Kow2 Observed These Myocardial Changes in Reaction to Celloidin Con- Taining Foreign Bodies

OCCURRENCE OF CATERPILLAR NUCLEI WITHIN NORMAL IMMATURE AND NORMAL APPEARING AND ALTERED MATURE HEART MUSCLE CELLS AND THE EVOLUTION OF ANITSCHKOW CELLS FROM THE LATTER

GEORGE E. MuRPiHY, M.D., and Cma G. BECKE, M.D. From the Department of Pathology, The New York Hospital-CorneU Medical Center, New York, N.Y. In I9OI von Oppell reported changes that occurred in rabbit myocardium in reaction to implantation of a sewing needle. Twenty-four hours after the implantation frank necrosis of muscle fibers was evident in the puncture canal. A little distant, less altered muscle fibers had lost striations and were coarsely granular. In their enlarged elliptic nuclei the chromatin had become distributed in granules or clusters of granules or condensed in a "single thread running in the long axis" of the nucleus. Among these degenerating muscle fibers, cells ("free cells") appeared with nuclei like those in the altered muscle fibers. Subsequently Anitsch- kow2 observed these myocardial changes in reaction to celloidin containing foreign bodies. He emphasized the condensation of chromatin into a "serrated stripe" in nuclei of some of the altered muscle fibers at a distance from the puncture canal and in nuclei of some of the cells ("myocytes") that appeared among the altered muscle fibers. These "free cells" or "myocytes" have come to be known as Anitschkow cells or myocytes. Because such elements are often prominent components of Aschoff bodies, the myocardial lesions characteristic of rheumatic heart disease, they are sometimes referred to as Aschoff cells. The chromatin structure of Anitschkow cells is known to occur in embryonic and postembryonic hearts in members of all classes of verte- brates,3'4 and is one of the most distinctive nuclear forms. From the central axial body of chromatin very fine fibrillar extensions radiate to- ward and sometimes to the nuclear membrane. In longitudinal section this chromatin pattern resembles the structure of a caterpillar. In transverse section the nuclear appearance has been likened to an owl eye. The cytoplasm of Anitschkow cells is usually scant and in many of the cells is barely or not detectable when they are observed with the light micro- scope, even when magnified as much as I,200 times. This investigation was supported by research grant HE-o0803 of the National Heart Institute of the National Institutes of Health, a grant of The Cross Foundation, and training grant 2G-78 of the Division of General Medical Sciences of the United States Public Health Service. Accepted for publication, February s, I966. 93' 932 MURPHY AND BECKER Vol. 48., No. 6 From experiment and histologic study conflicting opinions have arisen concerning the origin of Anitschkow cells.4'5 It is widely believed that they are histiocytes or reticuloendothelial cells never derived from muscle cells of the heart.8''7 It is also widely held that nuclei with structure like that characteristic of Anitschkow cells never occur in muscle cells of the heart. In contrast, such nuclei have been reported to occur in striated heart muscle cells5,8'9 and in fragments thereof and in smooth muscle cells of the heart.5'10 Others6'11 have interpreted these nuclei as residing not within striated muscle cells but within non- myogenic cells juxtaposed to the muscle cells. The purpose of the present communication is to demonstrate I) the occurrence of nuclei identical in structure to those characteristic of Anitschkow cells within striated muscle cells of the human heart, and 2) the evolution of Anitschkow cells from heart muscle cells. MATERIAL AND METHODS Sources and Treatment of Material Examined. Myocardia of 56 patients were examined. Fourteen were immature infants, 7 of which weighed between 390 and 699 gm and 7 between 700 and 999 gm at the time of death within several days after birth. Six were premature infants who weighed between I,ooo and 2,499 gm at the time of death i hour to 30 days after birth. In the majority of these immature and premature infants birth was preceded by spontaneous rupture of the fetal membranes or by premature separation of the placenta. In I premature there were multiple cardiac anomalies. In the remaining I9 prematures and all the immatures the hearts were anatomically normal. Ten patients were full-term infants who weighed between 2,900 and 3,600 gm at the time of death in the neonatal period. In 4 of these there were cardiac anomalies, in 2, anomalies of the brain or urinary tract, and in 2 others subdural hemorrhage occurred. Of the remaining 2 full-term infants one died of meningitis due to Hemophilus influenzae at 3 weeks of age. The other died with widespread petechial hemorrhages i day after birth. Six weeks after the beginning of gestation 2 other children of the mother of this infant developed rubella. The mother developed no signs or symptoms of rubella. From the myocardium and many other tissues of this infant rubella virus was cultured at necropsy. Sixteen of the patients died of active rheumatic heart disease; and numerous Aschoff bodies were found in the myocardium in every case. The ages of the rheumatic patients were as follows: 3 were from 14 months to 5 years, 6 from 6 to I0 years, 4 from 14 to 25 years, and 3 from 33 to 6o years. Another patient died at 29 years of age with bilateral salpingitis, pelvic abscess, and peritonitis; an anaerobic streptococci and E. coli were cultured from peritoneal exudate. An additional patient died at 37 years of age of cardiac failure caused by widespread degenerative changes in the heart muscle. The remaining 8 patients died of myocardial infarction one day to 3 weeks after onset of chest pain at ages ranging from 54 to 68 years. Two of this latter group suffered with angina pectoris for about 3 years before death. An additional 2 of this group had experienced myocardial infarction several years previously. Blocks of myocardium were fixed in Zenker's, 5 per cent formalin solution, and sections cut at approximately s,u were stained with hematoxylin and eosin or by the Masson trichrome technique. Optical Equipment. A Zeiss photomicroscope was used with a Zeiss iooX plana- chromat objective lens with oil for examination, photography at magnification of 400 to 1,260 X, for all measurements of striated heart muscle fibers and nuclei with lune, z966 CATERPILLAR NUCLEI 933 caterpillar-like intranuclear structure. We are informed by Carl Zeiss, Inc., that they have calculated the depth of field in focus to be less than i g (0.4 to 0.7 ,) when this lens is used for magnification in the range stated above. Measurement of Heart Muscle Fibers and Caterpillar Nuclei and their Axial Chromatin Bars. The diameters of striated cardiac muscle fibers and caterpillar nuclei and their axial chromatin bars were measured with a Zeiss ocular micrometer. Diameters were only measured when transverse striations of the muscle fibers and the membranes and chromatin bars of caterpillar nuclei were sharply and simultaneously in focus. Diameters of striated muscle fibers containing either caterpillar nuclei or ordinary nuclei were measured at a point 4 Ai to 8 ,u away from any nuclei of those muscle fibers. RESULTS In all of the hearts examined, except those with myocardial infarcts, caterpillar or owl eye nuclei identical to those characteristic of Anitsch- kow cells were present in immature or mature striated heart muscle cells or in fragments of the latter. Measurement was made of the transverse diameters of 40 of these nuclei and the transverse diameters of their axial chromatin bars in each of 2 hearts, I from a 14-month-old infant and the other from a 3-year-old child who died between 2 and 3 weeks after the first signs and symptoms of the first evident attack of rheumatic fever. Diameters were only measured when transverse striations of the muscle fiber, the membrane of the caterpillar nucleus, and the axial chromatin bar of the nucleus were all simultaneously and sharply in focus, as illustrated in Figures I9 to 2 2. The arithmetic mean of the diameters of the nuclei was 4.64,P (standard deviation, -+- o.86 u) and that of the axial chromatin bars was I.83 ,u (standard deviation, ± 0.25 ,). If these caterpillar nuclei resided outside the muscle fibers, then the average distance between the center of the nucleus (or center of the axial bar) and the surface of the muscle fiber would be at least 2.32 /. Because this distance is between 3 to 6 times the depth of field in focus (0.4 to 0.7 ) with the lens used, it would not be possible to have the membrane of the nucleus, the axial chromatin bar of the nucleus, and striations of the muscle fiber all sharply in focus in the same plane. Yet, all of these elements could be seen to be sharply in focus in the same plane (Figs. I9 to 22). The hypothesis that these nuclei resided outside the muscle fibers is, there- fore, not supported by the data. The alternative hypothesis, definitively supported by the above measurements, is that these caterpillar nuclei resided within the muscle fibers. The latter hypothesis is further supported by the occurrence of owl eye nuclei, which are transversely sectioned caterpillar nuclei, well within the cytoplasm of myoblasts in the myotube stage of differentiation in an immature infant (Fig. 5) and within heart muscle cells in a I4-month-old infant (Fig. 4I) who died of active rheumatic heart disease. 934 MURPHY AND BECKER Vol. 48, No. 6

Observations were made on the incidence and location of nuclei with fully formed caterpillar-like chromatin pattern in all of the 56 myocardia examined (Table I). One section of the left ventricular myocardium in each of the immature, premature and full-term infants, and two or more, usually several sections of the left ventricular myocardium, in each of the children and adults were examined many times in entirety. The myocardia were graded relatively to each other as to frequency of

TABLE I INCIDENCE AND LOCATION OF NUCLEI WITH FULLY FORM CATERPILLAR CHROMATIN PATTERN IN 56 HUMAN MYOCARDIA OF D RENT AGES AD CONDIIONS Number In myotubes * In fragments of or non-fragmented of In Age group Condition hearts striated striated Anitschkow of patients of patients examined muscle fibers muscle fibers cells Immature infants, Normal 14 +++++ 0 + 390-999 gms

Premature infants, Normal, pulmonary 6 ++++ o +-++ 1000-2499 gms hyaline membrane disease, or congenital heart disease

Full-term infants, Cardiac or other 8 +++ o +-++ 250o-3600 gms anomalies or subdural hemorrhage

Full-term infants, H. influenzae men- 2 +++++ +-++ ++-+++ 2710-3780 gms ingitis or congenital rubella

Infant and children, Active rheumatic 4 +++++ +++++ +++++ I4 mos-5 yrs heart disease

Children, Active rheumatic +++++ +++++ 6-io yrs heart disease

Adolescents and Active rheumatic 4 +++-o +++++ +++++ young adults, heart disease I4-25 yrs

Adults, Pelvic abscess with 2 +±+ +±++ 29-37 yrs peritonitis or widespread myocardial degeneration

Adults, Active rheumatic 0 +±+ +++ 33-6o yrs heart disease

Adults, Myocardial 8 0 0 54-78 yrs infarction * Myotube stage of heart muscle development in immature and premature infants only. Legend: +++++ - frequent, focally common; ++++ - frequent; +++ - moderate number; ++ - occasional; + - rare; o - not found. June, z966 CATERPILLAR NUCLEI 935 caterpillar nuclei within heart muscle cells, fragments thereof, and Anitschkow cells. Taking frequent, focally common occurrence as the upper standard for comparison, the incidence of caterpillar nuclei in these three sites in each heart was recorded as one of the following: frequent, focally common (+++++); frequent (++++); moderate number (+++); occasional (++); rare (+); not found (o). In the myocardia of all of the 14 immature infants caterpillar or owl eye nuclei were present frequently, in some foci commonly, in myoblasts in the myotube stage of development and in immature striated muscle cells (Figs. i to 8), and Anitschkow cells were found rarely. In the myocardia of the 6 prematures such nuclei were present frequently in immature striated muscle cells and striated muscle cells that appeared mature (Figs. 9 and io), and Anitschkow cells were encountered rarely or occasionally. Caterpillar or owl eye nuclei were present in moderate number in striated muscle cells and Anitschkow cells were only rarely or occasionally found in the myocardia of the 8 full-term infants with cardiac or other anomalies or subdural hemorrhage. In contrast, in the myocardia of the 2 full-term infants who died of meningitis due to H. influenzae and congenital rubella, respectively, such nuclei were present frequently, in some foci commonly, in striated muscle cells (Figs. II, 12, 14 to I6, and 53). In these 2 hearts rare or occasional myofiber fragments with such nuclei were found. Anitschkow cells were encountered occasionally or in moderate number. In the myocardia of the 4 patients with rheumatic fever who were I4 months to 5 years of age caterpillar or owl eye nuclei occurred frequently, in some foci commonly, in striated muscle cells that were apparently intact (Figs. I9 to 22) or degenerating (Fig. I7). Such nuclei were present in moderate number in striated muscle cells that were apparently intact (Fig. 13) or degenerating (Fig. i8) in the myocardia of the 5 children with rheumatic fever who were 6 to io years old. In all 9 of these young rheumatic patients caterpillar nuclei were present frequently, in some foci commonly, in fragments of muscle cells (Figs. 34 to 37, and 40), and Anitschkow cells were likewise present frequently, in some foci commonly. In the myocardia of the group of 4 patients with rheumatic fever who were I4 to 25 years of age the frequency of caterpillar or owl eye nuclei in apparently intact or degenerating striated muscle cells ranged from moderate number in a 14-year-old female and a 20-year-old male to absence in a I 7-year-old female and a 25-year-old female. In the myocardia of all of the 4 patients in this group, however, such nuclei in muscle fragments (Fig. 47) were present frequently, in some foci com- 936 MURPHY AND BECKER Vol. 48, No. 6 monly, and Anitschkow cells were likewise present frequently, in some foci commonly. A fully formed caterpillar or owl eye nucleus was found in only i heart muscle cell, obviously degenerating, in the 29-year-old woman with pelvic abscess and peritonitis (Figs. 32 and 33), and only very rarely in heart muscle cells, obviously degenerating, in the 37-year-old woman who died of cardiac failure due to widespread degenerative changes in heart muscle. Such nuclei were found in none of the heart muscle cells in the 3 patients with active rheumatic heart disease who were 33 to 6o years of age. Such nuclei were found in moderate number in fragments of muscle cells (Figs. 32, 33, 38, 39, 43 and 51), however, and Anitschkow cells were likewise present in moderate number in the myocardia of all these 5 patients. Numerous Aschoff bodies were present in the myocardia of all of the I6 patients who died with active rheumatic heart disease (Figs. 42, 45 and 48). In these lesions the frequency of caterpillar nuclei in muscle fragments, many of which had scant cytoplasm, appeared to be of about the same order as that of Anitschkow cells among striated muscle cells elsewhere in the myocardium. In contrast with the incidence of caterpillar or owl eye nuclei in the 48 myocardia described above, such nuclei were not found in either striated muscle cells or fragments thereof in or about infarcted or normal appearing myocardium in the 8 patients with myocardial infarction, some of whom had suffered protracted angina pectoris or previous myocardial infarction. These patients died of myocardial infarction i day to 3 weeks after the onset of chest pain. Furthermore, in these myocardia Anitschkow cells were present in normally rare number, irrespective of proximity to areas of infarction. The foregoing observations strongly suggest that certain injuries to heart muscle cells induce development of the caterpillar-like chromatin pattern characteristic of the Anitschkow cells in their nuclei and emer- gence of Anitschkow cells from the injured muscle cells. In testing this hypothesis measurement and comparison were made of the diameters of striated muscle fibers containing caterpillar nuclei and those containing ordinary nuclei in the hearts of a I4-month-old infant and a 3-year- old child who died within 2 to 3 weeks after the onset of a first known attack of rheumatic fever. The results are shown in Table II. The average diameter of striated muscle fibers containing ordinary nuclei was very significantly greater than the average diameter of striated muscle fibers containing caterpillar nuclei. This data indicates that the latter cells are either atrophic or budding myofibers. Progression of degenerative change to fragmentation of muscle cells with fragments containing June, z966 CATERPILLAR NUCLEI 937 A% A A A A 8.

0. a 0) U)

u4~ co 0*

tlC!5. I!Bi 0

0n S.0 11 0% +* _ Pm GUS Y X 1 0 0 I4 51) _+* Q @ 3 511) Co Hd > *Ii 0 z.. + o a oP. 4. soU)0 ot S 000 o~

5.) 5 Co II ax o) CX 54,X0)5. CGi.5 >

cd >s 0 8 00*

C6b0

0 0 El

o04 938 MURPHY AND BECKER Vol. 48, No. 6 caterpillar nuclei in these and many other hearts, and the concomitant evolution of Anitschkow cells from damaged heart muscle cells or their fragments are illustrated in Figures 23 to 49. Aschoff bodies in which some of the fragments of heart muscle cells contain owl eye or caterpillar nuclei are shown in Figures 42,45 and 48. Some of these fragments have scant cytoplasm and are apparently completing evolution into Anitschkow cells. In Figures 46 and 49 completion of evolution of An- itschkow cells from heart muscle cells is illustrated. DIscusSION In this investigation it has been demonstrated that nuclei with caterpillar-like chromatin pattern, identical to those characteristic of Anitsch- kow cells, occur within normal immature and in normal and altered mature striated heart muscle cells and fragments of the latter in man. It was further found that Anitschkow cells evolved from altered heart muscle cells. These observations are in harmony with conclusions reached many decades ago by von Oppel (i) and Anitschkow (2) that altered striated muscle fibers in rabbit hearts gave rise to "free cells" or "myocytes" in response to implantation of foreign bodies in nearby myocardium. Caterpillar nuclei occur frequently in the growing heart of the human fetus. They were found to be present frequently, in some foci commonly, in myoblasts in the myotube stage of muscle development and in immature striated muscle cells in the myocardia of all I4 immature infants examined. With frequency that gradually decreased with age such nuclei occurred in immature and mature appearing striated muscle cells in the myocardia of 6 premature infants and in mature appearing striated muscle cells in the myocardia of 8 full-term infants who died with cardiac or other anomalies or with subdural hemorrhage. In contrast, caterpillar nuclei were present with increased frequency in striated muscle cells in the myocardia of 2 full-term infants who died with meningitis and congenital rubella, respectively. Such nuclei also were found in rare or occasional myofiber fragments in these two hearts. Further- more, caterpillar nuclei occurred frequently, in some foci commonly, in striated muscle cells and in fragments thereof in the myocardium of a 4-month-old infant who died of active rheumatic heart disease. In our experience fully formed caterpillar nuclei have been encountered only rarely or not at all in heart muscle cells of children and adults who died of disease other than infection or active rheumatic heart disease. In contrast, such nuclei occurred frequently, in some foci commonly, in heart muscle cells that were apparently intact or degenerating and in fragments thereof in 3 children who died of active rheumatic heart disease at 3 to 5 years of age. The frequency of caterpillar nuclei in June.. I966 CATERPILLAR NUCLEI 939 striated heart muscle cells ofpatients with active rheumatic heart disease decreased progressively with age from 5 years into adulthood. Such nuclei were found in striated heart muscle cells in only the youngest I of the 5 rheumatic patients between 20 and 6o years of age. In contrast, striated heart muscle fragments containing caterpillar nuclei occurred in all of these 5 older rheumatic patients, although they occurred in those over 30 years of age less frequently than in the rheumatic children, adolescents and younger adults. The various stages of evolution of Anitschkow cells from altered heart muscle cells or their fragments were observed. In harmony with this demonstration is the close correlation shown to exist between the frequency of caterpillar nuclei in altered striated muscle fibers and their fragments taken together and the frequency of Anitschkow cells among striated muscle fibers or their fragments in the myocardia of the infant and children up to 6 years of age with active rheumatic heart disease. In the group of I2 rheumatic patients 6 to 6o years old this correlation gradually diminished as the frequency of caterpillar nuclei in nonfragmented striated muscle fibers gradually diminished with age. However, strik- ingly close correlation persisted between the frequency of caterpillar nuclei in fragments of muscle fibers and the frequency of Anitschkow cells among muscle fibers or their fragments. Caterpillar nuclei were not present in striated muscle fibers or their fragments either within, near, or far removed from myocardial infarcts of different ages in the 8 patients examined, and Anitschkow cells were present in normally rare number, irrespective of proximity to infarcted regions. These observations do not support the widely held hypothesis that Anitschkow cells are non-myogenic cardiac histiocytes or reticuloendothelial cells. From results of the investigation here reported it is reasonable to infer that in heart muscle fibers or their fragments in children, adolescents and adults, caterpillar-like chromatin pattern, here shown to occur in apparently normal muscle fibers of growing fetal and full-term infant hearts, represents a primitive attempt at muscle cell generation in reaction to certain injuries. In hearts of infants and young children the sometimes frequent development of caterpillar nuclear structure in the nonfragmented muscle fiber in response to certain evocative stimuli, e.g., as provided in rheumatic fever, may represent frequent attempts at budding from the muscle fiber as in the fetal heart; muscle fragments containing caterpillar nuclei may also represent generative attempts. In contrast, in hearts of adults the rare development or absence of caterpillar nuclear structure in the nonfragmented muscle fiber in response to the same stimulus may indicate the rarity or failure of attempt at 940 MURPHY AND BECKER VOI. 48, No. 6 budding from the whole muscle fiber, but, as in children, muscle fragments containing caterpillar nuclei may represent generative attempts. That caterpillar nuclei can divide is illustrated in Figures 50 to 53. We have never encountered mitotic figures of the known forms in caterpillar nuclei during many years of detailed microscopic examination of a large number of hearts in which these nuclei occurred. In a few children with severe rheumatic myocardial disease we have very occasionally encountered mitotic figures in cardiac muscle fragments like those found by others12"13 in cardiac muscle fibers and fragments in a few infants and children. These conclusions are in harmony with the evidence that myocardial Aschoff bodies are lesions of striated muscle fibers and comprise I) mono-, multi-, and non-nucleated fragments of muscle fibers, and 2) multinucleated myogenic masses that arise in tracks of disintegrating muscle fibers, and appear to represent a limited attempt at regeneration of muscle.5"4"15 It is noteworthy that in Aschoff bodies caterpillar or owl eye nuclei occur in muscle fragments with considerable cytoplasm, in those with scant cytoplasm (Anitschkow cells), and in the masses that develop in tracks of disintegrating muscle fibers and appear to represent a limited attempt at muscle regeneration.'4"5 Caterpillar nuclei have been shown to occur in nonstriated or smooth muscle cells of the heart5"0 as well as in striated muscle cells of the heart. Nonstriated or smooth muscle occurs in many parts of the heart, including the walls of blood vessels, mural endocardium, and valves. Thus, in these areas, smooth muscle cells can appear as Anitschkow cells or may, like cardiac striated muscle cells, give rise to Anitschkow cells. After caterpillar nuclei together with sarcoplasm emerge as Anitschkow cells from cardiac striated muscle fibers or their fragments, it is very possible that the latter perform a reparative function. Such reparative function may be similar to that which nonstriated or smooth muscle cells may perform in reaction to arterial or valvular injury. From such reparative function elastic or collagen fibers and capillaries may result. More than 6o years ago von Oppel 1 suggested that "free cells of muscular origin" play an active role in formation of "myogenic granulation tissue" in the myocardium. REFERENCES I. VON OPPEL, W. tYeber Veranderungen des Myocards unter der Einwirkung von Fremdkorpern. Virchow Arch. Path. Anat., I90I, I64, 406-436. 2. ANITSCHKOW, N. Experimentelle Untersuchungen iuber die Neubildung des Granulationsgewebes im Herzmuskel. Beitr. Path. Anat. Allg. Path., 19I3, 55, 373-415- 3. EHRLICH, J. C., and LAPAN, B. The Anitschkow "myocyte." Arch. Path. (Chicago), I939, 28, 36I-370. June, z966 CATERPILLAR NUCLEI 94I 4. RUMYANTZEV, P. P. [On the "myocyte" nature.] Arkh. Anat., 1957, 34, 50-55. S. MuRPHY, G. E. The characteristic rheumatic lesions of striated and of nonstriated or smooth muscle cells of the heart. Genesis of the lesions known as Aschoff bodies and those myogenic components known as Aschoff cells or as Anitschkow cells or myocytes. Medicine, I963, 42, 73-II8. 6. WENEZIANOWA-GRUSDKOWA, M. S. Zur Frage des Ursprungs der sogenannten "Myozyten" im Myokardium. Frankfurt. Z. Path., I929, 37, 538-549. 7. RUBENSTONE, A. I., and SAPHIR, 0. Myocardial reactions to induced necrosis and foreign bodies, with particular reference to the role of the Anitschkow cell. Lab. Invest., I962, II, 79I-807. 8. McDONALD, H. G. Origin of Anitschkow's myocytes from cardiac muscle fibers. Texas J. Med., I963, 59, IO62-IO67. 9. MIKAT, K. W. Anitschkow cell development in rat myocardium. Arch. Path. (Chicago), I964, 77, 47-52. IO. MINICK, C. R.; MURPHY, G. E., and CAMPBELL, W. G. Experimental induction of athero-arteriosclerosis by the synergy of allergic injury to arteries and lipid rich diet. I. Effect of repeated injections of horse serum in rabbits fed a dietary cholesterol supplement. J. Exp. Med., I966, 124. (In press) II. MURPHY, G. E. Experimental and Histological Investigation of the Nature of Rheumatic Heart Disease. Discussion. In: The Streptococcus, Rheumatic Fever, and Glomerulonephritis. UHR, J. W. (ed.). The Williams & Wilkins Co., Baltimore, I964, 209-237. 12. MACMAHON, H. E. Hyperplasia and regeneration of the myocardium in infants and in children. Amer. J. Path., 1937, I3, 845-854. 13. HUDSON, R.E.B. Cardiovascular Pathology. Williams & Wilkins Co., Balti- more, I965, Vol. I, P. 39. I4. MURPHY, G. E. Evidence that Aschoff bodies of rheumatic myocarditis develop from injured myofibers. J. Exp. Med., I952, 95, 3I9-332. I5. MURPHY, G. E. Nature of rheumatic heart disease with special reference to myocardial disease and heart failure. Medicine, I960, 39, 289-384. The technical assistance of Miss Judith Chesner, Mr. Leonard Konikiewicz, and Mr. Walter Lee is gratefully acknowledged.

[ Illustrations follow] 942 MURPHY AND BECKER Vol. 48, No. 6

LEGENDS FOR FIGURES The photographs that follow are of sections of left ventricular myocardium in 5 immature infants 540 to 85o gm, I premature infant 2,085 gm, 3 full-term infants i day to I4 months old, 7 children 3 to io years old, and 3 adults 25 to 33 years old. Of these i6 patients 9 died of active rheumatic heart disease with Aschoff bodies in the myocardium. Unless otherwise stated sections were stained with hematoxylin and eosin. FIG. I. Immature male infant (580 gm), 35 minutes after birth. Caterpillar nuclei are shown in myoblasts in myotube stage of development. X Sm FIG. 2. Patient referred to in Figure I. Caterpillar nuclei appear in myotubes. Masson trichrome stain. X SI2. FIG. 3. Immature female infant (630 gm), 6 days after birth. Caterpillar nuclei are evident in immature striated muscle fibers. Masson trichrome stain. X SI2. FIG. 4. Immature male infant (540 gin), 24 hours after birth. A caterpillar nucleus is shown in an immature striated muscle fiber. Masson trichrome stain. X 640. June, I966 CATERPILLAR NUCLEI 943 f#lEs4'ti'2st'Z ''t''X~~~~~~~~~~~~~1l 'A

iii .|

A4 944 MURPIIY AND BECKER Vol. 48, No. 6

FIG. 5. Immature male infant (68o gm), 4 days after birth. Perpendicularly or obliquely transected myotubes contain owl eye (transected caterpillar) or caterpillar nuclei. X 1,200. FIG. 6. Immature male infant (850 gm), i day after birth. Caterpillar nuclei appear in immature muscle fibers (other myofibers in this heart are shown in Figures 7 and 8). Masson trichrome stain. X I,200. FIG. 7. The infant referred to in Figures 6 and 8. Caterpillar nuclei are shown in immature muscle fibers. Masson trichrome stain. X I,536. FIG. 8. The infant referred to in Figures 6 and 7. A caterpillar nucleus lies in a myofiber. Masson trichrome stain. X I,920. FIG. 9. Premature female infant (2,085 gm), 30 days after birth. Patient had cardiac and other anomalies. A caterpillar nucleus lies in a muscle fiber. Masson trichrome stain. X I,920. FIG. io. The infant referred to in Figure 9. A caterpillar nucleus is shown in a muscle fiber. Masson trichrome stain. X I,920. June, I966 CATERPILLAR NUCLEI 945

. 4

10 946 MURPHY AND BECKER Vol. 48, No. 6

FIG. ii. Full-term male infant (2,7IO gm), dead of meningitis (Hemophilus influenzae) 3 weeks after birth. Caterpillar nuclei lie in muscle fibers (other myofibers in this heart are shown in Figures I2, I4, 50 and 53). Masson trichrome stain. X I,280. FIG. I2. The infant referred to in Figures II, I4, 50 and 53. A caterpillar-like chromatin pattern is evident in almost all of the muscle nuclei shown. Many nuclei occur in pairs. A higher mangification of the right upper portion of this figure is shown in Figure 53. Masson trichrome stain. X I,200. FIG. I3. Six-year-old girl, dead 3 weeks after the onset of the first known attack of rheumatic fever. One of a pair of muscle fiber nuclei has a well formed caterpillar-like chromatin pattern. X I,280. FIG. I4. The infant referred to in Figures II, I2, 50 and 53. A caterpillar-like chromatin pattern is well formed in one of the muscle fiber nuclei and is partially formed in the other. Masson trichrome stain. X I,965. FIG. I5. Full-term male infant (3,780 gm), dead I day after birth with widely distributed petechiae and corneal edema. Rubella virus was cultured from the myocardium and many other tissues. A couple of caterpillar nuclei appear in a muscle fiber. Figure I6 is also from this myocardium. Masson trichrome stain. X I,920. FIG. I6. The infant referred to in Figure I5. A caterpillar nucleus is shown in a muscle fiber. Masson trichrome stain. X I,920. FIG. I 7. Three-year-old boy, dead 2 weeks after the first known attack of rheumatic fever. A pair of caterpillar nuclei and a single one may be seen in muscle fibers. The muscle fiber on the right shows marked degenerative change in its lower portion (other myofibers in this heart are shown in Figures I9, 20, 22, 25, 28 and 52). Masson trichrome stain. X 2,350. FIG. I8. Ten-year-old boy, dead 3 months after the first known attack of rheumatic fever. Degenerative change is manifest in muscle fibers. In one there is a caterpillar nucleus (other myofibers or fragments thereof are shown in Figures 35, 45 and 48). Weigert-hematoxylin and eosin stain. X I,536. June, I966 CATERPILLAR NUCLEI 947 I

15 Aft 948 MURPHY AND BECKER Vol. 48, No. 6

FIG. ig. The child referred to in Figures I7, etc. Caterpillar nuclei appear in muscle fibers. Masson trichrome stain. X I,638. FIG. 20. The child referred to in Figures I7, etc. A pair of caterpillar nuclei and a single one lie in muscle fibers. Masson trichrome. X I,920. FIG. 2I. Five-year-old boy who died suddenly of active rheumatic heart disease while playing stick ball. A caterpillar nucleus appears in a muscle fiber. Masson trichrome stain. X 3,024. FIG. 22. A higher magnification of the muscle fiber nucleus shown in upper portion of Figure 20. X 3,900- June, I966 CATERPILLAR NUCLEI 949

..I Is

22 950 MURPHY AND BECKER Vol. 48, No. 6

FIG. 23. Six-year-old boy who died I month after the first known attack of rheumatic fever. Degenerative change is apparent in a muscle fiber; within this is a caterpillar nucleus (other myofibers or fragments of this heart are shown in Figures 26, 27, 29, 40, 46 and 49). X I,024. FIG. 24. Ten-year-old boy who died 3 months after a recurrent attack of rheumatic fever. Within a muscle fiber showing disintegration is a caterpillar nucleus (other myofibers or fragments of this heart are shown in Figures 34, 36 and 37). X I,024. FIG. 25. The child referred to in Figures I7, etc. In a degenerating muscle fiber is a caterpillar nucleus. X I,024. FIG. 26. The child referred to in Figures 23, etc. Degeneration of muscle fibers. Two fibers contain caterpillar nuclei. X I,024. FIG. 27. The child referred to in Figures 23, etc. A caterpillar nucleus is noted in a disintegrating portion of a muscle fiber. X I,024. FIG. 28. The child referred to in Figures I7, etc. A caterpillar nucleus is emerging from a damaged muscle fiber. X I,024. FIG. 29. The child referred to in Figures 23, etc. A caterpillar nucleus is being released from a disintegrating muscle fiber. X I,280. FIG. 30. Fourteen-month-old female infant, dead 3 weeks after first signs of a first attack of rheumatic fever. A caterpillar nucleus is being released from a disintegrating muscle fiber (other muscle fibers in this heart are shown in Figures 3I and 41). X I,024- FIG. 3I. The infant referred to in Figures 30 and 4I. A caterpillar nucleus is in process of release from a disintegrating muscle fiber. X 8oo. June, I966 CATERPILLAR NUCLEI 95I

.,..~ ~ ~~~~~~~~~~~~0

- -,f - - 952 MURPHY AND BECKER Vol. 48, No. 6

FIG. 32. Twenty-nine-year-old woman who died with bilateral purulent salpingitis, pelvic abscess, and peritonitis. Degenerative change is manifest in muscle fibers. Within one and emerging from the other are caterpillar nuclei shown at higher magnification in Figure 33 (caterpillar nuclei in muscle fragments in this heart are shown in Figures 38 and 43). X 512. FIG. 33. A higher magnification of the two caterpillar nuclei shown in Figure 32. One is within and the other is emerging from altered muscle fibers. X 8oo. FIG. 34. Child referred to in Figures 24, 36 and 37. Three caterpillar nuclei are shown. The one at the lower left and that near the center appear to be emerging from fragmenting muscle fibers. The third caterpillar nucleus is within a muscle fiber fragment. X I,024. FIG. 35. The child referred to in Figures i8, 45 and 48. Caterpillar nuclei appear in muscle fibers undergoing marked degenerative change. Weigert-hematoxylin and eosin stain. X i,536. FIG. 36. The child referred to in Figures 24, etc. A muscle fiber is undergoing fragmentation. The still striated fragment in the center contains a caterpillar nucleus. Close to this nucleus is another caterpillar nucleus, only partially in focus. This may have emerged from the same disintegrating muscle fiber. X 8oo. FIG. 37. The child referred to in Figure 36. Caterpillar nuclei appear in ragged sarcoplasmic fragments. X I,024. FIG. 38. The patient referred to in Figures 32, 33 and 43. A muscle fiber fragment contains a caterpillar nucleus. X I,024. FIG. 39. Thirty-three-year-old man with severe rheumatic heart disease who experienced obvious attacks of rheumatic fever at 9 and 2I years of age. Progressive congestive heart failure began i8 months before death and was associated with irregular low-grade fever. A muscle fiber fragment contains a caterpillar nucleus. X 1,280. FIG. 40. The child referred to in Figures 23, etc. A sarcoplasmic fragment contains a caterpillar nucleus. X I,024. June, I966 CATERPILLAR NUCLEI 953

32 K 33, 34

i ejii 2i*~v

F .3:3 k A.. K 36. 37

.III

38 39, 40 954 MURPHY AND BECKER Vol. 48, No. 6

FIG. 4I. The child referred to in Figures 30 and 3I. Nuclei with so-called owl eye appearance are shown in transected muscle fibers. X 8oo. FIG. 42. Three-year-old girl who died during the first attack of rheumatic fever. In the last two weeks of life prednisone, 8o mg per day, was given. A small Aschoff body is comprised of fragments of obliquely sectioned muscle fibers in several of which are owl eye or caterpillar nuclei. X 480. FIG. 43. The patient referred to in Figures 32, etc. On the left is a fragmenting muscle fiber. Caterpillar or owl eye nuclei appear in the fragments. In the lower central area is a caterpillar nucleus that has probably emerged from a disintegrating muscle fiber. X 8oo. FIG. 44. Ten-year-old girl who died 5 weeks after a second attack of rheumatic fever. The central portion of an Aschoff body contains mono-, multi-, and non- nucleated muscle fiber fragments. Some of the nuclei in these fragments have an owl eye appearance. X 8oo. FIG. 45. The child referred to in Figures i8, etc. An Aschoff body contains mono-, multi-, and non-nucleated muscle fiber fragments. Caterpillar or owl eye nuclei appear in some of these and in some adjacent muscle fragments. The Aschoff body is shown at higher magnification in Figure 48. Weigert-hematoxylin and eosin stain. X 300. FIG. 46. The child referred to in Figures 23, etc. Anitschkow cells are evolving from disintegrating muscle fibers. X 8oo. FIG. 47. Twenty-five-year-old woman with severe rheumatic heart disease who died about a week after an upper respiratory infection followed by marked dyspnea. An obliquely sectioned muscle fiber fragment contains an owl eye nucleus. X I,024. FIG. 48. A higher magnification of the Aschoff body shown in Figure 45. Small fragments of muscle fibers contain caterpillar or owl eye nuclei and scant cytoplasm (Anitschkow cells). X 6I5. FIG. 49. The child referred to in Figure 46. Anitschkow cells have evolved from disintegrating muscle fibers. x 8oo. June., z966 CATERPILLAR NUCLEI 955

r-..,

b.. ..je4 42, 43

a 45, 46

"in'irl". 4pi K

..,.- * ..s.: .: !", ".,oV %

lo.:,- ix

Al A .6 A dl"L.; I 47 A 48,49

A..X,

2-.-..71...! 956 MURPHY AND BECKER Vol. 48, Nu. 6

Stages of division of caterpillar nuclei. All magnifications are X 3,024. FIG. 50. The child referred to in Figures ii, etc. A dividing caterpillar nucleus appears within a striated muscle cell. Masson trichrome stain. FIG. 5I. Thirty-seven-year-old woman who died of cardiac failure due to widespread degenerative change in cardiac myofibers. Nearly complete division of a caterpillar nucleus has occurred within a large fragment of a muscle cell that has lost striations. FIG. 52. The child referred to in Figures I7, etc. A dividing caterpillar nucleus lies among degenerating heart muscle cells. FIG. 53. A higher magnification of the right upper portion of the area shown in Figure I 2. A contiguous pair of caterpillar nuclei lies within a striated muscle cell. It would appear that these nuclei resulted from recent nuclear division. Masson trichrome stain. June, I966 CATERPILLAR NUCLEI 957

...... ,, 5Um

i .I J

Zj_.M- . ' ij56.'

Qz~~~~~~~~~~~~~~.2#2 1'' .....