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Collagen and fibronectin production by human and rat liver cells
83
COLLAGENAND FIBRONECTIN PRODUCTION BY HUMANAND RAT LIVER CELLS. B. C l e m e n t * r H. E m o n a r d * * t FI. R i s s e l * ~ I,I. B o u r e l * and A. G u i l l o u z o * . * INSERM U ~9 - H ~ p l t a l
de P o n t c h a l l l o u
M. D r u g u e t * * P 3 . A .
Grimaud**p
- 3 5 0 1 1 R E N N E S C4dex - * *
D. H e r b a g e * * * ,
CNRS ERA 819 -
Instltut Pasteur - 77, r u e Pasteur - 69365 LYON C~dex - *~* Centre de Recherche Appliqu4e de Dermobiochimie - 51, rue C. Marot - 69007 LYON Cedex --IRANCE. Despite a number of studies showing the heterogenous composition of the normal and pathological liver connective biomatrix which includes several collagens and glycoproteins, the cellular sources of its different components remain unknown. We have investigated the intracellular distribution of variou~ collagen types and fibronectin in human and rat liver by indirect immunoperoxidase following paraformaldehyde fixation by perfusion and the use of saponin as a membrane permeabilizing agent. Similar observations weremadein
both human and
rat liver. With light microscopy) only fibronectin was clearly visualized in hepatocytes. At the ultrastroctural
level, hepatocytes stained for ftbronectin, types I and IV collagens
while fat storing cells showed strong labelling for types III and IV collagens and to a lesser extent for type I collagen and fibronectin. Endothelial cells contained few amounts of all these components. In all the cells) components were specifically located in the endoplasmic reticolum and the Golgi apparatus. These results demonstrate that in the liver) several cell types including hepatocytes) porticipate in the formation of the extracellular matrix components.
84
MONOCLONAL ANTI-IDIOTYPES HEPATITIS B VIRUS SURFACE
RECOGNIZE A BIOLOGICAL ANTIGEN (HBsAg)
RECEPTOR
ON
G. C o l u c c i * , S.D. W a k s a l D i v i s i o n of I m m u n o p a t h o l o g y , D e p t . o f P a t h o l o g y , M t . Sinai S c h o o l of Medicine New York,NY,USA-*Clinica M e d i c a III, S c h o o l of L i v e r di s e a s e U n i v e r s i t y o ~ Milan, Italy. To d e f i n e and c h a r a c t e r i z e the ~ o l y m e r i z e d a l b u m i n (po!y'A) r e c e p t o r on HBV and h e p a t o c y t e s we h a v e p r o d u c e d m o n o c l o n a l a n t i - I d i o t y p e s (anti-Id) w h i c h bea the i n t e r n a l i m a g e of p o l y A ( a n t i - a n t i - p o l y A) and t h e r e f o r e b i n d its recept0 We i m m u n i z e d B a l b / c m i c e w i t h an a f f i n i t y p u r i f i e d h u m a n IgG a n t i - p o l y A and f u s e d m i c e s p l e e n c e l l s w i t h the HAT s e n s i t i v e cell line S P 2 / 0 - A z - 1 4 . Six h~br d o m a s w h i c h i n h i b i t e d the b i n d i n g of p o l y A to the a n t i - p o l y A a n t i b o d y w e r e e p a n d e d a n d s u b c l o n e d , t h e n f u r t h e r a n a l y z e d w i t h R I A and E L I S A for t h e i r specif city. II d i f f e r e n t h u m a n m y e l o m a p r o t e i n s s e r v e d as c o n t r o l s and H B s A g was obt a i n e d from the H B V t r a n s f e c t e d c e l l line 4.10. Two a n t i - I d s , 63.14 a n d 70.F9, s t r o n g l y r e a c t e d w i t h (Fab') 2 of the i m m Q n o g e n as w e l l as w i t h HBsAg, b u t not w i t h the c o n t r o l s . The b i n d i n g w a s s p e c i f i c a l l y i n h i b i t e d by p o l y A b u t not by m o n o m e r i c A , B S A or a n t i - H B s . F u r t h e r m o r e , 63.14 and 70.F9, s t a i n e d 4.10 and P L C / P R F / 5 c e l l s w i t h a p a t t e r n i d e n t i c a l to a n t i - H B s . In t h e s e i s t a n c e s the sta n i n g was also b l o c k e d by p o l y A, but n o t by m o n o m e r i c A , B S A or a n t i - H B s . On the o t h e r h a n d , t h e two a n t i - I d s f a i l e d to s t a i n n o r m a l l i v e r s e c t i o n s or the heDat ma cell line G2. T h e s e a n t i - I d s w i l l be v a l u a b l e in i n v e s t i g a t i n g the o a t h o g e n e sis of v i r a l h e p a t i t i s and c o u l d be U t i l i z e d in the p r e p a r a t i o n of a v a c c i n e . -
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COLLAGENAND FIBRONECTIN PRODUCTION BY HUMANAND RAT LIVER CELLS. B. C l e m e n t * r H. E m o n a r d * * t FI. R i s s e l * ~ I,I. B o u r e l * and A. G u i l l o u z o * . * INSERM U ~9 - H ~ p l t a l
de P o n t c h a l l l o u
M. D r u g u e t * * P 3 . A .
Grimaud**p
- 3 5 0 1 1 R E N N E S C4dex - * *
D. H e r b a g e * * * ,
CNRS ERA 819 -
Instltut Pasteur - 77, r u e Pasteur - 69365 LYON C~dex - *~* Centre de Recherche Appliqu4e de Dermobiochimie - 51, rue C. Marot - 69007 LYON Cedex --IRANCE. Despite a number of studies showing the heterogenous composition of the normal and pathological liver connective biomatrix which includes several collagens and glycoproteins, the cellular sources of its different components remain unknown. We have investigated the intracellular distribution of variou~ collagen types and fibronectin in human and rat liver by indirect immunoperoxidase following paraformaldehyde fixation by perfusion and the use of saponin as a membrane permeabilizing agent. Similar observations weremadein
both human and
rat liver. With light microscopy) only fibronectin was clearly visualized in hepatocytes. At the ultrastroctural
level, hepatocytes stained for ftbronectin, types I and IV collagens
while fat storing cells showed strong labelling for types III and IV collagens and to a lesser extent for type I collagen and fibronectin. Endothelial cells contained few amounts of all these components. In all the cells) components were specifically located in the endoplasmic reticolum and the Golgi apparatus. These results demonstrate that in the liver) several cell types including hepatocytes) porticipate in the formation of the extracellular matrix components.
84
MONOCLONAL ANTI-IDIOTYPES HEPATITIS B VIRUS SURFACE
RECOGNIZE A BIOLOGICAL ANTIGEN (HBsAg)
RECEPTOR
ON
G. C o l u c c i * , S.D. W a k s a l D i v i s i o n of I m m u n o p a t h o l o g y , D e p t . o f P a t h o l o g y , M t . Sinai S c h o o l of Medicine New York,NY,USA-*Clinica M e d i c a III, S c h o o l of L i v e r di s e a s e U n i v e r s i t y o ~ Milan, Italy. To d e f i n e and c h a r a c t e r i z e the ~ o l y m e r i z e d a l b u m i n (po!y'A) r e c e p t o r on HBV and h e p a t o c y t e s we h a v e p r o d u c e d m o n o c l o n a l a n t i - I d i o t y p e s (anti-Id) w h i c h bea the i n t e r n a l i m a g e of p o l y A ( a n t i - a n t i - p o l y A) and t h e r e f o r e b i n d its recept0 We i m m u n i z e d B a l b / c m i c e w i t h an a f f i n i t y p u r i f i e d h u m a n IgG a n t i - p o l y A and f u s e d m i c e s p l e e n c e l l s w i t h the HAT s e n s i t i v e cell line S P 2 / 0 - A z - 1 4 . Six h~br d o m a s w h i c h i n h i b i t e d the b i n d i n g of p o l y A to the a n t i - p o l y A a n t i b o d y w e r e e p a n d e d a n d s u b c l o n e d , t h e n f u r t h e r a n a l y z e d w i t h R I A and E L I S A for t h e i r specif city. II d i f f e r e n t h u m a n m y e l o m a p r o t e i n s s e r v e d as c o n t r o l s and H B s A g was obt a i n e d from the H B V t r a n s f e c t e d c e l l line 4.10. Two a n t i - I d s , 63.14 a n d 70.F9, s t r o n g l y r e a c t e d w i t h (Fab') 2 of the i m m Q n o g e n as w e l l as w i t h HBsAg, b u t not w i t h the c o n t r o l s . The b i n d i n g w a s s p e c i f i c a l l y i n h i b i t e d by p o l y A b u t not by m o n o m e r i c A , B S A or a n t i - H B s . F u r t h e r m o r e , 63.14 and 70.F9, s t a i n e d 4.10 and P L C / P R F / 5 c e l l s w i t h a p a t t e r n i d e n t i c a l to a n t i - H B s . In t h e s e i s t a n c e s the sta n i n g was also b l o c k e d by p o l y A, but n o t by m o n o m e r i c A , B S A or a n t i - H B s . On the o t h e r h a n d , t h e two a n t i - I d s f a i l e d to s t a i n n o r m a l l i v e r s e c t i o n s or the heDat ma cell line G2. T h e s e a n t i - I d s w i l l be v a l u a b l e in i n v e s t i g a t i n g the o a t h o g e n e sis of v i r a l h e p a t i t i s and c o u l d be U t i l i z e d in the p r e p a r a t i o n of a v a c c i n e . -
$42