Mean central corneal thickness and corneal power measurementsin pigmented and white rabbits using Visante optical coherencetomography and ATLAS corneal topography

Xiaogang Wang,* Jing Dong† and Qiang Wu**Department of Ophthalmology, Affiliated Sixth People’s Hospital, Shanghai Jiao Tong University, No. 600 Yishan Road, Shanghai 200233, China; and

†Department of Ophthalmology, The First Hospital of Shanxi Medical University, Taiyuan, Shanxi 030001, China

Address communications to:

Q. Wu

Tel.: 13918261866(Cell)

Fax: 86-21-64369181-58367

e-mail: movie6521@gmail.com

AbstractObjective To document central corneal thickness (CCT) and power measurements

in pigmented and white rabbits using Visante anterior segment optical coherencetomography (AS-OCT) and the Humphrey ATLAS Corneal Topography System(HACTS).

Animal studied Fourteen female rabbits (seven pigmented and seven white rabbits)were involved in this study.

Procedure Twenty-eight eyes underwent AS-OCT and HACTS examination. Centralcorneal thickness, corneal power in the steepest and flattest meridians, astigmatism,

pupil diameter (PD), and white-to-white (WTW) were calculated.Results The CCT was 390 � 14.2 and 373 � 7.2 lm for pigmented and white rabbits,

respectively. The CCT values showed statistical difference between the two breeds(P = 0.017). The corneal power in the steepest meridian was 44.6 � 1.9 Diopter (D)

in pigmented rabbits and 47.8 � 1.5 D in white rabbits. The corneal power in theflattest meridian was 44.0 � 2.1 D in pigmented rabbits and 47.4 � 1.5 D in whiterabbits. The astigmatism, PD, and WTW values were 0.6 � 0.22 D, 7.9 � 0.9, and

14.5 � 0.1 mm in pigmented rabbits, respectively. The corresponding values in whiterabbits were 0.4 � 0.23 D, 7.5 � 0.5, and 13.5 � 0.2 mm. There was a statistically

significant difference in corneal power in the steepest and flattest meridians and forWTW. No such difference was observed for astigmatism or PD between them.

Conclusions Differences exist in the CCT, corneal power, and WTW between5-month-old pigmented and white rabbits. Such difference should be considered

when designing cornea-related experiments in rabbits.

Key Words: cornea, corneal topographer, optical coherence tomography, pigmentedrabbit, white rabbit

INTRODUCTION

Optical coherence tomography (OCT) was first intro-duced in the 1990s and is a safe and noninvasive methodthat provides high-resolution cross-sectional images of theanterior and posterior ocular tissues.1 Although posteriorsegment OCT is more popular than anterior segmentoptical coherence tomography (AS-OCT) in the clinic, thelatter can provide detailed information regarding anteriortissues, such as the cornea, the anterior chamber angle,the iris, and the lens. Such information can be veryuseful for refractive surgery and glaucoma diagnosis.2,3

The Humphrey ATLAS Corneal Topography System

(HACTS), which can provide corneal refractive para-meters, is helpful for certain clinical applications, such askeratoconus diagnosis and refractive surgery follow-up.4,5

As an important animal research species, rabbits playimportant roles in some kinds of ocular studies, especiallyin the refractive surgery–related experiments.6–9 However,normal data for rabbit ocular tissue are few, especially fordifferent breeds.10,11 Therefore, the aims of this studywere to investigate normal anterior ocular segment param-eters, such as 2-mm-diameter central corneal thickness(CCT), corneal power, pupil diameter (PD), and white-to-white (WTW, corneal diameter), measured with AS-OCTand HACTS in pigmented and white rabbits in vivo.

© 2013 American College of Veterinary Ophthalmologists

Veterinary Ophthalmology (2014) 17, 2, 87–90 DOI:10.1111/vop.12039

MATERIALS AND METHODS

MaterialsAll of the experiments were conducted in accordance withthe guidelines for animal experimentation of the AffiliatedSixth People’s Hospital Shanghai Jiao Tong University,and the Animals in Ophthalmic and Vision Research(ARVO) Statement. Fourteen healthy rabbits (female,seven pigmented and seven New Zealand white rabbits)were provided by the Animal Center of Affiliated SixthPeople’s Hospital, Shanghai Jiao Tong University. At thetime of study, the animals were 5 months old and weighedbetween 2.5 and 3.1 kg. The rabbits underwent a com-plete ophthalmologic examination to exclude any ocularabnormalities. Both of the eyes of 14 rabbits were sub-jected to AS-OCT corneal thickness measurements andHACTS corneal power measurements. The rabbits wereimmobilized in restraining boxes for all of the examina-tions, without general or topical anesthesia. The eyelidswere held open manually by an assistant.

Optical coherence tomography imagingThe Visante OCT-1000 (Carl Zeiss Meditec, Dublin, CA,USA) instrument is a noninvasive time-domain device witha 1310-nm superluminescent diode source based on lowcoherence interferometry. This system can provide high-quality cross-sectional images of the anterior segment withan axial resolution of approximately 18 lm and a scanningspeed of 2048 A-scans per second. The scan speed is lim-ited by the back-and-forth mechanical movement of a ref-erence mirror over a range of several millimeters.1 Apachymetric scan model with eight radial scan lines wasused to construct the corneal pachymetry map to measurethe average corneal thickness within the central 10-mmdiameter. The CCT values were recorded and were ana-lyzed. All the images in this study were captured by anexperienced and trained technician, and the images wereanalyzed automatically with the software (version 3.0.1.8)in Visante OCT-1000. During each scan, the techniciancaptured each cross-sectional corneal image with the lightbeam at the midpoint of the cornea to ensure a centralizedscan location. The tear film was included in corneal thick-ness calculations. Therefore, the assistant was asked toblink the rabbit’s eyes manually before measurements tominimize tear evaporation.

The Humphrey ATLAS Corneal Topographer-9000(Carl Zeiss Meditec, Dublin, California, USA) automati-cally provides mean keratometry values along the steepestand flattest meridians at the 3-mm zone of the central cor-nea with astigmatism, as well as PD and WTW measure-ments using the instrument’s software (version 3.0.0.39).The cornea is not a simple spherical surface with a singlecurvature but is rather a complex physiologic interfaceof various curvatures. Thus, the two principal meridians(i.e., the flattest and the steepest) are always used as thekeratometry values in the clinic and for research.12

All examinations in this study were performed by thesame operator under natural light conditions. Due to thelack of fixation, the assistant was asked to cooperate withthe operator to stabilize the eye in the proper position forimage capture.

Statistical analysisThe data, which included measurements of each eye, wereregistered in tables. One eye of each rabbit was randomlychosen for the final analysis. The statistical analyses wereperformed using commercial software (SPSS ver. 13.0;SPSS Inc., Chicago, IL, USA). The Mann–Whitney testwas used to analyze the difference between pigmented andwhite rabbits for each of the measured parameters. A Pvalue of <0.05 was considered to be significant.

RESULTS

The CCTs were 390 � 14.2 and 373 � 7.2 lm for thepigmented and white rabbits. The corneal power in thesteepest meridian was 44.6 � 1.9 D in pigmented rabbitsand 47.8 � 1.5 D in white rabbits. The corneal power inthe flattest meridian was 44.0 � 2.1 D in pigmented and47.4 � 1.5 D in white rabbits. The astigmatism, PD,and WTW values were 0.6 � 0.22 D, 7.9 � 0.9, and14.5 � 0.1 mm in pigmented rabbits. The correspondingvalues in white rabbits were 0.4 � 0.23 D, 7.5 � 0.5, and13.5 � 0.2 mm (Table 1). There were statistical differ-ences in CCT (P = 0.017), corneal power in the steepestand flattest meridians (P = 0.004; P = 0.005), and WTW(P < 0.001). No such differences were observed for astig-matism values (P = 0.213) or PD (P = 0.247).

DISCUSSION

Several studies have demonstrated that AS-OCT andHACTS have excellent intra-operator repeatability andreproducibility in measurements of anterior segmentparameters.12–15 As the rabbit cornea is much easier tomanipulate than the mouse cornea and as the rabbits are

Table 1. Normal anterior segment values (M � SD) of pigmented

and white rabbits

Pigmentedrabbit (n = 7)

White rabbit(n = 7) P*

CCT (lm) 390 � 14.2 373 � 7.2 0.017†

Steepest corneal power (D) 44.6 � 1.9 47.8 � 1.5 0.004†

Flattest corneal power (D) 44.0 � 2.1 47.4 � 1.5 0.005†

Astigmatism (D) 0.6 � 0.22 0.4 � 0.23 0.213PD (mm) 7.9 � 0.9 7.5 � 0.5 0.247WTW (mm) 14.5 � 0.1 13.5 � 0.2 0.000†

M = mean; SD = standard deviation; CCT = central corneal thick-ness; D = diopter; PD = pupil diameter; WTW = white-to-whitedistance.*Mann–Whitney test analysis.†P value, with significance at 0.05.

© 2013 American College of Veterinary Ophthalmologists, Veterinary Ophthalmology, 17, 87–90

88 wang , d ong and wu

much less expensive than monkeys, the use of rabbits ofvarious breeds is common in corneal research.16,17 There-fore, a database of rabbit eye characteristics is very useful.Compared with contact ultrasound techniques, OCT is anoncontact, noninvasive and reliable method of examina-tion.18 No iatrogenic injuries were observed in the presentstudy as a result of using this equipment. General anesthe-sia was not used to avoid reducing corneal metabolismand to limit any potential influence of the anesthetic onnormal corneal thickness measurements. Topical anesthe-sia was not used given that such anesthetics have beenreported to increase CCT when measured using ultrasonicpachymetry.19

In the present study, the pigmented rabbit CCT wasthicker than Emre’s reported values of 358.1 � 26.5 lmOD and 372.1 � 18.8 lm OS for 31 pigmented rabbits,which were measured using ultrasonic pachymetry.19 TheCCTs of white rabbits in the present study were less thanthe 407 � 20 lm that was reported in Chan’s study of 12adult New Zealand Albino rabbit eyes; this previous resultwas also based on ultrasonic pachymetric analyses.20 Thepossible reasons for these differences are as follows: (i)compared with AS-OCT, ultrasonic pachymetry may pro-vide a less precise determination of the central cornealocation due to the lack of any reference point and mayrely more on the ultrasound operator’s experience, (ii) dif-ferent breeds of rabbits may have different values for thesemetrics, (iii) the OCT system can provide an objectiveand stable CCT value through software analysis and is notsubjective or variable with respect to determining the cen-tral cornea location, as is the case for measurements usingultrasonic pachymetry (see reason 1), (iv) any topical anes-thesia that is used for contact measurements by ultrasonicpachymetry may influence CCT values. One possible rea-son for the different values that are reported in the presentstudy compared with Emre’s results may be the differencein the detection methods that were used (i.e., OCT andultrasonic pachymetry). Explanations for the clear differ-ence between the values reported in the present study andthose of Chan may be (i) the different rabbit breedsused, (ii) the ages of the rabbits, (iii) the technologyemployed. The corneal thickness measured by AS-OCTincludes the tear film thickness, which is approximately6.5–11.8 lm.21,22 That is to say, the real corneal thicknessvalues will be less than the auto-calculated values byAS-OCT.

Pigmented and white rabbit CCT values were clearlythinner than those of human central corneal thicknesses.23

The degree of cornea ablation should therefore be care-fully considered to avoid corneal ectasia when performingcornea refractive surgery or femtosecond laser-assistedposterior lamellar keratoplasty in rabbits.24

The PD values in both types of rabbit were similar tothose reported by Liu et al. in a study of seven adult maleNew Zealand albino rabbits. This study reported values ofapproximately 8.2 � 0.3 mm in the light phase, which

were clearly lower than the values that were observed(10.8 � 0.1 mm) in the dark phase.25 The average WTWvalues of white rabbits in the present study were compara-ble to the results of Bozkir et al., who examined approxi-mately 40 eyes of New Zealand White rabbits. Theseauthors reported corneal diameters of approximately13.41 � 0.34 mm in the horizontal meridian and13.02 � 0.30 mm in the vertical meridian; both of thesevalues are less than the values for pigmented rabbit thatwere observed in the present study.26 However, all ofthese previously reported values were greater than thevalues that have been observed for humans (averages:horizontal 11.75 mm, vertical 10.55 mm).

The precise measurement of corneal power is veryimportant for refractive surgery to correct corneal astig-matism. There are several types of equipment that can beused for such a measurement, which may return differentdiameters of the region measured, such as the Galilei dualScheimpflug system, the Pentcam system, the IOLMaster,the Orbscan system, and manual keratometry. Accordingto the formula for calculating corneal power from cornealcurvature that was described by Olsen, the corneal powerreported in the present study is similar to the valuesdescribed by Bozkir et al.26,27

There are several limitations to the present study: (i) weonly tested normal 5-month-old rabbit corneas, (ii) weused only one operator; and (iii) we did not assess the pos-terior corneal curvature. The precise measurement of theposterior corneal power is vital for corneas undergoingexcimer laser ablation given that the ablation changes theassumed ratio of the anterior-to-posterior curvatures.

In summary, we compared the difference between nor-mal pigmented and white rabbits with respect to certainanterior segment indexes. Most of the indexes that shouldbe considered in animal studies were significantly differ-ent, even in rabbits of the same age. We look forwardto further studies of anterior segment characteristics ofdifferent breeds of rabbits of various ages.

ACKNOWLEDGMENTS

We have no acknowledgements, meeting presentations,competing interests or funding to report.

REFERENCES

1. Huang D, Swanson EA, Lin CP et al. Optical coherence

tomography. Science 1991; 254: 1178–1181.2. Ramos JL, Li Y, Huang D. Clinical and research applications of

anterior segment optical coherence tomography – a review.

Clinical and Experimental Ophthalmology 2009; 37: 81–89.

3. Sakata LM, Deleon-Ortega J, Sakata V et al. Optical coherence

tomography of the retina and optic nerve – a review. Clinical and

Experimental Ophthalmology 2009; 37: 90–99.4. Barbero S, Marcos S, Merayo-Lloves J et al. Validation of the

estimation of corneal aberrations from videokeratography in

keratoconus. Journal of Refractive Surgery 2002; 18: 263–270.

© 2013 American College of Veterinary Ophthalmologists, Veterinary Ophthalmology, 17, 87–90

co rn e a l th i c kn e s s a nd p owe r o f p i gm ent e d and wh i t e r a b b i t s 89

5. Marcos S, Barbero S, Llorente L et al. Optical response to

LASIK surgery for myopia from total and corneal aberration

measurements. Investigative Ophthalmology and Visual Science 2001;42: 3349–3356.

6. Achari Y, Reno CR, Tsao H et al. Influence of timing (pre-

puberty or skeletal maturity) of ovariohysterectomy on mRNA

levels in corneal tissues of female rabbits. Molecular Vision 2008;

14: 443–455.

7. Almeida GC Jr, Faria e Souza SJ. Effect of topical dorzolamide

on rabbit central corneal thickness. Brazilian Journal of Medical

and Biological Research 2006; 39: 277–281.

8. Busin M, Yau CW, Yamaguchi T et al. The effect of collagen

cross-linkage inhibitors on rabbit corneas after radial keratotomy.

Investigative Ophthalmology and Visual Science 1986; 27: 1001–1005.9. Munnerlyn CR, Koons SJ, Marshall J. Photorefractive kera-

tectomy: a technique for laser refractive surgery. Journal ofCataract and Refractive Surgery 1988; 14: 46–52.

10. Riau AK, Tan NY, Angunawela RI et al. Reproducibility and

age-related changes of ocular parametric measurements in

rabbits. BMC Veterinary Research 2012; 8: 138.11. Wang X, Wu Q. Normal corneal thickness measurements in

pigmented rabbits using spectral-domain anterior segment optical

coherence tomography. Veterinary Ophthalmology 2013; 16: 130–

134.

12. Kobashi H, Kamiya K, Igarashi A et al. Comparison of corneal

power, corneal astigmatism, and axis location in normal eyes

obtained from an autokeratometer and a corneal topographer.

Journal of Cataract and Refractive Surgery 2012; 38: 648–654.13. Fukuda S, Kawana K, Yasuno Y et al. Repeatability and

reproducibility of anterior ocular biometric measurements with

2-dimensional and 3-dimensional optical coherence tomography.

Journal of Cataract and Refractive Surgery 2010; 36: 1867–1873.14. Mohamed S, Lee GK, Rao SK et al. Repeatability and

reproducibility of pachymetric mapping with Visante anterior

segment-optical coherence tomography. Investigative Ophthal-

mology and Visual Science 2007; 48: 5499–5504.15. Zhang Q, Jin W, Wang Q. Repeatability, reproducibility, and

agreement of central anterior chamber depth measurements in

pseudophakic and phakic eyes: optical coherence tomography

versus ultrasound biomicroscopy. Journal of Cataract andRefractive Surgery 2010; 36: 941–946.

16. Liu H, Zhu W, Jiang AC et al. Femtosecond laser lenticule

transplantation in rabbit cornea: experimental study. Journal of

Refractive Surgery 2012; 28: 907–911.17. Park CK, Kim JH. Comparison of wound healing after

photorefractive keratectomy and laser in situ keratomileusis in

rabbits. Journal of Cataract and Refractive Surgery 1999; 25:

842–850.18. Li EY, Mohamed S, Leung CK et al. Agreement among 3

methods to measure corneal thickness: ultrasound pachymetry,

Orbscan II, and Visante anterior segment optical coherence

tomography. Ophthalmology 2007; 114: 1842–1847.

19. Emre S, Akkin C, Afrashi F et al. Effect of corneal wetting

solutions on corneal thickness during ophthalmic surgery. Journal

of Cataract and Refractive Surgery 2002; 28: 149–151.20. Chan T, Payor S, Holden BA. Corneal thickness profiles in

rabbits using an ultrasonic pachometer. Investigative Ophthalmologyand Visual Science 1983; 24: 1408–1410.

21. Azartash K, Shy CJ, Flynn K et al. Non-invasive in vivo

measurement of the tear film using spatial autocorrelation in a

live mammal model. Biomedical Optics Express 2010; 1: 1127–1137.22. King-Smith PE, Fink BA, Hill RM et al. The thickness of the

tear film. Current Eye Research 2004; 29: 357–368.23. Bechmann M, Thiel MJ, Neubauer AS et al. Central corneal

thickness measurement with a retinal optical coherence

tomography device versus standard ultrasonic pachymetry. Cornea

2001; 20: 50–54.24. Abdelkader A, Esquenazi S, Shihadeh W et al. Healing process at

the flap edge in its influence in the development of corneal

ectasia after LASIK. Current Eye Research 2006; 31: 903–908.

25. Liu JH, Gallar J, Loving RT. Endogenous circadian rhythm of

basal pupil size in rabbits. Investigative Ophthalmology and Visual

Science 1996; 37: 2345–2349.26. Bozkir G, Bozkir M, Dogan H et al. Measurements of axial

length and radius of corneal curvature in the rabbit eye. ActaMedica Okayama 1997; 51: 9–11.

27. Olsen T. On the calculation of power from curvature of the

cornea. British Journal of Ophthalmology 1986; 70: 152–154.

© 2013 American College of Veterinary Ophthalmologists, Veterinary Ophthalmology, 17, 87–90

90 wang , d ong and wu