TF-IUPS160007 73..76


BIOGRAPHICAL ITEM

From phylogeny into ontogeny with Claes Hellerstr€om

Kjell Asplund

Riksstroke, Medicine, Department of Public Health and Clinical Medicine, Umeå University, Umeå, Sweden

My research career started with snakes. For a year, from 1965
to 1966, Claes Hellerstr€om and I worked together in herpetol-
ogy (the branch of zoology that deals with amphibians, includ-
ing frogs, toads, and reptiles). We joined forces with an SAS
pilot, Captain Florin, who, between the flights to Los Angeles,
was chairman of the Uppsala Herpetological Society. In his
spare time, he collected rattlesnakes in the Californian deserts,
transported them in the cockpit of his plane, and delivered
them to us. Our second supplier was a herpetologist located
close to Arlanda airport. Vipers on demand—as we needed
snakes, he went out into the forest and collected them for us.

Why snakes? In histology textbooks from the 1950s, the
pancreatic islets were described as being composed of three
cell types: alpha (A), beta (B), and delta (D). It had long been
known that the beta cells produce insulin. Although a sub-
stance with hyperglycemic properties—glucagon—had been
detected in pancreatic extracts as early as 1923 (1), it was not
until the early 1950s that it was discovered that alpha cells
produce glucagon (2).

In 1960, Claes Hellerstr€om and his PhD supervisor Bo
Hellman described a silver impregnation technique that made
it possible to distinguish between two types of alpha cells,
which they termed A1 and A2 cells (3,4). They made the case
that the A2 cells were the glucagon producers. The question
then remained: what is the function of the A1 cells? This is
where phylogenetic studies, including those on reptiles, came
into the picture.

The A1 cell enigma

Silver-positive A1 cells were found to be relatively uncommon
in mammals; beta cells outnumbered other islet cell types by
far. Hellman and Hellerstr€om had therefore been conducting
some of their early silver impregnations in pancreatic islets of
chickens and ducks, where alpha cells were more abundant
(3). However, even in birds only a minority of the alpha cell
population was silver-positive (A1 cells). Was it possible they
could be more prevalent in animals earlier on the evolution-
ary chain?

In Claes Hellerstr€om’s PhD thesis from 1963, he included
not only classical histological studies (on the two types of
alpha cells) but also a method to isolate pancreatic islets
from mice, a procedure that paved the way to cell physiology

studies—a more progressive and successful route (5) com-
pared to traditional histological studies.

Nevertheless, Hellerstr€om recruited me in 1964 as one of
his first two research aspirants (the other aspirant is now edi-
tor of this journal); I was to help him go back into phylogeny.
Hellerstr€om, together with his supervisor Bo Hellman, had
already plunged into herpetology. In 1962, they published a
paper on the pancreatic islets of bullfrogs (6). Now that
Hellerstr€om was on his own, he took on snakes and turtles. If
he could find a species where A1 cells were particularly com-
mon, he could get a glimpse of what was the function of
these mysterious cells. He could perhaps even isolate them
and study their function in vitro.

When we dissected the rattlesnakes and the vipers, we
were somewhat surprised to see how easy it was to find our
orientation. The location of organs was similar to that of
mammals, except in a more elongated, Giacometti-like style
(though I had not encountered Giacometti’s art at that time).
The second remarkable observation was that we did not
need a microscope to see a pancreatic islet. There was a sep-
arate, millimeter-sized organ, next to the spleen, a megastar
compared to islets in other species. In the microscope, we
saw islets dispersed throughout the pancreas but, as shown
in Figure 1, they appeared to accumulate both in number
and in size in the caudal direction, culminating in one or two
extremely large islets next to the spleen (7). It seemed that
this caudal movement had difficulties in stopping; in the
spleen, devoid of surrounding exocrine pancreatic tissue,
typical pancreatic islets were also present (Figure 2).

The hypothesis that A1 cells are more abundant in snakes
than in mammals was thereby confirmed (Figure 1). Half of
the islet cell population were alpha cells, most of which were
silver-positive, i.e. A1 cells (7). In stark contrast to islets in
mammals, only a minority of the islet cells in snakes were
beta cells. Staining for granules, we found distinct granulation
in both types of A cells. When we studied fresh squash prepa-
rations of microdissected islet tissue or frozen pancreatic sec-
tions in dark-field illumination, the islet cells exhibited a
distinct luminescence (7). Taken together, it seemed that the
A1 cells were producing a hormone different from insulin and
glucagon. What was it?

At the time, in 1966, we simply did not have the tools
available to proceed to explore the A1 cell enigma. Also, our

CONTACT Professor Kjell Asplund kjellasplund1@gmail.se Riksstroke, Medicine, Department of Public Health and Clinical Medicine, Umeå University,
SE-90185 Umeå, Sweden
� 2016 The Author(s). Published by Taylor & Francis. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.
org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited

UPSALA JOURNAL OF MEDICAL SCIENCES, 2016
VOL. 121, NO. 2, 1–000
http://dx.doi.org/10.3109/03009734.2016.1152332



enthusiasm for snakes had started to wane as an epidemic of
salmonella struck the animal quarters in the basement of our
institution. The culprits proved to be the rattlesnakes. Then,
Captain Florin was bitten by his puff-adder, hovered between
life and death, and was never able to fly again. This put a
decisive end to our interest in snakes.

Immunohistochemistry provided the answer:
somatostatin

Nearly a decade passed. Histology had become an academic
discipline that by the end of the 1960s seemed to have
entered a scientific cul-de-sac. Claes Hellerstr€om had arrived

at this conclusion early on; he turned from cell morphology
to cell physiology.

Then came immunohistochemistry. It had been concep-
tualized back in the 1940s. But it was not until the 1970s—
when the techniques to produce a wide range of specific
antibodies became more readily available—that immunohis-
tochemistry became a frontline technique. It vitalized, even
revolutionized, histology. It provided scientists with the tools
to reveal the secrets of cells with unknown or unclear func-
tions throughout the body, including A1 cells in pancreatic
islets. Researchers at Karolinska Institute found somatostatin-
containing cells in a number of endocrine tissues. In 1975,
in collaboration with Claes Hellerstr€om, they published an
article in which they reported that A1 cells contained som-
atostatin (8). The A1 cell enigma was thereby solved. In the
mid-1980s, a Canadian immunohistochemist demonstrated
that somatostatin was present in the pancreatic islets of
snakes as well (9).

Reflections on our phylogeny studies

Half a century has now passed since we published our snake
paper. Upon re-reading it, a few spontaneous thoughts come
to mind.

First, even though both A1 cells and snakes turned out to
be scientific dead ends, Claes Hellerstr€om found the ideal
beginner’s project for a researcher-to-be like me. The snakes
were thrilling, the project was well delineated and perfectly
feasible (the results were published within one-and-a-half
years), Hellerstr€om gave me a well-defined role in the project,
and we made some original (although not very sustainable)
observations.

Second, we published our article in Zeitschrift f€ur
Zellforschung und Mikroskopische Anatomie. In the mid-1960s,
a journal with a grandiose German name was still prestigious.
At the time our paper was published, traditional histology
remained tainted by its German cultural heritage. In Uppsala,
student textbooks in anatomy were in German. But Zeitschrift
f€ur Zellforschung und Mikroskopische Anatomie had started to
adapt to the cultural shift in medical sciences and published
articles not only in German but also in English (ours was in
English). It survived by changing its name to Cell and Tissue
Research Journal.

Third, it was fascinating to be part of a soon-to-be extinct
species of scientists—those involved in the upfront morph-
ology of pancreatic islets. In our 1966 article, we cited a num-
ber of papers dating all the way back to the nineteenth
century. When Hellerstr€om found out that that Vincenzo
Diamare, who had published a comprehensive study on the
comparative morphology of pancreatic islets in 1899 (10), was
still alive, we dedicated our paper to him on his 95th birth-
day. Just in time, as it turned out. Diamare died later in 1966
(11). Despite the fascinating structure of snake islets and the
ease with which endocrine tissue can be isolated—it can be
done with the naked eye—very few original articles on snake
islets have been published in recent decades (only one since
the year 2000, according to the Web of Science). Our paper
was last quoted in 1992.

Figure 1. Silver impregnation (Hellman–Hellerstr€om technique) of the pancreas
of a rattlesnake. A large islet containing numerous blackened Al cells is located
close to the spleen. Reprinted from reference 6, Hellman and Hellerstr€om, with
permission from Springer.

Figure 2. Islet from a viper imbedded in the spleen (aldehyde-fuchsin trichrome
staining). The insulin-producing beta cells are stained dark. Note the lack of sur-
rounding pancreatic acinar cells and the localization of beta cells in the periphery
of the islet. �400 in the original publication. Reprinted from reference 6,
Hellman and Hellerstr€om, with permission from Springer.

74 K. ASPLUND



A fourth note is that our findings on argyrophilic A1 cells
were ephemeral. The 1975 immunocytochemistry article
on somatostatin described above, with Hellerstr€om as a
co-author (8), solved a problem that had confused us when
we were working with the snakes. When Hellman and
Hellerstr€om first identified silver-positive A1 cells, they
described them as being distinctly different from D cells
(3,4). We apparently confirmed this in snakes, where we
found D cells to be agranular, hinting that this cell type did
not produce hormone. As immunohistochemistry came
along, it became apparent that A1 cells and D cells were
identical—both contained somatostatin. Since then, our
knowledge about the pancreatic islets as an endocrine
organ has grown. Not only are there alpha, beta, and delta
cells, but another distinct cell type that produces pancreatic
polypeptide has also been added to the list (12). Whether
or not there even are epsilon cells in the adult human pan-
creas (if so, producing ghrelin) seems to be controversial
(13). But the A1 cells have remained unheard of since 1980
(and they do not represent the only dead end in my long
publication list).

Interestingly, Lars Grimelius, an Uppsala pathologist,
worked on another silver impregnation method during the
1960s. In pancreatic islets, his method apparently impreg-
nated glucagon-producing A2 cells instead of A1 cells (14),
which is somewhat of a paradox. As the Grimelius method
was then found also to be useful for staining neuroendocrine
tissue in other organs, it became routine in histopathology.
Grimelius’s original description of the method was published
in the Upsala Journal of Medical Sciences (at the time Acta
Societatis Medicorum Upsaliensis) and is, by far, the most cited
article ever published in this journal (15).

Into ontogeny

During the early 1960s, Claes Hellerstr€om’s interest in phyl-
ogeny was paralleled by his curiosity in ontogeny. Together
with others in the Uppsala group, he published studies on
the morphology of pancreatic islets in human fetuses (16)
and in neonatal rats (17,18).

During the same period, Claes Hellerstr€om’s interest
turned from morphology to function. It is telling that the
same year that our morphological paper on the pancreatic
islets of snakes was published (1966), Hellerstr€om published
his first article on the metabolism of isolated mouse islets
(19). He saw the limitations of traditional histology and the
great potential in the emerging new cell physiology. The
turn from phylogeny to ontogeny and from morphology to
function was a strategic decision, which soon proved to be
rewarding.

So, ontogeny and function—how to combine these new
research directions? Hellerstr€om was a certified physician
and sought applications for his research that were more
practical than snakes could offer. In the 1960s, pregnancy
was often seen as a dreaded complication for women with
diabetes. There was a considerable mortality among preg-
nant mothers, there was a high fetal and neonatal mortality,
malformations were common, and delivery was complicated

by the excessive size of the babies. Early after birth, infants
born to mothers with diabetes were prone to develop
hypoglycemia.

This situation raised a number of basic questions. How do
insulin biosynthesis and release mechanisms mature during
fetal life? Is functional maturation affected by the mother’s
hyperglycemia? Could experimental studies in the offspring of
pregnant rats also provide some insights into how diabetic
pregnancy should be managed to avoid harm to the fetus
and the newborn? This became the themes of my PhD stud-
ies under Claes Hellerstr€om’s tutorship.

Now that I read those papers half a century later, I am less
sure that Hellerstr€om (and certainly not I) had these clinically
very important questions in mind when the project started.
Judging from what we wrote in our Introduction and
Discussion sections, we were entirely driven by curiosity in
the beginning. As the studies progressed, a shift can be dis-
cerned with more and more focus on clinical problems and
clinical lessons. By the time I left Uppsala in 1972, we had
found evidence that, in rats, the mechanisms for insulin bio-
synthesis and release normally do not mature until after birth
(20–24). This would provide protection not only against exces-
sive fetal growth in diabetic pregnancy but also against post-
natal hypoglycemia in the infant. In poorly regulated diabetes
during pregnancy, fetal exposure to hyperglycemia causes
early maturation of the insulin biosynthesis and release mech-
anisms with high insulin levels in the fetus. We concluded
that this would increase the risk of fetal macrosomia (and
ensuing complications during delivery) and a high risk of
hypoglycemia early after birth.

Reflections on our ontogeny studies

The clinical lesson appeared to be simple: to protect against
premature functional development of the pancreatic islets,
strict metabolic control must be maintained during diabetic
pregnancy. This is self-evident in today’s clinical practice, but
such was not the case in the early 1970s. Instead, a common
strategy was to advise young women with diabetes to avoid
becoming pregnant. Our observations were certainly not the
major contributor to the shift in the perception of how dia-
betic pregnancy should be managed, but they were one of
many pieces of evidence that promoted a shift in the thera-
peutic paradigm. When I left the Department of Histology for
clinical practice and research, Hellerstr€om recruited Ulf
Eriksson to study experimental diabetic pregnancy in more
detail; this is described in Eriksson’s contribution to this vol-
ume (25).

A lesson for me on a more personal level concerned Claes
Hellerstr€om’s tutoring style. I was the first PhD student to
graduate under his leadership. He must have had an intuitive
gift for scientific leadership, maintaining a careful balance
between giving the student an increasing level of independ-
ence and the need for scientific rigor. Revisiting my earliest
publications, it is telling that I was the sole author of several
of them. This illustrates not only the shift in publication prac-
tice that has occurred since then, but also Hellerstr€om’s mod-
esty and generosity as a tutor.

UPSALA JOURNAL OF MEDICAL SCIENCES 75



I did not fully realize the quality of my early scientific train-
ing under Claes Hellerstr€om’s leadership until I turned to clin-
ical research. In the clinic, there was an older generation of
researchers whose scientific training mostly had been learning
by doing. But, like me, several of my young colleagues in the
university hospital had a PhD in preclinical experimental
research. We clearly benefited from the elementary scientific
training that we had obtained; we came to represent a new
generation of clinical researchers.

A final reflection is that our work with pancreatic islets
from fetal and newborn rats signified yet another foresighted
shift in Claes Hellerstr€om’s scientific focus—from normal to
pathological states. In the last work I did under his tutorship,
we induced intermittent hyperglycemia in pregnant rats and
studied the effects on the fetuses (24). Hellerstr€om soon went
on to study an abundance of different animal models of
diabetes.

Summing up Claes Hellerstr€om’s years of transition

Thus, the years I worked with Claes Hellerstr€om in the 1960s
and early 1970s were transitional, both in scientific paradigms
and for Hellerstr€om as a researcher. We experienced the end
of a long era of German-oriented morphological research.
Hellerstr€om took a strategic decision to shift from morpho-
logical to functional studies and from traditional histology to
cell biology. He also abandoned his old interest in phylogeny
and focused on ontogeny. And he made a gradual transition
from studying normal to pathological states. It was a privilege
to accompany him on a small part of his innovative scientific
journey, now some 50 years ago.

Disclosure statement

The author report no conflicts of interest. The author alone is
responsible for the content and writing of the paper.

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76 K. ASPLUND


	From phylogeny into ontogeny with Claes Hellerström
	The A1 cell enigma
	Immunohistochemistry provided the answer: somatostatin
	Reflections on our phylogeny studies
	Into ontogeny
	Reflections on our ontogeny studies
	Summing up Claes Hellerström’s years of transition
	Disclosure statement
	References