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result(s) for
"Laboratory Study"
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Variation in rod and cone density from the fovea to the mid-periphery in healthy human retinas using adaptive optics scanning laser ophthalmoscopy
Purpose
To characterize the rod and cone photoreceptor mosaic at retinal locations spanning the central 60°
in vivo
using adaptive optics scanning laser ophthalmoscopy (AO-SLO) in healthy human eyes.
Methods
AO-SLO images (0.7 × 0.9°) were acquired at 680 nm from 14 locations from 30° nasal retina (NR) to 30° temporal retina (TR) in 5 subjects. Registered averaged images were used to measure rod and cone density and spacing within 60 × 60
μ
m regions of interest. Voronoi analysis was performed to examine packing geometry at all locations.
Results
Average peak cone density near the fovea was 164 000±24 000 cones/mm
2
and decreased to 6700±1500 and 5400±700 cones/mm
2
at 30° NR and 30° TR, respectively. Cone-to-cone spacing increased from 2.7±0.2
μ
m at the fovea to 14.6±1.4
μ
m at 30° NR and 16.3±0.7
μ
m at 30° TR. Rod density peaked at 25° NR (124 000±20 000 rods/mm
2
) and 20° TR (120 000±12 000 rods/mm
2
) and decreased at higher eccentricities. Center-to-center rod spacing was lowest nasally at 25° (2.1±0.1
μ
m). Temporally, rod spacing was lowest at 20° (2.2±0.1
μ
m) before increasing to 2.3±0.1
μ
m at 30° TR.
Conclusions
Both rod and cone densities showed good agreement with histology and prior AO-SLO studies. The results demonstrate the ability to image at higher retinal eccentricities than reported previously. This has clinical importance in diseases that initially affect the peripheral retina such as retinitis pigmentosa.
Journal Article
Local complement activation in aqueous humor in patients with age-related macular degeneration
Purpose
To investigate complement activation in aqueous humor and in plasma of patients with neovascular age-related macular degeneration (nAMD).
Patients and methods
Aqueous humor and EDTA-plasma of 31 nAMD patients and 30 age-matched controls was collected. The levels of the complement factor 3 (C3), the regulators factor H (FH), and factor I (FI), and of the complement activation products Ba, C3a, and the terminal complement complex (sC5b-9) were measured. Associations between complement levels and phenotype were determined using Mann–Whitney
U
-test.
Results
In plasma, no significant differences were found between the nAMD group and the control group. In aqueous humor, significantly increased levels of Ba (
P
=0.002), and C3a (
P
=0.002) indicate local complement activation in nAMD patients and a trend for a concomitant upregulation of the complement regulators FH (
P
=0.02) and FI (
P
=0.04).
Conclusions
Our findings provide strong evidence for a local complement dysregulation in nAMD patients.
Journal Article
Robot-assisted intraocular surgery: development of the IRISS and feasibility studies in an animal model
Purpose
The aim of this study is to develop a novel robotic surgical platform, the IRISS (Intraocular Robotic Interventional and Surgical System), capable of performing both anterior and posterior segment intraocular surgery, and assess its performance in terms of range of motion, speed of motion, accuracy, and overall capacities.
Patients and methods
To test the feasibility of performing ‘bimanual’ intraocular surgical tasks using the IRISS, we defined four steps out of typical anterior (phacoemulsification) and posterior (pars plana vitrectomy (PPV)) segment surgery. Selected phacoemulsification steps included construction of a continuous curvilinear capsulorhexis and cortex removal in infusion–aspiration (I/A) mode. Vitrectomy steps consisted of performing a core PPV, followed by aspiration of the posterior hyaloid with the vitreous cutter to induce a posterior vitreous detachment (PVD) assisted with triamcinolone, and simulation of the microcannulation of a temporal retinal vein. For each evaluation, the duration and the successful completion of the task with or without complications or involuntary events was assessed.
Results
Intraocular procedures were successfully performed on 16 porcine eyes. Four eyes underwent creation of a round, curvilinear anterior capsulorhexis without radialization. Four eyes had I/A of lens cortical material completed without posterior capsular tear. Four eyes completed 23-gauge PPV followed by successful PVD induction without any complications. Finally, simulation of microcannulation of a temporal retinal vein was successfully achieved in four eyes without any retinal tears/perforations noted.
Conclusion
Robotic-assisted intraocular surgery with the IRISS may be technically feasible in humans. Further studies are pending to improve this particular surgical platform.
Journal Article
A binocular iPad treatment for amblyopic children
Purpose
Monocular amblyopia treatment (patching or penalization) does not always result in 6/6 vision and amblyopia often recurs. As amblyopia arises from abnormal binocular visual experience, we evaluated the effectiveness of a novel home-based binocular amblyopia treatment.
Methods
Children (4–12 y) wore anaglyphic glasses to play binocular games on an iPad platform for 4 h/w for 4 weeks. The first 25 children were assigned to sham games and then 50 children to binocular games. Children in the binocular group had the option of participating for an additional 4 weeks. Compliance was monitored with calendars and tracking fellow eye contrast settings. About half of the children in each group were also treated with patching at a different time of day. Best-corrected visual acuity, suppression, and stereoacuity were measured at baseline, at the 4- and 8-week outcome visits, and 3 months after cessation of treatment.
Results
Mean (±SE) visual acuity improved in the binocular group from 0.47±0.03 logMAR at baseline to 0.39±0.03 logMAR at 4 weeks (
P
<0.001); there was no significant change for the sham group. The effect of binocular games on visual acuity did not differ for children who were patched
vs
those who were not. The median stereoacuity remained unchanged in both groups. An additional 4 weeks of treatment did not yield additional visual acuity improvement. Visual acuity improvements were maintained for 3 months after the cessation of treatment.
Conclusions
Binocular iPad treatment rapidly improved visual acuity, and visual acuity was stable for at least 3 months following the cessation of treatment.
Journal Article
MicroRNA-125b promotes tumor growth and suppresses apoptosis by targeting DRAM2 in retinoblastoma
Purpose
It is known that microRNAs (miRNAs) are a class of small, noncoding RNAs that act as key regulators in various physiological and pathological processes. However, the regulatory mechanisms involving miRNAs in retinoblastoma (RB) remain largely unknown. The miRNA miR-125b is dysregulated in various human cancers such as breast cancer, human hepatocellular carcinoma, ovarian cancer, and colorectal cancer. However, the significance of miR-125b in RB has not been sufficiently investigated. Our objective was to explore the role of the miR-125b in RB.
Methods
In this study, we measured miR-125b levels using real-time polymerase chain reaction in human RB cell lines, including HXO-Rb44, Y79, SO-RB50, and the normal human retinal pigment epithelial cell line ARPE-19; a total of 38 pairs of primary RB tissues and adjacent noncancerous tissues were also measured. In addition, overexpression of miR-125b in RB cell lines was performed to determine the role of miR-125b in RB.
Results
We found that miR-125b is significantly upregulated in RB, and closely associated with tumor cell proliferation and apoptosis. In addition, overexpression of miR-125b apparently promotes RB cell proliferation and migration
in vitro
. Gain-of-function
in vitro
experiments further showed that the miR-125b mimic significantly suppressed RB cell apoptosis. A subsequent dual-luciferase reporter assay identified the suppressor gene
DRAM2
as direct target of miR-125b.
Conclusions
Our data collectively demonstrate that miR-125b is a suppressor gene miRNA that can promote RB cell proliferation and migration by downregulating the suppressor gene
DRAM2
, indicating that miR-125b may represent a new potential diagnostic and therapeutic target for RB treatment.
Journal Article
Fibrin glue inhibits migration of ocular surface epithelial cells
Purpose
Fibrin glue has been used successfully in numerous ophthalmic surgical procedures. Recently, fibrin glue has been used in limbal stem cell transplantation to reduce both operative time and to negate the need for sutures. The aim of this study was to determine the effects of fibrin glue on epithelial cell migration
in vitro
.
Methods
Corneoscleral rims were split to retain the epithelial layer, Bowman’s layer, and anterior stroma. Rims were cut into eight equal-sized pieces and were placed directly on culture plates or affixed with fibrin glue. Rims were maintained in culture for 25 days and epithelial cell growth was monitored. Cells were photographed to measure area or growth and immunofluorescence staining of explants for fibrin was performed.
Results
Explants that were glued demonstrated significantly delayed epithelial cell growth and migration as compared with explants without glue. By day 16, all fibrin glue had dissolved and coincided with onset of cell growth from glued explants. Cell growth commenced between days 3 and 4 for control explants without glue and around days 14–16 for explants with fibrin glue.
Conclusions
Fibrin glue delays epithelial cell migration by acting as a physical barrier and can potentially interfere with explant-derived limbal epithelial cell migration on to the corneal surface. We propose that glue should be used to attach the conjunctival frill of the limbal explant but care should be taken to ensure that the glue does not wrap around the explant if used to secure the explant as well. Strategic use of glue, to attach the recessed conjunctiva, can be advantageous in delaying conjunctival cell migration and reducing the need for sequential sector conjunctival epitheliectomy.
Journal Article
A novel and less invasive technique to assess cytokine profile of vitreous in patients of diabetic macular oedema
PurposeA pilot study to validate the collection of vitreous reflux (VR) after intravitreal injection using Schirmers tear strips was carried out. We assessed its efficiency for proteomics studies by estimating the differential expression of 27 cytokines using multiplexed bead array in diabetic macular oedema and proliferative diabetic retinopathy. To set, validate and assess the efficacy of Schirmer tear strips for collecting VR in patients undergoing intravitreal injections for diabetic macular oedema (DME).Patients and methodsVR samples were collected from 11 eyes of DME patients after intravitreal injections using Schirmer tear strips. Undiluted vitrectomy samples were obtained from six eyes of non-diabetic patients with idiopathic macular hole and seven eyes of diabetic patients with high-risk proliferative diabetic retinopathy (Hr-PDR), which were also subsampled on the Schirmer tear strips. Tear sampling was done in a subset of the DME patients. Total protein concentration between VR and vitrectomy samples was compared. Levels of the set of 27 cytokines in Schirmer tear strips samples were measured. Inter-group comparison for cytokines was done using Mann-Whitney U-test.ResultsSimilar protein concentration in VR samples and vitrectomy samples (P<0.05) was obtained. Tear protein contamination was not detected in VR samples. In comparison with no-DR patients, 25 and 20 of the measured 27 cytokines were significantly elevated (P<0.05) in the Hr-PDR and DME patients, respectively. As compared with no-DR patients, vascular endothelial growth factor was only moderately elevated in DME patients (P>0.05), but significantly elevated in Hr-PDR patients (P<0.05). Interleukin 1 receptor antagonist/interleukin 1b (IL1RA/IL1b) ratio was 13 times higher in DME patients as compared with Hr-PDR group.ConclusionWe demonstrated a simple, safe method of VR sampling. This technique provides a pure, albeit small, vitreous sample for proteomics. IL1RA/IL1b ratio was found to be 13-fold higher in the DME group as compared to the Hr-PDR.
Journal Article
A hyaluronan hydrogel scaffold-based xeno-free culture system for ex vivo expansion of human corneal epithelial stem cells
Purpose
To develop a hyaluronan hydrogel scaffold-based xeno-free culture system for
ex vivo
cultivation of human corneal epithelial stem cells (CESCs).
Patients and Methods
CESCs were cultivated from donor limbal explants on the HyStem-C Hydrogel bio-scaffold in 12-well plates for 3 weeks. Group A used the traditional supplemented hormonal epidermal medium (SHEM) and group B used the defined SHEM (without fetal bovine serum and toxin A, adding 20% serum replacement). The growth and morphology of the cultured cells were assessed by phase contrast microscope. The expressions of specific cell markers were assessed by immunofluorescence staining and quantitative real-time PCR (qRT-PCR).
Results
Successful cultures of CESCs were obtained in both groups, resulting in multilayered stratified epithelia. Comparing to group A, the cells in group B was grown slightly slower and formed less cellular layers at the end of culture. The corneal specific cytokeratin (K) 12 and differentiation markers, involucrin, and connexin 43, were mainly expressed in the superficial cellular layers in both groups. Interestingly, certain basal cells were immune-positive to proposed stem cell markers such as K19, ABCG2, and integrin
β
1 in both groups. There was no significant difference between the two groups with regard to the gene expression levels of all these selected corneal markers (all
P
>0.05).
Conclusions
The hyaluronan hydrogel scaffold-based xeno-free culture system may support the expansion of regenerative CESCs without the risk of xeno component contamination. The regenerated epithelium maintains similar characteristics of native corneal epithelium.
Journal Article
Air pressure changes in the creation and bursting of the type-1 big bubble in deep anterior lamellar keratoplasty: an ex vivo study
PurposeTo measure the pressure and volume of air required to create a big bubble (BB) in simulated deep anterior lamellar keratoplasty (DALK) in donor eyes and ascertain the bursting pressure of the BB.Patients and methodsTwenty-two human sclera-corneal discs were used. Air was injected into the corneal stroma to create a BB and the pressure measured by means of a pressure converter attached to the system via a side port. A special clamp was designed to prevent air leak from the periphery of the discs. The pressure at which air emerged in the corneal tissue; the bursting pressure measured after advancing the needle into the bubble cavity and injecting more air; the volume of air required to create a BB and the volume of the BB were ascertained.ResultsType-1 BB were achieved in 19 and type-2 BB in 3 eyes. The maximum pressure reached to create a BB was 96.25+/- 21.61 kpa; the mean type-1 intrabubble pressure was 10.16 +/- 3.65 kpa. The mean bursting pressure of a type-1 BB was 66.65 +/- 18.65 kpa, while that of a type-2 BB was 14.77 +/- 2.44 kpa. The volume of air required to create a type-1 BB was 0.54 ml and the volume of a type-1 BB was consistently 0.1 ml.ConclusionsDua's layer baring DALK can withstand high intraoperative pressures compared to Descemet's membrane baring DALK. The study suggests that it could be safe to undertake procedures such as DALK-triple with a type-1 BB but not with a type-2 BB.
Journal Article