Deciphering Adult Neural Stem Cells with Single-Cell Sequencing.

Adult neural stem cells (NSCs) are restricted to the two neurogenic regions of the mammalian brain, where they self-renew and generate progenies of multiple lineages, including neurons, astrocytes, and oligodendrocytes. Single-cell RNA sequencing technology, which reconstructs high-resolution transcriptional landscapes, provides valuable insights into cellular heterogeneity and developmental dynamics. In this review, we overviewed recent progress in the single-cell analyses of both conventional and unconventional NSCs. We discussed the heterogeneity among the stem cell pool and characterized the transcriptional alterations in aging and brain tumors. A comprehensive understanding of NSCs in physiological and pathological settings will provide insights for the rejuvenation of the aged brain and restoration of normal brain function in multiple neurological disorders.

Lens nucleus dislocation in hypermature cataract: Case report and literature review.

INTRODUCTION: Hypermature cataract is a form of late-stage cataract progression that can lead to a variety of complications. Spontaneous capsular rupture with lens nucleus displacement in hypermature cataracts has rarely been reported. We describe 2 cases of spontaneous dislocation of the lens nucleus in a hypermature cataract and perform a review of the literature on this complication. PATIENT CONCERNS: We report 2 rural men aged 50 and 76 years with deteriorating vision. DIAGNOSIS: The final diagnosis was senile hypermature cataract with dislocation of the lens nucleus in both patients and secondary glaucoma for the second patient. INTERVENTIONS AND OUTCOMES: During admission, both patients complained of deteriorating vision. Slit-lamp examination showed lens nucleus dislocation into the anterior chamber. The 50-year-old patient exhibited a residual lens capsule and a turbid cortex, with a normal anterior chamber and intraocular pressure. The 76-year-old patient presented a shrunken and ruptured capsule and no cortex in the pupillary area, mild inflammation in the anterior chamber, and high intraocular pressure. Both patients underwent intracapsular cataract extraction combined with anterior vitrectomy and achieved good postoperative recovery. CONCLUSION: Lens nucleus dislocation in hypermature cataracts can be seen in clinical practice, particularly in underdeveloped areas. Early recognition and surgery can improve vision.

Pro370Leu myocilin mutation in a Chinese pedigree with juvenile-onset open angle glaucoma.

PURPOSE: To investigate the genotype and phenotype of juvenile-onset open angle glaucoma (JOAG) in a Chinese family (PN pedigree). METHODS: Each family member was comprehensively examined by an experienced ophthalmologist. The clinical characteristics of the family patients with JOAG were documented. Blood samples were obtained from 22 available participants from the PN pedigree. Linkage analysis was performed to identify the possible chromosome loci. The presence of gene mutation was ascertained by polymerase chain reaction amplification and subsequent direct sequencing. RESULTS: The affected members in the PN pedigree are characterized by early age of onset (mean age at diagnosis is 17 years old), severe clinical presentations, high intraocular pressure (mean IOP of 34.18±2.97 mmHg), and poor response to pharmacological treatment (87.5% of the patients required filtering surgery). The region on chromosome 1 between D1S3464 and D1S1619 was identified in this pedigree by linkage analysis. A Pro370Leu myocilin mutation resulting from a heterozygous C→T transition at the 1,109th nucleotide in exon 3 was detected by gene sequencing. The Pro370Leu mutation co-segregated among all affected individuals of PN pedigree. CONCLUSIONS: The GLC1A Pro370Leu mutation is firmly correlated with a severe POAG phenotype. These data provide clues for the severe disease-causing nature of the Pro370Leu allele. Gene screening may be a useful method for pre-symptom diagnosis and a forewarning to detect the at-risk individuals in familial open-angle glaucoma patients, especially in pedigrees of early-onset.

Anti-proliferation effects of Sirolimus sustained delivery film in rabbit glaucoma filtration surgery.

PURPOSE: To investigate the efficacy, safety, and mechanisms of Sirolimus sustained delivery film on prevention of scar formation in a rabbit model of glaucoma filtration surgery. METHODS: Sixty-four New Zealand white rabbits who underwent trabeculectomy in the right eye were randomly allocated to one of the four treatment regimens: Sirolimus sustained delivery film treatment group (Group A), or drug-free film treatment group (Group B), or 30 ng/ml Sirolimus-soaked sponge treatment group (Group C), or no adjunctive treatment group (Group D), and each group consists of 16 rabbits. Intraocular pressure (IOP), morphologic changes of bleb, anterior chamber flare, and corneal endothelial cell count and complications were evaluated over a 28-day period follow-up time. Aqueous humor samples were gathered from Group A, and the concentration of Sirolimus was measured regularly post-operation. Rabbits were sacrificed on the 7th, 14th, and 28th day post-operation separately, and the fibroblast hypertrophy, infiltration of inflammatory, and proliferation of new collagen fiber formation in each group were evaluated with HE and Masson staining. Proliferative cell nuclear antigen (PCNA) and fibroblast apoptosis were evaluated by immunohistochemistry and terminal deoxynucleotidyl transferasemediated dUTP nick end labeling (TUNEL) assay at the 28th day post-operation. RESULTS: Both Sirolimus sustained delivery film (Group A) and Sirolimus alone (Group C) were well tolerated in this model, and significantly prolonged bleb survival compared with no drug treatment group (Group B and D; p<0.001). Group A had the longest bleb survival time in comparison with other groups (p<0.001). There were significant differences in IOP readings between Group A and other groups at the last follow-up (p<0.05). The concentration of Group A maintained stable for over 2 weeks, drops from (10.56 ±0.05) ng/ml at day 3 to (7.74 ±0.05) ng/ml at day 14. The number of corneal endothelial cells of Group A was not statistically significant between pre and post-operation. Histologic examination demonstrated that eyes treated with Sirolimus, especially the Sirolimus sustained delivery film, showed an obvious reduction in subconjunctival fibroblast scar tissue formation compared with no drug treatment groups, and had minimal evidence of inflammatory cell infiltration and new collagen deposition in the subconjunctiva. Immunohistochemistry assay showed that PCNA-expression was lower in the Group A (16.25±3.24%) compared to other groups (p<0.01). TUNEL assay showed a significant increase in the number of apoptotic fibroblasts around the surgical area in Group A and Group C (9.75±1.71% and 8.50±1.92%) compared to the Group B and D (p<0.01). CONCLUSIONS: Sirolimus drug sustained delivery film can inhibit inflammatory cell activity, impede fibroblast proliferation activity, and induce fibroblast apoptosis in the filtration surgery sites in rabbit. The results indicate a safe and effective treatment strategy in anti-scaring treatment in glaucoma surgery.

[Clinical efficacy and safety of FP-7 Ahmed glaucoma valve implantation in neurovascular glaucoma patients].

OBJECTIVE: To evaluate the efficacy and safety of FP-7 Ahmed glaucoma valves (AGV) implantation in neurovascular glaucoma (NVG) as the first choice of surgery. METHODS: This retrospective, comparative case series study collected a total of 36 eyes of 36 patients with neurovascular glaucoma who underwent AGV implantation in Zhongshan Ophthalmic Center from January 2009 to June 2010. Change of intraocular pressure (IOP), the best corrected visual acuity, numbers of anti-glaucoma medication, success rate and postoperative complications were followed up at day 1, week 1, month 1, and every 3 months after surgery. Complete success of surgery was rated as reduction of IOP ≥ 30% without medication and those who failed to meet criteria was rated as partial success. Data were analyzed by paired Student t-test for IOP, rank sum test for paired non-parametric numbers of medication, and repeated measures analysis of variance for comparison of IOP between different time points using SPSS 13.0. RESULTS: Compared with pre-operation, IOP was significant (F = 9.26, P < 0.05) decreased after surgery with FP-7 AGV implantation (39.5 ± 9.7) mm Hg (1 mm Hg = 0.133 kPa) vs (9.2 ± 8.9), (11.8 ± 3.8), (13.7 ± 4.8), (16.9 ± 5.3), (16.9 ± 6.8) mm Hg at day 1, week 1, month 1, month 3 and the last following-up of post-operation, respectively. The numbers of anti-glaucoma medication were significantly (Z = 6.764, P < 0.05) reduced from 4.0 (1-6) of pre-operation to 1.0 (1-3) of post-operation. At the last following up, the complete success rate after FP-7 AGV implantation was 80.6%, and qualified success rate was 91.7%. The postoperative complications including occlusion of the drainage tube, exposure of the drainage tube, shallow anterior chamber and encapsulated cystic blebs around the plate were controlled with additional treatment. CONCLUSIONS: The clinical outcome indicated that the implantation of FP-7 AGV has a stable IOP lowering effect and fewer complications, which can be considered as one of the first choices for management of NVG.

Dexamethasone disrupts intercellular junction formation and cytoskeleton organization in human trabecular meshwork cells.

PURPOSE: Patients reproduce symptoms of primary open-angle glaucoma (POAG) when treated with glucocorticoids (GCs) topically on the eyes. Here we investigated the effects of GCs on junctional protein expression and cytoskeleton organization in primary human trabecular meshwork (TM) cultures to understand the molecular pathologies of POAG. METHODS: Human TM cells from POAG (GTM) and age-matched nondiseased (NTM) individuals were obtained by standard surgical trabeculectomy. Some of the cultures were treated with dexamethasone (DEX), a synthetic GC, at 1-5 x 10(-7) mol/l for 1-7 days. The expression levels of zonula occluden-1 (ZO-1) and connexin43 (Cx43) in TM cells with or without DEX treatment were measured using reverse transcription (RT)-PCR, immunocytochemistry, and western blot analysis. RESULTS: mRNA and proteins of ZO-1 and Cx43 were found in both NTM and GTM cells. ZO-1 and Cx43 were located on the plasma membrane, especially along the border of adjacent cells. ZO-1 had no marked changes in localization in NTM and GTM cells after treatment with 10(-7) mol/l DEX for 48 h, whereas Cx43 appeared to increase in the cytoplasm. mRNA of two ZO-1 isoforms, alpha+ and alpha-, were present in TM cells, and the former was expressed less than the latter. Only ZO-1 alpha- isoform protein was expressed in NTM cells, whereas proteins of both isoforms were found in GTM cells. DEX increased the protein levels of ZO-1 and Cx43 in both NTM and GTM cells. DEX also altered the F-actin architecture and promoted cross-linked actin network formation, the effects of which were more pronounced in GTM cells. CONCLUSIONS: Our findings not only provide molecular insights to the pathogenesis of GC-induced glaucoma but also suggest that junctional proteins ZO-1 and Cx43 as well as F-actin are targets for developing new modalities in glaucoma therapy.

[Long-term effects of non-penetrating trabecular surgery versus trabeculectomy for treating glaucoma].

OBJECTIVE: To evaluate clinical outcomes of non-penetrating trabecular surgery (NPTS) and trabeculectomy surgery (TS) in the treatment of primary open angle glaucoma (POAG). METHODS: It was a case-control study. A total of 63 patients (63 eyes) with POAG were observed retrospectively. Thirty one eyes and 32 eyes underwent NPTS and TS, respectively. Intraocular pressure (IOP), filtration bleb, visual field and post-operative complications were observed for 6-60 months. The CMH χ(2) test was used to analyse the difference of them. RESULTS: After operation, the IOP in the NPTS group were from (13.87 ± 4.88) mm Hg (1 mm Hg = 0.133 kPa) to (24.01 ± 6.55) mm Hg, the IOP in the TS group were from (11.90 ± 4.92) mm Hg to (19.10 ± 7.43) mm Hg. The IOP in the NPTS group was significantly higher than that in the TS group (F = 5.137, P < 0.05). The ratio of sustained filtration bleb of NPTS group after surgery was 25/31 (80.6%), while 6/31 were flat filtration bleb. There were statistically significant difference in the rate of disappearance of filtration bleb between these two groups (χ(2) = 8.129, P < 0.05). The difference of visual field loss postoperatively between these two groups was not statistically significant. The incidence rate of newly developed cataract after NPTS and TS was 6/31 and 12/32, respectively. The difference of rate of complication between these two groups was statistically non-significant (χ(2) = 3.797, P < 0.05). The successful rate after NPTS and TS was 61.54% (16/26) and 14.29% (4/28), respectively. The difference of successful rate between these two groups was statistically significant (χ(2) = 14.463, P < 0.05). CONCLUSIONS: Both NPTS and TS are effective methods for the treatment of POAG. Postoperative complications after NPTS are less than those of TS, But patients with TS could maintain a lower IOP than those with NPTS. Long-term efficacy of NPTS is uncertain, it's important to choose the suitable surgery to gain a high success rate.

Pro370Leu mutant myocilin impairs mitochondrial functions in human trabecular meshwork cells.

PURPOSE: Oxidative stress is a risk factor for the onset and progression of primary open-angle glaucoma (POAG), but the exact molecular basis remains unknown. Here, we investigated the mechanisms for Pro370Leu mutant myocilin to induce mitochondrial dysfunction and subsequent reactive oxygen species (ROS) generation in trabecular meshwork (TM) cells obtained from POAG individuals. METHODS: Primary non-diseased human TM cultures were transfected with pIRES-EGFP (Mock), pIRES-wild-type (WT), or pIRES-Pro370Leu mutant myocilin. Transfection efficiency and myocilin subcellular localization were determined by polymerase chain reaction (PCR), western blot analysis, and confocal microscopy. ROS levels as well as free Ca(2+) concentrations in cytoplasm ([Ca(2+)](c)) and mitochondria ([Ca(2+)]m) were examined by 2'7'-dichlorofluorescein diacetate (H(2)-DCF-DA), Fluo-3 acetoxymethyl ester (Fluo-3/AM), and Rhod-2 acetoxymethyl ester (rhod-2/AM), respectively, using flow cytometry. Mitochondrial functions were revealed by changes in mitochondrial membrane potential (DeltaPsim) and ATP production, which were found by fluorescent probe 5,5',6,6'-tetrachloro-1,1'3,3'-tetraethylbenzimid azolocarbocyanine iodide (JC-1) and a luciferin/luciferase-based ATP assay, respectively. RESULTS: Both WT and Pro370Leu mutant myocilin are localized in the mitochondria of TM cells as indicated using confocal microscopy and western blot analysis. Overexpression of WT myocilin decreases DeltaPsim, which is further reduced by Pro370Leu mutant myocilin. TM cells that overexpressed Pro370Leu mutant myocilin have greater cell death, higher endogenous ROS, [Ca(2+)](c), and [Ca(2+)](m) levels, and lower ATP production, and yet, these effects are not seen in the overexpression of WT myocilin. CONCLUSIONS: Our findings suggested that Pro370Leu mutant myocilin causes mitochondrial defects, which may lead to TM cell dysfunction and even cell death. Therefore, preventive measures targeting mitochondrial protection may delay the onset of glaucoma in individuals carrying the Pro370Leu myocilin mutation.

Suppression of IkappaBalpha increases the expression of matrix metalloproteinase-2 in human ciliary muscle cells.

PURPOSE: An increase of matrix metalloproteinase-2 (MMP-2) has been found to improve outflow through the uveoscleral pathway. This experiment was designed to test whether reduction of inhibitor of nuclear factor kappa B alpha (IkappaBalpha) levels could enhance MMP-2 expression in human ciliary muscle (HCM) cells in vitro. METHODS: The small interfering RNA (siRNA) targeting inhibitor of nuclear factor kappa B (IkappaBalpha) was transfected into HCM cells. The mRNA and protein levels of IkappaBalpha, nuclear factor-kappa B (NF-kappaB)p65, MMP-2, tissue inhibitor of metalloproteinase-2 (TIMP-2), and membrane-type 1 matrix metalloproteinase (MT1-MMP) in HCM cells were examined 24 h, 48 h, and 72 h after IkappaBalpha siRNA transfection by real-time reverse transcription polymerase chain reaction (RT-PCR) and western blot. The activation of NF-kappaBp65 was determined through the translocation of NF-kappaBp65 by fluorescence microscope. Gelatin zymography was used to detect the secretion and activity of MMP-2. RESULTS: Real-time RT-PCR and western blot showed that transfection of IkappaBalpha siRNA led to gradual downregulation of IkappaBalpha and TIMP-2 both at the mRNA and protein level after 24 h, 48 h and 72 h. The IkappaBalpha and TIMP-2 mRNA levels decreased 92.7%+/-1.6% and 70.3%+/-13.1%, respectively, and the protein levels were reduced 87.3%+/-2.0% and 62.9%+/-0.8% (p<0.01), respectively, when compared to the control 72 h after siRNA transfection. Conversely, the MMP-2 and MT1-MMP mRNA and protein levels increased in the time-dependent manner after IkappaBalpha siRNA transfection. The MMP-2 and MT1-MMP mRNA levels increased 178%+/-4.6% and 165%+/-8.2%, respectively, while protein levels were raised to 162%+/-3.7% and 157.6%+/-5.7% (p<0.01), respectively, when compared to the control 72 h after IkappaBalpha siRNA transfection. Although no obvious changes were seen in either mRNA or protein levels of total NF-kappaBp65 (p>0.05), the protein level of NF-kappaBp65 increased dramatically in the nucleus as revealed by western blot and fluorescence staining 24 h, 48 h, and 72 h after IkappaBalpha siRNA transfection. Moreover, gelatin zymography indicated that the secretion and activity of MMP-2 in treated cells were higher than those in the control cells. The maximum increases of pro-MMP-2 and active-MMP-2 were 172%+/-15% and 151%+/-14% (p<0.01), respectively, when compared to the control at the experiment's conclusion 72 h after siRNA transfection. CONCLUSIONS: Expression and activity of MMP-2 was enhanced by the IkappaBalpha siRNA in HCM cells through the activation of the NF-kappaB signaling pathway. Our results suggested that IkappaBalpha may therefore be a potential target for controlling the uveoscleral outflow pathway in glaucoma.

[The efficacy of surgical treatment for encapsulated cystic blebs around the plate after Ahmed glaucoma valve implantation].

OBJECTIVE: To evaluate the efficacy of surgical treatment for encapsulated cystic blebs around the plate after Ahmed glaucoma valve implantation. METHODS: This was a retrospective observational case series. Thirteen cases with elevated intraocular pressure (IOP) induced by fibrous proliferation around Ahmed valve were retrospectively studied. The intraocular pressure increased at 3 weeks to 5 months post-operation (2 months in average). The plates and tubes of Ahmed valve were in good position. No obstruction was seen in the tubes. Localized, encapsulated, cystic blebs were found in the equator of eyeball in which the plates were located. All cases underwent encapsulated cysts resection combined with mitomycin-C usage under conjunctival flaps. The excised tissues were sent for pathological examination. The changes of IOP and the outcome of the diseases were observed. RESULTS: IOP was normal one month after surgery in all cases. The success rate was 100%. In the 3 - 20 months follow-up period, the IOP level kept normal in 3 cases without any anti-glaucoma medications. The IOP level was in normal range in 6 cases using two or three different kinds of anti-glaucoma medications. Three cases needed further surgical treatment because of uncontrolled IOP even with anti-glaucoma medications. Among them, the complete success rate was 75% (3/4) in patients more than 57 years old, while the complete success rate was 0 (0/8) in patients less than 37 years old. CONCLUSIONS: It was considered an effective method of surgical treatment for encapsulated blebs around Ahmed glaucoma valve. The result was better in elderly patients than in younger patients relatively.

[Gene screening as the forewarning measure to predict glaucoma].

OBJECTIVE: To investigate the effect of gene screening on forewarning and monitoring of familial open-angle glaucoma pedigree. METHODS: Comprehensive ophthalmic examinations were performed in all available family members. The genomic DNA was extracted from peripheral blood. The myocilin gene was amplified and screened for mutations using direct sequencing. All family members were followed up. RESULTS: Among 12 family members, 5 individuals carry a C to T transition in exon 3 resulting in the substitution of proline to leucine (Pro370Leu), and the other 7 individuals did not carry this mutation. Ophthalmic examinations did not show any abnormality in the optic disc, the thickness of RNFL, and visual field parameters in mutation-carriers. During the follow-up, all carriers were diagnosed as open-angle glaucoma. The mean time of presentation of the defect of visual field was 21.6 months and 14.4 months after the changes in RNFL thickness. CONCLUSION: Genetic diagnosis was proven to be a method with high specificity and sensitivity; and can be used for presymptom diagnosis and forewarning in familial open-angle glaucoma pedigree.

[The inhibitive effects of 5-fluorouracil-loaded polylactic acid nanoparticles on human Tenon's capsule fibroblast in vitro].

OBJECTIVE: To investigate the inhibitive effects of 5-fluorouracil-loaded polylactic acid nanoparticles (5-FU-PLA-NPs) on human Tenon's capsule fibroblasts in vitro. METHODS: This paper was experimental study. MTT assay was performed to estimate the effects of 0.1 mg/L, 1.0 mg/L, 10.0 mg/L, 100.0 mg/L, 1000.0 mg/L unloaded polylactic acid nanoparticles (PLA-NPs) on fibroblasts proliferation on 48 h, 72 h. MTT assay was performed to estimate the effects of 0.1 mg/L, 1.0 mg/L, 10.0 mg/L, 100.0 mg/L, 1000.0 mg/L original 5-FU and 5-FU-PLA-NPs on fibroblasts' proliferation on 7 consecutive days respectively. The inhibitory rate of the two drugs against the fibroblasts was calculated. Cells were exposed to 100 mg/L original 5-FU and 5-FU-PLA-NPs respectively for 7 days. Semi-quantitative RT-PCR was used to observe the mRNA expression of type III procollagen at the 1st day, 5th day and 7th day. RESULTS: PLA-NPs had no cytotoxicity on the fibroblasts. Both 5-FU-PLA-NPs and original 5-FU could inhibit the proliferation of the fibroblasts and make the expression of type III procollagen mRNA lower in dose-dependent and time-dependent manner. The inhibitive effect of original 5-FU was more effective than 5-FU-PLA-NPs in the initial period, but the inhibitive effect of 5-FU-PLA-NPs was more effective than original 5-FU in long time. CONCLUSION: PLA has good biocompatibility and safety, and can be used as a carrier of ophthalmic drugs. 5-FU-PLA-NPs shows slow-releasing function. 5-FU-PLA-NPs can be proposed as a potential controlled and targeted ophthalmic delivery system for the treatment of antifibrosis after glaucoma filtering surgery.

[Clinical observation of trabeculectomy for primary angle closure glaucoma].

OBJECTIVE: To evaluate the effect of trabeculectomy on intraocular pressure (IOP) and complications in eyes with primary angle closure glaucoma (PACG). METHODS: It was retrospective clinical study. PACG was classified as acute primary angle closure glaucoma (APACG) group and chronic primary angle closure glaucoma (CPACG) group. APACG was then divided into acute attack phase and chronic phase, and CPACG was divided into chronic phase and late phase. IOP, best corrected visual acuity were compared before and after trabeculectomy in different subgroup of PACG. In addition, the incidence of complications of trabeculectomy was assessed. Statistical analyses were performed using SPSS 12.0 statistics software. Categorical variables such as best corrected visual acuity were compared using nonparametric test, continuous variables such as age and IOP were compared between the two groups using independent two-sample t-tests. Pre- and postoperative IOP were compared using one-way analysis of variance of repeated measures. RESULTS: 40 eyes (37 cases) of APACG and 56 eyes (45 cases) of CPACG were enrolled in this study. The median follow-up period was 24 and 25 months, respectively. IOP was significantly decreased from (53.6 +/- 17.9) mm Hg (1 mm Hg = 0.133 kPa, preoperation) to (10.5 +/- 4.9) mm Hg (postoperation) at time of discharging from hospital and (14.0 +/- 10.3) mm Hg at time of final follow up in APACG (F = 100.783, P < 0.01), respectively, and from (36.8 +/- 13.8) mm Hg to (11.7 +/- 4.2) mm Hg at time of discharging from hospital and (13.8 +/- 4.5) mm Hg at time of final follow up in CPACG (F = 54.383, P < 0.01), respectively. The IOP remained controlled (< or = 21 mm Hg) without antiglaucomatous medication in 38 eyes (95.0%) of APACG and in 50 eyes (89.3%) of CPACG. Visual acuity was significantly (H = 12.316, P < 0.01) decreased after trabeculectomy in all sub-types of PACG by Kruskal-wallis analysis. Shallow anterior chamber was commonly occurred after trabeculectomy. CONCLUSIONS: Trabeculectomy is an effective method to control IOP in APACG and CPACG. However, the high proportion of impaired vision was found following trabeculectomy in this study and warranted further clinical investigation.

Effects of chronic elevated intraocular pressure on parameters of optical coherence tomography in rhesus monkeys.

AIM: To determine the progression of parameters from optical coherence tomography (OCT) in chronic elevated intraocular pressure (IOP) monkeys. METHODS: A chronic elevated IOP model of rhesus monkeys was induced by laser photocoagulation. Representative OCT parameters, including the average and four-quadrant retinal nerve fiber layer (RNFL) thickness, and parameters from optic nerve head (ONH) analysis were collected before and after laser treatments biweekly for up to 28wk. The performance of each parameter for early progression detection was analyzed. The progressive trends toward elevated IOP were analyzed using a linear mixed-effects model. RESULTS: There were 10 successfully maintained high IOP eyes in 7 monkeys. The follow-up time was 24±5.37wk. With cumulative IOP elevation, the cup area, rim area and C/D area ratio were statistically significantly changed as early as 2wk after elevated IOP induction (P<0.05). The quadrant RNFL thickness changed at 6wk after high IOP induction, and the superior and inferior RNFL thicknesses exhibited more obvious reductions than other quadrants. The average RNFL thickness was the last one to show a significant decrease at 8wk. CONCLUSION: The parameters of ONH are most sensitive to elevated IOP in a primate glaucomatous model. These findings suggest that we should focus on those parameters instead of RNFL thickness in patients with elevated IOP, as they might present with earlier glaucomatous changes.

Pro370Leu MYOC gene mutation in a large Chinese family with juvenile-onset open angle glaucoma: correlation between genotype and phenotype.

PURPOSE: Glaucoma is the leading cause of irreversible blindness worldwide. Most of the cases are primary open angle glaucoma (POAG). POAG is a genetically heterogenous disease; autosomal dominance is the most frequent type of monogenic inheritance. In this study, we identified the genotype of a MYOC mutation and investigated the phenotype of a Chinese juvenile-onset open angle glaucoma (JOAG) pedigree (GZ.1 pedigree). METHODS: Blood samples were obtained from 24 participants. We performed sequence and gene linkage analysis in the GZ.1 pedigree retrospectively. Comprehensive ophthalmologic examinations were performed for each family member. Pharmacological treatment or filtering surgery was performed as needed according to the intraocular pressure (IOP) of each individual. RESULTS: A Pro370Leu myocilin mutation located in exon 3 of MYOC was identified in 24 members of the GZ.1 pedigree. Sixteen patients had juvenile-onset primary open-angle glaucoma (JOAG), and the others participating in the project had no such genotype. Analysis of polymorphic microsatellite markers indicated that the disease in GZ.1 is autosomal dominant inheritance. The patients in GZ.1 are characterized by early age of onset (before 35 years of age), severe clinical presentations, and high intraocular pressure unresponsive to pharmacological treatment; requiring 89.5% of the patients to undergo filtering surgery. Fortunately, the success rate of surgery was high. None of the patients required further medical treatment and only one demonstrated low IOP fundus changes. CONCLUSIONS: This is the first evidence of a founder effect for a Pro370Leu myocilin mutation in a Chinese POAG pedigree. The family with the Pro370Leu myocilin mutation presents with juvenile-onset glaucoma. After 10 years of follow-up, it is evident that the mutation is closely associated with the phenotype of the patients. Analysis of MYOC in JOAG patients may enable the identification of at-risk individuals and help prevent disease progression toward the degeneration of the optic nerve, and may also contribute to genetic counseling.

Analysis of MYOC gene mutation in a Chinese glaucoma family with primary open-angle glaucoma and primary congenital glaucoma.

BACKGROUND: Glaucoma is one of the leading causes of blindness in the world. Primary open-angle glaucoma (POAG) and primary congenital glaucoma (PCG) are subtypes of glaucoma. Myocillin is the first gene identified to be involved in POAG. Recently, myocillin mutation has been found in PCG. In this context, we reported a special glaucoma pedigree, which was composed of both PCG and POAG patients, and analyzed the mutation of myocillin in this pedigree. METHODS: The family was composed of the parents, a son and a daughter. All members of the family underwent the complete ophthalmologic examinations. All coding exons 1 - 3 and flanking introns of myocilin gene were screened for sequence alterations by polymerase chain reaction and direct DNA sequencing. RESULTS: The son was the proband, who was diagnosed as PCG in both eyes. The father was diagnosed as POAG in the right eye, the left eye was still normal. Both the sister and the mother of the proband had normal intraocular pressure without glaucomatous optic disc changes. The mutations in intron 2 of myocilin gene were detected in the family. While the proband and the father were homozygous, the mother and the sister were heterozygous for the mutation. CONCLUSIONS: Homozygous mutation in intron 2 of myocilin gene is involved in both POAG and PCG. It is suggested that the pathogenesis might be overlapping in POAG and PCG.

[The action of dexamethasone on trans epithelial electrical resistance and the expression of aquaporin-1 in immortalized trabecular cells].

OBJECTIVE: To study the effect of dexamethasone on trans epithelial electrical resistance (TEER) and the expression of AQP-1 in immortalized trabecular cells. METHODS: Immortalized human trabecular cells were cultured on PET membrane in 24 wells plate. The cells were treated with Dexamethasone in concentration of 10(-7) mol/L. After the cultured cell became confluence, TEER of the cells was detected to evaluate the resistance of outflow pathway at different time points. Western blot was used to analyze the expression of AQP-1 in the trabecular cells. RESULTS: The TEER and the expression of AQP-1 were much higher in the cells treated by Dexamethasone in comparison to control without dexamethasone exposure. CONCLUSIONS: The TEER and expression of AQP-1 in immortalized trabecular cells can be enhanced by Dexamethasone. AQP-1 upregulation induced by Glucocorticoid may relate with the increased resistance of outflow pathway in glaucoma.

[A preliminary study on gene mutation and its function in a large family (GZ.1 pedigree) with open-angle glaucoma].

OBJECTIVE: To determine the causative mutation of myocilin gene and to investigate its pathogenic function in a large Chinese pedigree (GZ.1) with familial open-angle glaucoma. METHODS: Genome-wide scanning was performed and the Lod scores were calculated. Candidate gene was amplified and screened for mutations using direct sequencing. To elucidate its expression, distribution and cytotoxicity of mutant myocilin, human trabecular cells (HTM) cells were transfected with pcDNA-wild-type and mutant myocilin vectors using liposomes. RESULTS: Mutation analysis of the myocilin gene showed a C-to-T transition at the 1, 109 th nucleotide in exon 3 resulting in a change of amino acid from proline to leucine (Pro370Leu). This mutation cosegregated with all affected individuals (16/16) and never presented in unaffected individuals (0/8). In transfected HTM cells, the mutant myocilin protein was not correctly processed in ER and accumulated as aggresome-like structures in the cytoplasma instead of being secreted. In addition, the expression of mutant protein also led to apoptosis of trabecular cells and the occurrence of. CONCLUSION: The mutation of Pro370Leu in myocilin gene could cause the accumulation of misfolding myocilin protein in HTM cells, which might lead to glaucoma in GZ.1 pedigree.

Role of mitochondria in the pathogenesis and treatment of glaucoma.

OBJECTIVE: To gain insight into the potential mechanism of mitochondria dysfunction in pathogenesis, progression and therapeutic management of glaucoma. DATA SOURCES: The data used in this review were mainly published in English from 2000 to present obtained from PubMed. The search terms were "mitochondria", "glaucoma" and "trabecular meshwork" or "retinal ganglion cells". STUDY SELECTION: Articles studying the mitochondria-related pathologic mechanism and treatment of glaucoma were selected and reviewed. RESULTS: Mitochondrial dysfunction or injury was demonstrated in different eye tissue of glaucoma. A variety of potential injuries (light, toxic materials, oxidative injury, mechanical stress, aging, etc.) and the inherent DNA defects are deemed to cause mitochondrial structural and functional destruction in trabecular meshwork cells, retinal ganglion cells, etc. of glaucoma. In addition, various new experimental and therapeutic interventions were used to preserve mitochondrial function, which may be useful for protecting against optic nerve degeneration or reducing the death of retinal ganglion cells in glaucoma. CONCLUSIONS: Mitochondria play an important role in the pathogenesis of glaucoma, various strategies targeting mitochondrial protection might provide a promising way to delay the onset of glaucoma or protect RGCs against glaucomatous damage.