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Actin in human renal cells
Biology of the Cell. Volume 63, 1988
STEREOLOGICAL APPROACH ON MACROPHAGES FROM HUMAN BRONCHOPULMONARY LAVAGE Frans VAN MEIR (I), Dietrich W. SCHEUERMANN (I), Marcel VAN DER STRAETEN (21, Romaln PAm¢ELS (21 and Marie H.A. DE GROODT-LASSEEL ( 1 ) . (11 I n s t i t u t e o f H i s t o l o g y and M i c r o s c o p i c Anatomy, U n i v e r s i t y of Antwerp (RUCAI, GroenenborgerlBan 171, B-2020 Antwerpen (Belgium), and (21 Department of Pneumology, U n i v e r s i t y H o s p i t a l , S t a t e U n i v e r s i t y of Ghent, De P t n t e l a a n 185, B-9000 Ghent (Belgium) To c h a r a c t e r i z e t h e p o p u l a t i o n o f a l v e o l a r maerophages in bronchopulmonary l a v a g e s , w i t h normal d i f f e r e n t i a l c e l l c o u n t s , ( a b o u t 90Z or more macrophages) o r i g i n a t i n g from p a t i e n t s w i t h a d i f f e r e n t c l i n i c a l h i s t o r y , a morphometrie a n a l y s t s o f t h e n u c l e o - c y t o p l a s m t c r a t i o and o f v a r i o u s c e l l e r g o h e l l a s is performed. A f t e r f i x a t i o n i n c o l d 2.5% N a - c a c o d y l a t e b u f f e r e d g l u t a r a l d e h y d e , p o s t f t x a t t o n i n 2% b u f f e r e d o s m t u m t e t r o x i d e and b l o c k s t a i n t n g i n 0.5% b u f f e r e d u r a n y l a c e t a t e , t h e p e l l e t i s embedded i n d u r c u pan-ACM r e s i n . A n u c l e a r b i a s e d sampling f o r macrophages i s used because i n t h i s way d i f f e r e n c e s i n the c e l l o r g a n e l l e s between t h e c e n t e r and p e r i p h e r y of t h e c e l l s a r e m i n i m i z e d . N e v e r t h e l e s s , i n about h a l f o f t h e c e l l s c u t t h r o u g h t h e n u c l e u s , no G o l g t a p p a r a t u s i s t r a n s s e c t i o n n e d . P r o f i l e a r e a s of c e l l s , n u c l e i and d i f f e r e n t c e l l o r g a n e l l e s , i . e . m i t o c h o n d r i a , lysosomes and lysosome l i k e b o d i e s , endoplasmtc r e t i c u l u m and Golgi a p p a r a t u s , a r e q u a n t i f i e d by p o i n t c o u n t i n g methods a p p l i e d on p r o j e c t e d p o s i t i v e f i l m s o f t h e macrophages a t a f i n a l m a g n i f i c a t i o n o f 25000 t i m e s . Our measurements of c e l l and n u c l e u s p r o f i l e - a r e a s p o i n t to a g r e a t v a r i e t y of c e l l forms i n t h e macrophage p o p u l a t i o n (87.8 + 36.9 sq ~m, r e s p . 14.2 + 6.6 sq ~m, nffi2481. To a s c e r t a i n w h e t h e r t h e s e k i n d s o f c e l l s a r e t h e e x p r e s s i o n o f d i f f e r e n t macrophage p o p u l a t i o n s or a r e o n l y an i n d i c a t i o n o f v a r i o u s s t a g e s o f macrophage a c t i v i t y , s p e c i a l a t t e n t i o n has been p a i d to t h e p r e s e n c e of p r i m a r y lysosomes and t h e v a c u o l a r s t r u c t u r e s t h a t are t h e s i t e of d i g e s t i v e a c t i v i t y ( s e c o n d a r y l y s o s o m e s ) . The f r e q u e n c y d i s t r i b u t i o n s o f l y s o s o m a l a r e a s per c e l l may a l l o w t o d i s t i n g u i s h between macrophages w i t h l e s s p h a g c c y t t c a c t i v i t y showing m a i n l y p r i m a r y lysosomes and macrophages which a f t e r an i n t e n s e p h a g o c y t i c a c t i v i t y c o n t a i n o b v i o u s l y more s e c o n d a r y l y s o s o m e s .
AMIODARONE-INDUCED INTRALYSOSOMAL LIPID STORAGE HIMIKING HEREDITARY LYSOSOMAL DISEASES. Edmondo GOVONI (2), Maria M. BRAGAGLIA (i), Bruno SEVERI (i) and Renzo LASCHI (i). (I) I ~ N ~ R of
Clinical Elect~on ~cro6copy, Univer6ity of Bologna, (2) I n s t i t ~ e of Pa~lology, UJ~v~rsity of ~lano, Italy. There are r e c e n t r e p o r t s of s e v e r e p e r i p h e r a l n e u r o p a t h y o c c u r r i n g in p a t i e n t s r e c e i v i n g amiodarone, a drug used b o t h as a c o r o n a r y d i l a t o r and to p r e v e n t s u p r a v e n t r i c u l a r and v e n t r i c u l a r a r r y t h m i a s . In t h i s r e s p e c t we s t u d i e d t h e muscle and p e r i p h e r a l nerve b i o p s i e s t a k e n from two a m i o d a r o n e - t r e a t e d patients with clinical and electrophysiological evidenc~ of a severe sensorymotor neuropathy. The neuromuscular symptoms occurred 28 days and 18 months respectively after the patients were started on a regime, of oral amiodarone L'ac~=py (600 rag/day). In the first patient a skin biopsy was also performed. In the second patient developing respiratory failure also a lung biopsy was carried out. Histological and histochemical study of the muscle biopsies disclosed features of denervation; electron microscopy showed subsarcolen~al clumps of hypertrophic lysosomes resembling myelinoid bodies. In nerve biopsies, c h a r a c t e r i s t i c changes of segmental d e m y e l i n a t i o n were v i s i b l e on s e m i t h i n s e c t i o n s a t l i g h t m i c r o s c o p y and on t e a s e d f i b r e p r e p a r a t i o n s , whereas f e a t u r e s of axonal d e g e n e r a t i o n were uncommon. E l e c t r o n microscopy c o n s t a n t l y showed a l o t of lysosomes c o n t a i n i n g g r a n u l a r a n d / o r m u l t i l a m e l l a r s t r u c t u r e s in the c y t o p l a s m of S c h w a n n c e l l s , f i b r o b l a s t s , e n d o t h e l i a l c e l l s and p e r i c y t e s . In the lung we o b s e r v e d numerous hypertrophi~ lysosomes loaded with densely packed lipid leaflets in the alveolar macrophages, type II pneumocytes, endothelial cells and septal histiocytes. Skin biopsy revealed large and numerous electrondense vacuoles in the cytoplasm of the endothelial cells of dermal capillaries as well as in the mesenchymal interstitial cells. These ultrastructural observations in the whole show that amiodarone-induTed lysosomal changes are quite llke those seen in hereditary lysosomal storage diseases.
ACTIN IN HI~iAN RENAL CELLS. Bruno SEVERI (I) , Edmondo GOVONI (2), Lucilla BADIALI (I) and
(1) IR6t.~J~e Of CIJ~R~.c~ EIRC~'LOnr~croseopy, u~,~vP.~t:.cj o~ Bolo~., (~) Z~.t:J.;Q~R of Pathologv, Univer~y of M~no, Itae V. Renzo LASCHI (1).
Bundles of f i l a m e n t s i n the c y t o p l a s m of r e n a l t u b u l e c e l l s have been so f a r d e s c r i b e d m a i n l y i n a n i m a l , more r a r e l y i n humans. T h e i r a c t l n n a t u r e i s p u r e l y d e d u c t i v e and p r o o f s about t h e i r m o l e c u l a r c o m p o s i t i o n a r e l a c k i n g . We employed an a n t i - a c t i n a n t i b o d y to t e s t the p r e s e n c e , a t the u l t r a s t r u c t u r a ] l e v e l (by the c o l l o i d a l gold t e c h n i q u e ) , of such a p r o t e i n w i t h i n tile f i l a m e n t b u n d l e s observed i n the k i d n e y c e l l s o f a case w i t h g l o m e r u l a r c r e s c e n t i c p r o l i f e r a t i o n . The same a n t i b o d y has been p r e v i o u s l y t e s t e d w i t h human myocardium as a c o n t r o l . Both t i s s u e s showed a c l e a r and s p e c i f i c d i s t r i b u t i o n o f the l a b e l . In the t u b u l a r e p i t h e l i u m the gold p a r t i c l e s m a i n l y d e c o r a t e d the b a s a l f i l a m e n t b u n d l e s , w h i l e o n l y a few d i s p e r s e d p a r t i c l e s were d e t e c t e d i n the n e i g h b o u r i n g c y t o p l a s m i c a r e a s . S i m i l a r f i l a m e n t o u s s t r u c t u r e s p o s i t i v e f o r a c t i n were found i n many e p i t h e l i a l c e l l s of t h e g l o m e r u l a r c r e s c e n t s . The c r e s c e n t i c c e l l s e x i b i t e d d i f f e r e n t t r a n s i t i o n a l s t a g e s from t y p i c a l e p i t h e l i a l p a r i e t a l c e l l s to myof i b r o b l a s t - l i k e c e ~ l s ( a c t i n p o s i t i v e ) and to f i b r o b l a s t - l l k e c e l l s ( i n t h e h i g h l y f i b r o t i c g l o m e r u l i ) . In o t h e r k i d n e y b i o p s i e s , we found s i m i l a r f i l a m e n t b u n d l e s i n d i f f e r e n t t u b u l a r segments and i n some e p i t h e l i a l c e l l s of the Bowman's c p a s u l e . Our o p i n i o n i s t h a t a c t i n f i l a m e n t s r e p r e s e n t a common f i n d i n g b o t h o f g l o m e r u l a r and t u b u l a r c e l l s and t h e i r number i n c r e a s e s d u r i n g impairment of t h e renal function.
6o
STEREOLOGICAL APPROACH ON MACROPHAGES FROM HUMAN BRONCHOPULMONARY LAVAGE Frans VAN MEIR (I), Dietrich W. SCHEUERMANN (I), Marcel VAN DER STRAETEN (21, Romaln PAm¢ELS (21 and Marie H.A. DE GROODT-LASSEEL ( 1 ) . (11 I n s t i t u t e o f H i s t o l o g y and M i c r o s c o p i c Anatomy, U n i v e r s i t y of Antwerp (RUCAI, GroenenborgerlBan 171, B-2020 Antwerpen (Belgium), and (21 Department of Pneumology, U n i v e r s i t y H o s p i t a l , S t a t e U n i v e r s i t y of Ghent, De P t n t e l a a n 185, B-9000 Ghent (Belgium) To c h a r a c t e r i z e t h e p o p u l a t i o n o f a l v e o l a r maerophages in bronchopulmonary l a v a g e s , w i t h normal d i f f e r e n t i a l c e l l c o u n t s , ( a b o u t 90Z or more macrophages) o r i g i n a t i n g from p a t i e n t s w i t h a d i f f e r e n t c l i n i c a l h i s t o r y , a morphometrie a n a l y s t s o f t h e n u c l e o - c y t o p l a s m t c r a t i o and o f v a r i o u s c e l l e r g o h e l l a s is performed. A f t e r f i x a t i o n i n c o l d 2.5% N a - c a c o d y l a t e b u f f e r e d g l u t a r a l d e h y d e , p o s t f t x a t t o n i n 2% b u f f e r e d o s m t u m t e t r o x i d e and b l o c k s t a i n t n g i n 0.5% b u f f e r e d u r a n y l a c e t a t e , t h e p e l l e t i s embedded i n d u r c u pan-ACM r e s i n . A n u c l e a r b i a s e d sampling f o r macrophages i s used because i n t h i s way d i f f e r e n c e s i n the c e l l o r g a n e l l e s between t h e c e n t e r and p e r i p h e r y of t h e c e l l s a r e m i n i m i z e d . N e v e r t h e l e s s , i n about h a l f o f t h e c e l l s c u t t h r o u g h t h e n u c l e u s , no G o l g t a p p a r a t u s i s t r a n s s e c t i o n n e d . P r o f i l e a r e a s of c e l l s , n u c l e i and d i f f e r e n t c e l l o r g a n e l l e s , i . e . m i t o c h o n d r i a , lysosomes and lysosome l i k e b o d i e s , endoplasmtc r e t i c u l u m and Golgi a p p a r a t u s , a r e q u a n t i f i e d by p o i n t c o u n t i n g methods a p p l i e d on p r o j e c t e d p o s i t i v e f i l m s o f t h e macrophages a t a f i n a l m a g n i f i c a t i o n o f 25000 t i m e s . Our measurements of c e l l and n u c l e u s p r o f i l e - a r e a s p o i n t to a g r e a t v a r i e t y of c e l l forms i n t h e macrophage p o p u l a t i o n (87.8 + 36.9 sq ~m, r e s p . 14.2 + 6.6 sq ~m, nffi2481. To a s c e r t a i n w h e t h e r t h e s e k i n d s o f c e l l s a r e t h e e x p r e s s i o n o f d i f f e r e n t macrophage p o p u l a t i o n s or a r e o n l y an i n d i c a t i o n o f v a r i o u s s t a g e s o f macrophage a c t i v i t y , s p e c i a l a t t e n t i o n has been p a i d to t h e p r e s e n c e of p r i m a r y lysosomes and t h e v a c u o l a r s t r u c t u r e s t h a t are t h e s i t e of d i g e s t i v e a c t i v i t y ( s e c o n d a r y l y s o s o m e s ) . The f r e q u e n c y d i s t r i b u t i o n s o f l y s o s o m a l a r e a s per c e l l may a l l o w t o d i s t i n g u i s h between macrophages w i t h l e s s p h a g c c y t t c a c t i v i t y showing m a i n l y p r i m a r y lysosomes and macrophages which a f t e r an i n t e n s e p h a g o c y t i c a c t i v i t y c o n t a i n o b v i o u s l y more s e c o n d a r y l y s o s o m e s .
AMIODARONE-INDUCED INTRALYSOSOMAL LIPID STORAGE HIMIKING HEREDITARY LYSOSOMAL DISEASES. Edmondo GOVONI (2), Maria M. BRAGAGLIA (i), Bruno SEVERI (i) and Renzo LASCHI (i). (I) I ~ N ~ R of
Clinical Elect~on ~cro6copy, Univer6ity of Bologna, (2) I n s t i t ~ e of Pa~lology, UJ~v~rsity of ~lano, Italy. There are r e c e n t r e p o r t s of s e v e r e p e r i p h e r a l n e u r o p a t h y o c c u r r i n g in p a t i e n t s r e c e i v i n g amiodarone, a drug used b o t h as a c o r o n a r y d i l a t o r and to p r e v e n t s u p r a v e n t r i c u l a r and v e n t r i c u l a r a r r y t h m i a s . In t h i s r e s p e c t we s t u d i e d t h e muscle and p e r i p h e r a l nerve b i o p s i e s t a k e n from two a m i o d a r o n e - t r e a t e d patients with clinical and electrophysiological evidenc~ of a severe sensorymotor neuropathy. The neuromuscular symptoms occurred 28 days and 18 months respectively after the patients were started on a regime, of oral amiodarone L'ac~=py (600 rag/day). In the first patient a skin biopsy was also performed. In the second patient developing respiratory failure also a lung biopsy was carried out. Histological and histochemical study of the muscle biopsies disclosed features of denervation; electron microscopy showed subsarcolen~al clumps of hypertrophic lysosomes resembling myelinoid bodies. In nerve biopsies, c h a r a c t e r i s t i c changes of segmental d e m y e l i n a t i o n were v i s i b l e on s e m i t h i n s e c t i o n s a t l i g h t m i c r o s c o p y and on t e a s e d f i b r e p r e p a r a t i o n s , whereas f e a t u r e s of axonal d e g e n e r a t i o n were uncommon. E l e c t r o n microscopy c o n s t a n t l y showed a l o t of lysosomes c o n t a i n i n g g r a n u l a r a n d / o r m u l t i l a m e l l a r s t r u c t u r e s in the c y t o p l a s m of S c h w a n n c e l l s , f i b r o b l a s t s , e n d o t h e l i a l c e l l s and p e r i c y t e s . In the lung we o b s e r v e d numerous hypertrophi~ lysosomes loaded with densely packed lipid leaflets in the alveolar macrophages, type II pneumocytes, endothelial cells and septal histiocytes. Skin biopsy revealed large and numerous electrondense vacuoles in the cytoplasm of the endothelial cells of dermal capillaries as well as in the mesenchymal interstitial cells. These ultrastructural observations in the whole show that amiodarone-induTed lysosomal changes are quite llke those seen in hereditary lysosomal storage diseases.
ACTIN IN HI~iAN RENAL CELLS. Bruno SEVERI (I) , Edmondo GOVONI (2), Lucilla BADIALI (I) and
(1) IR6t.~J~e Of CIJ~R~.c~ EIRC~'LOnr~croseopy, u~,~vP.~t:.cj o~ Bolo~., (~) Z~.t:J.;Q~R of Pathologv, Univer~y of M~no, Itae V. Renzo LASCHI (1).
Bundles of f i l a m e n t s i n the c y t o p l a s m of r e n a l t u b u l e c e l l s have been so f a r d e s c r i b e d m a i n l y i n a n i m a l , more r a r e l y i n humans. T h e i r a c t l n n a t u r e i s p u r e l y d e d u c t i v e and p r o o f s about t h e i r m o l e c u l a r c o m p o s i t i o n a r e l a c k i n g . We employed an a n t i - a c t i n a n t i b o d y to t e s t the p r e s e n c e , a t the u l t r a s t r u c t u r a ] l e v e l (by the c o l l o i d a l gold t e c h n i q u e ) , of such a p r o t e i n w i t h i n tile f i l a m e n t b u n d l e s observed i n the k i d n e y c e l l s o f a case w i t h g l o m e r u l a r c r e s c e n t i c p r o l i f e r a t i o n . The same a n t i b o d y has been p r e v i o u s l y t e s t e d w i t h human myocardium as a c o n t r o l . Both t i s s u e s showed a c l e a r and s p e c i f i c d i s t r i b u t i o n o f the l a b e l . In the t u b u l a r e p i t h e l i u m the gold p a r t i c l e s m a i n l y d e c o r a t e d the b a s a l f i l a m e n t b u n d l e s , w h i l e o n l y a few d i s p e r s e d p a r t i c l e s were d e t e c t e d i n the n e i g h b o u r i n g c y t o p l a s m i c a r e a s . S i m i l a r f i l a m e n t o u s s t r u c t u r e s p o s i t i v e f o r a c t i n were found i n many e p i t h e l i a l c e l l s of t h e g l o m e r u l a r c r e s c e n t s . The c r e s c e n t i c c e l l s e x i b i t e d d i f f e r e n t t r a n s i t i o n a l s t a g e s from t y p i c a l e p i t h e l i a l p a r i e t a l c e l l s to myof i b r o b l a s t - l i k e c e ~ l s ( a c t i n p o s i t i v e ) and to f i b r o b l a s t - l l k e c e l l s ( i n t h e h i g h l y f i b r o t i c g l o m e r u l i ) . In o t h e r k i d n e y b i o p s i e s , we found s i m i l a r f i l a m e n t b u n d l e s i n d i f f e r e n t t u b u l a r segments and i n some e p i t h e l i a l c e l l s of the Bowman's c p a s u l e . Our o p i n i o n i s t h a t a c t i n f i l a m e n t s r e p r e s e n t a common f i n d i n g b o t h o f g l o m e r u l a r and t u b u l a r c e l l s and t h e i r number i n c r e a s e s d u r i n g impairment of t h e renal function.
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