Andrographolide suppresses epithelial mesenchymal transition by inhibition of MAPK signalling pathway in lens epithelial cells.

Epithelial mesenchymal transition (EMT) of lens epithelial cells (LECs) may contribute to the development of posterior capsular opacification (PCO), which leads to visual impairment. Andrographolide has been shown to have therapeutic potential against various cancers. However, its effect on human LECs is still unknown. The purpose of this study is to evaluate the effect of andrographolide on EMT induced by growth factors in the fetal human lens epithelial cell line (FHL 124). Initially the LECs were treated with growth factors (TGF-β2 and bFGF) to induce EMT. Subsequently these EMT-induced cells were treated with andrographolide at 100 and 500 nM concentrations for 24 h. Our results showed that FHL 124 cells treated with growth factors had a significant decrease in protein and m-RNA levels of epithelial markers pax6 and E-Cadherin. After administering andrographolide, these levels significantly increased. It was noticed that EMT markers α-SMA, fibronectin and collagen IV significantly decreased after treatment with andrographolide when compared to the other group. Treatment with andrographolide significantly inhibited phosphorylation of ERK and JNK. Cell cycle analysis showed that andrographolide did not arrest cells at G0/G1 or G2/M at tested concentrations. Our findings suggest that andrographolide helps sustain epithelial characteristics by modulating EMT markers and inhibiting the mitogen-activated protein kinase (MAPK) signalling pathway in LECs. Hence it can prove to be useful in curbing EMT-mediated PCO.

Estrogen mediated protection of cytoskeleton against oxidative stress.

Background & objectives: Cytoskeletal proteins are deregulated during oxidative stress and cataract formation. However, estrogen which protects against cataract formation and harmful effects of oxidative stress has not been tested on the cytoskeleton of lens epithelial cells (LECs). The current study was undertaken to assess if the protection rendered to LECs by estrogen was mediated by preserving the cytoskeletal proteins. Methods: Oxidative stress was induced by 50 μM of H2O2 in cultured goat LECs (gLECs) and effect of 1 μM 17β-estradiol (E2) was tested. After treatment, morphological analysis of cells was carried out using haematoxylin-eosin staining and cell density was also quantified. Cell viability was determined using Hoechst (Ho), YO-Pro (YP) and propidium iodide (PI). F-actin and vimentin were localized using phalloidin and anti-vimentin antibody, respectively, and viewed under fluorescence microscopy. Vimentin was further analysed at protein level by Western blotting. Results: H2O2 led to increased condensation of nucleus, cell death and apoptosis but these were prevented with pre- and co-treatment of E2 with increase in cell viability (P<0.001). E2 also prevented H2O2 mediated depolymerization of cytoskeleton but was not able to reverse the changes when given after induction of oxidative stress. Interpretation & conclusions: Our findings showed that E2 helped in preventing deteriorating effect of H2O2, inhibited cell death, apoptosis and depolymerisation of cytoskeletal proteins in LECs. However, the exact mechanism by which estrogen renders this protection to cytoskeleton of lens epithelial cells remains to be determined.

Cx43, ZO-1, alpha-catenin and beta-catenin in cataractous lens epithelial cells.

Specimens of the anterior lens capsule with an attached monolayer of lens epithelial cells (LECs) were obtained from patients (n052) undergoing cataract surgery. Specimens were divided into three groups based on the type of cataract: nuclear cataract, cortical cataract and posterior subcapsular cataract (PSC). Clear lenses (n011) obtained from donor eyes were used as controls. Expression was studied by immunofluorescence, real-time PCR and Western blot. Statistical analysis was done using the student’s t-test. Immunofluorescence results showed punctate localization of Cx43 at the cell boundaries in controls, nuclear cataract and PSC groups. In the cortical cataract group, cytoplasmic pools of Cx43 without any localization at the cell boundaries were observed. Real-time PCR results showed significant up-regulation of Cx43 in nuclear and cortical cataract groups. Western blot results revealed significant increase in protein levels of Cx43 and significant decrease of ZO-1 in all three cataract groups. Protein levels of alpha-catenin were decreased significantly in nuclear and cortical cataract group. There was no significant change in expression of beta-catenin in the cataractous groups. Our findings suggest that ZO-1 and alpha-catenin are important for gap junctions containing Cx43 in the LECs. Alterations in cell junction proteins may play a role during formation of different types of cataract.

Analysis of single nucleotide polymorphisms of CRYGA and CRYGB genes in control population of western Indian origin.

Aim: Polymorphisms in gamma-crystallins ( CRYG ) can serve as markers for lens differentiation and eye disorders leading to cataract. Several investigators have reported the presence of sequence variations within crystallin genes, with or without apparent effects on the function of the proteins both in mice and humans. Delineation of these polymorphic sites may explain the differences observed in the susceptibility to cataract observed among various ethnic groups. An easier Restriction Fragment Length Polymorphism (RFLP)-based method has been used to detect the frequency of four single nucleotide polymorphisms (SNPs) in CRYGA / CRYGB genes in control subjects of western Indian origin. Materials and Methods: A total of 137 healthy volunteers from western India were studied. Examination was performed to exclude volunteers with any ocular defects. Polymerase chain reaction (PCR)-RFLP based method was developed for genotyping of G198A (Intron A), T196C (Exon 3) of CRYGA and T47C (Promoter), G449T (Exon 2) of CRYGB genes. Results: The exonic SNPs in CRYGA and CRYGB were found to have an allele frequency 0.03 and 1.00 for ancestral allele respectively, while frequency of non-coding SNP in CRYGA was 0.72. Allele frequency of T90C of CRYGB varied significantly ( P = 0.02) among different age groups. An in-silico analysis reveals that this sequence variation in CRYGB promoter impacts the binding of two transcription factors, ACE2 (Member of CLB2 cluster) and Progesterone Receptor (PR) which may impact the expression of CRYGB gene. Conclusions: This study establishes baseline frequency data for four SNPs in CRYGA and CRYGB genes for future case control studies on the role of these SNPs in the genetic basis of cataract.

Comparison between Ringer's lactate and balanced salt solution on postoperative outcomes after phacoemulsfication: A randomized clinical trial.

Aim: To compare the effects of balanced salt solution (BSS) and Ringer's lactate (RL) on corneal thickness, endothelial morphology, and postoperative anterior chamber inflammation in eyes undergoing phacoemulsification. Setting: Iladevi cataract and IOL research center, Ahmedabad, India. Materials and Methods: This prospective randomized study comprised 90 consecutive patients with age-related cataract who were randomly assigned to either Group 1 (n = 45) with BSS or Group 2 (n = 45) with RL. Observations made included measurement of central corneal thickness (CCT), presence of anterior chamber flare and cells, endothelial cell loss, and change in coefficient of variation (CV). Data was analyzed using Mann Whitney test and test of proportion. Results: Mean increase in CCT on postoperative Day 1 was 58microm and 97microm in Groups 1 and 2 respectively ( P = 0.01). Increase in CCT at one month was 10microm and 11microm in Groups 1 and 2 respectively ( P = 0.99); increase in CCT at three months was 3microm and 6microm in Groups 1 and 2 respectively ( P = 0.86). Number of eyes with flare grades in a range of 0 to 3 was statistically higher in Group 2 on postoperative Day 1 ( P = 0.004, 0.016, < 0.001, 0.047 for Grade 0, 1, 2 and 3 respectively). Number of eyes with cells of Grade 3 on first postoperative day was significantly higher in Group 2 as compared to Group 1 ( P = 0.004). Three months postoperatively, endothelial cell loss was 5.5% and 7.8% in Groups 1 and 2 ( P = 0.21) and change in CV was 3 and 5.4 in Groups 1 and 2 ( P = 0.20) respectively. Conclusion: BSS offers a significant advantage over RL in terms of increase in corneal thickness and postoperative inflammation on the first postoperative day in patients undergoing phacoemulsification.

The probable associations for corticocapsular adhesions in patients undergoing cataract surgery: A clinic-based observational study.

Background: Corticocapsular adhesions (CCA) are frequently seen between lens capsule and adjacent cortical layer. During cataract surgery, in the presence of CCA, excessive efforts to rotate the nucleus can result in zonular damage. To reduce morbidity, identification of associations with CCA can be helpful in appropriately modifying the surgical procedure. Aim: To investigate probable associations with CCA in patients undergoing cataract surgery. Setting and Design: Iladevi Cataract and IOL research center. Case-control study. Materials and Methods: A single eye of 600 patients, 200 patients with CCA (cases) and 400 patients without CCA (controls) were considered. A CCA diagnosis was based on: (i) preoperative presence of CCA on slit-lamp examination with visualization of furry surface of cortex during surgery; (ii) preoperative absence of CCA on slit-lamp examination but intraoperative visualization of furry surface of cortex. Variables such as age, gender, type of cataract, grade of cataract, high myopia, diabetes mellitus and hypertension were studied. Statistical Analysis: Multivariate logistic regression was done. Results were presented as odds ratio (OR) with 95% CI. Results: Mean age was 64.71 +/- 9.10 years in cases and 59.27 +/- 8.79 years in controls. Presence of CCA increased with age from 22% (n = 59) in 45 to 49 years to 70% (n = 110) in 70 to 79 years. An increase in age was associated with CCA by 3.3% (OR = 3.3%, P = 0.028). The odds of CCA for females were 83% higher ( P = 0.027). Presence of anterior cortical cataract increased odds of CCA by 9.5 times ( P = 0.001), while posterior cortical cataract increased odds by 3.3 times ( P = 0.001). Conclusion: Corticocapsular adhesions were strongly associated with cortical cataracts, increased age and female gender.

Phacoemulsification in total white cataract with Stevens-Johnson syndrome.

Cataract surgery with Stevens-Johnson syndrome (S-J) is associated with a high incidence of complications and can worsen the primary disease. We report a case of phacoemulsification of a total, white cataract barely visible through the vascularized, keratinized cornea in the only seeing eye of a patient with S-J syndrome. We describe the intra-operative difficulties encountered during phacoemulsification and the surgical tools and techniques employed to overcome the surgical difficulties. The postoperative course was uneventful and the patient regained useful, navigational vision.

Preoperative prediction of posterior capsule plaque in eyes with posterior subcapsular cataract.

AIM: To determine whether the plaque on the posterior capsule can be predicted preoperatively, in patients with posterior subcapsular cataract (PSC), undergoing cataract surgery. MATERIALS AND METHODS: A prospective study of 140 consecutive eyes with PSC, who underwent cataract surgery, was conducted. The prediction of preoperative presence or absence of plaque within the PSC was noted on slit lamp examination, in dilated pupils. A single observer made the observations under oblique illumination, where the slit lamp was placed at an angle of 30 degrees to 45 degrees. Evaluation of the plaque through slit lamp examination was standardized in terms of illumination and magnification. The observations were recorded using a video camera (Image archiving system, Carl Zeiss, Jena Germany) attached to a slit lamp (Carl Zeiss, SL 120 Jena, Germany), keeping the illumination at 100%. The prediction of plaque was noted in terms of its presence or absence on the posterior capsule. All the patients received counseling regarding the presence of plaque. Capsule polishing of the posterior capsule in Cap Vac mode, was done in all cases. The posterior capsule was examined for presence or absence of plaque, either on the first postoperative day, or within a week, with maximal mydriasis. The observer's results were tabulated and later analyzed to judge the incidence of predictability of plaque in PSC. RESULTS: The mean age of the patients was 45+/-6.2 years (range 32-61 years); 104 (74.3%) were males. One hundred and eight (77.1%) patients were under 50 years. The presence or absence of plaque was predicted correctly in 124 (88.6%) eyes. The prediction of plaque was incorrect in 16 (11.4%) eyes. CONCLUSION: The prediction of presence or absence of plaque was accurate in 88.6% cases. We believe that counseling patients with posterior capsule plaque before the surgery is the key to avoiding unpleasant surprises.

Management of congenital bilateral subluxated cataract with Cionni ring.

Use of Cionni ring helped in the successful intraocular lens (IOL) implantation in a 30-year-old patient with bilateral congenital subluxated cataracts. The IOL was stable, well-centered without any iridodonesis or pseudophakodonesis.

Corneal hydration intra-operatively during phacoemulsification.

PURPOSE: To evaluate incidence and risk factors for corneal hydration intra-operatively during phacoemulsification (CHIP). METHODS AND MATERIALS: This was a randomised, clinical trial, where 240 eyes of 240 patients with senile cataracts undergoing phacoemulsification were prospectively randomised to receive one of three possible types of incision architecture - single, two or three planes. Viscoelastic injection through paracentesis, before instruments were withdrawn from the main incision, was randomly done. Incision length was grouped into 1.5, 2 and 2.5 mm. CHIP was graded as nil, mild, moderate or severe: nil - no visible CHIP, mild-CHIP involving pillars of the incision, moderate - CHIP involving pillars and lateral extension, severe - extension of CHIP in front into the clear cornea. Moderate and severe categories were combined as 'significant' CHIP. Statistical analysis was done by logistic regression. RESULTS: The incidence of significant CHIP was 17.9 % (43 eyes out of 240). Five out of 80 eyes (6.25%) single plane, 8 out of 80 (10%) two plane and 30 out of 80 (37.5%) three- plane incisions developed significant CHIP (p = 0.005). Fifteen out of 120 (12.5%) with and 28 out of 120 (23.3%) without injection of viscoelastic developed significant CHIP (p = 0.044). Eighteen out of 43 (42.8%) and 25 out of 43(58.1%) with incision lengths 2 and 2.5 mm developed significant CHIP respectively (p<0.001). CONCLUSIONS: The incidence of significant CHIP was 17.9 %. Three-plane incisions, not injecting viscoelastic prior to retraction of instruments and incision length 2 mm or more are risk factors for significant CHIP.

Intraoperative performance and longterm outcome of phacoemulsification in age-related cataract.

PURPOSE: To evaluate intraoperative performance and longterm surgical outcome after phacoemulsification of age-related cataracts. METHODS: Prospective, observational, non-comparative study of 165 consecutive eyes undergoing phacoemulsification with nuclear sclerosis Grade I to III (Scale I to V). Preoperative evaluation included specular microscopy. Phacoemulsification was performed by a single surgeon using a standardised surgical technique under topical anaesthesia. Intraoperatively, effective phaco time (EPT), wound site thermal injury (WSTI), serious complications (eg. vitreous loss, posterior capsule rupture, zonulolysis) and intraoperative posterior capsule opacification (plaque) were evaluated. Postoperatively, posterior capsule opacification (PCO), Neodymium:YAG (Nd:YAG) laser posterior capsulotomy rate, corneal endothelial count, best corrected visual acuity and cystoid macular oedema were evaluated. Eyes were examined at 6 months and then yearly for 3 years. RESULTS: Mean ages of 78 males and 87 females were 59.12 +/- 8.56 and 58.34 +/- 7.45 years respectively. EPT was 36 +/- 19 seconds and WSTI occurred in 7 eyes (4.7%). No serious intraocular complications occurred. Intraoperative posterior capsule opacification (plaque) was present in 21 eyes (13.93%). Postoperatively, PCO occurred in 8 eyes (4.84%) and Nd:YAG laser posterior capsulotomy was performed in 3 eyes (1.8%). Endothelial cell loss was 7.1% at 3 years follow-up. At the end of 3 years follow-up, 146 eyes (88.89%) maintained a best corrected visual acuity of > or = 6/12. Cystoid macular oedema did not occur in any eye at 1 and 6 months' follow-up. CONCLUSION: PCO rates and endothelial cell loss were acceptable. Consistent and reproducible outcome can be obtained after phacoemulsification of age related cataracts (grade I to III).

Corneal endothelial safety of intracameral preservative-free 1% xylocaine.

PURPOSE: To evaluate the effect of intracameral preservative-free 1% xylocaine on the corneal endothelium as an adjuvant to topical anaesthesia during phacoemulsification and Acrysof foldable IOL implantation. MATERIAL & METHODS: This is a prospective, controlled, randomised, double-masked study. 106 patients with soft to moderately dense (Grade 1-3) senile cataract and corneal endothelial cell density of >1500/mm2 were randomised to the xylocaine group (n=53) and control group(n=53). Central endothelial specular microscopy and ultrasound corneal pachymetry were performed preoperatively. On the first postoperative day the eyes were evaluated for corneal oedema and Descemet's folds. Ultrasound corneal pachymetry was performed at 1, 3 and 12 months. Specular microscopy was performed at 3 and 12 months. Cell loss was expressed as a percentage of preoperative cell density. Six patients could not complete one year follow-up. Chi-square and paired t test (2 tail) statistical tests were applied for analysis. RESULTS: Four (7.54%) patients in the xylocaine group and 5 (9.43%) in the control group had a few Descemet's folds associated with mild central stromal oedema. Corneal thickness increased from 549.3micro +/- 37.2micro to 555.5micro +/- 36.5micro in the xylocaine group and from 553.1micro +/- 36.2micro to 559.3micro +/- 40.5micro in the control group at the one-month postoperative visit. Thickness returned to the preoperative level in xylocaine group 549.6micro +/- 34.5micro and control group 554.7micro +/- 41.1micro at three months. (P=0.484) The percentage of cell loss was 4.47 +/- 2.53% in the xylocaine group and 4.49 +/- 3.09% in the control group at one year. (P=0.97) CONCLUSION: Intracameral preservative-free 1% xylocaine does not appear to affect corneal endothelium adversely during phacoemulsification.

Sir Nicholas Harold Ridley. He changed the world, so that we might better see it.

Cataract surgery with intraocular lens (IOL) implantation has become the most common and most successful of all operations in medicine. Sir Harold Ridley's first cataract extraction with implantation of an IOL marked the beginning of a major change in the practice of ophthalmology. Millions of patients worldwide have benefited from Sir Ridley's invention, and are likely to continue to derive benefit from this device. However, the development of the IOL was not without its share of ups and downs. Sir Harold Ridley, the inventor of IOL, died at the age of 94, on 25 May 2001, and ophthalmology lost one of its greatest and most influential practitioners. We are happy that he lived to enjoy the fruits of his labour--to see the amazing improvements and the expansive growth that evolved in the cataract-IOL technique, from early and unsatisfactory operations in previous decades, to the superb results attainable today. The invention of the IOL has not been just the addition of one new form of treatment, but rather, Sir Harold's tiny disc-shaped sliver of plastic has changed the world so that our patients may better see it. This article presents a brief biographical sketch of Sir Harold and lists his major inventions and contributions to ophthalmology.

Phacoemulsification in subluxated cataract.

PURPOSE: To evaluate the outcome of phacoemulsification in eyes with subluxated cataract. MATERIALS AND METHODS: This retrospective study comprised 22 eyes of 20 consecutive patients with subluxated cataracts of varying aetiology operated between March 1998 and March 2001. Detailed preoperative assessment included visual acuity (VA), slitlamp examination, presence of vitreous in anterior chamber extent of subluxation, intraocular pressure (IOP) and detailed fundus examination. Phacoemulsification was done to retain the natural bag support and all patients had acrylic foldable Acrysof IOL implantation either in-the-bag or by scleral fixation. Postoperative observations included best-corrected visual acuity (BCVA), IOP, pupillary reaction and the IOL position. RESULTS: The aetiology of the subluxation was traumatic in 11 patients and non-traumatic in 9 patients. Fifteen were males and 5 were females. Mean follow-up was 11.7 +/- 9.71 months (range 4-39). The average age was 39.15 +/- 16.33 (range 5-74). A 2-port anterior chamber vitrectomy was performed in 6 eyes (27.2%). Capsule tension ring (CTR) was implanted in 15 eyes (68.18%). Twelve eyes (54.5%) had in-the-bag implants, while 5 (22.72%) had scleral fixation. The remaining 5 eyes (22.72%) had one haptic in-the-bag and another sutured to sclera. No major intraoperative complications were noted. Twelve eyes (54.5%) had clinically and geometrically well centered IOLs while 9 eyes (40.9%) had geometrically decentered IOLs. One patient was lost to follow-up. Fifteen eyes (55.55%) had postoperative BCVA of 6/12 - 6/6 while 2 eyes (7.40%) had BCVA of 6/18. The remaining 4 eyes (14.81%) had less than 6/24 BCVA due to pre-existing posterior segment pathology. Postoperative complications included rise in IOP in 1 eye (4.54%), pupillary capture of the IOL optic in 2 eyes (9.09%); the same 2 eyes (9.09%) required redialing of IOL. One eye (4.54%) had to undergo refixation (one haptic was fixed to sclera) year after cataract surgery. Postoperative retinal detachment was noticed in one patient after a month of phacoemulsification. CONCLUSION: In subluxated cataracts it is essential to have appropriate parameters depending on the grade of cataract. This contributes to a safe and predictable outcome in subluxated cataract surgery.