Okajimas Fol. anat. jap., 42 : 63-89, 19E6 Electron Microscopic Studies of the Follicle Cells and

22 Pages·2012·8.27 MB·English

Okajimas Fol. anat. jap., 42 : 63-89, 19E6
Electron Microscopic Studies of the Follicle Cells
and Parafollicular Cells in the Thyroid
Gland of the Primates
By
Tsuneto Aoi
Department of Anatomy and Third Department of Internal Medicine,
Nagoya University School of Medicine, Nagoya, Japan
(Directors : Prof. Dr. Shooichi Sugiyama
and Prof. Dr. Kozo Yamada)
Electron microscopic observations of the thyroid gland have
been repeated in many mammals such as dogs and rodents, to study
its unique and complicated situation of secretion and it is well-known
that (1) the follicle cells line the follicle cavity by their apical surface
and have an intimate relation with the capillaries at their basal
surface, and (2) hormone substance secreted into the follicle cavity
and stored there is kept in readiness to be released into the circu-
lation. Notwithstanding this as far as the author is aware of, there
exists little description of electron microscopic observations of the
thyroid gland of primates (M o n r o e, '53). Recently, another kind of
cell in the follicles—parafollicular cells (light cells)—have been re-
ceiving electron microscopic interest. Some have regarded them to be
modified follicle cells and others to be an independent type. The main
purpose of the present study is to elucidate electron microscopically
these problems and at the same time to compare the findings with
previous results known of lower mammals, by using primates.
Materials and Methods
Materials consisted of thyroid glands taken from 9 healthy adult
male monkeys of 3 species, Macaca fuscata yakui, Macaca mulatta and
Macaca irus, bred in the Japan Monkey Center in Inuyama City, Aichi
Prefecture.
Immediately after blood letting from the left carotid artery under
nembutal anaesthesia, the thyroid glands were removed and the left
lobes were cut into small pieces for electron microscopy. The pieces
63
64 T. Aoi
were fixed in 1% osmium tetroxide solution buffered at pH 7.5 with
veronal acetate (P .a 1 a d e, '52) or in the same solution buffered at
pH 7.5 with sodium phosphate (M illoni g, '61). After fixation for
2 hours the pieces were rapidly dehydrated in a graded series of
alcohol and embedded in Epon 812 (L u f t, '61). Ultrathin sections
were cut with a Porter-Blum microtome by using glass knives and
were mounted on grids without formvar or collodion film. The sec-
tions were stained singly with lead hydroxide (W a t so n, '58) or
combining saturated uranil acetate (W a t so n, '58) and lead citrate
(R eynold s, '63). Stained sections were examined with an electron
microscope HU-11 A type. Electron micrographs were made at initial
magnifications of 2000 to 10000 and subsequently enlarged photogra-
phically to the desired sizes.
Observations
Electron microscopical differences were not found between the
follicle cells of the three monkey species examined.
From the apical surface, finger-like microvilli bounded by the
plasma membrane were seen protruding into the colloid (figs. 1, 3, 4
and 5). They were of different sizes. The lengths ranged from 0.1
to 0.8p and the widths from 0.1 to 0.2p. They increased in number
and length near the terminal bars. Their matrix showed almost the
same density as that of the superficial cytoplasmic zone but contained
no special structure such as secretory granules and vesicles. The
plasma membrane found between the microvilli, especially around
their bases, formed sometimes tiny indentations or short invagina-
tions. It could not be determined whether they are related to pino-
,cytosis or to secretion (fig . 1).
On the lateral surface of the follicle cells, the plasma membranes
•were separated by a narrow intercellular space of almost the same
width and of low electron density. The two adjacent lateral plasma
membranes had distinct terminal bars along the apical margin, and
.a few desmosomes somewhat deep below the terminal bars (figs. 1, 2
-and 5). The two adjacent lateral plasma membranes, together with
the cytoplasmic matrix, formed interdigitations, simple and compli-
cated. The simple interdigitations were found near the desmosomes
and the complicated interdigitations in places where more than two
follicle cells were in contact (figs. 3 and 5). The complicated inter-
digitations were sometimes transformed into microvilli, protruding
into the intercellular spaces dilated near the interstitium.
EM of Follicle Cell and Parafollicular Cell in Primate Thyroid 65
On the basal surface, the plasma membrane often formed invagi-
nated caves of different sizes, towards which short and twisted
microvilli projected (figs. 2 and 5). The basement membrane was
moderately electron dense and covered usually simply the basal sur-
faces limited by the plasma membranes of the follicle cells. It did
not extend into the caves and dilated intercellular spaces near the
interstitium but rather bridged over them (figs. 1 and 3). The caves
and spaces were continuous with the ordinary intercellular spaces but
were not found communicating with the follicle cavity.
The nuclei were round to oval and situated chiefly in the basal
part of the cell body (fig. 1). They were limited by a double mem.-
brane. The outer membrane was studded unevenly with ribonnucle-
°protein particles. Along the inner membrane 'there were localized
aggregations of granules of high density. The nucleolus was found
as usual.
Just below the apical plasma membranes, the superficial cyto-
plasmic zone was seen as a narrow zone, free of mitochondria Land
rough-surfaced endoplasmic reticulum but contained more or less
numerous small vesicles. The small vesicles were limited by a limiting
membrane and were similar in appearance to the Golgi vesicles. In
the deeper part, rough-surfaced endoplasmic reticulum, mitochondria,
Golgi complex, secretory granules and other inclusions were found.
The rough-surfaced endoplasmic reticulum was well-developed
and seen around the nucleus, especially basal or lateral to it. It
consisted of dilated round to oval cisterns and often dilated irregular-
shaped canals, which were limited by a single membrane and contain-
ed material of less electron density. Its outer surface was associated
with ribonucleoprotein particles. The ribonucleoprotein particles
were more abundantly distributed in a part where the mitochondria
were in contact (figs. 1 to 5).
The mitochondria were distributed almost evenly throughout the
cell body, but slightly abundant in the basal half of the cell body.
They were generally oval to rod-shaped, sometimes more ,elongated.
The outlines were relatively smooth. They were limited by a double
membrane and the cristae were perpendicular to the long axis of the
mitochondrion, but sometimes oblique to it or ruffled. The matrix
was less electron dense and contained no other structures (figs. 2, 3,
6 and 6).
The Golgi complex was always located just above or lateral to
the nucleus. It consisted of abundant small vesicles, a few vacuoles
and several arcuated lamellae. The Golgi complex was found closely
66 T. Aoi
intermingled with cisterns of the rough-surfaced endoplasmic reticu-
lum and numerous small secretory granules (figs. 1 and 5).
Secretory granules were found numerously in the cytoplasm.
They were always bounded by a limiting membrane and were gen-
erally homogeneous. They were divided into three groups with the
some intermediate variability according to the electron density (fig.
1). The first group were 0.1 to 0.4,u in diameter, less dense, and found
chiefly in the apical part of the cell body and in the Golgi zone. The
second were 0.4 to 0.7p in diameter and moderate dense. They were
observed chiefly lateral to the nucleus but some in or near the Golgi
zone. The third were dense and were 0.3 to 1.3p in diameter. Some
dense granules were large and inhomogeneous and contained vesicles
and vacuoles of different size. A few of them had whorled paired
membranes and a double limiting membrane. They were chiefly seen
in the apical or basal part of the cell body. As other inclusions, ag-
gregates of vesicles or multivesicular bodies were found. Pseudo-
podium-like projections were very rarely found and projected merely
as a single but not paired irregular-shaped formation towards the
cavity. They contained rarely secretory granules (fig. 4) and appeared
rather like an apocrine projection.
As observed in other animals (Z i m m e r m a n, 1898 ; K a n o, '52,
in man ; Muramot o, '64, in pigeons ; T a shir o et al., '64, in dogs)
a flagellum was rarely seen projecting from the apical surface to-
wards the follicle cavity (fig. 2).
In flattened follicle cells, the microvilli were fewer and short (figs.
3 and 5). The superficial cytoplasmic zone was very thin and con-
tained no vesicles.. Two adjacent lateral plasma membranes and
intercellular spaces in between ran partly oblique or almost parallel
to the apical or basal surface. The Golgi complex was located lateral
to the nucleus and consisted of several arcuated lamellae and a
number of small vesicles. Rough-surfaced endoplasmic reticulum and
secretory granules appeared to be reduced in number and size.
Parafollicular cells were almost alike in appearance in the three
monkey species. Parafollicular cells were always in direct contact
with the follicle cells but did not line the follicle cavity directly.
They were generally ellipsoidal in shape but partly deformed when
in groups (fig. 7). The plasma membrane showed the same density
as that of the follicle cells. The intercellular spaces between the
parafollicular cells and follicle cells were found limited by two
adjacent plasma membranes (figs. 8 and 9). In the two plasma
membranes, desmosomes rarely but simple interdigitations sometimes
EM of Follicle Cell and Parafollicular Cell in Primate Thyroid 67
were found (fig. 7). No complicated interdigitations were formed
here. The basement membranes were continuous directly with that
of the follicle cells.
The nuclei were situated centrally or eccentrically in the cell
body and oval in shape (fig. 7).
Rough-surfaced endoplasmic reticulum associated with abundant
ribonucleoprotein particles was found as a small number of vesicles
and/or a few to several long paired thin lamellae. The thin lamellae
were usually arranged in parallel rows and found in a corner of the
cell body (figs. 7, 9 and 10). These lamellae were not dilated—poor
development of rough-surfaced endoplasmic reticulum. This was one
of the characteristics of the parafollicular cells.
Mitochondria were also somewhat -different from those of follicle
cells. Firstly, they were slightly smaller in number and size than
those of the follicle cells. Secondly, they were generally round to
oval but rarely rod-shaped. Their outlines were sometimes ruffled
rather than smooth. The cristae were very irregularly arranged and
often twisted. This was the second characteristic (figs. 7, 8 and 10).
The Golgi complex was well-developed and appeared as a horse-
shoe-shaped structure consisting of numerous small vesicles, a few
vacuoles and several long thin lamellae (fig. 8).
The parafollicular cells were further characterized by the pre-
sence of round to oval vesicles which were distributed in abundance
and evenly throughout the cell body. They were bounded by a limiting
membrane, approximately 0.3 to 0.5p in diameter and different in
electron density (figs. 8 and 10).
The secretory granules were of two types (figs. 7, 8 and 10), one
dense and the other moderate dense. The dense secretory granules
were smaller in number and approximately 0.9p in diameter. Some
appeared like lysosomes and contained vacuoles. The moderately
dense secretory granules were smaller in diameter. The two types
of secretory granules were also found near the Golgi zone and some
were indistinguishable from the Golgi vesicles. These secretory gran-
ules were sometimes in contact with the basal plasma membrane.
Very rarely, a flagellum was found near the Golgi complex and
showed the same transverse section as that of the usual flagellum
found in other kinds of cell (fig. 7).
Perifollicular capillaries were composed of endothelial cells. The
endothelial cells were extremely attenuated in many regions except
in the nuclear region and its neighbourhood. Discontinuities or fenet-
rations which were bridged by a single membrane—the diaphragms
68 T. Aoi
were often found. Further, pinocytotic vesicles were very often
found. The basement membranes were of moderate electron density
and were approximately 0.1tc in width. The basement membrane
surrounded the perifollicular capillaries completely.
Discussion
Monroe ('53, in rats and monkeys) and Br a u n stein e r et
al. ('53, in rats and guinea pigs) pointed out that the production of
thyroid hormone is carried out in the rough-surfaced endoplasmic
reticulum of the follicle cells. Dempsey et al. ('55, in rats) stated
similarly that the secretory product is accumulated and condensed in
the sacs of rough-surfaced endoplasmic reticulum and are transformed
into secretory droplets by losing ribonucleoprotein particles from their
surface. The concept has been supported by several other authors
(W a n g, '58, in rats ; F u j i t a et al., '58a and b, in chicks ; S u g a-
w a r a, et al., '59, in man ; I r i e, '60, in mammals ; Roos, '60. in
rats).
Herman ('60, in salamanders) and Fuji t a et al. ('63, in rats)
suggested a possibility that both rough-surfaced endoplasmic reti-
culum and Golgi complex can produce hormone proteins. Harrison
et al. ('62, in seals) observed that secretory droplets of different size
and density are seen extending from the Golgi area towards the apex
of the follicle cells. W i s s i g ('60, in rats) considered that homo-
geneous material contained in the rough-surfaced endoplasmic reti-
culum represents a precursor of the secretory granules and empha-
sized that the secretory granules are derived from the Golgi complex
because of the marked similarlity to vesicles of this organelle. H e
('63, in rats) demonstrated the colloid droplets to be formed from large
vesicles of the Golgi complex and to release their content into the
follicle cavity in stimulated thyroid glands. Contrary to this. Nadler
and his colleagues ('62 and '64) repeatedly suggested that the colloid
droplets are a result of pinocytosis and originate in the follicle colloid.
Stein et al. ('64) agreed with this by radioautography of radioiodine
1125. By electron microscopic radioautography, Nadler et al. ('64)
elucidated that the follicle cells synthesize two kinds of proteins,
sedentary and exportable, in the rough-surfaced endoplasmic reti-
culum and that the exportable protein migrates via the cisternae of
the rough-surfaced endoplasmic reticulum, the Golgi zone and apical
vesicles to be added to the follicle colloid, while the sedentary remains
relegated to intracellular structure.
EM of Follicle Cell and Parafollicular Cell in Primate Thyroid
A survey of the literature above seems to reveal that the follicle
cells can contain two kinds of secretory granules, excretory and it-
cretory, which are morphologically indistinguishable, and that the.
excretory are primarily matured in the Golgi apparatus and migrate
as those like Golgi vesicles and partly changing secondarily in quality
and quantity by unknown factors towards the apical zone, while the.
incretory originate in the follicle colloid. There is no doubt that the
three kinds of secretory granules with some intermediate variability
observed in the present study can be more or less included in these
excretory and incretory secretory granules (figs. 1 to 6). M u r a-
m o to ('64, in tortoises and pigeons) and T a s h i r o et al. ('64, in
dogs) could not explain the significance of secretory granules of dif-
ferent electron density and sizes in the follicle cells, but suggested
that their density and size may depend upon the molecular weight
of the changing hormone substance which contains thyroxine or its
precursor as a core and that less dense granules of low molecular
weight move to the apical plasma membrane. The present author
agrees in major part with their opinion, considering another possi-
bility of transfer of secretory granules through the basal plasma
membrane.
Some dense granules observed here appear like lysosomal gran-
ules and are different from those observed by the two authors
(M u r a m o to and T a s h i r o) in the following points-1. frequent
inhomogeneity and inclusion of vacuoles and vesicles and 2. whorled.
paired membranes and double limiting membrane which may be sup-
posed to be produced by degeneration of the organellae, probably
mitochondria (figs. 1, 3, 4, 5 and 6). M u r a m o to ('64) found them
rarely in tortoises.
Electron microscopic studies combined with histochemistry sum-
marized that the follicle cells have numerous adielectronic granules.
which contain acid phosphatase and esterase and these granules.
hydrolyze the colloid droplets (L arse n, '65). In the Ambystoma
follicle cells homogeneous granules have been found to contain acid.
hydrolase (W e t z el et al., '63 ; W o 11 m a n et al., '64). The origin
of the lysosomes in other glandular cells has been elucidated to be in
the Golgi apparatus by demonstrating the presence of acid phos-
phatase here (Essner and Novikoff, '60; Osinchak, '64).
This has been histochemically identified in the Golgi area of the
follicle cells (S o b e 1, '62, in rats). On the other hand, the other
origin has been demonstrated to be in the process of autophagy
salamander follicle cells (L a r se n, '65). The origin of granules like_
70 T. Aoi
lysosomes observed here seems to be not only in the Golgi apparatus
but also in the cellular debris.
A few words should be mentioned about other formations. The
microvilli formed in the caves of the basal surface of the follicle
cells may be comparable with the apical microvilli and may be
merely significant as augmenting the effective surface of function
because they contain no minute structures such as vesicles, vacuoles
and secretory granules (figs. 3 and 5). The dilations and the basal
caves do not appear to communicate further with the follicle cavity
as an intercellular canal (K a n o, '52 ; Y o s h i m u r a et al., '59) or
as an intracellular canal (Y o s h i m u r a et al., '65, in rats) which
serves the transfer into the circulation of the follicle colloid.
Pseudopodium-like projections occur very rarely also in normal
primates and some appear like an apocrine projection (fig. 4) but
remain still unknown. Wi s s i g ('63, in rats) has described the
pseudopodia which are utilized in releasing the content of mature
droplets into the follicle cavity in stimulated glands. P o n s e ('38)
reported in stimulated glands that the pseudopodia serve rather to
take up the follicle colloid into the cell body. Williams ('38)
confirmed this by observing living follicles implanted in transparent
chambers installed in rabbits' ears. T a s h i r o et al. ('64, in dogs)
described that the pseudopodium-like projections occur relatively
often in thyroid glands stimulated by overdose of thyrotrophin and
suggested a possibility reported by William s.
The second cell type referred to as "parafollicular cell " in this
paper has repeatedly been studied in different mammals and has
been given different names, such as protoplasmareiche Z e 11 e n
(H U r t h l e, 1894), parafollicular cells (N o n i d e z, '32, in dogs),
gray cells (G o d w i n, '37, in dogs), Makrothyreocyten (L u d w i g,
'54
, in rats), and light cells (S t u x et al., '61, in rats). Their origin
also has been one of the main subjects of investigation. Until today,
two different origins have been considered ; 1. They are derived
from the follicle cells and 2. originate from the ultimobranchial
body. Wilson ('27/'28, in man) and Ludwig ('53, in rats, guinea
pig and dogs) considered them merely as tangential sections of
follicles, and later, by Ludwig ('54, in rats and dogs) as follicle
.cells beginning mitosis. I s e n s c h m i d ('10
, in infants) found that
the interfollicular cells occur through depletion of colloid in the
follicles. B e r n a r d ('27, in dogs) stated that they are cells of the
follicles compressed after depletion of the colloid. Sander so n-
_D a m b e r g ('11, in man) and F e y r t e r ('53, in man) reported
EM of Follicle Cell and Parafollicular Cell in Primate Thyroid 71
that they develop by the budding process of the follicle walls.
N o n i d e z ('32, in dogs ; '33, in mammals) and Raymond ('32, in
rabbits) found that the parafollicular cells develop in the follicle wall
and migrate from here towards the interstitium. Sarker et al. ('64,
in rats) confirmed this experimentally by observing the increase of
light cells after treatment with growth hormone or after hypophy-
sectomy. This origin from the follicle cells has been further sup-
ported by many investigators (0 h k u b o, '35, in dogs ; S u g i y am a,
'39
, in rats ; '50, in rabbits ; G a b e, '59, in dogs ; S tux et al., '61,
in rats ; Yoshimur a et al., '62, in rats ; Id elma n, '63, in
domestic mammals ; I t o, '65, in mammals).
Godwin ('37, in dogs) found that parafollicular cells develop
from the ultimobranchial body. Van Dyke ('45) found two kinds
of interfollicular cell in sheep and pointed out that one originate
from the follicle cells of broken follicles and the other from the
ultimobranchial body. Recently, Dumont ('56 and '58, in rabbits)
suggested that the parafollicular cells are distributed around the
remnants of the thyroglossal duct from which they take their origin,
and derived from the 4th branchial cleft. Similarly, Sato ('59, in
hamsters) supported this origin by observing them more frequently
in and near the residual cysts of ultimobranchial origin.
These views are also suported by electron microscopic observa-
tions. Some suggest that reversible development between the two
kinds of cell is possible because of similarity in cytoplasmic com-
ponents (L u c i a n o et al., '64, in rats) or that parafollicular cells
are follicle cells which have lost contact with the colloid and are
unable to discharge their secretion due to degeneration of secretory
vesicles (Y o u n g et al., '64, in rats).
The parafollicular cells observed in monkeys are distinguishable
in fine structure from the follicle cells by the following points-1.
the non-dilated and poorly developed rough-surfaced endoplasmic
reticulum consisting of a small number of vesicles and/or a few to
several thin lamellae arranged in parallel rows, 2. smallness and
paucity of the mitochondria, 3. well-developed horse-shoe-shaped
Golgi complex and 4. abundant distribution of smooth-surfaced
vesicles, with no intermediate transitional forms in between. W i s-
s i g ('62, in rats) observed that the parafollicular cells do not develop
from the follicle cell but rather are a second, independent class of
endocrine cells. T a s h i r o ('63 and '64, in dogs) also suggested
that the parafollicular cells resemble the epithelial cells of the
ultimobranchial cyst, with the common cardinal characteristics of
72 T. Aoi
two organellae—poor development and paucity of rough-surfaced
endoplasmic reticulum, and paucity and smallness of mitochondria, and
confirmed that they are completely different in fine structure from
follicle cells and produce no transitional forms in normal and
thyrotrophin-stimulated glands. I s h i k a w a (p65, in rats) demon.
strated by electron microscopy that the parafollicular cells develop
in late embryonic life from the ultimobranchial body, maintaining
common cardinal characteristics of the two organellae.
The problem of the significance of the parafollicular cells has
received some interest. Most of the authors (0 h k u b o, '35, in
dogs ; Arimitsu, '37, in rats ; S u g i y am a, '50, in rabbits ; S a t o,
'59
, in hamster ; Y o s h i m u r a et al., '62, in rats ; Luciano et
al., '64, in rats) have described them to be secretory cells. Previ-
ously, the parafollicular cells have been considered to be secreting
cells which discharge their secretion into the follicle cavity (T a k a-
g i, '22, in dogs). N o n i de z ('31/'32, in dogs) assumed that argyro-
phile granules contained in the parafollicular cells represent the
antecedant of an endocrine secretion poured directly into the vessels.
Luciano et al. ('64) regarded them to be active secreting elements
by observing the presence of numerous vesicles in the basal zone
and in the well-developod G o l g i complex. A z z a l i ('62, in rats,
homster and bats ; '64, in virginia opossum, three fingered slothes,
flying foxes, hamsters and rats) pointed out that the parafollicular
cells possess an activity of their own by observing the difference
of granulations after TSH treatment.
Two kinds of secretory granules observed here (figs. 7, 8 and
10) may correspond to the vesicles of Young et al. ('64, in rats
and Luciano et al. ('64, in rats). The origin of these secretory
granules seems to be from the G o l g i complex because some of
them are found near this organellae and indistinguishable from its
vesicles. The content of the granules in contact with the basal
plasma membrane may suggest a possibility of release of their
content towards the interstitium. The presence of the lysosomal
granules observed here may correspond to the reaction of acid
phosphatase activity demonstrated in Makrothreocytes by Gabe
('59, in dogs). The lipid granules found by Luciano et al. ('64,
in rats) are not found in the parafollicular cells of monkeys. Numer-
ous vesicles characteristic of this kind of cell remain undetermined
in significance and origin.