Effect of SMILE-derived decellularized lenticules as an adhesion barrier in a rabbit model of glaucoma filtration surgery.

BACKGROUND: To investigate the effects of small incision lenticule extraction (SMILE)-derived decellularized lenticules on intraocular pressure (IOP) and conjunctival scarring in a rabbit model of glaucoma filtration surgery. METHODS: Trabeculectomy was performed on both eyes of New Zealand rabbits. A decellularized lenticule was placed in the subconjunctival space in one eye of the rabbits (the decellularized lenticule group), and no adjunctive treatment was performed in the fellow eye (the control group). The filtering bleb features and IOP were evaluated 0, 3, 7, 14, 21, and 28 days after surgery, and histopathologic examination was performed 28 days after surgery. RESULTS: Decellularized lenticules significantly increased bleb survival and decreased IOP postoperatively in the rabbit model with no adverse side effects. The histopathologic results showed a larger subconjunctival space and less subconjunctival fibrosis in the decellularized lenticule group. CONCLUSIONS: Decellularized lenticules can prevent postoperative conjunctiva-sclera adhesion and fibrosis, and they may represent a novel antifibrotic agent for trabeculectomy.

Alpha-melanocyte stimulating hormone suppresses the proliferation of human tenon's capsule fibroblast proliferation induced by transforming growth factor beta 1.

Alpha-melanocyte stimulating hormone (alpha-MSH) is a proopiomelanocortin derivative and a multi-function neuropeptide, well know for its pigment-inducing capacity, inhibitory action on proinflammatory cytokines and chemoattractant cytokines, and suppressive action on collagen synthesis. Human Tenon's capsule fibroblasts (HTF) are the main effector cells in the initiation and mediation of wound healing and fibrotic scar formation after trabeculectomy. In this study the effects of alpha-MSH on proliferation of HTF stimulated by transforming growth factor beta1 (TGF-beta1), have been investigated and discussed. Fibroblasts were cultured in Dulbecco's modified Eagle's medium (DMEM) in the control group, and in DMEM with TGF-beta1 at concentration of 10(-12) M in the TGF-beta1 group, and DMEM with 10(-12) M TGF-beta1 and alpha-MSH ranging from 0, 10(-8) to 10(-4) M in the TGF-beta1/alpha-MSH groups. Cell proliferation was assessed 48 h later by the CellTiter 96 Aqueous One Solution Cell Proliferation Assay. After administration of TGF-beta1 at a concentration of 10(-12) M, or TGF-beta1 at 10(-12) M plus alpha-MSH at 10(-6) M, the mRNA level of type I (alpha1) collagen, fibronectin, TNF-alpha, intercellular cell adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), MMP-1, MMP-2, TIMP-1, and TIMP-2 in HTF were analyzed using the real time reverse transcription polymerase chain reaction. Alpha-MSH demonstrated an inhibitory effect on the proliferation of HTF induced by TGF-beta1 in a dose-dependent manner, when the concentration was lower than 10(-5) M, and a suppressive effect on the mRNA expression of type I (alpha1) collagen, TNF-alpha, ICAM-1 and VCAM-1, which were up-regulated by TGF-beta1. Our results showed a reverse effect of alpha-MSH on the imbalance between MMPs and TIMPs compared with TGF-beta1. Based on all these results, we conclude that alpha-MSH could effectively suppress HTF proliferation and modulate correlative genes in collagen synthesis stimulated by TGF-beta1, which implies that alpha-MSH could be exploited in the treatment of conjunctival fibrotic scar disorder.

[Clinical investigation of medication adherence of glaucoma patients].

OBJECTIVE: To investigate the medication adherence and modulation factors of medication adherence in long-term treated glaucoma patients. METHODS: It was a cross-sectional study. One hundred glaucoma patients who have been followed-up for more than 6 month in the Second Hospital affiliated to Medical College of Zhejiang University were investigated with questionnaires and interview. The data was analyzed by binary logistic regression and χ(2) test. RESULTS: In the 100 glaucoma patients being investigated, the medication compliance of patients is 61(61.0%), the medication non-compliance of patients is 39 (39.0%). Age, gender, habitat, glaucoma types, surgical history, ingesting patient before writing out a prescription and informing them the methods for application of eyedrops were not significantly co-related with medication adherence (χ(2) = 0.718, 0.537, 0.338, 3.797, 0.338, 0.297, 0.157; P > 0.05). Intraocular pressure before treatment, types of eyedrops, instillation of eye-drops to the eyelids, request for help during application of eyedrops, rechecking regularly and confidence in glaucoma treatment were found significantly related to medication adherence (P = 0.013, 0.005, 0.005, 0.012, 0.011, 0.004). However, drug side effect, forgetting application of eyedrops and inconvenience to see a doctor was considered the most important factors by patients in the interview. CONCLUSIONS: Glaucoma physicians should discuss with patients about the methods for the application of glaucoma medication. This can improve the medication adherence of glaucoma patients and promote the therapeutic effect.

Inhibition by brimonidine of forskolin-induced nitric oxide synthase expression in human ciliary bodies in vitro.

PURPOSE: To investigate the mRNA and protein expression of nitric oxide synthase (NOS) in human ciliary bodies in vitro. The effect of the adenylcyclase activator forskolin and/or the alpha2-adrenergic agonist brimonidine (an ocular hypotensive agent that inhibits aqueous humor formation) on NOS mRNA or protein expression was also studied. METHODS: Frozen human ciliary bodies obtained from local eye bank were thawed and incubated with 0.1 mM forskolin for 24 h in the absence or in the presence of 100 muM brimonidine. The mRNA and protein expression of three NOS isoforms (neuronal NOS or nNOS, inducible NOS or iNOS, endothelial NOS or eNOS) were assessed by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis, respectively. RESULTS: mRNA and protein expression of three NOS isoforms were detected in human ciliary bodies. Forskolin significantly up-regulated the mRNA and protein expression of nNOS, but not that of iNOS or of eNOS. In the presence of brimonidine, the forskolin-induced up-regulation of nNOS mRNA or protein expression was significantly inhibited. CONCLUSIONS: In human ciliary body (where aqueous humor is produced), brimonidine inhibits the up-regulation of nNOS expression induced by forskolin.

Novel mutations of the FRMD7 gene in X-linked congenital motor nystagmus.

PURPOSE: Congenital motor nystagmus (CMN) is a relatively common oculomotor disorder characterized by bilateral uncontrollable ocular oscillations. Recently, the FRMD7 gene mutation has been identified as the genetic cause of CMN. The purpose of this study was to identify mutations of the FRMD7 gene in Chinese patients with CMN. METHODS: Clinical data and genomic DNA of three Chinese CMN families were collected after informed consent. Genescan by two-point linkage analysis combined with haplotype analysis was performed and mutation screening of the FRMD7 gene was conducted by direct sequencing. RESULTS: Maximum two-point LOD scores of 2.00, 1.76, and 1.16 at theta=0.00 were obtained with markers in proximity to the FRMD7 gene on chromosome Xp26 in the three CMN families. Mutation screening in the FRMD7 gene identified two novel missense mutations (c.781C>G and c.886G>C) and one reported nonsense mutation (c.1003C>T). These nucleotide alterations were not seen in unaffected members of the families or in 100 unrelated control subjects. CONCLUSIONS: This study widens the mutation spectrum of the FRMD7 gene.

[Therapeutic effect of laser photocoagulation of ciliary processes in aphakic glaucoma].

OBJECTIVE: To evaluate the therapeutic effect of laser photocoagulation of ciliary processes after pars plana vitrectomy in aphakic glaucoma. METHODS: Twenty patients (20 eyes) of aphakic glaucoma underwent vitrectomy plus laser treatment. During the surgery, after conventional 3-incision pars plana vitrectomy, a probe of semi-conductor diode laser (532 nm) was inserted through the sclera incision, the ciliary processes were then photocoagulated under direct visualization for 180 degree range. Before and after the surgery, the visual acuity, the intraocular pressure (IOP) as well as the outer appearance of the anterior segment were evaluated. The mean follow-up period was 13 months. RESULTS: The IOP at the last visit was (21.35 +/-2.52) mmHg, which was significantly lower than that before the surgery [(39.75 +/-6.27) mmHg, P=0.000]. Atrophy of the ciliary processes was observed 1-3 months after the surgery. CONCLUSION: Pars plana vitrectomy combined with laser coagulation of ciliary processes reduces the IOP in patients with aphakic glaucoma effectively.

[Indocyanine green and fluorescein angiography study of glaucomatous peripapillary atrophy].

OBJECTIVE: To investigate the incidence of glaucomatous peripapillary atrophy (PPA) and choroidal circulation of glaucomatous patients. METHODS: The subjects of glaucoma groups included 43 (43 eyes) glaucomatous patients including 16 (16 eyes) chronic angle-closure glaucoma (CACG), 27 (27 eyes) open-angle glaucoma. Control group included 33 (33 eyes) cases. Indocyanine green angiography and fundus fluorescein angiography were performed on glaucoma groups and control group with Heidelberg retinal angiography (HRA) to observe alpha zone and beta zone of PPA. RESULTS: The frequency of beta zone in CACG, open-angle glaucoma and control group was 31.3%, 77.8% and 21.9%, respectively, with statistically significant difference between these groups (chi(2) = 19.915, P = 0.000). Beta zone showed ICG hypofluorescence in all eyes. There were no statistically significant difference in the frequency of alpha zone between glaucoma groups and control group. However, there were two different types of indocyanine green filling in alpha zone: normal or decreased choroid filling. Decreased choroid filling in alpha zone was observed in 53.8% of eyes with chronic angle-closure glaucoma, 66.7% of open-angle glaucoma and 32.1% of in control, the difference was significant (chi(2) = 6.648, P = 0.036). CONCLUSIONS: The frequency of zone beta with no choroidal filling in open-angle glaucoma group is the highest among these three groups. Zone alpha in glaucomatous eyes show decreased choroidal circulation, which may be the early sign for the extension of beta zone.

Idiopathic Choroidal Neovascularization: Intraocular Inflammatory Cytokines and the Effect of Intravitreal Ranibizumab Treatment.

Idiopathic choroidal neovascularization (ICNV) is a disorder that primarily affecting patients younger than 50 years and can cause severe loss of vision. Choroidal abnormalities, especially choroidal inflammation, have been thought to be involved in the pathophysiology of ICNV. However, the exact pathogenesis of ICNV remains unclear. The aim of our study was investigate the levels of 27 inflammatory cytokines in the aqueous humor of eyes with ICNV, and to determine the effect of intravitreal injection of ranibizumab (IVR) on cytokine levels. Significantly higher levels of IL-2, IL-10, IL-15, IL-17, basic FGF, and GM-CSF were observed in patients with ICNV compared with controls. However, only IL-17 levels were significantly higher in patients with ICNV compared with controls after adjusting for axial length. Furthermore, there were significant correlations between the levels of IL-10, IL-17, GM-CSF, and VEGF and the lesion area. Significant changes in visual acuity and central retinal thickness were observed after IVR. Besides VEGF, IVR also significantly reduced the levels of IL-2, IL-10, basic FGF, and IL-12, however, the IL-6 levels were significantly increased. Our results suggest that there may be an involvement of IL-17-related inflammatory processes in the etiology of ICNV.

Transcorneal electrical stimulation promotes survival of retinal ganglion cells after optic nerve transection in rats accompanied by reduced microglial activation and TNF-α expression.

Microglial activation plays a crucial role in the pathological processes of various retinal and optic nerve diseases. TNF-α is a pro-inflammatory cytokine that is rapidly upregulated and promotes retinal ganglion cells (RGCs) death after optic nerve injury. However, the cellular source of TNF-α after optic nerve injury remains unclear. Thus, we aimed to determine the changes of retinal microglial activation in a rat model of optic nerve transection (ONT) after transcorneal electrical stimulation (TES). Furthermore, we assessed TNF-α expression after ONT and evaluated the effects of TES on TNF-α production. Rats were divided into 2 control groups receiving a sham surgery procedure, 2 ONT+Sham TES groups, and 2 ONT+TES groups. The rats were sacrificed on day 7 or 14 after ONT. RGCs were retrogradely labelled by Fluorogold (FG) 7 days before ONT, one TES group and corresponding controls were stimulated on day 0, 4, and the second were stimulated on day 0, 4, 7, 10. Whole-mount immunohistofluorescence, quantification of RGCs and microglia, and western blot analysis were performed on day 7 and 14 after ONT. TES significantly increased RGCs survival on day 7 and 14 after ONT, which was accompanied by reduced microglia on day 7, but not 14. TNF-α was co-localized with ameboid microglia and significantly increased on day 7 and 14 after ONT. TES significantly reduced TNF-α production on day 7 and 14 after ONT. Our study demonstrated that TES promotes RGCs survival after ONT accompanied by reduced microglial activation and microglia-derived TNF-α production.

[Treatment of open angle glaucoma with deep sclerectomy combined with laser trabecular puncture].

OBJECTIVE: To study the effect of deep sclerectomy combined with laser trabecular puncture for the treatment of open angle glaucoma (OAG). METHODS: Deep sclerectomy was performed in 32 eyes of 32 patients with moderate to advanced stages of OAG. After the exposure of external wall of Schlemm's canal and the remaining trabecula, the intraocular optic fiber of diode laser photocoagulator was directed to the trabecula-limbus near the anterior edge of the scleral spur. Two or three punctures, approximately 0.5 mm in diameter, were performed with laser beam under direct visualization until aqueous humor was extravasated slowly. The scleral flap was closed with one or two sutures. Postoperative observation included intraocular pressure (IOP), intraocular tissue reaction, the appearance of filtering bleb, gonioscopy and ultrasound biomicroscopy. The follow-up period was 4 to 24 months (7.94 +/- 6.20) months. RESULTS: The anterior chamber was recovered shortly after the operation and the visual acuity remained the same in all eyes. At the end of follow-up period, the postoperative IOP (14.86 +/- 4.15) mm Hg was lower than the preoperative IOP (29.68 +/- 5.76) mm Hg, the difference was statistically significant (t = 7.415, P < 0.001). Diffuse and obvious filtering blebs were found in all eyes after the operation, including 27 functional blebs and 5 blebs that disappeared 2 to 3 months later. Iris was burned slightly in 2 cases, pigment was released and appeared in the anterior chamber temporarily. No laser injury occurred in the cornea and the lens. High IOP was found in 2 cases after long-term follow-up and could be controlled by antiglaucoma medication. CONCLUSIONS: Deep sclerectomy with laser trabecular puncture can effectively reduce the IOP, without the occurrence of serious complications that commonly seen after the trabeculectomy. It is an effective surgical method for the treatment of OAG.

Microcoaxial phacoemulsification combined with viscogoniosynechialysis for patients with refractory acute angle-closure glaucoma.

PURPOSE: To evaluate the safety and efficacy of microcoaxial phacoemulsification combined with viscogoniosynechialysis in managing refractory acute angle-closure glaucoma (AACG) unresponsive to conventional therapy. METHODS: Seventeen consecutive eyes of patients with AACG>270 degrees peripheral anterior synechiae were treated with microcoaxial phacoemulsification combined with viscogoniosynechialysis. After intraocular lens implantation, a heavy viscoelastic agent was used to deepen the anterior chamber, and then injected near the angle for 360 degrees without touching any ocular structure, to release the peripheral anterior synechiae under gonioscopy. The viscoelastic agent was then removed. RESULTS: Intraocular pressure was reduced from a median of 45.0 mm Hg initially to 15.0 mm Hg after the combined procedures at the final follow-up visit (P<0.001). All angles showed exposure of the trabecular meshwork over 360 degrees postoperatively without residual synechiae. The mean best corrected visual acuity improved from 0.7 to 0.18 logMAR at 6-month follow-up (P<0.001). Surgically induced astigmatism was 0.29 at 6-month follow-up. There was a 6.4% reduction in the mean corneal endothelial cell count. No severe complications were observed in any patient. CONCLUSIONS: Microcoaxial phacoemulsification combined with viscogoniosynechialysis is an effective and safe treatment for managing refractory AACG. Using a 2.2-mm incision caused less surgically induced astigmatism and also improved visual quality.

Molecular analysis and phenotypic study in 14 Chinese families with Bietti crystalline dystrophy.

PURPOSE: To investigate the clinical features and cytochrome P450 family 4 subfamily V polypeptide 2 (CYP4V2) gene mutations in 14 Chinese families with Bietti crystalline dystrophy (BCD). METHODS: Seventeen patients from 14 unrelated Chinese families with BCD were recruited for complete clinical ophthalmic examination and genetic study. The 11 exons of CYP4V2 were amplified from genomic DNA of all patients and their family members by polymerase chain reaction (PCR) and then sequenced. Exons of TIMP3 were also sequenced in BCD patient associated with choroidal neovascularization (CNV). One hundred and seventy unrelated healthy Chinese subjects were screened for mutations in CYP4V2. RESULTS: All 17 patients with BCD had mutations in CYP4V2; one of these mutations was novel (c.219T>A, p.F73L) and four other mutations had been reported. The p.F73L mutation was a commonly detected mutation in our study (seven out of 34 alleles), either in the homozygous state or in the heterozygous state. Among the patients, considerable phenotypic variability was detected, both within and between families. Screening of TIMP3 did not find any mutation in the BCD patient associated with CNV. CONCLUSION: The novel CYP4V2 c.219T>A (p.F73L) mutation may be another recurrent mutation in Chinese patients with BCD. Our study expands the mutation spectrum of CYP4V2 and characterizes novel genotype-phenotype associations in Chinese patients with BCD.

Identification of a novel frameshift mutation in PITX2 gene in a Chinese family with Axenfeld-Rieger syndrome.

OBJECTIVE: Axenfeld-Rieger syndrome (ARS) is phenotypically and genetically heterogeneous. In this study, we identified the underlying genetic defect in a Chinese family with ARS. METHODS: A detailed family history and clinical data were recorded. The ocular phenotype was documented using slit-lamp photography and systemic anomalies were also documented where available. The genomic DNA was extracted from peripheral blood leukocytes. All coding exons and intron-exon junctions of paired-like homeodomain transcription factor 2 (PITX2) gene and the forkhead box C1 (FOXC1) gene were amplified by polymerase chain reaction (PCR) and screened for mutation by direct DNA sequencing. Variations detected in exon 5 of PITX2 were further evaluated with cloning sequencing. The exon 5 of PITX2 was also sequenced in 100 healthy controls, unrelated to the family, for comparison. Structural models of the wild type and mutant homeodomain of PITX2 were investigated by SWISS-MODEL. RESULTS: Affected individuals exhibited variable ocular phenotypes, whereas the systemic anomalies were similar. After direct sequencing and cloning sequencing, a heterozygous deletion/insertion mutation c.198_201delinsTTTCT (p.M66Ifs*133) was revealed in exon 5 of PITX2. This mutation co-segregated with all affected individuals in the family and was not found either in unaffected family members or in 100 unrelated controls. CONCLUSIONS: We detected a novel frameshift mutation p.M66Ifs*133 in PITX2 in a Chinese family with ARS. Although PITX2 mutations and polymorphisms have been reported from various ethnic groups, we report for the first time the identification of a novel deletion/insertion mutation that causes frameshift mutation in the homeodomain of PITX2 protein.

Assessment of macular function by multifocal electroretinogram in diabetic macular edema before and after vitrectomy.

The purpose of this study was to assess the effects of vitrectomy on retinal function in macular and paramacular areas in patients with diabetic macular edema (DME). Ten eyes of 9 patients with successful vitrectomy for DME were evaluated by multifocal electroretinogram (mfERG), optical coherence tomography (OCT), and visual acuity preoperatively and 1, 2, 3, 4, 5 and 6 months postoperatively. Compared with pretreatment values, the response of the positive wave (P1) in macular and paramacular areas tended to decrease in latency from the second postoperative month, and increase in amplitude at the third postoperative month. There was no significant change in response of the negative wave (N1). The tendency in the change of macular function is coincident with that of macular morphology. Therefore, The multifocal ERG may provide objective criteria for the functional evaluation of DME before and after vitrectomy.