DNA and oncoprotein in cervical specimens 581 20Hann SR, Thompson CB, Eisenman RN. C-myc oncogene protein 23Makino R, Hayashi KA, Sugimura T. C-myc is induced in rat liver is independent of the cell cycle in human and avian cells. Nature at a very early stage of regeneration or by cycloheximide treat- 1985;314:366-9. ment. Nature 1984;310:697-8. Kelly K, Cochran BH, Stiles CD, Leder P. Cell specific regulation of the c-myc gene by lymphocyte mitogens and platelet derived growth factor. Cell 1983;35:603-10. Kelly K, Cochran BH, Stiles CD, Leder P. The regulation ofc-myc Requests for reprints to: Dr JV Watson, MRC Clinical by growth signals. Curr Tops Microbiol Immunol Oncology and Radiotherapy Unit, Medical Research 1984;133:1 17-26. Council Centre, Hills Road, Cambridge CB2 2QH, England.

Letters to the Editor ation. In systemic primary and myeloma an endocrine organ, and peptides arising associated amyloid the fibrils contain pro- from the atria have been isolated and char- tein derived from immunoglobulin light acterised. They have potent natriuretic, Identification of imnunoreactive atrial chains. In secondary amyloid the fibrils con- diuretic, and vasodilating actions and may natriuretic peptide in atrial amyloid tain a protein related to the acute phase play an important part in the homeostasis of reactant serum amyloid A. In hereditary or body fluids.89 In this report we describe the Amyloidosis is a disorder characterised by familial amyloidosis and in cerebral amyloid immunohistochemical localisation of the deposition of an abnormal pro- of Alzheimer's disease they contain pre- human atrial natriuretic peptide to amyloid teinaceous material in the extracellular tis- albumin. Amyloid localised to endocrine fibrils in human atrial appendage, indicating sues.1 2 It may occur as a primary disease organs may contain hormone related that some cardiac amyloid is analogous to process or secondary to a variety of peptides-for example, immunoreactive cal- that seen in other endocrine organs. conditions characterised by chronic citonin is found in amyloid associated with inflammation. It has been described in asso- medullary carcinoma of the thyroid.3 s Material and methods ciation with tumours and occurs in heredo- Amyloid affects the heart as part of sys- familial forms. It occurs in a localised or sys- temic amyloidosis or in a localised form as a A fresh piece of right atrial appendage temic distribution. manifestation of aging.6 Two types occur in removed at coronary bypass surgery was Amyloid has characteristic staining reac- the aging heart: that affecting both the ven- fixed in paraformaldehyde lysine sodium tions, but is best defined by its appearance tricles and atria and that affecting the atria periodate'° for 24 hours. It was then divided on electron microscopy where fine non- alone, known as isolated atrial amyloid.7 to provide blocks for routine paraffin histol- branching fibrils about 10 nm in diameter Isolated atrial amyloid has been reported in ogy and electron microscopy. Paraffin sec- can be identified. Despite the similarity of 78% of patients over the age of 80 years: its tions were cut at 4 um and stained with appearance the chemical structure of the origin is unknown. haematoxylin and eosin, sulphated alcian fibrils varies with site and disease associ- Recent evidence suggests that the heart is blue, and alkaline Congo red for amyloid. 1I

Fig. Ia Ekectron micrograph ofhwnan atrial appendage Fig. Ib Electron micrograph ofhuman atrial appendage showing labelling ofamyloidfibrilsfor human atrial showing labelling ofneurosecretary granulesfor human atrial natriuretic peptide (Immunogoldl uranyl acetate lead citrate natriuretic peptide. (Immunogold/uranyl acetate lead citrate, lSnmgoldballs.) x 56000. 15 nm gold balls.) x 112000. 582 Letters

They were also stained by the indirect peptide in atrial amyloid fibrils, which can the thyroid: an immunogold technique. Arch immunoperoxidase technique for atrial Pathol Lab Med (in press). be seen on electron microscopy, and have 6Pomerance A. Senile cardiac amyloidosis. Br natriuretic peptide.12 Tissue for electron also confirmed its presence within neu- microscopy was embedded in LR white Heart J 1965;27:711-19. rosecretory granules in atrial cardiocytes.5 7Cornwell GB III, Murdoch WL, Kyle RA, West- resin, and ultrathin sections were stained by These findings support the view that some ermark P, Pitkanen P. Frequency and distri- indirect immunochemistry for atrial atrial amyloid is of an endocrine type and bution of senile cardiovascular amyloid. Am J natriuretic peptide, using goat antirabbit suggest that atrial natriuretic peptide, or an Med 1983;75:618-23. immunoglobulin conjugated to 15 nm collo- atrial natnuretic related peptide, may be its 8Cantin M, Genest J. The heart and atrial na- idal gold (Janssen Pharmaceutical) as the primary constituent. triuretic factor. Endocrine Review second antibody.5 The antibodies to atrial 1985;6:107-27. The amyloid was only visualised ultra- 9Maack T, Camargo MJF, Kleinert HD, Laragh natriuretic peptide were raised in rabbits structurally and was not evident on light using synthetic atrial natriuretic peptide JH, Atlast SA. Atrial natriuretic factor: struc- microscopy. This phenomenon is not ture and functional properties. Kidney Int conjugated to bovine thyroglobulin as the uncommon if deposits are small."6 In this 1985;27:607-15. immunogen. case it was therefore impossible to confirm °0McLean I, Nakane PK. Periodate-lysine- the ultrastructural observations at light parformaldehyde: a new fixative for immuno- Results microscopy level, although there was a little electron microscopy. J Histochem Cytochem perivascular positivity for atnal natriuretic 1974;22: 1077-83. Ultrastructural examination of the atrial tis- Pomerance A, Slavin G, McWatt JJ. Experience peptide on indirect immunoperoxidase with the sodium sulphate-alcian blue stain for sue showed the presence of fine deposits of staining. As deposits of isolated atrial amy- amyloid in cardiac pathology. J Clin Pathol amyloid fibrils 7-10 nm in diameter around loid increase with age and the patient stud- 1976;29:22-263. small blood vessels and adjacent to muscle ied was only 64 years old, it may be possible 12Bums J. Immunohistological methods and their cells. The amyloid fibrils clearly and consis- to show amyloid atrial natnuretic peptide application in the routine laboratory. In: An- tently labelled for immunoreactive atrial on light microscopy more readily in older thony PP, Woolf N, eds. Recent advances in natriuretic peptide (Fig. la), whereas back- patients. Further studies are currently being histopathology. Edinburgh: Churchill Liv- ground labelling of the surrounding con- undertaken. ingstone, 1978:337-50. nective tissue was negligible. The presence of 13Pearse AGE, Ewen SWB, Polak JM. The gene- atrial natriuretic peptide was also shown in GC KAYE* sis of apudamyloid in endocrine polypeptide MG BUTLER tumours: histochemical distinction from im- neurosecretory granules within atrial muscle munoamyloid. Virchows Arch (Pathol Anat) cells (Fig. lb). Immunoperoxidase staining AJ D'ARDENNE 1972;10:93-107. for atrial natriuretic peptide by light micros- SJ EDMONDSON* Westermark P, Johansson B, Natvig JB. Senile copy was positive in atrial muscle cells and AJ CAMM* cardiac amyloidosis: evidence of two different focally positive around small blood vessels. G SLAVIN amyloid substances in the ageing heart. Scand Reaction with preabsorbed antiatrial Department of Histopathology J Immunol 1979;10:303-8. natriuretic peptide was consistently nega- and *Cardiology, Chapeau C, Gutkowska J, Schiller PW, Milne tive. The amyloid was not detected by light St Bartholomew's Hospital, RW, Thibault G, Garcia R, Genest J, Cantin West Smithfield, M. Localisation of immunoreactive synthetic microscopy, and the Congo red stain was atrial natriuretic factor (ANF) in the heart of negative. London ECIA 7BE various animal species. J Histochem Cyto- chem 1985;33:541-50. Discussion 16Pomerance A. Infiltrations and storage diseases. In: Pomerance A, Davies MJ, eds. The pathol- Amyloid has been reported in various ogy of the heart. Oxford: Blackwell Scientific endocrine tumours, including medullary Publications, 1975:256. carcinomas of the thyroid, carcinoids, phae- ochromocytomas, carotid body tumours, References insulinomas, and gastrinomas.13 Endocrine associated amyloid differs from other types Skinner M, Cohen AS. Amyloidosis: clinical, pa- in that histochemical staining for tryp- thologic and biochemical characteristics. In: tophan is negative. Moreover, in some cases Wagner BM, Fleischmajor R, Kaufman N, of endocrine associ- eds. Connective tissue diseases. Baltimore: the constituent protein Williams and Wilkins, 1983:97-119. ated amyloid has been shown to be related 2 Kisilevsky R. Amyloidosis; a familiar problem in Manual or radiometric method for detection to the native hormone product.3 the light of current pathologic developments. of bacteraemia Studies on the cardiac amyloid associated Lab Invest 1983;49:381-90. with aging have subdivided it into two types, 3Sletten K, Westermark P, Natvig JB. Character- We read with interest the paper by Ganguli which differ both in chemical composition isation of amyloid fibril protein from medul- et ali that a manual rather than a radio- and distribution.14 Isolated atrial amyloid is lary carcinoma of the thyroid. J Exp Med metric method was more rapid for the similar to endocrine associated amyloid in 1976:143:993-8. detection of bacteraemia. In this laboratory lacks demonstrable 4Dammwich J, Ormanns W, Schaffer K. Electron we compared the radiometric that it histochemically microscopic demonstration of calcitonin in technique with tryptophan, whereas senile cardiac amyloid human medullary carcinoma of the thyroid by the standard method of blood culture exam- found in both atria and ventricles contains the immunogold staining method. Histo- ination.2 No difference between the twc readily demonstrable tryptophan. chemistry 1984;81:369-72. techniques was found. The isolation rate In this report we describe the presence of 'Butler MG, Khan S. Immunoreactive calcitonin was 4% for each method when 5216 blood immunoreactive human atrial natriuretic in amyloid fibrils of medullary carcinoma of cultures were examined by the manual