• Journal of Life Science
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• v.32 no.9
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• pp.712-720
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• 2022

The purpose of this study was to investigate the efficacy of rat corneal-derived epithelial cells as an in vitro model to evaluate the harmfulness of the cornea caused by particulate matter 2.5 (PM_2.5). To establish an experimental model for the effect of PM_2.5 on corneal epithelial cells, it was confirmed that primary cultured cells isolated from rat eyes were corneal epithelial cells through pan-cytokeratin staining. Our results showed that PM_2.5 treatment reduced cell viability of primary rat corneal epithelial (RCE) cells, which was associated with the induction of apoptosis. PM_2.5 treatment also increased the generation of reactive oxygen species due to mitochondrial dysfunction. In addition, the production of nitric oxide and inflammatory cytokines was increased in PM_2.5-treated RCE cells. Furthermore, through heatmap analysis showing various expression profiling between PM_2.5-exposed and unexposed RCE cells, we proposed five genes, including BLNK, IL-1RA, Itga2b, ABCb1a and Ptgs2, as potential targets for clinical treatment of PM-related ocular diseases. These findings indicate that the primary RCE cell line is a useful in vitro model system for the study of PM_2.5-mediated pathological mechanisms and that PM2.5-induced oxidative and inflammatory responses are key factors in PM2.5-induced ocular surface disorders.

• Journal of Veterinary Clinics
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• v.18 no.4
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• pp.418-423
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• 2001

To investigate the modulation of nerve growth factor (NGF) during corneal epithelial wound healing and the effect of topical NGF on corneal epithelial wound healing in dogs. An axial epithelial defect was created in the right eye using 6mm axial corneal mechanical debridement while the left served as an unwounded control. The tears were collected from both eyes during 1 week and the corneal epithelium was processed for the measurement of NGF at day 0 and 7. The NGF content of tears and corneal epithelium was determined by enzyme-linked immunosorbent assay. In another experiment, the animals were divided into 3 groups. The right eyes in each group were treated every six hours with 200 ug/ml of recombinant human (rh) NGF, murine NGF, or 600 ug/ml of anti-NGF blocking antibody. The left eye of each animal was treated with bovine serum albumin (BSA) to serve as controls. Wound healing was analyzed using NIH image software. Tear NGF was markedly increased in the wounded eyes, relative to tears from control eyes during the early healing period. The NGF content of the corneal epithelium was elevated in the wounded eye (p=0.024). Time to wound closure and rate of epithelial migration were not significantly different between the NGF treated or the NGF antibody treated, and the control BSA treated eyes. Corneal epithelial wounding increased NGF content only on the wounded side during the early healing period. Neither topical recombinant human or murine NGF affected corneal epithelial wound healing in the normal dog.

• Journal of Korean Ophthalmic Optics Society
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• v.13 no.4
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• pp.65-73
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• 2008

Purpose: The purpose of this study is to investigate the effect of age in the response to long-term overnight orthokeratology (OK) lens wearing. Methods: Among volunteers, ninety-five healthy subjects who had no eye diseases and could wear OK contact lens at least for 8 hours every day were divided into three groups children, youngsters and young adults. Unaided logMAR visual acuity, refractive error, apical corneal radius, corneal asphericity and central corneal thickness were measured with different period; before and after one day, one week, two weeks, one month, three months and six months of OK lens wear. Paired student t-test, ANOVA analysis and Pearson correlation were used with a critical p value of 0.05 for statistical analysis. Results: All groups showed statistically significant (p<0.001) improvement in unaided visual acuity, a trend for flattening in the apical corneal radius, decrease in central corneal thickness and less prolate after OK lens wear. The child group showed significantly rapid change (p<0.001) in visual acuity, and apical corneal radius showed that they reached the targeted refractive change earlier compared with youngster and adult groups. The visual effect of OK lens was significantly related with the change in central corneal thickness after long-term OK lens wear, especially in child and youngster group, and central corneal thickness were highly correlated with the targeted refractive change. Conclusions: Visual acuity change is statistically correlated with the central corneal thickness change, which is highly correlated with targeted refractive change in the long-term orthokeratology and younger lens wearers showed a rapid response to OK lens wear, suggesting a reduced epithelial response with increasing age. The results found this study extends our understanding and development in the long-term orthokeratology.

• Applied Microscopy
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• v.36 no.3
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• pp.227-234
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• 2006

To investigate the comparative effect of spherical and aspherical RGP lens were worn during 3 weeks on rabbit's cornea. Four white rabbits were worn right eyes with spherical lens and 4 white rabbits were worn right eyes with aspherical RGP lens. Left eyes were served as control. The rabbits were sacrificed at 3 weeks after fitting and observed morphological changes by scanning electron microscopy and also investigate proliferation rate of the corneal epithelium with RGP wearing. After spherical RGP lens wearing, the epithet layer damaged compared to aspherical lens. The superficial cell layer strip off seriously, cell size significantly changed abnormal. Both spherical and aspherical RGP lens fitting group showed so many bacteria and back surface of lens was found like a fern shape. The aspherical RGP lens original material type was some formal than spherical lens. We thought that these pattern was significantly altered with spherical lens by prohibited transmitter oxygen from atmosphere therefore the epithelium shape was changed. This suggested wearing the aspherical lens might be less physiologic than shperical lens fitting.

• Journal of Korean Ophthalmic Optics Society
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• v.12 no.3
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• pp.19-25
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• 2007

It was studied how using soft contact lens multi-purpose solution (MPS), often used for medical treatment, effects the inhibition on cell growth, and how using the MPS demages eye cells, on rabbit eye's corneal epithelium and endothelium tissue. $ReNu^{(R)}$ (Baush & Lomb, USA), Opti-free $express^{(R)}$ (Alcon, USA), Free-sol $plus^{(R)}$ (Hanamedicon, Korea) were used as MPS. After culturing Clone 1-5C-4 cell lines (Human conjunctival cell lines), cell growth inhibition rate was measured by MIT assay. By making Hematoxylin and Eosin stain specimen, the morphology was observed by optical microscope. In the In vivo experiment, 9 white rabbit eyes (18 eyes) were classified into 3 groups. The experiment group is left eyes (9 eyes) of rabbit, and MPS were dropped; however the control group, the right eyes (9 eyes), were only used a saline solution without a preservatives. After the dropping within the period, the cornea surface of rabbit endothelium tissue was observed using scanning electron microscopy (SEM).

• Journal of Korean Ophthalmic Optics Society
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• v.3 no.1
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• pp.93-101
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• 1998

The present study was conducted to investigate the structural changes in rat cornes. Sixty eyes from one-day-old uneyed rats, fourty eyes from 10-weeks-old adult rats were used. With the increase of age, the epithelial layer was thickened by the addition of new successive cellular layers. Then, the new-born rat's epithelial cells formed a pentagonal shape, and the quality of decidual cells showed a high electron-density, although the boundary between cells was distinctive. The newly produced cells showed a low electron-density so that there was the distinctive difference between light and darkness. In Bowman's layer, collagen fibrils demonstrate a regularly arranged structure along with the age. In stroma's layer, the density of keratocytes was decreased and thereby progressively flattened during the development. The collagenous layer of the adult rats was more distinctive than that of the new-born rats in a form of vertical and horizontal parallel alignment running vertically and horizontally. Descemet's membrane changed its structure significantly along with the age. It changed the shape from "banded-layer" to "non-banded layer" gradually. The thickness was also increased along with the age. Regardless of developmental stages, the endothelium is usually monolayer. During the developmental process, endothelial cells disappeared, so the density of endothelium was also decreased. The empty spaces were replaced by the expansion of adjacent cells.

• Applied Microscopy
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• v.36 no.3
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• pp.141-153
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• 2006

The purpose of this study is to find out how soft contact lens multi-purpose solution (MPS), often used for medical treatment, effects the inhibition on cell growth and research the result of using MPS, suspected to demage eye cells, on rabbit eye's corneal epithelium and endothelium tissue. In this treatise, $ReNu^{(R)}$ (Baush & Lomb, USA), Opti-free $express^{(R)}$ (Alcon, USA), Free-sol $plus^{(R)}$ (Hanamedicon, Korea) had been selected among the MPS. After culturing L929 roil line, cell growth inhibition rate was measured by MTT assay, and by making Hematoxylin and Eosin stain specimen. the morphology was observed by optical microscope. In the In vivo experiment,9 white rabbit eyes (18 eyes) were classified into 3 groups. The experimental group is left eyes (9 eyes) of rabbit, and MPS were dropped; however. the control group, the right eyes (9 eyes), were only used a saline solution without preservatives. After the dropping within the period, the cornea surface of rabbit eyes were stained by Rose bengal and observed. To figure out the changes of the corneal epithelium and endothelium tissue scanning electron microscopy (SEM) has been used. As the result, the rate of cell growth inhibition was 54%. 73% and 36%, respectively. Morphological changes represented that the shape has been changed into oval or round shape and those are not considered as a common formation of L929 cell line. When it comes to staining Rose bengal, each experiment group was stained red which is not shown in controls. The polygonal mosaic pattern of a corneal epithelium was disturbed in the picture taken by SEM; furthermore, the shape of the corneal endothelium was irregular. In conclusion, as we consider antimicrobial effect and the safety on living cells, it is necessary that we should improve concentration of preservatives and study continuously to develop a new preservatives without a toxic effect on the cornea surface.

• Applied Microscopy
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• v.28 no.4
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• pp.491-502
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• 1998

The present study was conducted to investigate the structural changes in rat cornea. Sixty eyes from one-day-old uneyed rats, fourty eyes from 4-weeks-old rats, and foully eyes from 10-weeks-old adult rats were used. With the increase of age, the epithelial layer was thickened by the addition of new successive cellular layers. Then, the new-born rat's epithelial cells formed a pentagonal shape, and the quality of decidual cells showed a high electron-density, although the boundary between cells was distinctive. The newly produced cells showed a low electron-density so that there was the distinctive difference between light and darkness. In Bowman's layer, collagen fibrils demonstrate a regularly arranged structure along with the age. In stroma's layer, the density of keratocytes was decreased and thereby Progressively flattened during the development. The collagenous layer of the adult rats was more distinctive than that of the new-born rats in a form of parallel alignment running vertically and horizontally.

• Korean Chemical Engineering Research
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• v.43 no.1
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• pp.103-109
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• 2005

Isolation and primary culture technique of rabbit oral keratinocytes, and the study for effect of environmental factors on the cell growth were carried out in T75-flask. $1.92{\pm}0.59{\times}10^6$ viable cells were isolated by trypsin enzymatic digestion method from $0.25cm^2$ biopsy of rabbit oral mucosa. Primary culture with 10 mL of K-SFM containing 50 mg/L BPE, $5.0{\mu}g/L$ EGF and 0.15 mM $Ca^{2+}$ showed confluence after 8 days and doubling time was 2.54 days. Effect of medium types, medium volume and supplement types on the cell growth was investigated after the cultured keratinocytes had been harvested from primary confluence. Serum addition showed adverse effect and the increase of serum concentration didn't have an effect on the cell growth. The increase of medium volume decreased the cell growth. The increase of calcium concentration increased the cell growth and 2.0 mM was optimum value. In conclusion, when rabbit oral keratinocytes was cultured in T75-flask, the most effective conditions was to use 10 mL of K-SFM containing 50 mg/L BPE, $5.0{\mu}g/L$ EGF and 2.0 mM $Ca^{2+}$, and doubling time was 1.32 days. This study can provide the useful informations to develop a process and design a bioreactor for the culture of keratinocytes in human body like skin and cornea, as well as mucosa.

• Analytical Science and Technology
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• v.34 no.6
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• pp.275-283
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• 2021

This study presents the development of cell-based microfluidic chips for the performance of acute eye irritation tests due to chemicals and examined some of their applications. Microfluidic chips were fabricated by photolithography and soft lithography, and they had three compartments with different areas for cell culture. Rabbit corneal epithelial cells were used for the eye irritation test. The death of cells cultured inside the chip was monitored at regular time intervals after treatment with an aqueous solution of chemicals, and the cell death rate constants were calculated based on the viability curve. The performance of the microfluidic chip was verified by examining the effects of cell-cell junctions, cell-substrate adhesion, and initial cell numbers compared to cell death rates. Eye irritation tests were performed at various concentrations of an aqueous solution of sodium dodecyl sulfate (SDS), a standard substance for the eye irritant test. The cells were exposed to the SDS aqueous solution for 300 s, and the resulting eye irritation was assessed by cell viability. Finally, the equation for calculating the toxicity score (TS) was derived based on the weighting factor for each compartment in the chip. The cell-based microfluidic chip developed in this study may be used for eye irritation tests from chemicals used in cosmetics and pharmaceuticals.