limbal microenvironment maintained limbal epithelial stem cell stemness and proliferation.

Quantitative Measurements of the Ciliary Body in Eyes with Malignant Glaucoma after Trabeculectomy Using Ultrasound Biomicroscopy

PURPOSE To evaluate and compare the structural differences of the ciliary body in eyes with and without malignant glaucoma. DESIGN Cross-sectional study. PARTICIPANTS Twenty-seven consecutive patients diagnosed with malignant glaucoma in 1 eye after trabeculectomy were recruited. They were all originally diagnosed with primary angle closure (PAC) or PAC glaucoma (PACG). Twenty-seven PAC/PACG eyes of 27 patients who had undergone uneventful trabeculectomy in the same period were also recruited. They were comparable with the fellow eyes of the malignant glaucoma patients in terms of surgical type, glaucoma type, and stage. METHODS A-scan ultrasonography and ultrasound biomicroscopy measurements were performed on the eyes with malignant glaucoma, the fellow eyes of the patients with malignant glaucoma, and the matched eyes. MAIN OUTCOME MEASUREMENTS Ciliary body parameters included maximum ciliary body thickness (CBTmax), ciliary body thickness at the point of the scleral spur (CBT0) and 1000 μm from the scleral spur (CBT1000), anterior placement of the ciliary body (APCB), and the trabecular-ciliary process angle (TCA). Biometric measurements including axial length, central anterior chamber depth (ACD), pupil diameter (PD), anterior chamber width, and lens vault (LV) were also recorded. RESULTS Average CBTmax were 0.545±0.088 (mean ± standard deviation), 0.855±0.170, and 0.960±0.127 mm in eyes with malignant glaucoma, their fellow eyes, and the matched eyes, respectively. Average APCB were 0.860±0.176, 0.608±0.219, and 0.427±0.139 mm, respectively. Average TCA were 18.49±4.12, 41.79±17.27, and 48.53±10.38 degrees, respectively. The CBTmax, CBT0, CBT1000, and TCA were smaller, whereas APCB was larger in eyes with malignant glaucoma compared with their fellow eyes (P < 0.01). The fellow eyes had larger APCB and smaller CBTmax and CBT0 than the matched eyes (P < 0.05). The ACD, anterior chamber width, and PD were smaller, whereas LV was larger in eyes with malignant glaucoma compared with their fellow eyes (P < 0.05). No differences were found in the ACD, anterior chamber width, PD, or LV between the fellow eyes of malignant glaucoma and matched eyes (P > 0.1). CONCLUSIONS The ciliary bodies were thinner and more anteriorly rotated in eyes with malignant glaucoma as well as in their fellow eyes, which may be the predisposing factor for malignant glaucoma.

Roles of limbal microvascular net and limbal stroma in regulating maintenance of limbal epithelial stem cells.

Knowledge of the microenvironment (niche) of stem cells is helpful for stem-cell-based regenerative medicine. In the eye, limbal epithelial stem cells (corneal epithelial stem cells) provide the self-renewal capacity of the corneal epithelium and are essential for maintaining corneal transparency and vision. Limbal epithelial stem cell deficiency results in significant visual deterioration. Successful treatment of this type of blinding disease requires studies of the limbal epithelial stem cells and their microenvironment. We investigate the function of the limbal microvascular net and the limbal stroma in the maintenace of the limbal epithelial stem cell niche in vivo and examine the regulation of limbal epithelial stem cell survival, proliferation and differentiation in vivo. We assess the temporal and spatial changes in the expression patterns of the following markers during a six-month follow-up of various rabbit limbal autograft transplantation models: vascular endothelial cell marker CD31, corneal epithelium differentiation marker K3, limbal epithelial stem-cell-associated markers P63 and ABCG2 and proliferating cell nuclear marker Ki67. Our results suggest that limbal epithelial stem cells cannot maintain their stemness or proliferation without the support of the limbal microvascular net microenvironment. Thus, both the limbal microvascular net and the limbal stroma play important roles as components of the limbal epithelial stem cell niche maintaining limbal epithelial stem cell survival and proliferation and the avoidance of differentiation. The limbal stroma constitutes the structural basis of the limbal epithelial stem cell niche and the limbal microvascular net is a requirement for this niche. These new insights should aid the eventual construction of tissue-engineered cornea for corneal blind patients in the future.

Human amniotic epithelial cell niche enhances the functional properties of human corneal endothelial cells via inhibiting P53-survivin-mitochondria axis

The particular microenvironment or niche plays an important role in determining the fate of stem cells and adult cells. The objective of this study was to explore the potential role of the niche of human amniotic epithelial cells in enhancing the functional properties of human corneal endothelial cells (HCECs). The HCECs were cultured in different media, including corneal endothelium medium (CEM), 20% human amniotic epithelial cell culture medium (20% HAEC-Me), and 20% human amniotic epithelial cell-conditioned medium (20% HAEC-CM). We observed that the HCECs cultured in the 20% HAEC-CM had an increased proliferative capacity, higher colony-forming efficiency (CFE), fewer apoptotic cells, and similar cell-junction formation capabilities and pump functionality compared with primary HCECs. Compared with CEM and 20% HAEC-Me, the 20% HAEC-CM system enhanced the functional properties of HCECs by reducing the generation of reactive oxygen species (ROS), maintaining the membrane potential (Δψm) at higher levels, reducing the expression of the p53 protein, and increasing the level of survivin protein expression. This study may shed light on the expansion of HCECs and the clinical applications of these cells in regenerative medicine, especially in corneal tissue engineering.

A modified symblepharon ring for sutureless amniotic membrane patch to treat acute ocular surface burns.

The objective of this study is to evaluate a sutureless technique by using a modified symblepharon ring to fix an amniotic membrane (AM) patch on the ocular surface to treat acute ocular burns. Seventy-five patients with acute ocular burns of total 75 eyes graded III to VI were enrolled in this study. They were randomly divided into two groups. Thirty-nine eyes received the sutureless AM patch with a modified symblepharon ring, and the other 36 eyes underwent the conventional sutured AM patch as control. The time and the rate of epithelialization, corneal neovascularization, and complications were recorded. Both the operation time and the time to epithelial closure in the sutureless group were much shorter than that in the suture group (P < .01). The rate of reepithelialization in the sutureless group was higher than in the suture group (P < .05). The rate of the vascularization and symblepharon were lower in the sutureless group than in the suture group (P < .05). The conjunctival sac contraction occurred only in the eyes with grade V and VI in the sutureless group and was later than in the suture group (P < .05). This modified method is simple, minimally invasive, free from trauma, and more effective compared with controls.

Novel air-injection technique to locate the medial cut end of lacerated canaliculus

Locating the medial cut end of the severed canaliculus is the most difficult aspect of canalicular repair, especially in patients with more medial laceration, severe oedema, persistent errhysis and a narrow canaliculus. Irrigation is a widely used technique to identify the cut end; however, we found that air injected through the intact canaliculus with a straight needle failed to reflux when the common canaliculus or lacrimal sac was not blocked. We describe a simple, safe and efficient air-injection technique to identify the medial cut edge of a lacerated canaliculus. In this method, we initially submersed the medial canthus under normal saline, then injected filtered air through the intact canaliculus using a side port stainless steel probe with a closed round tip. The tip was designed to block the common canaliculus to form a relatively closed system. The efficiency of this novel air-injection technique was equivalent to the traditional technique but does not require the cooperation of the patient to blow air. Using this technique, the medial cut end was successfully identified by locating the air-bubble exit within minutes in 19 cases of mono-canalicular laceration without any complication.

Two novel mutations in the bestrophin-1 gene and associated clinical observations in patients with best vitelliform macular dystrophy

The purpose of the current study was to investigate the 11 bestrophin-1 (BEST1) exons in patients with best vitelliform macular dystrophy (BVMD), and to characterize the associated clinical features. Complete ophthalmic examinations were conducted on two families, and two family members were diagnosed with BVMD. Genomic DNA was extracted from the leukocytes of peripheral blood collected from the patients and their family members, in addition to 100 unrelated control subjects recruited from the same population. The polymerase chain reaction was used to amplify a total of 11 exons of the BEST1 gene, which were directly sequenced. Ophthalmic examinations, including best-corrected visual acuity, slit-lamp examination, fundus examination, fundus photography and fluorescein angiography imaging, as well as anterior segment analysis with Pentacam and optical coherence tomography, were conducted. The patients exhibited yellowish lesions in the macular area. A heterozygous mutation c.910_912delGAT (p.304del Asp) in exon 7 was identified in Case 1. A heterozygous BEST1 missense mutation c.685T>G (p.Trp229Gly) in exon 5 was identified in Case 2, but not in any of the unaffected family members or normal controls. Although BEST1 gene mutations and polymorphisms have previously been reported in various ethnic groups, the current study identified, for the first time to the best of our knowledge, two novel BEST1 gene mutations in patients with BVMD.

A novel modified soft probe for identifying the distal cut end in single canalicular laceration

Canalicular laceration, especially lower canaliculus laceration, is the most common type of trauma of the lacrimal system. The primary aim of management of these cases is to restore the canalicular system both anatomically and functionally. Previously, we have successfully developed a modified soft probe to treat lacrimal passage obstruction.1 ,2 Herein, we reported a novel and simple procedure to identify the distal cut end of the lower or upper canalicular laceration by intubation of a modified soft probe through the uninjured ipsilateral canaliculus. This study was a non-comparative, interventional case series. Thirty-six eyes with lower or upper canalicular lacerations (lower, 33 eyes, and upper, three eyes) as a result of trauma were involved in the study. A modified soft probe system was developed for this procedure including a segmental epidural catheter with the blind tip (with a self-made side aperture 2.0 cm from the tail end), a stainless steel acupuncture needle (needle point was cut-off and the tip was burnished) which was inserted into the lumen of epidural catheter to increase its rigidity, and a syringe which was used to inject oculentum (figure 1A). Under an operating microscope, the …

A Novel Method for Measuring Anterior Segment Area of the Eye on Ultrasound Biomicroscopic Images Using Photoshop

Purpose To describe a novel method for quantitative measurement of area parameters in ocular anterior segment ultrasound biomicroscopy (UBM) images using Photoshop software and to assess its intraobserver and interobserver reproducibility. Methods Twenty healthy volunteers with wide angles and twenty patients with narrow or closed angles were consecutively recruited. UBM images were obtained and analyzed using Photoshop software by two physicians with different-level training on two occasions. Borders of anterior segment structures including cornea, iris, lens, and zonules in the UBM image were semi-automatically defined by the Magnetic Lasso Tool in the Photoshop software according to the pixel contrast and modified by the observers. Anterior chamber area (ACA), posterior chamber area (PCA), iris cross-section area (ICA) and angle recess area (ARA) were drawn and measured. The intraobserver and interobserver reproducibilities of the anterior segment area parameters and scleral spur location were assessed by limits of agreement, coefficient of variation (CV), and intraclass correlation coefficient (ICC). Results All of the parameters were successfully measured by Photoshop. The intraobserver and interobserver reproducibilities of ACA, PCA, and ICA were good, with no more than 5% CV and more than 0.95 ICC, while the CVs of ARA were within 20%. The intraobserver and interobserver reproducibilities for defining the spur location were more than 0.97 ICCs. Although the operating times for both observers were less than 3 minutes per image, there was significant difference in the measuring time between two observers with different levels of training (p<0.001). Conclusion Measurements of ocular anterior segment areas on UBM images by Photoshop showed good intraobserver and interobserver reproducibilties. The methodology was easy to adopt and effective in measuring.