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The electron microscopic diagnosis of tumours - workshop
3
ANATOMICAL PATHOLOGY
RADIATICN-INDUCED SMALL BOWEL DIVEHTICLILA: A CASE REFOF3 AN UNUSUAL MANIFESTATION OF SElrERE RADIATION EWERITIS.
OF
J . Scurry*, T-_ Weerasiri, -~
R . Planner, P. Grant, S_ kung, Departments of Anatomical Pathology and Cncology, Mercy Hospital for Women, E a s t Melbourne, Vic 3002. A 43-year-old m n with stage IVa cervical c a r c i n m was treated by pelvic radiotherapy followed by t o t a l pelvic exenteration a f t e r a vesicovaginal f i s t u l a developed. Seven weeks post exenteration, an enterccutaneous fistula occurred, necessitating laparotany and p a r t i a l i l e a l resection. The resected seqnent of ileum s h m d dense adhesions, p a r t i a l obstruction and dilated thinned areas forming a s e r i e s of bubbles on the antimesenteric border. Microscopically, t h e "bubbles" were diverticula due to herniations of attenuated rnucosa and sutmucosa through transversely ruptured muscularis propria. ?he frayed ends of the muscularis shaved myocytolysis, microinfarcts and c e l l swelling, changes consistent with radiation damage. Following s m l l bowel resection, new diverticula and a jejunocolic f i s t u l a were readily identified on a b a r i m s m a l l bowel series. Massive gastrointestinal haemorrhage led t o death shortly thereafter.
THE ELECTRON MICROSCOPIC DIAGNOSIS OF TUMOURS
- WORKSHOP
B.A. Warren. Department of Pathology, University of New South Wales, Sydney, NSW Before examining a sample of tumour under the electron microscope the usual special and immunoperoxidasestains should be performed so as to arrive at a short differential diagnosis. Toluidine stained "thick" sections of the EM cell blocks are examined by the pathologist to see what has actually been taken. The indications for electron microscopic examination of tumours are: (1) to aid in elucidation of the diagnosis when modalities such as light microscopy and irnmunoperoxidase stains have failed to provide a definitive diagnosis; (2) to identify any distinguishing cytoplasmic organelles which are not resolvable under the light microscope; (3) to identify the nature and density of cytoplasmic organelles; (4) to identify specializations of the plasmalemmal membrane, especially the nature of cell junctions and filiform processes and (5) identification of melanosomes in obscure malignant melanomas. For adequate study appropriate selection of the region of the turnour to be taken is essential, i.e. preferably from the zone of the solid non-necrotic tumour. and not from the necrotic area of the tumour. The diagnosis of turnours is based on the nature of the cell of origin of the tumour. Knowledge of the other populations of cells likely to be encountered is necessary. Identification of malignant cells is basically based on cytological criteria for malignancy found in the nucleus. Once that is done the specific ultrastructural features of the cell of origin are examined. Structures which may be found on electron microscopy of tumour cells are tonofilaments. desmosomes. intracellular mucin, melanosomes, granules of the APUD turnours. granules of phaeochromocytoma. Birbeck granules, long filiform processes on the surface of the tumour cell, etc. There are characteristic cytoplasmic organelles for specific types of tumours.
Presenter: D r . J. S c u r r y , A n a t h c a l Pathology, MHW.
STUDY OF PCNA / CYCLIN EXPRESSION I N BENIGN, BORDERLINE L MALIGNANT EPITHELIAL TUMOURS OF OVARY Pathology, *Khoo JJ L Pathmanathan R., D e p a r t m e n t o f F a c u l t y o f M e d i c i n e , U n i v e r s i t y o f Malaya, 59100 K u a l a Lumpur, M a l a y s i a . C y c l i n , o r P r o l i f e r a t i n g c e l l n u c l e a r a n t i g e n (PCNA) i s a c e l l c y c l e r e l a t e d n u c l e a r p r o t e i n expressed i n inc r e a s e d l e v e l s during t h e l a t e G 1 a n d S phases o f DNA synthesis. P r e v i o u s workers have c o r r e l a t e d t h e degree of PCNA p o s i t i v i t y w i t h g r a d e and m i t o t i c a c t i v i t y i n s o l i d carcinomas, p a r t i c u l a r l y c a r c i n o m a o f t h e b r e a s t a n d c o l o n . We a t t e m p t e d t o c o r r e l a t e t h e d e g r e e o f PCNA a c t i v i t y i n malignant, b o r d e r l i n e and benign e p i t h e l i a l tumours o f t h e o v a r y w i t h a v i e w t o d e t e r m i n i n g i f t h i s might prove u s e f u l i n categorising these lesions. A t o t a l o f 51 c a s e s s e r o u s a n d mucinous o v a r i a n t u m o u r s d i a g n o s e d i n our d e p a r t m e n t were s t u d i e d . These comp r i s e d 17 s e r o u s ( b e n i g n = 5 ; b o r d e r l i n e = 5 : malignant = 7) and 34 m u c i n o u s ( b e n i g n = 5; b o r d e r l i n e = 15: m a l i g n a n t = 1 4 ) tumours. A l l l e s i o n s were r e v i e w e d typed a n d g r a d e d a c c o r d i n g t o e s t a b histologically, l i s h e d c r i t e r i a . PCNA e x p r e s s i o n was d e t e r m i n e d u s i n g a s t a n d a r d PAP t e c h n i q u e d e s c r i b e d p r e v i o u s l y . Slides w e r e e v a l u a t e d f o r PCNA p o s i t i v i t y a t a m a g n i f i c a t i o n o f X 160. The p e r c e n t a g e p o s i t i v i t y was d e r i v e d b y c o u n t i n g a t l e a s t 1000 e p i t h e l i a l c e l l s i n e a c h c a s e studied. PCNA p o s i t i v i t y was s i g n i f i c a n t l y h i g h e r i n m a l i g n a n t tunours o f o v a r y (X = 27.7%) when compared w i t h b o r d e r l i n e ( % = 2.2%) a n d b e n i g n (X = 1.0%) t u m o u r s . The r e s u l t s show t h a t PCNA a c t i v i t y i s a u s e f u l c r i t e r i o n i n d i f f e r e n t i a t i n g b e t w e e n b o r d e r l i n e and m a l i g n a n t e p i t h e l i a l tumours o f t h e ovary, especially in i n s t a n c e s when t h e h i s t o l o g i c a l d i a g n o s i s i s e q u i v o c a l .
AN OVERVIEW OF TRANSMISSION ELECTRON MICROSCOPY: PROCESSING, SECTIONING, TRANSMISSION ELECTRON MICROSCOPE OPERATION AND PHOTOGRAPHY S.E. Zimmerman. EM Unit, Anatomical Pathology, Prince Henry Hospital, Little Bay, NSW 2036. Electronmicroscopy has become an important adjunct in routine surgical diagnosis in many large laboratories.Many pathologists.particularly those who have no access to electron microscopy, are not familiar with the techniques involved in electron microscopy. This paper IS therefore aimed at providing the basic information for routine procedures for the preparation of materials for transmission electron microscopy in surgical pathology and research. The pathologistand the technician must accept equal responsibilityin obtaining the best possible result from the surgical material for the management of the patient There are a number of things that are to be followed in electron microscopic work. Chemicals used in EM are generally very toxic, many are carcinogenic and therefore must be handled safely and with care. Materials and equipment for EM work are comparatively more expensive than for routine histology and should only be used by experienced and trained staff. Tissues for examination are precious and may be rendered useless if improperly processed. Proper archival and retrieval records and logs must be kept to ensure a higher standard of practice. The consequences of poor practice may be an Incorrect diagnosis. The ability to make a correct diagnosis depends on the quality of the TEM section and the experience of the pathologist. It is thereforevitally important that all correct procedures for the preparation of good TEM sections be observed. This paper describes some of the technical procedures and has also highlighted some of the pitfalls associated with TEM. A diagnosis can not be made from poor sections, no matter how clever or experienced the pathologist is. Research material should be treated with equal care, as an enormous amount of work is involved in each step from processing to photography. Since good quality work is considered for publication, care must be taken when performing each of the technical steps involved in transmission electron microscopy. In the future, immunoelectron microscopy (an extension of TEM) will add a new dimension to our understanding of the pathogenesis of many diseases.
ANATOMICAL PATHOLOGY
RADIATICN-INDUCED SMALL BOWEL DIVEHTICLILA: A CASE REFOF3 AN UNUSUAL MANIFESTATION OF SElrERE RADIATION EWERITIS.
OF
J . Scurry*, T-_ Weerasiri, -~
R . Planner, P. Grant, S_ kung, Departments of Anatomical Pathology and Cncology, Mercy Hospital for Women, E a s t Melbourne, Vic 3002. A 43-year-old m n with stage IVa cervical c a r c i n m was treated by pelvic radiotherapy followed by t o t a l pelvic exenteration a f t e r a vesicovaginal f i s t u l a developed. Seven weeks post exenteration, an enterccutaneous fistula occurred, necessitating laparotany and p a r t i a l i l e a l resection. The resected seqnent of ileum s h m d dense adhesions, p a r t i a l obstruction and dilated thinned areas forming a s e r i e s of bubbles on the antimesenteric border. Microscopically, t h e "bubbles" were diverticula due to herniations of attenuated rnucosa and sutmucosa through transversely ruptured muscularis propria. ?he frayed ends of the muscularis shaved myocytolysis, microinfarcts and c e l l swelling, changes consistent with radiation damage. Following s m l l bowel resection, new diverticula and a jejunocolic f i s t u l a were readily identified on a b a r i m s m a l l bowel series. Massive gastrointestinal haemorrhage led t o death shortly thereafter.
THE ELECTRON MICROSCOPIC DIAGNOSIS OF TUMOURS
- WORKSHOP
B.A. Warren. Department of Pathology, University of New South Wales, Sydney, NSW Before examining a sample of tumour under the electron microscope the usual special and immunoperoxidasestains should be performed so as to arrive at a short differential diagnosis. Toluidine stained "thick" sections of the EM cell blocks are examined by the pathologist to see what has actually been taken. The indications for electron microscopic examination of tumours are: (1) to aid in elucidation of the diagnosis when modalities such as light microscopy and irnmunoperoxidase stains have failed to provide a definitive diagnosis; (2) to identify any distinguishing cytoplasmic organelles which are not resolvable under the light microscope; (3) to identify the nature and density of cytoplasmic organelles; (4) to identify specializations of the plasmalemmal membrane, especially the nature of cell junctions and filiform processes and (5) identification of melanosomes in obscure malignant melanomas. For adequate study appropriate selection of the region of the turnour to be taken is essential, i.e. preferably from the zone of the solid non-necrotic tumour. and not from the necrotic area of the tumour. The diagnosis of turnours is based on the nature of the cell of origin of the tumour. Knowledge of the other populations of cells likely to be encountered is necessary. Identification of malignant cells is basically based on cytological criteria for malignancy found in the nucleus. Once that is done the specific ultrastructural features of the cell of origin are examined. Structures which may be found on electron microscopy of tumour cells are tonofilaments. desmosomes. intracellular mucin, melanosomes, granules of the APUD turnours. granules of phaeochromocytoma. Birbeck granules, long filiform processes on the surface of the tumour cell, etc. There are characteristic cytoplasmic organelles for specific types of tumours.
Presenter: D r . J. S c u r r y , A n a t h c a l Pathology, MHW.
STUDY OF PCNA / CYCLIN EXPRESSION I N BENIGN, BORDERLINE L MALIGNANT EPITHELIAL TUMOURS OF OVARY Pathology, *Khoo JJ L Pathmanathan R., D e p a r t m e n t o f F a c u l t y o f M e d i c i n e , U n i v e r s i t y o f Malaya, 59100 K u a l a Lumpur, M a l a y s i a . C y c l i n , o r P r o l i f e r a t i n g c e l l n u c l e a r a n t i g e n (PCNA) i s a c e l l c y c l e r e l a t e d n u c l e a r p r o t e i n expressed i n inc r e a s e d l e v e l s during t h e l a t e G 1 a n d S phases o f DNA synthesis. P r e v i o u s workers have c o r r e l a t e d t h e degree of PCNA p o s i t i v i t y w i t h g r a d e and m i t o t i c a c t i v i t y i n s o l i d carcinomas, p a r t i c u l a r l y c a r c i n o m a o f t h e b r e a s t a n d c o l o n . We a t t e m p t e d t o c o r r e l a t e t h e d e g r e e o f PCNA a c t i v i t y i n malignant, b o r d e r l i n e and benign e p i t h e l i a l tumours o f t h e o v a r y w i t h a v i e w t o d e t e r m i n i n g i f t h i s might prove u s e f u l i n categorising these lesions. A t o t a l o f 51 c a s e s s e r o u s a n d mucinous o v a r i a n t u m o u r s d i a g n o s e d i n our d e p a r t m e n t were s t u d i e d . These comp r i s e d 17 s e r o u s ( b e n i g n = 5 ; b o r d e r l i n e = 5 : malignant = 7) and 34 m u c i n o u s ( b e n i g n = 5; b o r d e r l i n e = 15: m a l i g n a n t = 1 4 ) tumours. A l l l e s i o n s were r e v i e w e d typed a n d g r a d e d a c c o r d i n g t o e s t a b histologically, l i s h e d c r i t e r i a . PCNA e x p r e s s i o n was d e t e r m i n e d u s i n g a s t a n d a r d PAP t e c h n i q u e d e s c r i b e d p r e v i o u s l y . Slides w e r e e v a l u a t e d f o r PCNA p o s i t i v i t y a t a m a g n i f i c a t i o n o f X 160. The p e r c e n t a g e p o s i t i v i t y was d e r i v e d b y c o u n t i n g a t l e a s t 1000 e p i t h e l i a l c e l l s i n e a c h c a s e studied. PCNA p o s i t i v i t y was s i g n i f i c a n t l y h i g h e r i n m a l i g n a n t tunours o f o v a r y (X = 27.7%) when compared w i t h b o r d e r l i n e ( % = 2.2%) a n d b e n i g n (X = 1.0%) t u m o u r s . The r e s u l t s show t h a t PCNA a c t i v i t y i s a u s e f u l c r i t e r i o n i n d i f f e r e n t i a t i n g b e t w e e n b o r d e r l i n e and m a l i g n a n t e p i t h e l i a l tumours o f t h e ovary, especially in i n s t a n c e s when t h e h i s t o l o g i c a l d i a g n o s i s i s e q u i v o c a l .
AN OVERVIEW OF TRANSMISSION ELECTRON MICROSCOPY: PROCESSING, SECTIONING, TRANSMISSION ELECTRON MICROSCOPE OPERATION AND PHOTOGRAPHY S.E. Zimmerman. EM Unit, Anatomical Pathology, Prince Henry Hospital, Little Bay, NSW 2036. Electronmicroscopy has become an important adjunct in routine surgical diagnosis in many large laboratories.Many pathologists.particularly those who have no access to electron microscopy, are not familiar with the techniques involved in electron microscopy. This paper IS therefore aimed at providing the basic information for routine procedures for the preparation of materials for transmission electron microscopy in surgical pathology and research. The pathologistand the technician must accept equal responsibilityin obtaining the best possible result from the surgical material for the management of the patient There are a number of things that are to be followed in electron microscopic work. Chemicals used in EM are generally very toxic, many are carcinogenic and therefore must be handled safely and with care. Materials and equipment for EM work are comparatively more expensive than for routine histology and should only be used by experienced and trained staff. Tissues for examination are precious and may be rendered useless if improperly processed. Proper archival and retrieval records and logs must be kept to ensure a higher standard of practice. The consequences of poor practice may be an Incorrect diagnosis. The ability to make a correct diagnosis depends on the quality of the TEM section and the experience of the pathologist. It is thereforevitally important that all correct procedures for the preparation of good TEM sections be observed. This paper describes some of the technical procedures and has also highlighted some of the pitfalls associated with TEM. A diagnosis can not be made from poor sections, no matter how clever or experienced the pathologist is. Research material should be treated with equal care, as an enormous amount of work is involved in each step from processing to photography. Since good quality work is considered for publication, care must be taken when performing each of the technical steps involved in transmission electron microscopy. In the future, immunoelectron microscopy (an extension of TEM) will add a new dimension to our understanding of the pathogenesis of many diseases.