Cell and Tissue Research

Cell Tissue Res. 197, 295-312 (1979)

9 by Springer-Verlag 1979

The Relation Between Solid Cell Nests and C Cells of the Thyroid Gland An Immunohistochemical and Morphometric Investigation R.C. Janzer, E. Weber, and Chr. Hedinger Institute of Pathology, University of Z~irich, Ziirich, Switzerland (Chairmen: Prof. Chr. Hedinger and Prof. J.R. Riittner)

Summary. Thyroid tissue of 300 routine autopsies was processed in a standardized manner. So-called solid cell nests (SCN) were found in 21 patients (7 %). These cases were investigated carefully by serial step sectioning. In order to explore the correlation of SCN to the C-ceU system, the sections were stained by silver impregnation and the immunoperoxidase method. Morphometric analyses revealed a significant increase in the density of C cells in the proximity of the SCN. With progressive distance from the SCN, the C-cell density decreased and reached normal values. In 30 % of the cases argyrophilic and calcitonin-positive cells were found lying within the SCN. Occasionally, mixed follicles could be discerned: These were lined on the one side by a multilayered squamous epithelium, on the other side by normal monolayered cubic follicular epithelium, and contained a peculiar granular material. In one case, SCN were associated with intrathyroid portions of the parathyroids and adult adipose tissue, in a second case with adipose tissue only. Most probably SCN are vestiges of the ultimobranchial body and should be interpreted as such, despite the fact that other authors have expressed different views. The lack of disturbances in the calcium metabolism of the patients and the absence of medullary carcinoma in their family histories led us to interpret locally confined C-cell hyperplasia not as reactive nor premalignant, but rather as normal. Key words: Thyroid gland - C cells - Ultimobranchial body - Immunocytoc h e m i s t r y - Morphometry.

The histological appearance of the normal thyroid gland shows a remarkable diversity. This applies in particular to its epithelial structures. One finds, in addition to the normal follicles and the calcitonin-producing C cells, solid intra- and parafollicular cell clusters, nests and cords of squamous epithelium, falsely referred to as "yon Ebner'sche Zellstriinge" in the German literature, cysts with various Send offprint requests to: Dr. R.C. Janzer, Institute of Pathology,University-Hospital,CH-8091 Ziirich,

Switzerland

0302-766X/79/0197/0295/$03.60

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epithelial linings (single-layered to stratified, cubic, cylindrical or squamous epithelia) and intrathyroidal portions of the parathyroids, of the thymus or of the laryngeal paraganglia. Despite a uniform morphological description of these elements, there is a wide diversity of opinion in the literature as to their significance. Nevertheless, it appears probable since the studies of Sugiyama (1967, 1971), Welsch (1972) and Nadig et al. (1978) that some of these structures are vestiges of the ultimobranchial body (UBB), especially the solid nests and cords of squamous epithelium, as well as the cysts lined by the same epithelium. The C cells, being of neural crest origin (Le Douarin and Le Li6vre, 1970), migrate into the thyroid gland with the UBB (Pearse and Carvalheira, 1967). Nadig et al. (1978) have succeeded in demonstrating isolated calcitonin-containing cells in the wall structures of the solid cell nests (SCN) by means of immunofluorescent methods. The present authors intended to check the latter finding in a larger sampling of material. At the same time, the question of whether increased numbers of C cells can be demonstrated in the vicinity of the SCN was examined. Increased numbers of C cells and their arrangement in larger groups is regarded by Wolfe et al. (1973), De Lellis et al. (1977) and Schiirch et al. (1977) as nodular or diffuse hyperplasia of the C cells and interpreted as the obligatory precursor of a medullary carcinoma, provided that the presence of a disturbance of calcium metabolism does not imply a reactive phenomenon (Kracht, 1977).

Materials and Methods The present study was carried out on thyroid tissue of 300 unselected routine autopsies from the year 1977. Highly enlarged goiters were excluded.

Processing of the Material The thyroid glands were carefully freed from the trachea, and one of the lateral lobes was cut macroscopically in accordance with Fig. 1. It has been known since the studies of Roediger (1973a), Wolfe et al. (1974, 1975) and Okamoto et al. (1979) that the C cells are normally distributed along a diffusely defined central axis with posterolateral accentuation. The material was fixed for 6 to 20 h in Bouin solution and embedded in paraffin. Routinely, four serial sections of 5-6 ~tm thickness were cut.

Histological Investigation The first section was stained with hematoxylin-eosin and systematically examined at x 128 magnification for the presence of SCN, branchiogenic cysts and intrathyroidally located portions of parathyroid tissue or portions of the thymus. The second section was silver-stained according to the method of De Grandi (1970) or slightly modified according to Fernandez-Pasqual (1976) in order to demonstrate the C cells. In the method of Fernandez-Pasqual the second reduction step was omitted, and the tissue section was finally fixed with 5 % thiosulphate. The advantage of the method according to Fernandez-Pasqual as opposed to that of De Grandi is firstly the even staining of the preparation (even at the edges) and secondly the simpler and shorter processing. The C cells are clearly revealed using this latter method by virtue of the presence of brown-black granules. The danger of non-specific deposits is somewhat greater because of the use of higher concentrations, but is never so extensive as to interfere with evaluation of the preparation.

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Immunoperoxidase Method In cases in which the silver staining revealed a high n u m b e r of C ceils (more than 10 per visual field at x 32 magnification), the third and fourth sections were used for demonstration o f C cells by m e a n s of the slightly modified immunoperoxidase method according to Sternberger et al. (1970). In a first step, the endogenous peroxidase was blocked in deparaffinated sections (1 ml 30~o H202 dissolved in 50ml methanol was applied to the preparation for 30rain at r o o m temperature). In a second step, anticalcitonin serum from goats immunized against h u m a n calcitonin (obtained from a patient with medullary carcinoma) a was applied to the section (dilution 1 : 1280 in phosphate buffer, p H 7.4 for 45 min at 37 ~ C). In the third step, rabbit anti-goat IgG (Cappel Laboratories, Cochranville, Pa., USA) was applied for 30 min (dilution 1 : 2 0 in phosphate buffer containing 1 : 2 0 rabbit serum) at room temperature. In the fourth step, the sections were incubated with goat peroxidase-antiperoxidase complex (Cappel Laboratories, Cochranville, Pa., USA) for 30 rain at r o o m temperature (dilution 1 : 50 in phosphate buffer containing 1 : 20 rabbit serum). The fifth and final step was the demonstration of peroxidase by m e a n s of diaminobenzidine ( 5 m g diaminobenzidine-HCl in 10ml phosphate buffer + 0.015 ~ HzOz) for 5 min at room temperature. After each of the steps mentioned above, the sections were washed 2 • with phosphate buffer (pH 7.4). The calcitonin-containing C cells are demonstrated by the presence of dark brown granules. To check the specifity of the antiserum, instead of the goat a n t i - h u m a n calcitonin of the second step, an incubation with goat immunoglobulin was performed; the procedure was otherwise identical. A n o t h e r control was performed by absorption of the antiserum with synthetic h u m a n calcitonin.

Histometry In order to identify the C cells unequivocally, an 80fold magnification was employed. No counterstaining was used. By insertion of an integration plate into the microscope, a square visual field was demarcated which comprised 1400 btm 2 after calibration by means o f an objective micrometer. The solid cell nests (SCN) were projected into the middle of the visual field and the C cells counted. This area was defined as sector I (see Fig. 2). Exact displacement of the preparation by m e a n s of a cross-slide table ensured the complete examination of the entire sectional surface and obviated overlap of visual fields. The eight visual fields contiguous to sector I were defined as sector II; here again the C-cell density par visual field was evaluated at 80fold magnification and was compared with the value o f sector I. The further 16 visual fields following sector II on the outside were defined as sector III; the m e a n C-cell density was also calculated here.

Examination of Serial Step Sections All the 21 cases revealing SCN were studied by serial step sectioning at intervals of 500 ~tm. At every new level, serial sections were prepared with hematoxylin-eosin and the immunoperoxidase method. In the 21 patients SCN could thereby be identified in sections at 57 different levels. In slides o f 17 levels SCN occurred more than once and had to be excluded from the study, since one and the same visual field would have been accounted to different sectors according to the above definition o f the sectors I to III. Sections o f 40 levels were thus available for calculation of the m e a n C-cell density as a function o f the distance from a SCN.

Statistics The assumption of a normal distribution was reinforced by accessory tests. The significance of the difference in m e a n values was investigated by m e a n s o f the Student t-test. 1 The anti-calcitonin serum was kindly provided Universit/itsklinik, Ziirich

by Prof. J.A.

Fischer,

Orthop~idische

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Table 1. Clinical-pathological data of 21 patients with SCN Case No.

Sex

Age

Clinical diagnosis

Density of C cells Sectors a II

III

7 83 28 18

4 16 20 18

1 5 4 4

4 23 21 18

54 0 38 11 2 0 2 45

32 0 14 7 0 0 1 26

9 1 15 1 0 0 1 11

33 0 17 9 0 0 1 28

17 14 96 28 104 51

4 3 46 9 45 6

2 2 12 5 6 0

6 4 56 11 54 I1

I

1. 2. 3. 4. 5.

AZ AZ AZ AZ AZ

178/77 219/77 230/77 249/77 259/77

Q Q d d Q

76 56 71 39 43

6. 7. 8. 9. 10. 11. 12. 13.

AZ AZ AZ AZ AZ AZ AZ AZ

289/77 380/77 439/77 500/77 536/77 549/77 553/77 650/77

~ c~ c~ c~ c~ ~ c~ c~

76 46 60 67 66 48 42 78

14. 15. 16. 17. 18. 19. 20. 21.

AZ AZ AZ AZ AZ AZ AZ AZ

665/77 775/77 859/77 863/77 870/77 360/77 369/77 393/77

c~ ~ Q c~ Q c~ Q d

77 70 70 74 53 62 80 45

a

For definition of sectors I, II and III, see text

Pulmonary embolism Mammary carcinoma Carcinoma of the bladder Glioblastoma multiforme Carcinoma of the mammary gland and uterus Generalized atherosclerosis Hairy cell leukemia Phenacetin nephritis Pleural mesothelioma Hepatocellular carcinoma Intestinal infarction Delirium tremens Anaplastic carcinoma of unknown origin Carcinoma of the bladder Myocardial infarction Cor pulmonale Chronic lymphocytic leukemia Mammary carcinoma Plasmocytoma ] Stroke syndrome / Aspiration pneumonia

I+II

multiple SCNs

Results

Frequency of Solid Cell Nests (SCN) P r o c e e d i n g in a c c o r d a n c e w i t h t h e d e s c r i p t i o n g i v e n a b o v e , S C N w e r e f o u n d in 21 p a t i e n t s o u t o f 300 cases ( = 7 %). A s c a n be seen f r o m T a b l e 1, 15 p a t i e n t s w e r e m e n ( = 7 1 % ) a n d 6 w e r e w o m e n ( = 29 %) w i t h a n a v e r a g e age o f 58.7 a n d 59.7 years, r e s p e c t i v e l y . T w e l v e p a t i e n t s d i e d f r o m m a l i g n a n c i e s , five f r o m diseases o f the c a r d i o v a s c u l a r s y s t e m a n d f o u r f r o m o t h e r causes. I n t h r e e cases, m u l t i p l e (up to 13) S C N w e r e f o u n d in o n e a n d the s a m e section. Serial step sections at i n t e r v a l s o f 500 ~tm r e v e a l e d S C N at 57 levels, in 17 o f w h i c h the o c c u r r e n c e o f m u l t i p l e S C N was noted.

Location of Solid Cell Nests (SCN) C o r r e s p o n d i n g t o the s t a n d a r d i z e d s a m p l i n g o f m a t e r i a l in a c c o r d a n c e w i t h Fig. 1, the S C N w e r e f o u n d in a c e n t r a l axis o f the l a t e r a l lobes. I n the 40 sections w i t h single S C N , a slight t e n d e n c y to a d o r s o - c r a n i a l p o s i t i o n c o u l d be d e t e c t e d (Fig. 3). A s illustrated in Fig. 4, n o site o f p r e d i l e c t i o n c o u l d be d e m o n s t r a t e d for m u l t i p l e occurrence.

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~

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I

I

Fig. 1. Diagram of tissue sampling: The middle part o f the upper two thirds of a lateral thyroid lobe was removed for histological investigation

Fig.2. Definition of sectors for histometry: The basic unit of area is a visual field o f 1400pro 2 ( = sector I). In the center of sector I a solid cell nest (SCN) is projected. Sector II consists of the eight visual fields contiguous to sector I, while the 16 area units surrounding sector II comprise sector III

Fig. 3. Schematic localization o f the singly occurring SCN: Every circle marks a SCN on a slide. The numbers correspond to those of Table 1

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Fig. 4. Topography of multiply occurring SCN: case no. 19, 6 step sections at 500 lam intervals Fig. 5. Example of a solid cell nest (SCN) (case no. 3, hematoxylin-eosin, x 400)

Histology The S C N are embedded in a more or less dense connective tissue stroma and are in m o s t cases clearly demarcated from the thyroid gland follicles by a silver stainable basal lamina. They are most frequently organized as solid cell nests (Fig. 5). Two distinct cell types are always recognized in hematoxylin-eosin-stained sections. The majority o f cells appears polygonal to elongated-oval in shape, measuring 1418 ~tm in diameter with a h o m o g e n e o u s slightly basophilic cytoplasm and welldefined cell boundaries. The centrally located cell nuclei are oval to spindle-shaped, show a p r o n o u n c e d nuclear m e m b r a n e and a loosely organized, fine chromatin structure. A clear, r o u n d nucleolus can be discerned. The second cell type is preferentially but not exclusively basal in location, and always shows a round,

Solid Cell Nests and C Cells in Thyroid Glands

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Fig. 6. Example of a cystic SCN (case no 17, hematoxylin-eosin, x 320)

compact, dark nucleus. The cells are likewise polygonal, but vary in size, although they usually appear smaller than the first cell type. The cell borders are clearly recognizable, but the cytoplasm is optically empty in some cases and highly vacuolized in others; occasionally, it is strongly eosinophilic and finely granular. The behavior of these cell types in histochemical studies is further described below. In addition to solid nests, cystic structures sometimes resembling follicles and lined by the cells mentioned above are revealed (Fig. 6). The content of these cysts either cannot be distinguished from normal colloid, or consists of a peculiar granular material which is weakly basophilic. Occasionally, mixed follicles can also be discerned. These are lined on one side by a rnultilayered squamous epithelium and on the other side by normal monolayered cubic follicular epithelium (Fig. 7b). These mixed follicles mainly contain the granular material mentioned above. All three manifestation forms of the SCN are mostly elongated-oval in shape; the average diameters are 80 x 110 ~tm, although the variation is great. The largest

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e

Fig. 7. a Association of SCN, intrathyroidaUylocated parathyroid tissue and adult adipose tissue (case no. 5, hematoxylin-eosin, x 130). b Mixed folliclelined partly by stratified squamous epithelium, partly by single layer of cubic epithelium and containing basophilic granular material. (Case no. 5, hematoxylin-eosin, • 320, detail of Fig. 7a). e Association of SCN with adult adipose tissue interpreted as tissue of thymic origin (case no. 19, hematoxylin-eosin, x 130)

d i a m e t e r f o u n d o n a section was 110 x 170 pm. However, since in five step sections at intervals o f 500 ~tm parts of p r o b a b l y the same S C N could be found, it m u s t be a s s u m e d that S C N c a n a t t a i n a l o n g i t u d i n a l extension u p to 2000 tam.

Histochemistry Both silver staining a n d the i m m u n o p e r o x i d a s e m e t h o d showed argyrophilic or c a l c i t o n i n - c o n t a i n i n gcells situated within the S C N in one third of the cases (6 of 21)

Solid Cell Nests and C Cells in Thyroid Glands

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Fig. 8a-c. C cellswithin SCN: Case no 15, serial sections, Bouin fixation, paraffin embedding, x 250. a Immunoperoxidase method, b Hematoxylin-eosin. c Argyrophilia and levels (18 of 57) (Fig. 8). These cells corresponded to the type described above with dark, compact cell nuclei. Furthermore, it was noticeable in the immunoperoxidase method that cells located on the periphery of the SCN not unambiguously containing calcitonin nevertheless stained much more darkly than the cells in the central regions (Fig. 9).

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Fig. 9 a and b. Case no 5, serial sections, • 400: a Hematoxylin-eosin. b Immunoperoxidase method. Note the darkly stained cells at the periphery of the SCN. In addition, a few scattered calcitonin-positive cells can be recognized by their intensely stained, black cytoplasm

Solid Cell Nests and C Cells in Thyroid Glands

305

Fig. 10 a and b. C cell density as a function of the distance from the SCN (marked with a circle). Sections are taken at an interval of 500 lain, each point corresponding to a C cell. Note the high C cell density in the vicinity o f the SCN and its decrease with progressive distance from it. a Case no. 1. h Case no. 7

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19urcttion of autolysis ( hours ) Fig. 11. C cell density as a function of duration of postmortem autolysis

Density of the C Cells Around the SCN In the calculation of the mean C-cell density as a function of the distance from SCN, the sectors I to III were defined arbitrarily (see Materials and Methods). The average values per case are presented in Table 1. F o r exact statistical calculation, the sections of 40 levels with a single occurrence of an SCN were chosen. It was shown that the mean C-cell density in sector I was higher compared to that in sector II by a factor of 2.37 (I : II = 2.37 : 1 ; t = 3.40, p < 0.0025). The mean cell density in sectors I + I I as compared to that in sector III was 4.02 times greater ( I + II : III = 4.02 : 1 ; t--- 4.89, p < 0.001) (Fig. 10). The influence of autolysis on the absolute number of C cells in sector I is shown in Fig. 11. However, this autolytic effect does not play a role in the present study, since it can be assumed that the autolytic alterations affect all parts of the thyroid gland evenly so that the relative ratio of C-cell density as a function of the distance from the SCN remains unchanged.

Intrathyroid Portions of the Thymus and Parathyroids, Branchiogenic Cysts Thymus tissue was not found in any of the 300 cases investigated. Nevertheless, the association of SCN with adult adipose tissue could be demonstrated in three cases (Fig. 7a, c). The latter does not communicate with the pericapsular adipose tissue, but appears as an isolated focus of fatty tissue in an otherwise completely intact thyroid gland. Intrathyroid parathyroid tissue was found in 2 out of 300 cases. In one case, the parathyroid tissue was associated with a SCN and fatty tissue (Fig. 7a). In one of the 300 cases, cysts lined with several layers of ciliated epithelium were demonstrated; these were located near the isthmus and immediately outside the thyroid gland tissue. N o paraganglionic tissue could be demonstrated.

Solid Cell Nests and C Cells in Thyroid Glands

307

Discussion

Solid intra-acinar cell clusters in the form of single-layered cords or loose groups of cells were first described in human thyroid gland by W61fler in 1880. Various authors (Miiller, 1896; von Ebner, 1902; Lobenhoffer, 1909; Wegelin, 1926; Marine, 1928) also described these structures and interpreted them as embryonic vestiges of the immature thyroid gland follicles, or as budded portions of thyroid gland follicles in hyperactivity. In general, these authors attributed the solid cell cords with the capacity to form new follicles. As a result of studies on serial sections, other authors (Staudacher-Dalle Aste, 1941 a, b; Ludwig, 1953) postulated that no interfollicular epithelial structures exist in the thyroid gland, but that the solid cell clusters are identical with tangentially cut follicular epithelia. The C cells found in a parafollicular location light microscopically are shown in electron microscopic studies to be situated within the follicular basal lamina, i.e. intraepithelial. Small nests of squamous epithelia, which are situated partly in the septa and partly within the lobes, were described in the human thyroid gland for the first time by Kloeppel in 1910. Wegelin (1926) confirmed this finding as normal and regarded them as vestiges of the ductus thyreoglossus or as dispersed pharyngeal pouch epithelium. Additionally, he described instances of squamous epithelium in degeneratively altered adenoma nodes, which he interpreted as being ofmetaplastic origin. Without designating them as squamous epithelial nests, Getzowa described as early as 1907 and 1911 solid, occasionally cystic cell clusters that displayed two distinct cell types. However, these are not identical with the solid cell cords described by W61fler. Getzowa interpreted her cell clusters as being of ultimobranchial origin, because she could also demonstrate the same cells in a case of thyroid gland aplasia. Erdheim (1904) also communicated a case of thyroid aplasia with cystic structures at the site of the thyroid, which were interpreted to be of branchiogenic origin. Identical cystic structures in animals were interpreted by Prenant (1894) as the central canal of the ultimobranchial body, and designated as ultimobranchial cysts in man by Hermann and Verdun (1899). Most modern .authors also came to this conclusion (Meeker, 1925; Van Dycke, 1945; Lindsay et al., 1968; Sugiyama, 1971; N~ve and Wollman, 1971; Lietz, 1971; Jordan et al., 1973; Christov et al., 1973; Calvert, 1975; Petko, 1975). Despite the convincing study of Sugiyama (1971), some authors (e.g. Roediger, 1973b) still believe that the origin of these squamous epithelial nests from the ultimobranchial body is not yet unequivocally proven. In particular, many authors (Jaffe, 1937; Klinck and Menk, 1951; Goldberg and Harvey, 1955; Dube and Joyce, 1971) regard a metaplastic genesis as being the most probable, especially since Van Dyke (1955) observed increased frequency of squamous epithelial nests (partly keratinized) in vitamin A deficiency; Krupp (1972) confirmed these observations. Some authors (Fukunaga and Lockett, 1971) regard SCN, irrespective of their origin, as being precursors of squamous cell carcinoma. The data given in the literature (Yamaoka, 1973) concerning the frequency of SCN in adults based on a single section per thyroid gland (0-3 ~) are significantly lower than the value found in the present study (7 %). We explain this difference as being due to our standardized sampling of material from the upper 2/3 of the lateral lobes (Fig. I). Thereby, predilection sites of SCN were included in our study (Sugiyama, 1971; Yamaoka, 1973; Kirkeby, 1976). The studies of Yamaoka (1973)

308

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show that the frequency with which SCN are found rises with increasing number of sections examined per thyroid gland. Investigation by serial sections revealed the presence of SCN in all cases. Kirkeby (1976) also found "cyst-like structures" with the characteristics of SCN in all investigated cases of serially sectioned guinea pig thyroid glands. Sugiyama (1971) found structures resembling SCN in 87.5 ~ of 98 serially sectioned prenatal thyroid glands. Concerning the description of the location and morphology of SCN, our data agree with the results of most authors, who have already been quoted. The study of Kirkeby (1977) provides an indication as to the relationship of SCN to C cells. In the ultimobranchial body cysts of the guinea pig, he found a markedly raised esterase activity compared to the follicular epithelium; this property can also be demonstrated for C cells (Pearse, 1966). Our results confirm the presence of isolated C cells within the SCN (Nadig et al., 1978) and show a significant clustering of C ceils around the SCN. This finding is consistent with a common ultimobranchial origin of the SCN and C cells and raises the question of whether the ability of human ultimobranchial bodies to mature and release C cells in the prenatal dissolution stage of the UBB (Sugiyama, 1971 ; Chan and Conen, 1971) is restricted to the embryonic period. In the adult mouse, Sato et al. (1966) were able to demonstrate in an electron microscopic study the formation of parafollicular cells from the wall of UBB cysts. The possibility remains that the SCN are a probable source for a proliferation of C cells also in adult man, in addition to the functions discussed by Calvert (1975). The density of C cells demonstrated in the vicinity of the SCN markedly exceeds the norm as evaluated by Wolfe et al. (1974, 1975). The interpretation of this finding is difficult. It should be kept in mind that we used another antiserum than Wolfe et al. The normal value found by these authors may not necessarily correspond to our normal values. Clinical conditions, especially disturbances of calcium metabolism that might explain the hyperplasia as reactive were not present in our material. Moreover, as none of our patients' family histories disclosed the occurrence of medullary carcinoma, the interpretation of local C cell hyperplasia as a premalignant lesion seems unlikely. One must further be aware that the factors which bring about a stimulation of secretion and/or production of calcitonin do not necessarily affect the rate of division or formation of C cells. The tendency to find fewer C cells with increasing duration of autolysis is consistent with the observations of many authors on the influence of autolysis on cells with endocrine activity. The significance of the fact that 15 out of the 21 patients were men remains elusive; Heath and Sizemore (1977) found a significantly higher basal calcitonin level in men than in women. Twelve of the 21 patients bore a malignancy, roughly corresponding to the percentage in our autopsy material as a whole. Since Coombes et al. (1974) demonstrated ectopic calcitonin production in 50 ~o of all tumors and Becker et al. (1975) even in 60 ~ of all pulmonary tumors, our findings appear to indicate that an ectopic calcitonin production does not exert any demonstrable effect on the SCN and the number of C cells. However, a hypercalcitoninemia has not been proven in any of the cases of our series. Delimination of SCN from epithelial nests which have arisen metaplastically is possible in many cases. SCN are regularly found in normal or only slightly altered thyroid glands, whereas metaplasias can mostly be seen on the edge of nodules in

Solid CellNests and C Cellsin ThyroidGlands

309

goiters with degenerative lesions. Onion-like arrangement of the cells, formation of horn pearls or clear intercellular bridges only occur in metaplasias. The SCN are variable in form, but always exhibit the two cell types described and a basal lamina, which can be stained with silver. Electron microscopically, the ultimobranchial bodies exhibit two distinct cell types in the rat (N~ve and Wollman, 1971 ; Wollman and N~ve, 1971). The cells of the outer or basal layer are characterized by hemidesmosomes, tonofibrils (which in some cases irregularly penetrate the cytoplasm and in other cases radiate in bundles into the desmosomes), irregular cell surfaces, pinocytotic vesicles on the basal plasma membrane and an abundance of free ribosomes. The more centrally or apically situated cells differ from the basal cells by the absence of hemidesmosomes, by the mainly dispersed arrangement of the tonofibrils and by the lower content of free ribosomes. Cells in contact with the lumen exhibit microvilli. The methods applied for demonstration of SCN and C cells unfortunately do not permit us to go into the interesting question of the so-called mixed ultimobranchial follicles (cf. Calvert, 1975; Petko, 1975). We nevertheless found, in juxtaposition to the SCN, thyroid follicles which were coated on the one side by a stratified squamous epithelium and on the other side by thyreocytes of normal appearance. The follicular content partly corresponded to the typical homogeneous and eosinophilic colloid, but was also at certain sites peculiarly granular and slightly basophilic. This finding is consistent with the ultrastructural studies of N~ve and Wollman (1971) in the rat; besides "cell debris", these authors also found acellular material which partly corresponded to the colloid, but which was in part less electron dense and exhibited a reticular character. Thyroid follicles with the same granular material could be found especially frequently in the vicinity of SCN. Figure 13 of Bargmann (1939b) is worth mentioning in that it shows an intrathyroidally-located parathyroid gland in the immediate vicinity of which two cysts can be seen partly appearing to possess a multilayered lining and a granular content. In many animals branchiogenic organs (parathyroid glands, thymus, ultimobranchial body) form a complex within the thyroid gland (Bargrnann, 1939a), which may, in some species, include even the carotid body (Bargrnann, 1939 b). In his studies of serial sections of neonatal human thyroid glands, Sugiyama (1971) found thymic tissue intrathyroidally in 18 % of the cases, half of which were associated with SCN. The fact that we found no intrathyroid thymic tissue at all in our 300 adult cases may be explained on the basis of physiologic involution. Nevertheless, we found adult adipose tissue in two cases in direct association with SCN; this can be regarded as a residue of the thymus. Bargmann (1939a) described, however, adipose tissue of the mature type lying in the interfollicular space as a normal finding in elderly humans. Sugiyama (1971) demonstrated parathyroid glands intrathyroidally in 9.1%of the cases. Taking into consideration the different methods of investigation, our finding of only two parathyroids in the thyroid tissue, one of them associated with a SCN and adult adipose tissue, is comparable with the above cited percentage. References

Bargmann,W.: Die Schilddriise.In: M611endorff,W. v., Handbuchder mikroskopischenAnatomiedes Menschen, Bd. 6, Teil 2, pp. 1-136. Berlin: Springer 1939a

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Solid Cell Nests and C Cells in Thyroid Glands

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Accepted December 3, 1978