Current Eye Research
ISSN: 0271-3683 (Print) 1460-2202 (Online) Journal homepage: http://www.tandfonline.com/loi/icey20
Zymosan induced epithelial cells
45
Ca uptake by retinal pigment
Rocío Salceda 45
To cite this article: Rocío Salceda (1992) Zymosan induced Ca uptake by retinal pigment epithelial cells, Current Eye Research, 11:3, 195-201, DOI: 10.3109/02713689209001770 To link to this article: http://dx.doi.org/10.3109/02713689209001770
Published online: 02 Jul 2009.
Submit your article to this journal
Article views: 2
View related articles
Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=icey20 Download by: [University of Saskatchewan Library]
Date: 03 April 2016, At: 19:33
Current Eye Research
Volume 1 I numbcr 3 1992. 19.5-201 -
Zymosan induced 45Ca uptake by retinal pigment epithelial cells
Rocio Salccda
Downloaded by [University of Saskatchewan Library] at 19:33 03 April 2016
Departamento de Neurociencias, Instituto de Fisiologia Celular, UNAM, Apartado Postal 70-600, 04510 Mexico, Mexico
ABSTRACT 45Ca u p t a k e was s t u d i e d i n i s o l a t e d f r o g r e t i n a l pigment e p i t h e l i a l c e l l s i n r e s p o n s e t o t h e p h a g o c y t i c s t i m u l i , zymosan. 45Ca u p t a k e was s t r o n g l y s t i m u l a t e d immediately i n t h e p r e s e n c e of zymosan p a r t i c l e s . Calcium u p t a k e was p r o p o r t i o n a l t o t h e zymosan c o n c e n t r a t i o n s . A f t e r 60 min i n t h e p r e s e n c e of zymosan a c i d p h o s p h a t a s e and ( ? J - g l u c u r o n i d a s e a c t i v i t i e s showed a 25% and 50% i n c r e a s e , r e s p e c t i v e l y . Rod o u t e r segments induced a similar i n c r e a s e o f t h e s e enzyme a c t i v i t i e s . The zymosan-induced l y s o s o m a l enzyme a c t i v i t i e s was i n h i b i t e d by c y t o c h a l a s i n B and ruthenium r e d . The i o n o p h o r e A23187 produced a r e m a r k a b l e i n c r e a s e i n 45Ca u p t a k e b u t d i d n o t a f f e c t t h e l y s o s o m a l enzyme a c t i v i t i e s . T h e s e r e s u l t s s u g g e s t t h a t i n v i t r o TPE c e l l s a r e a b l e t o respond t o zymosan a s p h a g o c y t o s a b l e s t i m u l i and t h a t c a l c i u m m e d i a t e t h a t r e s p o n s e .
o t h e r c e l l s y s t e m s ( 1 3 , 1 4 ) i s i n f l u e n c e d by e x t e r n a l calcium, t h i s study w a s undertaken t o examine t h e p o s s i b l e r o l e of c a l c i u m f l u x e s d u r i n g zymosan i n d u c e d p h a g o c y t o s i s i n i s o l a t e d f r o g RPE c e l l s .
P r e l i m i n a r y s t u d i e s h a v e been
p r e v i o u s l y r e p o r t e d i n a b s t r a c t form ( 1 5 ) .
METHODS The r e s e a r c h w a s done i n a c c o r d a n c e w i t h t h e ARVO R e s o l u t i o n on t h e u s e of Animals i n R e s e a r c h . Frog RPE c e l l s were i s o l a t e d by a f i c o l - d e n s i t y g r a d i e n t ( 1 6 ) . The i s o l a t e d c e l l s were r e s u s p e n d e d i n a R i n g e r ' s K r e b s b i c a r b o n a t e medium c o n t a i n i n g (mM):
N a C 1 , 118; K C 1 , 4 . 7 ; KH2P04, 1.2;MgS04,
INTRODUCTION
1 . 1 7 ; CaC12, 2.5;
The r e t i n a l pigment e p i t h e l i u m (RPE) i s a u n i q u e
b i c a r b o n a t e , 3 5 ; pH 7.4. E x p e r i m e n t s were c a r r i e d
n e u r o e p i t h e l i a l t i s s u e t h a t forms a f u n c t i o n a l
o u t u n d e r dim r e d l i g h t .
u n i t witii t h e p h o t o r e c e p t o r c e l l of t h e n e u r a l r e t i n a . RPE p a r t i c i p a t e s among o t h e r f u n c t i o n s ,
g l u c o s e , 5 . 6 ; sodium
F o r measurements of 45Ca u p t a k e , RPE c e l l s (0.2-0.4
mg p r o t e i n ) w e r e i n c u b a t e d a t 37" C i n
i n t h e d e z r a d a t i o n of p h o t o r e c e p t o r o u t e r
1 m l of K r e b s medium c o n t a i n i n g 0 . 2 f l i
segment membranes (ROS) t h a t a r e shed from t h e
45CaC12 (New England N u c l e a r , B o s t o n , MA) and
t i p s of rod and cone c e l l s ( 1 ) . I n a d d i t i o n t o
d i f f e r e n t zymosan c o n c e n t r a t i o n s .
of
Zymosan A from
(Sigma Chem. Co. S t . L o u i s Mo, USA)
X O S , t h e RPE c e l l s h a v e been r e p o r t e d t o b i n d
S.cerevisiae
a n d / o r i n g e s t l a t e x b e a d s , c a r b o n p a r t i c l e s and
was p r e p a r e d a c c o r d i n g t o T a f f e t a n d R u s s e l l
r e d blood c e l l s ( 2 - 6 ) .
( 1 7 ) . A t s p e c i f i c p e r i o d s of t i m e c e l l s were
The p h a g o c y t i c p r o c e s s i s
s i m i l a r t o p h a g o c y t o s i s by o t h e r c e l l s s u c h a s
c e n t r i f u g e d i n a Beckman m i c r o f u g e f o r 20 s e c .
monocytes and macrophages ( 7 ) . The h i s t o l o g i c a l
The p e l l e t w a s r i n s e d s u p e r f i c i a l l y w i t h c o l d
c h a r a c t e r i s t i c s of p h o t o r e c e p t o r s h e d d i n g and
medium a n d t i s s u e s o l u b i l i z e d w i t h 0.1 % sodium
p h a g o c y t o s i s h a v e been s t u d i e d e x t e n s i v e l y ( 8 , 9 )
d o d e c y l s u l f a t e (SDS). The a c c u m u l a t e d
b u t t h e b i o c h e m i c a l mechanisms a r e n o t w e l l
r a d i o a c t i v i t y w a s d e t e r m i n e d a f t e r a d d i t i o n of
defined.
5 ml T r i t o s o l i n a Beckman l i q u i d s c i n t i l l a t i o n
S i n c e changes i n e x t r a c e l l u l a r c a l c i u m c a n a l t e r t h e a d h e s i v e n e s s of v a r i o u s c e l l t y p e s (10) and s i n c e p h a g o c y t o s i s i n t h e RPE (11,12)
Received on September 6, 1991; accepted
on
and
s p e c t r o p h o t o m e t e r . The v a l u e s were c o r r e c t e d f o r 45Ca b i n d i n g a t c e r o t i m e i n c u b a t i o n . F o r 45Ca e f f l u x , RPE c e l l s were l o a d e d w i t h
February 28, 1992
~
0 Oxford University Press
195
Current Eye Research 45Ca f o r 10 min a t 37" C . They w e r e c e n t r i f u g e d , and r e s u s p e n d e d i n normal medium. A f t e r s u i t a b l e i n c u b a t i o n p e r i o d a t 37" C t h e c e l l s were c e n t r i f u g e d and t h e 45Ca r e m a i n i n g i n RPE c e l l s c o u n t e d a s d e s c r i b e d above. 14C-Sucrose a c c u m u l a t i o n by RPE c e l l s was t e s t e d i n c u b a t i n g t h e RPE c e l l s a t 37" C i n 1 m l K r e b s medium c o n t a i n i n g l p C i of 14C-sucrose (New England N u c l e a r , Boston, MA.) Accumulated r a d i o a c t i v i t y was d e t e r m i n e d a s d e s c r i b e d above.
Downloaded by [University of Saskatchewan Library] at 19:33 03 April 2016
Acid p h o s p h a t a s e and p - g l u c u r o n i d a s e a c t i v i t i e s
were t e s t e d a f t e r f r e e z i n g and thawing RPE c e l l s , by t h e h y d r o l i s i s o f p - n y t r o p h e n y l
1
I
I
1
10
30
60
p h o s p h a t e and
Time (min)
p h e n o p h t h a l e i n mono b - g l u c u r o n i c a c i d , r e s p e c t i v e l y , u s i n g a commercial k i t (Sigma Chem. Co., S t . L o u i s , Mo.). The p h a g o c y t o s i s of p h o t o r e c e p t o r o u t e r segments (ROS) was t e s t e d f o l l o w i n g t h e g e n e r a l p r o c e d u r e of Goldman e t a1 (18). R e t i n a s i n K r e b s
F i g u r e 1. Time c o u r s e of 14C-sucrose u p t a k e by RPE. Cells were i n c u b a t e d i n K r e b s medium c o n t a i n i n g 1 y C i of 14C s u c r o s e i n t h e p r e s e n c e ( ) o r a b s e n c e ( A ) of zymosan (250 p g / m l ) . Zymosan (250 r g / m l ) + r u t h e n i u m r e d ( l O O p M ) ( 0 ) . Each p o i n t r e p r e s e n t s t h e mean o f f o u r e x p e r i m e n t s f SEM.
medium were g e n t l y p i p e t e d up and down s e v e r a l times w i t h a wide-bore P a s t e u r p i p e t t e . T h e s u s p e n s i o n was t h e n f i l t e r e d t h r o u g h t e f l o n s c r e e n and ROS were c o l l e c t e d from t h e f i l t r a t e
c o n t r o l l e v e l a f t e r 1 0 min and c o n t i n u e d f o r
by low s p e e d c e n t r i f u g a t i o n (900 xg 1 0 min a t 4"
a b o u t 30 min b e f o r e l e v e l i n g o f f ( F i g . 1 ) .
C) and r e s u s p e n d e d i n K r e b s medium. RPE c e l l s
I n c u b a t i o n of RPE c e l l s i n t h e p r e s e n c e of
were i n c u b a t e d i n K r e b s medium f o r d i f f e r e n t
zymosan b r i n g s a b o u t a n a l m o s t i m m e d i a t e i n c r e a s e
p e r i o d s of t i m e i n t h e p r e s e n c e of ROS. A t t h e
i n 45Ca u p t a k e . E x p o s u r e t o zymosan i n c r e a s e d
end of t h e i n c u b a t i o n , RPE c e l l s were s e p a r a t e d
45Ca a c c u m u l a t i o n by a l m o s t t h r e e f o l d o v e r t h e
from t h e ROS by 1 5 min c e n t r i f u g a t i o n a t 1000 rpm
c o n t r o l l e v e l a f t e r two min, r e a c h i n g a p e a k a t
i n a d i s c o n t i n u o us f i c o l d e n s i t y g r a d i e n t (bottom
1 0 min. T h e r e a f t e r t h e r a t e of u p t a k e of l a b e l
l a y e r 20%, m i d d l e l a y e r 10% and u p p e r l a y e r 5 % ) .
d e c r e a s e d r e a c h i n g t h e c o n t r o l l e v e l s a t 60 min
The RPE c e l l s a t t h e bottom of t h e t u b e w e r e
( F i g . 2 ) . The u p t a k - of 45Ca i n c r e a s e d w i t h
washed w i t h c o l d medium a n d l y s o s o m a l enzyme
i n c r e a s i n g zymosan l e v e l s i n t h e e x t e r n a l medium,
a c t i v i t i e s were d e t e r m i n e d .
r e a c h i n g s a t u r a t i o n a t 500 p / m l ( F i g . 3 ) .
P r o t e i n s w e r e d e t e r m i n e d by t h e p r o c e d u r e
I n t h e p r e s e n c e of l O O p M of t h e c a l c i u m b l o c k e r , r u t h e n i u m r e d (RR) , t h e zymosan i n d u c e d 45Ca
d e s c r i b e d by Lowry e t a 1 ( 1 9 ) .
uptake w i t h i n t h e c e l l s w a s reduced t o c o n t r o l RESULTS
levels, b u t i t was n o t i n f l u e n c e d by t h e p r e s e n c e
I n o r d e r t o e s t a b l i s h zymosan a s a p h a g o c y t o s a b l e
of t h e c a l c i u m c h a n n e l b l o c k e r v e r a p a m i l
p a r t i c l e by RPE, w e f o l l o w e d t h e u p t a k e of 14C-
( T a b l e 1). While e x p o s u r e of RPE t o l O O p M R R ,
s u c r o s e and t h e a c t i v i t y of t h e l y s o s o m a l enzymes
p a r t i a l l y r e d u c e d t h e zymosan i n d u c e d 14C-sucrose
a c i d phosphatase and 6-glucuronidase.
u p t a k e ( F i g . 1).
Under normal
c o n d i t i o n s RPE c e l l s a c c u m u l a t e d l o w l e v e l s of
I n c o n t r a s t t o 45Ca u p t a k e , zymosan a d d i t i o n
14C-sucrose. Exposure of RPE t o zymosan i n c r e a s e d
t o RPE c e l l s d i d n o t c h a n g e t h e amount of 45Ca
t h e u p t a k e of 14C-sucrose by t h r e e f o l d o v e r t h e
r e l e a s e d f r o m p r e l a b e l e d c e l l s ( F i g . 4), which
196
Current Eye Research T a b l e 1. E f f e c t o f d i f f e r e n t d r u g s on t h e 45Ca u p t a k e by RPE Addition
45Ca u p t a k e (?moles /mg p r o t e i n )
--
0
0
Downloaded by [University of Saskatchewan Library] at 19:33 03 April 2016
0.73 f 0.12 1 . 8 2 f 0.30 1 . 3 8 2 0.20
Zymosan 250 ,ug/ml A23187 ( 5 ,ug/ml) Zymosan (250 p g / m l ) + Ruthenium r e d ( 1 0 0 p M ) Zymosan (250 p g / m l ) + C y t o c h a l a s i n ( 5 +g/ml) Zymosan (250 y g / m l ) + V e r a p a m i l (10 fM)
T i m e (min) F i s u r e 2 . T i m e c o u r s e of zymosan provoked 45Ca u p t a k e by RPE, ( A ) c o n t r o l c e l l s ; ( 6 ) i n t h e p r e s e n c e of zymosan (250 ,ug/ml). 45Ca and zymosan were added a t z e r o t i m e . The d a t a a r e mean of s i x e x p e r i m e n t s f SEM.
0.94
2 0.09
1.84
2
0.26
1.65
2
0.24
RPE c e l l s were i n c u b a t e d f o r 10 min i n a Krebsb i c a r b o n a t e medium i n t h e d a r k . 45Ca w a s added a t t h e same t i m e a s t h e d r u g . Each v a l u e i s t h e mean 2 SEM of 4 t o 7 e x p e r i m e n t s .
I
I T
*
I
1
I
1
500
1000
I
1
I
10
20
30
Time (min)
Zymosan ( y g / m l ) F i g u r e 3. Dependence of 45ca2+ u p t a k e on t h e amount of zymosan. The i n c u b a t i o n t i m e i n t h e p r e s e n c e of 45Ca and zymosan w a s 10 min t h r o u g h o u t . Each p o i n t r e p r e s e n t s t h e mean of f o u r e x p e r i m e n t s f SEM.
F i g u r e 4. T i m e c o u r s e of 45Ca e f f l u x from RPE. C e l l s were l o a d e d w i t h 45Ca f o r 1 0 min a t 37OC. They w e r e c e n t r i f u g e d a n d r e s u s p e n d e d i n n o r m a l medium i n t h e p r e s e n c e ( 0 ) o r a b s e n c e ( & ) of zymosan ( 2 5 0 p g / m l ) . A t t h e i n d i c a t e d t i m e s , c e l l s were r a p i d l y c e n t r i f u g e d a n d t h e remaining r a d i a c t i v i t y counted. The d a t a are mear! of t h r e e e x p e r i m e n t s SEM.
s u g g e s t s t h a t zymosan i n d u c e d a n e t c a l c i u m
were i n c r e a s e d 25% a n d 50% r e s p e c t i v e l y , i n RPE
accumulation.
cells a f t e r one hour i n c u b a t i o n i n t h e presence
I n o r d e r t o e s t a b l i s h a r e l a t i o n s h i p between 45Ca u p t a k e and p h a g o c y t o s i s by RPE c e l l s w e
of zymosan ( F i g . 5 ) . A t s h o r t e r p e r i o d s of
examined t h e e f f e c t of zymosan on t h e l y s o s o m a l
i n c u b a t i o n , t h e s e enzymes showed a s l i g h t
enzymes a c i d p h o s p h a t a s e a n d + - g l u c u r o n i d a s e .
The
a c i d p h o s p h a t a s e and Q - g l u c u r o n i d a s e a c t i v i t i e s
i n c r e a s e . The zymosan i n d u c e d l y s o s o m a l enzyme a c t i v i t i e s w e r e reduced considerably i n t h e
197
Current Eye Research d e t a c h e d f o r s u b s e q u e n t p h a g o c y t o s i s , and d e g r a d a t i o n by a d j a c e n t p i g m e n t e p i t h e l i a l c e l l s
(1) By a n a l o g y w i t h t h e model p u t f o r t h f o r t h e macrophage, p h a g o c y t o s i s of ROS d i s c s i s o f t e n
T
considered t o be a multistep process, encompassing b i n d i n g , i n g e s t i o n a n d d e g r a d a t i o n (3.20).
I t is w e l l known t h a t t a r g e t p a r t i c l e s
i n t e r a c t w i t h macrophage c e l l membranes t h r o u g h s p e c i f i c r e c e p t o r l i g a n d bonds, and t h a t t h i s
Downloaded by [University of Saskatchewan Library] at 19:33 03 April 2016
binding i n t u r n t r i g g e r s engulfment through t h e
+ .g
t
e x t e n s i o n of
pseudopodial processes (21,22).
P r e v i o u s s t u d i e s h a v e shown t h a t l i g h t - e v o k e d d i s c s shedding r e q u i r e s e x t r a c e l l u l a r calcium
Acid phosphatase
)-glucuronidose
(11,12). E x p o s u r e t o RPE c e l l s t o zymosan b r i n g s a b o u t a dose-dependent
F i g u r e 5. E f f e c t of d i f f e r e n t compounds and ROS on t h e a c i d p h o s p h a t a s e and (5-glucuronidase a c t i v i t i e s of RPE c e l l s . Cells were i n c u b a t e d i n Krebs medium d u r i n g 60 min a t 37" C . A f t e r i n c u b a t i o n s c e l l s were c e n t r i f u g e d and t h e enzyme a c t i v i t i e s measured a s d e s c r i b e d i n Methods. Zymosan, 250 p g / m l ; c y t o c h a l a s i n B , 5 pglrnl; A23187, 5pg/ml; r u t h e n i u m r e d , 100pM. Values a r e t h e mean f SEM ( v e r t i c a l b a r s ) of 4-7 e x p e r i m e n t s .
-
i n c r e a s e i n 45Ca- u p t a k e
( F i g . 3,4). T h i s i n c r e a s e d i n f l u x p r e c e d e s t h e o n s e t of 14C-sucrose u p t a k e and enhancement of l y s o s o m a l enzyme a c t i v i t i e s ( F i g . 1 , 5 ) . 14C-Sucrose u p t a k e r e q u i r e s 1 0 t o 30 min a f t e r i n i t i a l e x p o s u r e of t h e c e l l s t o t h e zymosan p a r t i c l e s , w h e r e a s 30-60 min a r e r e q u i r e d f o r t h e zymosan a n d ROS-induced
i n c r e a s e of t h e lysosomal
enzyme a c t i v i t i e s t e s t e d . T h e s e r e s u l t s a r e i n
p r e s e n c e of c y t o c h a l a s i n B , a d r u g which d i s r u p t s
a g r e e m e n t w i t h t h o s e d e s c r i b e d t h a t i n macrophages
a c t i n filaments (Fig. 5), without a f f e c t i n g
and L c e l l s , where e n d o c y t i c v a c u o l e s f u s e w i t h
45Ca a c c u m u l a t i o n ( T a b l e 1 ) . On t h e o t h e r h a n d ,
l y s o s o m e s w i t h i n 5-15 min a n d w h e r e by 60 min
t h e a d d i t i o n of 100 p M RR t o t a l l y i n h i b i t e d t h e
most of t h e a d s o r b e d p a r t i c l e s were i n l o c a t e d
zymosan i n d u c e d a c i d p h o s p h a t a s e a n d
l y s o s o m e s ( 2 3 , 2 4 ) . F u r t h e r m o r e , t h e k i n e t i c s of
(3-glucuronidase a c t i v i t i e s , s u g g e s t i n g t h a t
p h a g o c y t o s i s r e p o r t e d h e r e f o l l o w similar
calcium uptake mediates such response (Fig. 5 ) .
p a t t e r n s t o t h o s e r e p o r t e d by o t h e r i n v e s t i g a t o r s
S i m i l a r r e s u l t s were o b s e r v e d i n RPE c e l l s
(8,18,25,26).
exposed t o t h e n a t u r a l p h a g o c y t o s a b l e s t i m u l i ,
The i n v o l v e m e n t of membrane i n t e r n a l i z a t i o n
ROS. I n c u b a t i o n of RPE c e l l s i n t h e p r e s e n c e of
d u r i n g 45Ca u p t a k e i n d u c e d by zymosan, was
i s o l a t e d ROS l e a d s t o a n i n c r e a s e of 20% a n d 40%
examined by e x p o s i n g RPE c e l l s t o zymosan
on t h e a c i d p h o s p h a t a s e a n d ( b - g l u c u r o n i d a s e
p a r t i c l e s i n t h e p r e s e n c e of c y t o c h a l a s i n B ,
a c t i v i t i e s , respectively (Fig. 5).
which i n h i b i t s p h a g o c y t o s i s and i n t e r n a l i z a t i o n
I n t e r e s t i n g l y , e x p o s u r e of RPE c e l l s t o t h e
of t h e p l a s m a membrane b u t d o e s n o t a f f e c t
c a l c i u m i o n o p h o r e A23187, which produced a
a t t a c h m e n t of p a r t i c l e s t o t h e plasma membrane.
r e m a r k a b l e i n c r e a s e i n 45Ca u p t a k e ( T a b l e l), d i d
W h i l e c y t o c h a l a s i n B d i d n o t a f f e c t 45Ca u p t a k e ,
n o t a f f e c t t h e l y s o s o m a l enzyme a c t i v i t i e s ( F i g , 5 ) .
i t b l o c k e d t h e i n c r e a s e i n l y s o s o m a l enzyme
a c t i v i t i e s , suggesting t h a t phagocytosis and/or DISCUSSION
phagosome-lysosomal
Rod p h o t o r e c e p t o r d i s c s h e d d i n g i s t h e p r o c e s s by
i n v o l v e m e n t i s f u r t h e r s u p p o r t e d by t h e f a c t t h a t
which d i s c s from t h e o u t e r segment d i s t a l t i p a r e
t h e r e i s a n a b u n d a n c e of a c t i n f i l a m e n t s i n t h e
198
fusion was inhibited. Actin
Current Eye Research a p i c a l p r o c e s s e s o f t h e RPE c e l l s ( 2 7 , 2 8 ) , and
i n d u c e d similar i n c r e a s e on t h e l e v e l s of 45Ca,
t h a t a c t i n h a s been l o c a l i z e d underneath t h e
b u t l y s o s o m a l enzyme a c t i v i t i e s w e r e n o t
plasma membrane a t t h e p h a g o c y t i c s i t e s i n RPE
i n c r e a s e d . C o n s e q u e n t l y , a s s o c i a t i o n o f zymosan
c e l l s ( 5 , 2 8 ) . F u r t h e r m o r e , c y t o c h a l a s i n B and D
with
b l o c k b o t h l i g h t - e v o k e d and dark-primed d i s c s
calcium
shedding (29,30).
s p e c i f i c c e l l u l a r compartments.
Downloaded by [University of Saskatchewan Library] at 19:33 03 April 2016
The most i n t e r e s t i n g f e a t u r e i s t h a t e x p o s u r e
specific surface receptor uptake
mechanism d i r e c t i n g
study
indicates
to
RPE
r e c o g n i z e s zymosan as a p h a g o c y t i c s t i m u l u s .
The
of i n c r e a s e i n l y s o s o m a l enzyme a c t i v i t i e s w i t h i n
observation
RPE c e l l s . The mechanism by which RPE r e c o g n i z e s
block
and p h a g o c y t i z e s ROS t i p s i s n o t w e l l u n d e r s t o o d .
blocks
O t h e r p h a g o c y t i c c e l l s s e l e c t i v e l y t a k e up
activities
p a r t i c l e s by s p e c i f i c l i g a n d - r e c e p t o r i n t e r a c t i o n s
uptake, suggests t h a t calcium p a r t i c i p a t e s i n the
t h o u g h t t o i n v o l v e r e c o g n i t i o n of g l y c o c o n j u g a t e s
mechanism of p h a g o c y t o s i s i n RPE c e l l s .
RR,
in
calcium
vitro,
that
that
activate
of RPE c e l l s t o ROS i n d u c e d a similar p a t t e r n
(21,22,31-33).
This
might
a n i n o r g a n i c dye know
t h e c a l c i u m u p t a k e t h r o u g h membranes the
increase
of
lysosomal
i n d u c e d by zymosan,
to (42)
enzyme
as w e l l as
45Ca
RPE p h a g o c y t o s i s of ROS h a s a l s o
been s u g g e s t e d t o i n v o l v e g l y c o c o n j u g a t e l i g a n d -
ACKNOWLEDGMENTS
r e c e p t o r i n t e r a c t i o n ( 3 , 3 4 , 3 5 ) and t h e p r e s e n c e
This
of a mannose s p e c i f i c r e c e p t o r h a s been
P228CCX891591 from CONACyT.
work
was
supported
in
part
by
grant
d e m o n s t r a t e d (36-38). CORRESPONDING AUTHOR
The zymosan and ROS i n d u c e d l y s o s o m a l enzyme a c t i v i t i e s may b e d u e b o t h t o i n t e r n a l i z a t i o n and s y n t h e s i s of l y s o s o m a l enzymes. The RPE r e l e a s e s and r e c a p t u r e s t h e l y s o s o m a l enzyme N
-
acetyl
-4
D r . Roclo S a l c e d a , Departamento d e N e u r o c ie n c ia s , I n s t i t u t o de F i s i o l o g I a C e l u l a r ,
UNAM.
P o s t a l 70-600,
MEXICO.
04510 Mzxico, D.F.
Apartado
g l u c o s a m i n i d a s e by way of a mannose 6-phosphate r e c e p t o r ( 3 9 ) . I t seems l i k e l y t h a t o t h e r enzymes
REFERENCES
may a l s o f o l l o w t h i s pathway. On t h e o t h e r h a n d ,
1.
p h a g o c y t o s i s of r e d b l o o d c e l l s and p i n o c y t o s i s of serum p r o t e i n s i n d u c e t h e s y n t h e s i s of
2.
l y s o s o m a l enzymes i n mouse macrophages a n d l e a d s to a tenfold increase i n t h e i r intracellular content (23). Those r e s u l t s i n d i c a t e t h a t t h e i n c r e a s e i n
3.
45Ca u p t a k e may p a r t i c i p a t e i n c o u p l i n g t h e i n i t i a l i n t e r a c t i o n o f t h e zymosan p a r t i c l e s w i t h
4.
s u r f a c e r e c e p t o r s t o t h e cytoplasmic phase of p h a g o c y t o s i s . T h a t i d e a a g r e e s w i t h t h e r a p i d and t r a n s i e n t e l e v a t i o n of i n t e r n a l c a l c i u m o b s e r v e d d u r i n g c o n t a c t of a macrophage w i t h a zymosan
5.
p a r t i c l e ( 4 0 ) , as w e l l a s w i t h t h e s i n g l e c a l c i u m p u l s e s o b s e r v e d d u r i n g zymosan p h a g o c y t o s i s by
6.
macrophages ( 4 1 ) . T h a t t h i s i n c r e a s e i s n o t simply a non-specific
e f f e c t on t h e c y t o p l a s m i c
c a l c i u m a s s o c i a t e d w i t h t h e o n s e t of p h a g o c y t o s i s
i s shown by t h e 45Ca u p t a k e r e s u l t i n t h e p r e s e n c e of t h e c a l c i u m i o n o p h o r e A23187. T h i s i o n o p h o r e
7.
Young, R. W. and Bok, D . (1969) P a r t i c i p a t i o n of t h e r e t i n a l pigment e p i t h e l i u m i n t h e r o d o u t e r segment r e n e w a l p r o c e s s . J . C e l l B i o l . 42, 392-402. Edwards, R.B. and S z a m i e r , R.B. (1977) D e f e c t i v e p h a g o c y t o s i s of i s o l a t e d r o d o u t e r segments by RCS rat retinal pigment e p i t h e l i u m i n c u l t u r e . S c i e n c e , 197, 10011003. Feeney, L . and Mixon, R.M. (1976) An i n v i t r o model of p. h a g o c .y t o s i s i n b o v i n e and human r e t i n a l pigment e p i t h e l i u m . Exp. Eye Res. 22, 533-548. J.G. and Ward, A. (1974). Hollyfield, P h a g o c y t i c a c t i v i t y i n t h e r e t i n a l pigment I. e p i t h e l i u m of t h e f r o g Rana p i p i e n s . Uptake of polystyrene spheres. J. U l t r a s t r u c t . Res. 46, 327-338. M.O. (1983) D e f e c t i v e C h a i t i n , M.H. a n d ? a l l , i n g e s t i o n of r o d o u t e r s e g m e n t s by c u l t u r e d dystrophic r a t pigment e p i t h e l i u m c e l l s . 812-820. I n v e s t . Ophthalmol. V i s . S c i . 6, P.L. a n d H a l l , M.O. (1986) R a t Mayerson, retinal pigment epithelium cells show s p e c i f i c i t y of p h a g o c y t o s i s i n v i t r o . J . C e l l B i o l . 103, 299-308. H.L. and S t o s s e l , T.P. (1982) The Yin, mechanism of p h a g o c y t o s i s . I n " P h a g o c y t o s i s M.L. and P a s t and F u t u r e " (Ed. Karnovsky, B o l i s , L.) Pp. 1 S 2 7 . Academic P r e s s , New York.
199
Downloaded by [University of Saskatchewan Library] at 19:33 03 April 2016
Current Eye Research 8. LaVail, M.M. (1976) Rod outer segment disk shedding in rat retina: relationship to cyclic lighting. Science,=, 1071-1073. 9. Matsumoto, B., Defoe, D.M. and Besharse, J.C.(1987) Membrane turnover in rod photoreceptors:ensheathment and phagocytosis of outer segment distal tips by pseudopodia of the retinal pigment epithelium. Proc. R. SOC. Lond. B, 230, 339-354. 10. Ginell, D., Garod, D.R. and Palmer, J.F. (1970) Divalent cations and cell adhesion. In "Calcium and Cell Function" (Ed.Cuthbert, A.W.) Pp. 50-59. St. Martin's Press, New York. 11. Greenberger, L.M. and Besharse, J.C. (1983) Photoreceptor disc shedding eye cups. Inhibition by deletion of extracellular divalent cations. Invest. Ophthalmol. Vis. Sci. 24, 1456-1464. 12. Besharse, J.C. Spratt, G. and Forestner, D.M. (1986) Light-evoked and kainic-acidinduced disc shedding by rod photoreceptors: differential sensitivity to extracellular calcium. J. Comp. Neurol. 251, 185-197. 13. Hoffstein, S.T., Korchak, H.M., Smolen, J.E. and Weissmann, G.(1982) Early consequences of neutrophil activation and their association with degranulation. In "Phagocytosis-Past and Future" (Ed. Karnovsky, M.C. and Bolis, L.) Pp. 47-65, Academic Press, New York. (1983) Cazf sensitive 14. Oliveira-Castro, G . h . K+ channels in phagocytic cell membranes. Cell Calcium, 5, 475-492. K. (1990) Zymosan induced 45Ca 15. Salceda, accumulation by retinal pigment epithelium. 278. Proc. Int. Eye Res. 16. Salceda, R. (1986) Isolation and biochemical characterization of frog retinal pigment epithelium cells. Invest. Ophthalmol. Vis. 1172-1176. Sci. 17. Taffet, S.M. and Russell, S.W. (1981) Identification of mononuclear phagocytes by ingestion of particulate materials, such as erythrocytes, carbon, zymosan, or latex. In "Methods for Studying Mononuclear phagocytes" (Ed. Adams, D.O., bdelson, P.J. and Koren, H.) Pp. 289-293. Academic Press, New York. 18. Goldman, A.I. O'Brien. P.L., Materson, E., Israel, P., Teirstein, P. and Chader, G. (1979) A quantitative system for studying phagocytosis in pigment epithelium tissue culture. Exp. Eye Res. 28, 455-467. 19. Lowry, O.H., Rosebrough, N.J., Farr, A.L. and Randal, K.J. (1951) Protein measurement with the Folin-phenol reagent. J. B i o l . Chem. 193, 265-275. 20. Burnside, B. and Laties, A.M. (1979) Pigment movement and cellular contractility in the retinal pigment epithelium. In "The Retinal Pigment Epithelium" (Ed. Zinn, K.M. and Marmor, M.F.) Pp. 175-191. Harvard University Press, Cambridge. MA. 21. Griffin, F.M., Griffin, J.A., Leider, J.E. and Silverstein, S.C. (1975) Studies on the mechanism of phagocytosis. I. Requirements for circumferential attachment of particle bound ligands to specific receptors on the
z,
I,
200
macrophage plasma membrane. J. Exp. Med. 142, 1263-1282. 22. Tietze, C., Schlesinger, P. and Stahl, P. (1982) Mannose-specific endocytosis receptor of alveolar macrophages: Demonstration of two functionally distinct intracellular pools of receptor and their roles in receptor recycling. J. Cell Biol. 92, 417-424. 23. Silverstein, S.C., Steinman, R.M. and Cohn, Z.A. (1977) Endocytosis. Annu. Rev. Biochem. 46, 669-722. 24. Steinman, R.M., Mellman, I . S . , Muller, W.A. and Cohn, Z.A. (1983) Endocytosis and the recycling of plasma membrane. J. Cell Biol. 96, 1-27. 25. Esinger, S., Hoffman, R. and Matthes, M. (1976) Photoreceptor shedding is initiated by light in the frog retina. Science, 194, 10741077. 26. Immel, J.H. and Fisher, S.K. (1985) Cone photoreceptor shedding in the tree shrew (Tupaia belangerii). Cell Tissue Res. 239, 667-675. 27. Burnside, M.B. (1976) Possible roles of microtubules and actin filaments in retinal 257pigmented epithelium. Exp. Eye Res. 23, 275. Flood, M.T., Gouras, P. and 28. Haley, J . E . , Kjeldbye, H.M. (1983) Proteins from human retinal pigment epithelium cel1s:evidence that a major protein is actin. Invest. Ophthalmol. Vis. Sci. g,803-811. 29. Besharse, J.C. and Dunis, D.A. (1982) Kod photoreceptor disc shedding in vitro: Inhibition by cytochalasins and activation by colchicine. In "The Structure of the Eye". (Ed. Hollyfield, J.G.). Pp. 85-96. Elsevier, New York. 30. Defoe, D.M., Matsumoto, B. and Besharse, inhibits LJ.C. (1989) Cytochalasin D glutamate-induced disc shedding without altering L-glutamate-induced increase in adhesiveness. Exp. Eye. Res. 48, 641-652. 31. Warr, G.A. (1980) A macrophage receptor (mannane/glucosamine)-glycoproteins of for potential importance in phagocytic activity. Biochem. Biophys. Res. Commun. 93, 737-745. 32. Ashwell, G. and Harford, J. (1982) Carbohydrate specific receptors of the liver. Annu. Rev. Biochem. 2,531-554. 33. Sung, S.S.J., Nelson, R.S. and Silverstein, S.C. (1983) Yeast mannans inhibit binding and phagocytosis of zymosan by mouse peritoneal macrophages. J. Cell Biol. 96, 160-166. 34. O'Brien, P.J. (1976) Khodopsin as a glycoprotein: A possible role for the oligosaccharide in phagocytosis. Exp. Eye 127-137. Res .G, M.O. (1978) Phagocytosis of light-and 35. Hall, dark-adapted rod outer segments by cultured pigment epithelium. Science, 202, 526-528. 36. Tarnowski, B.I., Shepherd,. V.L. and McLaughlin, B.J. (1988) Expression of mannose receptors for pinocytosis and phagocytosis GII rat retinal pigment epithelium. Invest. 742-748. Ophthalmol. Vis Sci. 9, C.A. and Bernstein, M.H. (1991) 37. Heth,
Current Eye Research Mannose-sensitive HRP endocytosis by the retinal pigment epithelium. Exp. Eye Res. 52, 75-82. 38. Boyle,
D., Tien, L., Cooper, N.G.F., Shepherd, V. and McLaughlin, B.J. (1991) A mannose receptor is involved in retinal phagocytosis. Invest. Ophthalmol. V i s Sci.
32, 1464-1470. 39. E l c o x , D.K. (1987) Extracellular release of
acid hydrolases from cultured retinal pigment epithelium. Invest. Ophthalmol. Vis. Sci. 2 ,
Downloaded by [University of Saskatchewan Library] at 19:33 03 April 2016
76-82. 40. Jaconi,
M.E.E., Lew, D.P., Carpentier, J.L., Magnusson, K.E., Sjogren, M. anit Stendahl, 0 . (1990) Cytosolic free calcium elevation mediates the phagosome-lysosome fusion during phagocytosis in human neutrophils. J. Cell. Biol. 110, 1555-1564. 41. Meagher, L.C., Moonga, B.S., Haslett, C., Huang, C.L.H. and Zaidi, M. (1991) Single pulses of cytoplasmic calcium associated with phagocytosis of individual zymosan particles by macrophages. Biochem. Biophys. Res. Commun. 177, 460-465. 42. Rahamimoff,
R. and Alnes, E. (1973) Inhibitory action of ruthenium red on neuromuscular transmission. Proc. Natl. 70, 3613-3616. Acad. Sci. USA, -
20 1