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Histological observations upon an adult human pancreas (autofluorescence fat and pigment).

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HISTOLOGICAL OBSERVATIONS UPON AN ADULT
HUMAN PANCREAS (AUTOFLUORESCENCE,
FAT AND PIGMENT)
ALLAN L. GRAFFLIN
Department of Anatomy, Harvard Medical School, Boston, Massachusetts
The object of this note is to present briefly some observations which have been made upon an apparently normal human
pancreas from an adult white male, 45 years of age. Study
of frozen sections demonstrates quite numerous fatty inclusions in the cells of both the exocrine and the endocrine portions of this pancreas. In the exocrine portion many of these
inclusions exhibit a natural yellow color and likewise a yellow
fluorescence. I n the islets of Langerhans the inclusions are
uniformly colorless, but exhibit a yellowish-brown fluorescence
of sufficient intensity to demarcate the islets quite sharply
from the surrounding tissue. I n ordinary histological preparations (paraffin sections, hematoxylin and eosin), these fatty
inclusions have completely disappeared, and one would never
suspect that they had originally been present in such
abundance.
The observation has been repeatedly made that fatty inclusions may be present in considerable numbers in the various
glandular organs of man, and that in general these inclusions
increase in number with increasing age. These studies have
been made primarily by pathologists upon autopsy material.
However, from the available data, there is every reason to
believe that this deposition of f a t is an entirely normal process.
I n the widely used textbooks of histology, this process is
almost wholly ignored, so that the presence of considerable
amounts of fat comes as a complete surprise to the student,
and erroneously suggests the presence of pathological change.
207
208
ALLAN L. GRAFFLIN
This circumstance, plus the fact that the material for the
present study was obtained under rather ideal conditions,
constitutes the real justification for this report.
MATERIAL AND METHODS
The individual from whom the pancreatic tissue was obtained came under observation on Dr. Soma Weiss's Medical
Service at the Peter Bent Brigham Hospital. A diagnosis of
islet cell adenoma was made, and the tumor, which was small,
was found and removed at operation by Dr. Elliott Cutler.
At the same time a portion of the tail of the pancreas was
resected, and blocks of this tissue were fixed a t once in 10%
neutral formalin. The specimen is known as Brigham
5-39-1946. The operation was performed on December 6,1939;
frozen sections at 10 to 20 p were cut on December 9th, and
mounted in glycerine either unstained or after staining with
Sudan I11 (saturated solution in 70% alcohol containing 1%
KOH). All of the following observations were completed
within 24 hours after the sections were cut.
Formalin has proved to be the fixative of choice f o r studies
of fluorescence, but even with formalin considerable alterations
in fluorescence may result from long immersion (Haitinger,
'34 ; Hamperl, '34). The fluorescence microscope employed
was the Leitz Ultropak (23X water immersion objective, 6X
and 12X oculars), with a n intense electric arc as a light source,
and a polished Corning filter, Corex (3986, supplemented by
a cooling cell containing a strong solution of copper sulfate,
to filter out the rays of the visible spectrum. Frozen sections,
unstained and stained with Sudan 111, were studied with the
ordinary microscope at x 900 (oil immersion). For studies
with polarized light the Zeiss attachment was used, with
magnifications of from X 80 to X900 (oil immersion). For
ordinary histological preparations blocks were embedded in
paraffin, sectioned at 5 p, and mounted unstained or stained
with hematoxylin and eosin.
HISTOLOGY O F ADULT H U M A N PANCREAS
209
OBSERVATIONS
S t u d y of m s t a i n e d frozest sectiovts with the ordinary
microscope
Eaocriine portion o f pancreas. The serozymogenic cells appear entirely normal. Zymogen granules are present in large
numbers, and an abundance of long, filamentous mitochondria
is usually quite well delineated in the basal half of the cells.
Clear, colorless droplets, variable in size and of low refractility,
are frequently noted in variable numbers (never abundant)
in the basal half of the cells. In addition, droplets and larger
masses (some of them spherical, some mulberry-shaped, some
irregular in shape), exhibiting from a pale to a distinct yellow
color, are seen at times in this region.
The cells of intercalated ducts sometimes contain, in addition to clear, colorless droplets of variable size, masses which
exhibit a distinct yellow color of variable intensity. These
masses differ widely in number and size. They are spherical,
oval or irregular in shape, and the larger ones frequently
appear to have been formed by the incomplete fusion of
numerous smaller masses. The cells of the interlobular ducts
contain numerous droplets, aggregates of droplets and mulberry-shaped masses. Although many of these inclusions are
colorless, the larger ones are often yellow in color. I n the
connective tissue of the gland, which appears normal in
amount, yellow droplets and masses are nowhere observed.
Endocrine portion of p a w r e a s . The islets of Langerhans
are readily identifiable, and appear t o be normal. A very fine
granular (ground glass) appearance of the cytoplasm is commonly noted. The cells contain, in variable numbers, clear
colorless droplets of variable size and of low refractility.
These droplets are distributed at random in the cytoplasm;
a t times they seem to have an almost imperceptible pale
yellow color, but of this one cannot be certain.
210
ALLAN
L. GRAFFLIN
Study o f frozes sections staimed with szcdan IIT
Esocrine portion of pancreas. The great bulk of the serozymogenic cells contain fat. Some of them contain only one
or several small droplets, and there are all intermediates up
t o cells containing abundant fat. Irrespective of amount, this
fat is usually confined to the basal half of the cells, but at
times droplets are either supranuclear or still more apical in
position (actually among the zymogen granules). Not infrequently one observes rather large, mulberry-shaped, sudanophile masses, which appear to have been formed by the
incomplete fusion of numerous smaller masses. The cells of
the intercalated and interlobular ducts contain a variable
number of small fat droplets, and frequently rather large
droplets or mulberry-shaped masses are present. Except for
the zymogen granules, no non-sudanophile droplets or masses,
colorless or yellow, can be identified; in other words, all of
the colorless and yellow droplets and masses observed in
unstained sections have been stained with the sudan 111.
Emdocriqae portion of pancreas. The islet cells with great
uniformity contain within their cytoplasm a variable, and at
times quite large, number of small fat droplets of variable
size. The distribution of these droplets is so variable that
there is no suggestion of a cellular polarity. None of the
colorless droplets observed in unstained sections can now be
detected; i.e., they have all been stained with the sudan 111,
and are fatty in nature.
All of the sudanophile material in this pancreas shows a
surprising uniformity in its staining properties. It is orange
in color, without the red quality exhibited by the ordinary fat
cells.
Study of unstained frozen sections with the polarixi.ng
microscope
All of the fatty inclusions in the serozymogenic cells, duct
cells and islet cells are uniformly non-birefringent, as are
the zymogen granules and the islet cell granules.
HISTOLOGY OF ADULT HUMAN PANCREAS
211
Study of unstaiaed froxeln sectioas with the
$uo resceace micros cope
Exocrine portioii of pamcreas. The accumulations of
zymogen granules exhibit a homogeneous violet fluorescence
of low intensity. The basal portions of the acinar cells are
essentially devoid of fluorescence. Scattered throughout the
exocrine portion of the pancreas one observes rather numerous
granules and masses, variable in size, which exhibit a brownishyellow to golden-yellow fluorescence of moderate intensity.
They are more numerous in some regions than in others, and
their fluorescent intensity varies considerably. With few
exceptions they are definitely less numerous than the sudanophile droplets and masses observed in the seroeymogenic
cells and in the ducts. On the other hand, they are on the
whole larger than the majority of these sudanophile inclusions. Under the circumstances, the conclusion is drawn that
a large number of the sudanophile droplets in the exocrine
portion of the pancreas are devoid of fluorescence ; and that
it is primarily the larger yellow droplets and masses which
exhibit the brownish-yellow to golden-yellow fluorescence here
observed. As noted above, in sections studied with the ordinary microscope these yellow masses are of more frequent
occurrence in the cells of the intercalated and interlobular
ducts than in the serozymogenic cells.
The delicate peri-aeinar connective tissue is devoid of
fluorescence ; the interlobular connective tissue exhibits a
violet or bluish-violet fluorescence of moderate intensity ; and
the walls of the larger vessels exhibit a bluish-greenish-white
fluorescence of considerably greater intensity. A number
of yellow or golden-yellow fluorescent granules are seen in
the interlobular connective tissue.
Endocrine portion of pancreas. The islets of Langerhans
of appreciable size are always sharply demarcated from the
surrounding tissue by virtue of the presence in them of numerous small granules exhibiting a yellowish-brown fluorescence
of moderate intensity. The number and distribution of these
granules is such as to leave no doubt that they represent
212
ALLAN L. GRAFFLIN
the abundant sudanophile droplets previously described. The
nuclei of the islet cells are non-fluorescent, and the basic cytoplasm exhibits a barely perceptible violet fluorescence. There
is no evidence of different cell types characterized by particular
fluorescent properties.
Study of ulzstaimed amd stailzed parafilz sectiom
I n unstained paraffin sections (embedded via chloroform,
deparaffinized with xylol), there is absolutely no trace to be
found of the yellow droplets and masses observed in unstained frozen sections in the serozymogenic cells and in the
ducts. I n paraffin sections stained with hematoxylin and
eosin, the histological structure of the pancreas appears to
be entirely normal; and one would never suspect that the
abundant fatty inclusions described above had originally been
present in the gland.
DISCUSSION
The presence of numerous small fatty inclusions in the
cells of the islets of Langerhans was first described, in a
“quite fresh” human pancreas, by Dogie1 (1893). Stangl ( ’01)
subsequently reported a similar finding in the pancreas of a
30-year-old executed man. I n addition, he described the presence of numerous fatty inclusions, variable in shape and size
and at times quite large, in the acinar cells, where they were
confined primarily to the basal zone; and similar inclusions
were likewise observed, in small numbers, in the centro-acinar
cells and in the epithelium of the intercalated and interlobular ducts. I n a supplementary study of the human pancreas
in all age poups, Stangl observed that fatty inclusions appeared quite early in life in both the exocrine and endocrine
portions of the pancreas, and increased in numbers with increasing age, although the increase appeared to be more rapid
in the first 20 years of life than in the ensuing decades. He
concluded that this fat deposition was an entirely physiological
process. Contrary to the observations of Stangl, Symmers
(’09) concluded that the presence of fat in appreciable amounts
HISTOLOGY O F ADULT H U M A N PANCREAS
213
in the islets was invariably pathologic, and bore an essential
relationship to the prolonged use of alcohol. I n this connection,
it should be stated that the individual from whom the pancreatic tissue was obtained for the present study has always
been a total abstainer.
Nakamura ( '24) could not demonstrate the presence of fatty
inclusions in the pancreas of all of the children he examined,
but did find that the incidence of positive findings increased
with age, as did the amount of demonstrable fat. In agreement with Stangl, he was unable to establish a relationship
between the presence of fat and certain disease processes.
Seki ('28) observed the more or less constant presence of fatty
inclusions, from birth to old age, in both acinar and islet tissue,
the amount of fat increasing in general with increasing age.
While the inclusions in the acinar cells were quite variable,
those in the islet cells were rather uniform in both size and
distribution, in agreement with the findings of Stangl, of
Nakamura, and of the present study.
That the numerous small fatty inclusions in the islet cells
of man exhibit sufficient fluorescence to demarcate the islets
from the surrounding tissue has previously been described
by Hamperl ( '34). He likewise noted the presence of yellow
fluorescent granules in the exocrine portion of the pancreas,
and the present observations are entirely analogous to his.
From his wide experience with human tissues, Hamperl concluded that the yellow fluorescence of the fatty inclusions in
the cells of the acini, ducts and islets of the pancreas is to be
attributed to the presence in them of so-called Abnutzungspigment. Abnutzungspigment is variously referred to in the
English and American literature as wear-and-tear pigment,
old-age pigment and lipofuscin. It is usually considered not
to contain iron, and, according to Hamperl, is regularly characterized by a yellow fluorescence of variable intensity and
quality. It usually appears in increasing amount with increasing age in the ordinary fat depots and in fatty inclusions
in the cells of the various organs and tissues of the body.
214
ALLAN L. GRAFFLIN
It should be emphasized that hemoglobin and its iron-containing breakdown products are devoid of fluorescence, as is
melanin ; and that neutral fat is likewise non-fluorescent
(Haitinger, '34; Hamperl, '34). The occurrence and distribution of fat and pigment in the parenchymal cells of the
thyroid and parathyroid glands have recently been reviewed
(Brafflin, '39), and offer an interesting comparison with the
present observations upon the pancreas.
SUMMARY
Observations are reported upon an apparently normal adult
human pancreas, with particular reference to autofluorescence,
fat and pigment. The cells of the exocrine portion contain a
rather large number of fatty inclusions, many of which are
yellow in color and exhibit a yellow fluorescence. The islet
cells contain abundant small fatty inclusions, which exhibit
a yellowish-brown fluorescence of sufficient intensity to demarcate the islets quite sharply from the surrounding tissue.
The yelIow fluorescence of the fatty inclusions, in both the
exocrine and the endocrine portions of the gland, is attributed
to the presence in them of wear-and-tear pigment (Abnutzungspigment).
LITERATURE CITED
DOGIEL,A. 8. 1893 Zur Frage iiber die Ausfiihrungsgange des Pankreas des
Menschen. Arch. f. Anat. u. Entwickelungsgesch., S. 117-122.
GRAFFLIN,A. L. 1939 The thyroid and parathyroid glands of the Barasingha
deer, with particular reference to autofluorescence, f a t and pigment.
J. Morph., vol. 65, pp. 297-321.
HAITINGER,
M. 1934 Die Methoden der Fluoreszenzmikroskopie. I n : Abderhaldens Handbuch der biologischen Arbcitsmethoden, Abt. 11, T. 3,
S. 3307-3337.
HAMPERL,
H. 1934 Die Fluoresccnzmikroskopie menschliclier Gewebe. Virehows
Arch., Bd. 292, S. 1-51.
NAKAMURA,
N . 1924 Untersuehungen iiber das Pankreas bei FGten, Neugeborenen, Kindern und im Pubertiitsalter. (Mit einem Anhang: Falle
mit Diabetes und Glykosurie.) Virchows Arch., Bd. 253, pp. 286-349.
SEKI,M. 1928 Morphologisehe Studien betreff end Pankreasfette. Trans. Jap.
Path. Soe., vol. 18, pp. 402-411.
STANGL,E. 1901 Zur Histologie des Pankreas. Wien klin. Wochenschr., Jahrgang 14, S. 964-968.
SYMMEW,
D. 1909 The occurrenee of f a t in the islands of Langerhans. Arch.
Int. Med., vol. 3, pp. 279-285.
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