Light Damage in Detached Retina

Light Damage in Detached Retina

Light Damage in Detached Retina John D. Zilis, M.D., and Robert Machemer, M.D. We i n j e c t e d h o m o l o g o u s fibroblasts o v e r t h e r e ...

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Light Damage in Detached Retina

John D. Zilis, M.D., and Robert Machemer, M.D.

We i n j e c t e d h o m o l o g o u s fibroblasts o v e r t h e r e t i n a l v a s c u l a r w i n g i n five r a b b i t s t o i n d u c e a fractional retinal detachment. Eleven days later, a focal area of d e t a c h m e n t was exposed to 30 m i n u t e s of visible light by a n i n t r a o c u l a r fiberoptic probe. Histologic damage to the detached retina exposed to light was demon­ strated. T h e o u t e r retina was affected m o s t severely.

L I G H T D A M A G E IN A T T A C H E D R E T I N A h a s

been

demonstrated with various ophthalmic light sources, including the intraocular fiberoptic p r o b e . T h i s damage primarily involves the o u t e r r e t i n a a n d p i g m e n t e p i t h e l i u m , ^ * b u t it c a n a l s o affect t h e i n n e r r e t i n a i f e x p o s u r e is i n t e n s e a n d o f l o n g duration.* T h e e x a c t m e c h a ­ nisms of such retinal damage and the role of the melanin-containing retinal pigment epitheli­ u m " r e m a i n u n k n o w n . We e v a l u a t e d l i g h t d a m a g e in e x p e r i m e n t a l l y d e t a c h e d r a b b i t r e t i ­ n a that h a d b e e n s e p a r a t e d from t h e r e t i n a l pigment epithelium.

Material and Methods Five p i g m e n t e d r a b b i t s w e i g h i n g b e t w e e n 2 . 5 a n d 3 . 5 kg w e r e h o u s e d u n d e r 5 0 f o o t - c a n d l e (ft-c) i l l u m i n a t i o n on a 1 2 - h o u r , l i g h t - d a r k c y ­ c l e . At l e a s t t w o w e e k s b e f o r e t h e e x p e r i m e n t s , the rabbits were given anesthesia with an intra­ muscular injection of k e t a m i n e h y d r o c h l o r i d e (30 m g / k g of body weight) and xylazine hydro-

Accepted for publication Sept. 12, 1990. From the Devers Eye Institute, Portland, Oregon (Dr. Zilis), and Department of Ophthalmology, Duke Univer­ sity, Durham, North Carolina (Dr. Machemer). This study was supported by National Eye Institute grant F32 EY06046-01, the Helena Rubinstein Foundation, New York, New York, and Research to Prevent Blindness, Inc. Reprint requests to Robert Machemer, M.D., Duke University Eye Center, Box 3802, Durham, NC 27710.

c h l o r i d e (5 m g / k g o f b o d y w e i g h t ) . P u p i l s w e r e d i l a t e d b y a 1:1 m i x t u r e o f 0 . 2 5 % t r o p i c a m i d e and 5 % phenylephrine hydrochloride. The pe­ r i p h e r a l r e t i n a w a s t r e a t e d for 3 6 0 d e g r e e s at a distance o f 4 . 5 m m from the c o r n e o s c l e r a l limbus with transscleral cryopexy, under direct v i s u a l o b s e r v a t i o n , in p r e p a r a t i o n for future sclerotomy sites. At l e a s t t w o w e e k s l a t e r , t h e v i t r e o u s c a v i t i e s were injected with 2 5 0 , 0 0 0 h o m o l o g o u s tissuec u l t u r e d fibroblasts. F i b r o b l a s t s w e r e o f a s i n g l e l i n e a n d h a d b e e n h a r v e s t e d from r a b b i t r u m p dermis and tissue cultured in D u l b e c c o s modi­ fied E a g l e m e d i u m w i t h 1 0 % f e t a l b o v i n e s e r ­ um, antibiotics, and a n t i m y c o t i c s , a n d main­ t a i n e d at 3 7 C in a n i n c u b a t o r w i t h a n a t m o s p h e r e o f 5 % c a r b o n d i o x i d e in a i r . T h e s e w e r e h a r v e s t e d for i n j e c t i o n in a m a n n e r p r e v i ­ ously described.'" At t h e t i m e o f i n j e c t i o n , t h e a n i m a l s w e r e again given anesthesia, and the pupils were dilated. Under direct visualization by the indi­ r e c t o p h t h a l m o s c o p e , t h e c e l l s , s u s p e n d e d in 0.1 m l o f p h o s p h a t e - b u f f e r e d s a l i n e , w e r e i n ­ jected slowly through the previously cryotreated peripheral retina into the vitreous body over the vascular wing o f the retina. T h e ani­ m a l s w e r e p l a c e d on t h e i r s i d e s for o n e h o u r t o allow settling of cells to occur on the vascular­ ized retina. Indirect o p h t h a l m o s c o p y was per­ f o r m e d at D a y 1 a n d D a y 7 a f t e r i n j e c t i o n t o monitor progression of the proliferative re­ sponse and resulting fractional retinal detach­ ment. E l e v e n days after fibroblast i n j e c t i o n , t h e a n i ­ mals were again given anesthesia, and the pu­ pils of the injected eyes were dilated. R e t r o ­ bulbar injections were not given. Rectal temperatures were monitored, and warming b l a n k e t s w e r e u s e d to m a i n t a i n b o d y t e m p e r a ­ t u r e as n e e d e d . S c l e r o t o m i e s w e r e m a d e 3 . 0 m m p o s t e r i o r to the corneoscleral limbus through previously cryotreated peripheral retina, and cannulas w e r e p l a c e d t o m a i n t a i n an o p e n i n g . T w o a d d i ­ t i o n a l s c l e r o t o m i e s w e r e p l a c e d for i n f u s i o n and outflow. Midvitreous temperature was ob-

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tained before beginning infusion and moni­ tored during i n f u s i o n b y a t e m p e r a t u r e p r o b e . L a c t a t e d R i n g e r ' s infusion fluid w a s w a r m e d to 3 9 . 5 ± 0 . 5 C in a w a t e r b a t h that w a s e l e v a t e d 2 6 i n c h e s a b o v e the l e v e l o f t h e e y e . I n t r a o c u l a r p r e s s u r e was m a i n t a i n e d b e t w e e n 14 a n d 2 1 m m Hg, a n d i n t r a o c u l a r t e m p e r a t u r e w a s m a i n ­ t a i n e d at 3 8 . 3 ± 0 . 3 C b y c o n s t a n t p e r f u s i o n o f lactated Ringer's solution. A g l a s s fiberoptic p r o b e a t t a c h e d to a l i g h t source with a 1 5 0 - W , 2 1 - V tungsten-halogen projector bulb had a measured light output of 4 0 . 0 ± 0 . 8 ft-c. To confirm c o n s t a n t l i g h t out­ put, t h e l i g h t m e a s u r e m e n t w a s p e r f o r m e d b e ­ fore a n d after t h e e x p e r i m e n t b y a r a d i o m e t e r / photometer with integrating sphere. T h e s c l e r o t o m y c r e a t e d for i n t r o d u c t i o n o f the fiberoptic p r o b e w a s m a d e i n f e r i o r l y . T h e p r o b e was a d v a n c e d s l o w l y b y a m i c r o m a n i p u ­ lator to a d i s t a n c e o f 1.0 m m from t h e d e t a c h e d r e t i n a , w h i c h c r e a t e d an a p p r o x i m a t e c i r c l e o f light m e a s u r i n g 2 . 0 m m in its w i d e s t d i a m e t e r as m e a s u r e d b y t h e o c u l a r m i c r o m e t e r of t h e surgical m i c r o s c o p e . T h e a r e a o f t h e s m o o t h l y d e t a c h e d r e t i n a e x p o s e d to l i g h t w a s c e n t e r e d 1.5 m m b e l o w t h e e d g e o f t h e m y e l i n a t e d n e r v e layer a n d 1 disk d i a m e t e r t e m p o r a l to t h e disk in a v a s c u l a r r e t i n a . T h e c a l c u l a t e d r e t i n a l irra­ diance was 0 . 3 4 W / c m ^ which correlated well w i t h p r e v i o u s in v i t r o m e a s u r e m e n t s m a d e b y a s i m i l a r e x p e r i m e n t a l set-up."* L i g h t e x p o s u r e w a s m a i n t a i n e d for 3 0 m i n u t e s . T h e p o s i t i o n o f the fiberoptic p r o b e w a s c h e c k e d f r e q u e n t l y during the procedure. After 3 0 m i n u t e s , the i n s t r u m e n t s a n d c a n n u ­ las w e r e r e m o v e d a n d the s c l e r o t o m i e s w e r e closed with 7-0 Vicryl sutures. The conjuncti­ v a e w e r e c l o s e d , a n d 5 m g of g e n t a m i c i n sulfate w a s i n j e c t e d s u b c o n j u n c t i v a l l y . A t r o p i n e 1% o i n t m e n t w a s p l a c e d on t h e c o r n e a s . P o s t o p e r a t i v e l y , the a n i m a l s w e r e e x a m i n e d daily by t h e i n d i r e c t o p h t h a l m o s c o p e . O n t h e third p o s t o p e r a t i v e day, the a n i m a l s w e r e k i l l e d , a n d the e y e s w e r e i n j e c t e d w i t h 3 % g l u t a r a l d e h y d e in 0.1 m o l / l of c a c o d y l a t e buff­ er i m m e d i a t e l y b e f o r e e n u c l e a t i o n . A d d i t i o n a l glutaraldehyde was injected through 3 . 0 - m m radial i n c i s i o n s b i s e c t i n g the i n f e r i o r c o r n e o ­ scleral limbus, and the eyes were refrigerated a n d fixed for 4 8 h o u r s . P o s t e r i o r eye c u p s w e r e p r e p a r e d , a n d 3 . 0 x 3 . 0 - m m s e c t i o n s o f r e t i n a c e n t e r e d in t h e a r e a of l i g h t e x p o s u r e w e r e r e m o v e d . A d j a c e n t 3 . 0 x 3 . 0 - m m a r e a s of d e t a c h e d r e t i n a w e r e a l s o s e c ­ tioned and examined. T h e s p e c i m e n s w e r e p r o c e s s e d in 2 % buff­

ered osmium tetroxide through graded alcohol a n d e m b e d d e d in e p o x y r e s i n p l a s t i c . S e c t i o n s w e r e s t a i n e d w i t h m e t h y l e n e b l u e a n d a z u r e II for l i g h t m i c r o s c o p y , a n d t h i n s e c t i o n s w e r e prepared and stained with uranyl acetate and l e a d c i t r a t e for t r a n s m i s s i o n e l e c t r o n m i c r o ­ scopy.

Results Tractional retinal detachments developed w i t h i n s e v e n days o f t h e fibroblast i n j e c t i o n in all r a b b i t s . T h e d e t a c h m e n t w a s h i g h e s t in t h e a r e a o f t h e v a s c u l a r m e d u l l a r y w i n g from w h i c h a p r o l i f e r a t i v e fibroblast s t r a n d e x t e n d e d to t h e a r e a of t h e i n j e c t i o n site ( F i g . 1 ) . T h e a v a s c u l a r retina b e l o w the wing was elevated with a defined i n f e r i o r d e m a r c a t i o n l i n e s e p a r a t i n g t h e r e m a i n i n g a t t a c h e d r e t i n a from t h e g r a y white detached retina. Subretinal white precip­ i t a t e s w e r e s o m e t i m e s n o t e d in t h e d e p e n d e n t portion of the d e t a c h m e n t . Rabbits that developed rhegmatogenous, bullous detach­ ments were excluded. A total of five p i g m e n t e d r a b b i t s fulfilled t h e experimental criteria. Daily postoperative indi­ rect o p h t h a l m o s c o p i c examinations were per­ f o r m e d . O n t h e first p o s t o p e r a t i v e day, all e y e s d i s p l a y e d s i g n i f i c a n t p o s t e r i o r i n t r a o c u l a r in-

Fig. 1 (Zilis and Machemer). Tractional retinal detachment of vascular medullary wings in a rabbit. Note proliferative fibroblast strand superiorly.

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flammation that o b s c u r e d r e t i n a l d e t a i l . P r o ­ gressive clearing, however, occurred by Day 3, which allowed observation of the area of light exposure. Focal retinal edema was not found, although slight retinal thinning was suspected in two a n i m a l s . H i s t o l o g i c a l l y , t h e e y e s of all five a n i m a l s h a d m u c h m o r e s e v e r e d a m a g e in t h e a r e a o f l i g h t e x p o s u r e t h a n in o t h e r a r e a s o f r e t i n a l d e t a c h ­ m e n t . In t h e a r e a o f r e t i n a l d e t a c h m e n t n o t e x p o s e d to l i g h t , p a t c h y l o s s o f p h o t o r e c e p t o r o u t e r s e g m e n t s a n d o c c a s i o n a l m a c r o p h a g e in­ filtration w e r e n o t e d . G e n e r a l l y , c e l l u l a r o r g a ­ n i z a t i o n o f r e t i n a l layers w a s w e l l m a i n t a i n e d (Fig. 2 ) . C o n v e r s e l y , t h e a r e a s of d e t a c h e d r e t i n a e x ­ p o s e d to light d i s p l a y e d e x t r e m e d e s t r u c t i o n (Fig. 3 ) . T h e p h o t o r e c e p t o r e l e m e n t s w e r e v i r ­ tually e l i m i n a t e d w i t h o n l y s m a l l d e g e n e r a t e d inner segments remaining. Macrophages were found a l o n g t h e o u t e r l i m i t i n g m e m b r a n e . T h e o u t e r n u c l e a r layer d e m o n s t r a t e d n u c l e a r d r o p ­

Fig. 2 (Zilis and Machemer). Photomicrograph of detached retina without light exposure ( x 4 0 0 ) .

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o u t a n d p y k n o s i s . T h e i n n e r r e t i n a l layers w e r e relatively well preserved, although small cystic spaces were observed.

Discussion R e t i n a l d a m a g e c a u s e d b y v i s i b l e l i g h t is k n o w n to o c c u r w h e n an i n t r a o c u l a r fiberoptic l i g h t s o u r c e is used.^'' In a t t a c h e d r e t i n a , h i s t o ­ logic damage has been demonstrated primarily in t h e o u t e r r e t i n a a n d p i g m e n t epithelium.^* Noell and a s s o c i a t e s ' speculated that the dam­ a g e is c a u s e d b y p h o t o c h e m i c a l e v e n t s t h a t are s e n s i t i v e t o c h a n g e s in t e m p e r a t u r e . The melanin-containing retinal pigment epi­ thelium demonstrated damage when exposed t o v i s i b l e l i g h t , a l t h o u g h i t s r o l e in m e d i a t i n g

Fig. 3 (Zilis and Machemer). Photomicrograph of detached retina with 30 minutes of light exposure. Note massive destruction of outer retina and loss of photoreceptors ( x 4 0 0 ) .

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d a m a g e to t h e o v e r l y i n g r e t i n a r e m a i n s u n ­ known. Ham and a s s o c i a t e s " originally sug­ g e s t e d that t h e site of injury in t h e r h e s u s m o n k e y may b e t h e m e l a n i n g r a n u l e s o f t h e m e l a n o s o m e s o f the r e t i n a l p i g m e n t e p i t h e l i ­ u m . D u r i n g the first 4 8 h o u r s after l i g h t e x p o ­ sure, d a m a g e a p p e a r e d c o n f i n e d to t h i s layer, whereas overlying photoreceptor outer seg­ ments remained intact. W o r k i n g w i t h t h e rat, h o w e v e r , R a p p a n d Williams* s h o w e d a l m o s t e q u a l r e t i n a l s e n s i t i v ­ ity to light d a m a g e in b o t h p i g m e n t e d a n d albino animals. Hoeppler and associates' found essentially no difference w h e n m o r p h o l o g i c d a m a g e w a s c o m p a r e d in p i g m e n t e d a n d a l b i n o r a b b i t s , w h i c h l e d t h e m to p o s t u l a t e t h a t m e l a ­ nin may n o t b e c r i t i c a l l y i n v o l v e d in l i g h t d a m ­ age to t h e r e t i n a . We d e s i g n e d t h i s s t u d y to a s s e s s l i g h t d a m a g e in r e t i n a that w a s d e t a c h e d from t h e r e t i n a l p i g m e n t e p i t h e l i u m . To m a x i m i z e l i g h t d a m ­ age, a relatively long exposure (30 minutes) of v i s i b l e light w a s p l a c e d on a s m a l l a r e a ( 2 . 0 - m m d i a m e t e r ) o f d e t a c h e d , a v a s c u l a r r a b b i t r e t i n a at b o d y t e m p e r a t u r e . T h i s r e s u l t e d in s e v e r e h i s ­ t o l o g i c d a m a g e to t h e r e t i n a (Fig. 3 ) , w h i c h w a s much more profound than the usual outer reti­ nal d e g e n e r a t i o n e x p e c t e d in d e t a c h e d r e t i ­ na.'^'^ I n d e e d , t h e d a m a g e w a s d r a m a t i c w h e n c o m p a r e d to a d j a c e n t a r e a s o f d e t a c h e d r e t i n a n o t e x p o s e d to t h e fiberoptic l i g h t (Fig. 2 ) . In t h e l i g h t - e x p o s e d d e t a c h e d r e t i n a , d a m a g e w a s m o s t a p p a r e n t in t h e o u t e r r e t i n a w i t h resulting destruction of photoreceptor inner and outer segments and their nuclei. T h e same p a t t e r n o f d a m a g e w a s n o t e d in l i g h t e x p o s u r e to a t t a c h e d retina.'*' In a t t a c h e d r e t i n a , t h e o u t e r r e t i n a is e x t r e m e l y a c t i v e , m e t a b o l i c a l l y , a n d w e l l o x y g e n a t e d from t h e u n d e r l y i n g c h o ­ roid. B e c a u s e o f t h i s , o n e p r o p o s e d m e c h a n i s m o f l i g h t d a m a g e to t h e r e t i n a i n v o l v e s p h o t o o x i dative a n d p h o t o d y n a m i c r e a c t i o n s i n v o l v i n g the retinal pigment epithelium with generation of d e s t r u c t i v e o x y g e n - d e r i v e d free r a d i c a l s . ' " Because we found a similar pattern of damage in the o u t e r d e t a c h e d r e t i n a t h a t w a s d e v o i d o f m e l a n i n a n d p r e s u m e d to b e i s c h e m i c , a d d i ­ t i o n a l u n d e f i n e d m e c h a n i s m s or m e d i a t o r s m a y play a r o l e . W e a l s o c o n c l u d e from o u r e x p e r i ­ m e n t s that light d a m a g e in d e t a c h e d r e t i n a m a y o c c u r w i t h light s o u r c e s c o m m o n l y u s e d b y clinicians, particularly during lengthy vitreo­ retinal surgical procedures.

References 1. Calkins, J. L., Hochheimer, B. P., and D'Anna, S. Α.: Potential hazards from specific ophthalmic devices. Vision Res. 20:1039, 1 9 8 0 . 2. Fuller, D., Machemer, R., and Knighton, R. W.: Retinal damage produced by the intraocular fiber optic light. Am. J. Ophthalmol. 85:519, 1 9 7 8 . 3. Noell, W., Walker, V. S., Kang, B. S., and Berg­ man, S.: Retinal damage by light in rats. Invest. Ophthalmol. Vis. Sei. 5:450, 1966. 4. Rinkoff, ] . , Machemer, R., Hida, T., and Chan­ dler, D.: Temperature-dependent light damage to the retina. Am. J. Ophthalmol. 102:452, 1986. 5. T'so, M. O. M., Fine, B. S., and Zimmerman, L. E.: Photic maculopathy produced by the indirect ophthalmoscope. I. Clinical and histopathologic study. Am. J. Ophthalmol. 73:686, 1972. 6. Lawwill, T.: Three major pathologic processes caused by light in the primate retina. A search for mechanisms. Trans. Am. Ophthalmol. Soc. 80:516, 1982. 7. Ham, W. T., Ruffolo, J. J . , and Mueller, H. Α.: The nature of retinal radiation damage. Dependence on wave length, power level and exposure time. Vision Res. 20:1105, 1980. 8. Rapp, L. M., and Williams, T. P.: The role of ocular pigmentation in protecting against retinal light damage. Vision Res. 20:1127, 1 9 8 0 . 9. Hoeppler, T., Hendrickson, P., Dietrich, C , and Reme, C : Morphology and time course of defined photochemical lesions in the rabbit retina. Curr. Eye Res. 7:849, 1988. 10. Chandler, D. B., Quansah, F. Α., Hida, T., and Machemer, R.: A refined experimental model for proliferative vitreoretinopathy. Graefes Arch. Clin. Exp. Ophthalmol. 224:84, 1986. 11. Ham, W. T., Ruffolo, J. J . , Mueller, H . A . , Clarke, A. M., and Moon, M. E.: Histologic analysis of photochemical lesions produced in the rhesus retina by short wavelength light. Invest. Ophthal­ mol. Vis. Sei. 17:1029, 1 9 7 8 . 12. Machemer, R.: Experimental retinal detach­ ment in the owl monkey. II. Histology of retina and pigment epithelium. Am. J. Ophthalmol. 6 6 : 3 9 6 , 1968. 13. Kroll, A. J . , and Machemer, R.: Experimental retinal detachment in the owl monkey. III. Electron microscopy of retina and pigment epithelium. Am. J. Ophthalmol. 66:410, 1 9 6 8 . 14. Ham, W. T., Mueller, H. Α., Ruffolo, J. J . , Millen, J. E., Cleary, S. F., Guerry, R. K., and Guerry, D.: Basic mechanisms underlying the production of photochemical lesions in mammalian retina. Curr. Eye Res. 3:165, 1 9 8 4 .