Anterior lens capsule: biomechanical properties and biomedical engineering perspectives.

Anterior lens capsule, as the thickest basement membrane in the body, has its unique physiology characteristics. In ophthalmology, many attempts have been made to culture different kinds of cells including iris pigment epithelial cells, retinal pigment epithelial cells, corneal epithelium and endothelium cells, trabecular meshwork cells etc and anterior lens capsule has been confirmed to be served as an excellent scaffold for the growth and expansion of different ocular cells. Furthermore, anterior lens capsule also has unique potential in gestation evaluation and the treatment of various ocular diseases, including corneal ulcer, glaucoma, age-related macular degeneration and macular hole, etc. Here, we provide an overview of the biomechanical properties and biomedical engineering perspectives of anterior lens capsule.

Decorin inhibits glucose-induced lens epithelial cell apoptosis via suppressing p22phox-p38 MAPK signaling pathway.

PURPOSE: To determine the effect of decorin on oxidative stress and apoptosis of human lens epithelial (HLE) cells under high glucose condition. METHODS: HLE cell line (HLEB3) was incubated in normal glucose (5.5 mM) or high glucose (60 mM) medium. Decorin (50 nM) was applied 2 hours before high glucose medium was added. Apoptosis detection was executed by flow cytometry and western blotting (analysis of bcl-2 and bax). Oxidative stress level was measured by the generation of reactive oxygen species (ROS), glutathione peroxidase (GSH) and superoxide dismutase (SOD). P38 mitogen-activated protein kinase (MAPK) phosphorylation, the expression of p22phox of HLE cells and human lens anterior capsules were detected by western blotting. Small interfering RNA transfection to p22phox and p38 MAPK was also carried out on HLEB3. RESULTS: High glucose caused HLE cells oxidative stress and apoptosis exhibiting the increase of apoptotic cells and ROS production and decrease of bcl-2/bax ratio, GSH/GSSG ration and SOD activity. P22phox and phospho-p38 MAPK were upregulated in high glucose treated HLEB3 cells. Knocking down p22phox or p38 by siRNAs can reduce high glucose induced cell apoptosis and oxidative stress level. Silencing p22phox by siRNA can downregulate the phosphorylation of p38 MAPK. Decorin can inhibit the apoptosis, oxidative stress level and the induction of p22phox and phospho-p38 of HLEB3 induced by high glucose. Furthermore, the expression of p22phox and p38 were found significantly increased in lens anterior capsules of diabetic cataract patients compared to that of normal age-related cataract patients. CONCLUSIONS: Results showed that p22phox-p38 pathway may be participated in high glucose induced lens epithelial cell injury, decorin may inhibit the high glucose induced apoptosis and oxidative stress injury by suppressing this pathway in part.

Evaluation of collagen derived antiangiogenic factors and matrix metalloproteinases in anterior lens epithelial cells of pediatric eyes with persistent fetal vasculature.

Purpose: To measure levels of collagen-derived antiangiogenic factors (arresten, canstatin, tumstatin, endostatin) and matrix metalloproteinases (MMP-2 and MMP-9) in anterior lens epithelial cells (LECs) and anterior capsules of children with cataract and persistent fetal vasculature (PFV) as cases and cataract without PFV as controls. Methods: Anterior capsules harboring LECs were collected from pediatric cataract patients with (n = 13) and without PFV (n = 13) during surgery. Samples were immediately subjected to RNA extraction and cDNA preparation. Quantitative real time PCR was performed to determine the mRNA levels of antiangiogenic factors and matrix metalloproteinases. GAPDH (Glyceraldehyde 3-Phosphate Dehydrogenase) and ß Actin were used as the housekeeping control. The mRNA levels were expressed as a ratio, using the delta-delta method for comparing the relative expression results between controls and cases. The non-parametric Mann-Whitney U test was applied for statistical evaluation. P values < 0.05 were statistically significant. Results: The relative mRNA levels of arresten, canstatin, tumstatin, endostatin, MMP-2 and MMP-9 in cases were 6.20E-03 ± 0.003, 1.49E-01 ± 0.02, 1.70E-01 ± 0.007, 3.20E-03 ± 0.003, 1.11E-03 ± 0.0009 and 3.72E-04 ± 0.0001. The mRNA levels of arresten was 1.6 times lower (P = 0.01) while mRNA levels of MMP-2, tumstatin and canstatin were 4, 2.5, and 2.3 times higher in cases than in controls. No change was observed in mRNA levels of MMP-9 and endostatin (P = 0.82). Conclusion: A significant difference in the levels of arresten, canstatin, tumstatin, and MMP-2 was found in LECs with PFV.

Relationship between SIRT1 gene expression level and disease in age-related cataract cases

Background/aim: Age-related cataract is the most important visual impairment all over the world. Epigenetic modifications, especially overexpression of histone deacetylases, have become the focus of interest for cataract development in recent years. Sirtuin 1 (SIRT1), a class II histone deacetylase and a member of the sirtuin family, is one of the best-characterized histone deacetylases and has a pivotal role in age-related diseases. However, the association of SIRT1 with age-related cataracts has not yet been fully elucidated. Therefore, we aimed to determine the expression of SIRT1 in age-related cataract patients. Materials and methods: Expressions of SIRT1 were evaluated by quantitative polymerase chain reaction (qPCR) in patients and healthy controls. RNA samples were collected from the anterior capsule and peripheral blood samples of age-related cataract patients. Human lens epithelial cell line B3 and peripheral blood samples of healthy subjects were used as controls. Results: We determined that the expression of SIRT1 in blood and anterior capsule samples increased significantly compared to the control group (P < 0.05). Conclusion: The expression level of SIRT1 plays a vital role in the development of age-related cataract and it can be used as a biomarker. Thus, SIRT1 inhibitors can be used in the treatment of age-related cataract disease.

Anterior lens capsule and epithelium thickness measurements using spectral-domain optical coherence tomography.

BACKGROUND: To investigate the anterior lens capsule and epithelium thickness (defined as anterior lens capsular complex: ALCC) in normal Chinese subjects using spectral-domian optical coherence tomography (SD-OCT) and examine the factors that may influence the ALCC, such as age, gender, pupil diameter (PD) and signal strength index (SSI). METHODS: A prospective observational case series. One-hundred-thirty-four normal subjects (134 eyes) were included. The ALCCs were determined manually via SD-OCT. Using the pupil center as a reference position, the central ALCC (CALCC), nasal 1-mm ALCC (NALCC), temporal 1-mm ALCC (TALCC) and PD were measured manually. RESULTS: The mean CALCC, NALCC and TALCC were 33 ± 6 µm, 36 ± 7 µm and 34 ± 6 µm, respectively. The NALCC was significantly thicker than the CALCC (P < .001) and TALCC (P < .001). Moreover, CALCC was significantly thinner than TALCC (P = 0.013). Age was positively correlated with the CALCC (r = 0.292, P < .001), NALCC (r = 0.400, P < .001) and TALCC (r = 0.521, P < .001). PD, gender and SSI were not significantly correlated with the three ALCC parameters. CONCLUSIONS: The SD-OCT can be used to demonstrate the ALCC thickness, and age is positively correlated with the ALCC in the central, nasal and temporal sides.

Serum and aqueous xanthine oxidase levels, and mRNA expression in anterior lens epithelial cells in pseudoexfoliation.

PURPOSE: The aim of this study was to determine serum and aqueous xanthine oxidase (XO) levels, and mRNA expression in anterior lens epithelial cells in pseudoexfoliation (PEX). METHODS: In this prospective study, serum, aqueous and anterior lens capsules were taken from 21 patients with PEX and 23 normal subjects who had undergone routine cataract surgery. Serum and aqueous XO levels were analyzed using the colorimetric method. mRNA expression of XO in anterior lens epithelial cells was evaluated using reverse transcription polymerase chain reaction analysis. RESULTS: Serum XO levels (means ± standard deviations) were 207.0 ± 86.1 IU/mL and 240.6 ± 114.1 IU/mL in the normal and PEX groups, respectively (p = 0.310). Aqueous XO levels (means ± standard deviations) were 65.5 ± 54.3 IU/mL in the normal group and 130.5 ± 117.4 IU/mL in the PEX group (p = 0.028). There was a 2.9 fold decrease in mRNA expression in anterior lens epithelial cells of PEX, which is significantly lower than the normal group (p = 0.01). CONCLUSIONS: Higher aqueous XO levels lacking associated different serum XO suggests higher oxidative stress in the aqueous. Higher aqueous XO levels in PEX with decreased mRNA expression in anterior lens epithelial cells indicate possible overexpression of XO in other structures related to the aqueous.

The effect of gentian violet on human anterior lens epithelial cells.

PURPOSE: To evaluate whether the gentian violet staining of the anterior lens capsule during the cataract surgery is cytotoxic for the human lens epithelial cells, as an indirect indication of possible toxicity towards the corneal endothelium and the safety of gentian violet application. MATERIALS AND METHODS: Two groups of anterior lens capsules obtained during the cataract surgery, gentian violet stained and non-stained, were incubated with fluorescent dye Fura-2. Their fluorescence, upon excitation at 360 and 380 nm, was imaged to monitor changes in free intracellular calcium concentration ([Ca(2+)]i) in response to pharmacological stimulation by acetylcholine. The [Ca(2+)]i homeostasis is the indicator of cellular function. The changes in [Ca(2+)]i were compared between the two groups. RESULTS: Epithelial cells responded to acetylcholine in both groups of capsules - gentian violet stained (n = 17) and non-stained ones (n = 33). No significant differences of the elicited responses were found in rise time (p = 0.89), decay time (p = 0.61) or amplitude of [Ca(2+)]i (p = 0.96 for 63× and p = 0.26 for 40× objectives) between the two groups of capsules (Student t test). CONCLUSIONS: The staining of the anterior lens capsule with gentian violet during phacoemulsification in concentration of 0.01%, does not have detectable cytotoxic effects, which would affect the [Ca(2+)]i homeostasis in lens epithelial cells. The data, if extrapolated to corneal endothelium, exposed to the same concentration, suggest that gentian violet in concentration of 0.01% is safe as an adjunct for capsule visualization in cataract surgery.

Light microscopy and scanning electron microscopy analysis of rigid curved interface femtosecond laser-assisted and manual anterior capsulotomy.

PURPOSE: To study the microanatomic edge structures of anterior lens capsule specimens derived from manual and femtosecond laser-assisted capsulotomies. SETTING: Department of Ophthalmology, Goethe-University, Frankfurt, Germany. DESIGN: Experimental study. METHODS: Of 60 eyes with lens removal and intraocular lens implantation, 30 received a manual capsulotomy and 30 received a femtosecond laser-assisted capsulotomy (Lensx, rigid curved interface, pulse energy 15 µJ, spot separation 4 µm, layer separation 3 µm). After anterior capsule removal, tissues were immediately fixed in 4.5% formalin. Approximately 30 minutes after fixation, the tissues were removed from the fixation containers and air dried for at least 2 hours. Fifteen capsules in each group had further staining for light microscopy (LM). The surface of the capsulotomy edge was the primary focus of LM and scanning electron microscopy (SEM). Cell configuration, capsule shape, and abnormalities were evaluated. RESULTS: Subjective LM and SEM analysis showed smooth edges at all magnifications, no cell destruction, and cells up to the cutting edge in the manual capsulotomy group. Light microscopy demonstrated almost continuous anterior capsule incisions of the femtosecond laser-assisted capsulotomy, a prominent demarcation line along the cutting edge, and several tags and bridges. Scanning electron microscopy showed microgrooves and valley- and mountain-like structures as signs of the photodisruption process. CONCLUSION: Compared with manual procedures, curved, rigid interface femtosecond laser-assisted capsulotomy specimens using 15 µJ pulse energy showed tags, bridges, rougher edges, and demarcation lines on the capsulotomy edges on SEM but subjectively estimated a more round shape on LM. FINANCIAL DISCLOSURES: Mr. Klaproth received travel reimbursements and/or lecture fees from Alcon Laboratories, Inc., Rayner Intraocular Lenses Ltd., and Oculus GmbH. Dr. Kohnen received travel reimbursements, grant support, and/or lecture fees from Alcon Laboratories, Inc., Abbott Medical Optics, Inc., Bausch & Lomb, Carl Zeiss Meditec AG, Neoptics AG, Rayner Intraocular Lenses Ltd., and Schwind eye-tech-solutions GmbH and Co. KG; he is a consultant to Alcon Laboratories, Inc., Carl Zeiss Meditec AG, Rayner Intraocular Lenses Ltd., and Schwind eye-tech-solutions GmbH and Co. KG. No other author has a financial or proprietary interest in any material or method mentioned.

Human anterior lens capsule serving as a substrate for human trabecular meshwork cells cultivation.

To investigate if human anterior lens capsule is a suitable substrate for the culture of primary human trabecular meshwork (HTM) cells. Trabecular meshwork cells derived from four human donors were seeded on anterior lens capsules that were prepared from the lenses of donor eyes. Cell morphology and viability were examined at 1, 3, 5 and 7 days. Cell viability was measured based on a two-colour fluorescence assay (membrane-impermeable propidium iodide and membrane permeable Hoechst 33342). Immunocytochemistry studied Zonula occludens-1 (ZO-1), vimentin, tissue transglutaminase (tTgase) and Na(+)/K(+)-adenosine triphosphatase (Na(+)/K(+)-ATPase). Morphology of the cultivated cells followed a typical model while their viability was > 95% in all cases. ZO-1 was found at the cell boundaries of the HTM-AC complex. Vimentin was located at the lateral membranes of the HTM cells. Na(+)/K(+)-ATPase was found at the basolateral membrane of the HTM cells. tTgase was also identified. Anterior lens capsule can be considered as a suitable alternative substrate for cultivation of HTM cells and assist the expansion of existing knowledge about glaucoma pathophysiology and therapy.

Human anterior lens capsule epithelial cells contraction.

PURPOSE: Human anterior lens epithelial cells, attached to surgically isolated capsules, were found to contract upon stimulation. The purpose of this study was to characterize these contractions, which create gaps between cells, and to assess the underlying physiological mechanisms and their possible association with cataract formation. METHODS: Lens capsules obtained during cataract surgery were stained with fluorescent dye Fura-2. Its fluorescence, upon excitation at 360 and 380 nm, was imaged to monitor changes in cell morphology and cytosolic free Ca(2+) concentrations ([Ca(2+) ](i) ) in response to pharmacological stimulation by acetylcholine (ACh) and to mechanical stimulation by flow of saline or direct contact. RESULTS: Epithelial cells contracted in approximately a third of preparations when stimulated by either ACh application, fluid movement or direct mechanical contact. Contractions started either before or at best simultaneously with the rise in [Ca(2+) ](i). Contractions also occurred when there was hardly any change in [Ca(2+) ](i) upon application of physiological saline alone. The probability of contractions occurring did not differ significantly among cortical, nuclear and combined cortical + nuclear cataract. CONCLUSIONS: This study provides the evidence that contractions of the anterior lens epithelial cells take place in significant portion of human lens anterior capsule postoperative preparations after non-specific stimulation. Contractions are at least partially independent of changes in [Ca(2+) ](i). They can be mechanically induced, are localized and reversible and have a fast response and did not differ among different types of cataract. Physiological and clinical significance of this phenomenon remains to be elucidated.