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Original Paper
Acta Anatómica 1992; 143:41—47
Department of Anatomy. Abel Salazar
Institute for the Biomedical Sciences, and
Metallurgic and Material Sciences Center
of the Engineering Faculty.
University of Porto, Portugal:
Department of Anatomy.
Faculty of Medicine.
University of Agostinho Neto.
Luanda. Angola
Key Words
Scanning electron microscopy
Arterial capillaries
Popliteal lymph node
Tracheobronchial lymph node
Paratrachéal lymph node
Blood Vasculature of the Lymph
Node in the Dog:
Anatomical Evidence for
Participation of Extrahilar Arterial
Vessels in the Blood Supply
of the Cortex
Abstract
The organization of the arterial vessels of dog lymph nodes (LN) was studied
using methods of visualization of the vasculature by systemic injection of differ­
ent tracers (colloidal carbon. Micropaque® resin and methylmethacrylate) fol­
lowed by observation of the samples by light microscopy (after clearing of the
thick sections of LN) or scanning electron microscopy (corrosion casts). LN
from all of the three groups of nodes studied (tracheobronchial, paratracheal
and popliteal) showed an extensive network of arterial vessels encircling the cap­
sule of the organ. We found that branches of these capsular arteries penetrated
deeply into the cortical domain of LN. The capsule-originating vessels appeared
to have a significant participation in the blood supply of the LN parenchyma at
the cortical domains of the organs. Our findings are in contrast with current
views on the angiology of the LN that consider that virtually all of the arterial
capillaries of the LN parenchyma come from hilar arteries. We propose, there­
fore. that important segments of the LN cortex receive their blood supply from
capsular arteries rather than from hilar vessels.
Introduction
The lymph nodes (LN) play a central role in host
defence of mammals against foreign infectious agents and
particles [ 1-5]. These lymphoid organs contain all the cellu­
lar elements of the immune system and are capable of trig­
Received:
June 15. 1991
Accepted:
August 20. 1991
gering specific responses directed at antigenic components.
The LN vasculature shows characteristic anatomical
arrangements that are known to modulate the physiology
of the organ, namely lymphocyte traffic and recirculation
[6,7]. Current prevalent views on the micro-anatomy of the
LN vascular system consider that virtually all arteries enter
Nuno Rodrigues Grande. MD, Phi)
Professor and Chairman. Department of Anatomy
Abel Salazar Institute for the Biomedical Sciences
P-4000 Porto (Portugal)
© 1992 Kargcr A G . Basel
0001-5180/92/1431-0041
S 2.75/0
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A.C. Salvador, a , b
A.S. Pereira"
C.M. de Sdc
N.R. Grande1
Materials and Methods
Animals
Nine male and female aduli mongrel dogs were used in this study.
The animals were housed in individual kennels, fed standard commer­
cial dog food and had unrestrained access to water. The dogs were
anaesthetized by intravenous injection of sodium pentobarbital (-40
mg/kgof weight), the abdominal cavity was surgically exposed and the
animals were bled by cutting the inferior cava vein. Before death of
the animals, a cannula was introduced into the abdominal aorta, and
the vascular system of the animals was washed with warm saline. This
was immediately followed by injection of different tracers (see below)
into the arterial system of the dogs to visualize LN vessels.
Injection o f Colloidal Carbon and Micropaque
Colloidal carbon (Pelikan China ink: 30% in gelatin) or 5% gelati­
nated Micropaque were injected either in the external iliac arteries (to
visualize the popliteal LN) or in the upper portion of the abdominal
aorta (to visualize the tracheobronchial and paratrachcal LN).
Injection o f Methylmethacrylate
Methylmethacrylate (MMA: tensol cement No. 70; ICL Chemical
and Polimcrs Ltd.. Darwin. Lanes, UK) was injected into the distal
portion of the femoral arteries (to visualize the popliteal LN) or in the
thoracic aorta at the origin of the bronchial arteries (to visualize the
tracheobronchial and paratrachcal LN).
Fig. 1,2. General views of whole vascular casts of a popliteal (1)
and a tracheobronchial (2) LN from dogs perfused with MMA resin.
The nodes show a high density of vessels coating all of the outer sur­
face of organ in a hair-like fashion. 1 Notice that several large arterial
vessels are scattered around all of the node surface without a precise
indication of the positioning of the Itilus. The vessels shown in the
lower half of figure l are depicted in higher magnification in figure 6.
x 10.
the node through the hilus and that the blood supply of the
organ’s parenchyma comes from branches of hilar arteries
[8-12], According to this view, the few arterial vessels that
reach the LN independently of the hilar arteries are
directed to the capsule and have no or very limited partici­
pation in the blood supply of the lymphoid tissue.
In this work we have investigated the micro-anatomy of
the arterial blood vasculature of dog LN. We integrated
information retrieved by several methods of filling the
lumen of the vessels: resin casts of the LN arteries, sections
of nodes perfused with gelatinated colloidal carbon or
Micropaque® resin, and scanning electron microscopy of
resin casts. We show here that LN have elaborated net­
works of extrahilar arteries. The striking development and
the topography of these arteries suggest that extrahilar
arteries may form an independent blood supply system of
the cortical domain of LN, rather than being restricted just
to the capsular tissue of the node.
42
Processing o f LN
All nodes were surgically removed and processed according to the
method used to trace their blood vasculature. The LN injected with
gelatinated colloidal carbon were fixed in 10% forntol. The organs
were either sectioned into 1-mm layers and xylol cleared or serially fro­
zen sectioned (with widths of 60. 150 and 300 pm) and cleared in xylol.
The tissue sections were mounted on glass slides anil studied by light
microscopy. Routine histological sections were also made and stained
with hacmatoxylin and eosin. The nodes injected with MMA were
digested in HCI and washed in water with the help of sonication. The
MMA samples were then mounted on metal plates to be further
treated for scanning electron microscopy observation.
Scanning Electron Microscopy
The MMA casts were coated by Au/Pt under vacuum and studied
in a JEOL ISM-35C scanning electron microscope. The electron
micrographs were derived from secondary electrons. For identifica­
tion of the vessels we followed Rhodin's criteria 113. I4| in what con­
cerns dimensions and general morphology of the vessels and those of
Lametshwandtner et al. 115] regarding their anatomical features by
scanning electron microscopy.
Results
We studied the micro-anatomy of the arterial vascula­
ture of the dog LN using two types of nodes: popliteal LN
that contain mostly primary follicles, and tracheobronchial
and paratracheal LN that predominantly present second­
ary follicles.
Salvador/Pereira/de Si/Grande
Lymph Node Blood Vasculature
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2
43
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(For legends, see p. 45.)
44
Salvador/Pereira/de Sa/Grande
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Lymph Node Blood Vasculature
Sections o f LN Perfused with Colloidal Carbon
or Micropaque
Scrutiny of these light microscopy preparations con­
firmed the observations derived from MMA corrosion casts
and further illustrated the penetration of the LN cortex by
branches of capsular arteries in tracheobronchial LN
(fig. 7-9). The sections showed multiple examples of corti-
Fig. 3-5. Scanning electron micrographs of vascular casts of dog
LN perfused with MMA resin. The capsule is supplied by multiple
medium-size arteries (top in 3) that, on reaching the outer surface of
the node, originate an elaborated network of anastomotic vessels that
encircle the surface of the nodes. The arrangement of this anastomotic
arterial network is best seen in face views of the node surface (4. 5).
3 Arrowheads show penetration of the cortical domain of the node by
branches coming from capsular arteries. 3 x 80. 4 x 105. 5 x 120.
Fig. 6. Scanning electron micrograph of vascular cast of dog
popliteal LN perfused with MMA resin. This is a higher magnification
of an area of the same node shown in figure I. The large arteries seen
on the right side of the figure penetrate deeply into the node: they may
correspond to hilar vessels. Notice, however, that in figure 1 several
other arteries, with similar large calibres, enter the node in other areas
of its surface. x70.
cal regions that appeared to receive their blood supply from
branches of capsule-associated arteries (arrowheads in
fig.7-9). We found that elements from all of the three
groups of dog LN presented some areas of their cortical
domains receiving their arterial vessels directly from capsu­
lar arteries.
Discussion
Current views on the organization of the arterial supply
of the LN consider that virtually all of the node arteries
enter the organ through its lulus [8-12]. It is. therefore,
assumed that cortical and medullary regions of the organ
are irrigated by branches of the hilar vessels. Only small
calibre vessels would pass through the lulus and go directly
to the LN capsule; these vessels are considered not to have
a significant role in the arterial blood supply of the LN
parenchyma [12. 16-18).
In this report we offer anatomical evidence that in differ­
ent groups of LN of the dog. the capsular arteries make up
a well-developed surface vascular network. These vessels
were so numerous that it was difficult to identify precisely
which arteries did enter the nodes through the hilar invagi­
nation. Some of the capsular vessels penetrated deeply into
the LN cortex. This topography indicated that they contrib­
uted to the blood supply of at least part of the cortical
region and could play a significant role, albeit not com­
pletely understood as yet. in the circulation physiology of
the LN parenchyma.
It has been shown before that the total occlusion of the
hilar blood supply of the popliteal LN did not result in
interruption of the physiological activity of the LN [19],
This surprising result was interpreted as an indication of the
capacity of the lymphatic supply of the LN to take over by
itself all the nutritional needs of the lymphoid organ. Our
anatomical data suggest an alternative explanation: the
occlusion of the hilar arteries may not result in the com­
plete interruption of all of the arterial blood supply of the
node, since, in our preparations, it was clear that the corti­
cal tissue received extrahilar vessels which originated from
capsular arteries. We suggest, therefore, that the arterial
branches coming from capsular vessels may have a signif­
icant participation in the blood supply of the LN paren­
chyma. In conclusion, we propose that the parenchyma of
the LN receives its arterial blood supply from two vascular
systems, the hilar and the capsular arteries. We believe that
the plasticity of these two arterial systems of LN deserves to
be investigated in well-defined pathophysiological events
of the organ.
45
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MMA Resin Casts o f LN Vessels
These preparations offered surface views of the LN and
of their peripheral arterial blood supply. Because of the
high viscosity of this resin. MMA perfusion stopped at the
level of arterial capillaries, and the casts, therefore, did not
include any venous vessels. In low-magnification views of
these corrosion casts, we found an abundant number of
arterial vessels around the outer surface of all of the LN
studied (fig. 1.2). These vessels formed a continuous hair­
like coating that completely encircled the LN. The striking
development of this peripheral vascular layer did not allow
an easy identification of the hilar pole in a good number of
LN by observing a concentration of arteries in the hilar
region of the nodes (fig. 1.2). Higher-magnification views
of the MMA casts were obtained by scanning electron
microscopy (fig.3. 4). Here, it was clear that the predom­
inant direction of the arterial vessels seen on the node sur­
face was parallel to the capsule (fig. 3-5). The capsular arte­
rial vessels made up a very elaborated network that was
best seen in face views (fig.4, 5). In several areas of the
extrahilar regions of the LN surface, vessels originating
from this encircling arterial network penetrated the capsu­
lar layer and entered deeply into the cortical domain of the
organ (see arrowheads in fig.3 and compare fig.6 with its
lower magnification illustrated in fig. I).
46
Fig. 8, 9. Light micrographs of sections of paratrachéal (8) and
popliteal (9) LN of the dog after arterial perfusion of the animal with
the resin Micropaquc. Examples of penetration of the cortex of the
two nixies by branches of capsular arteries are indicated by arrow­
heads. x 35.
Salvador/Pereira/de S¿¡/Grande
Lymph Node Blood Vasculature
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Fig. 7. Light micrograph of xylol-cleared section of dog LN after
arterial perfusion of the animal with colloidal carbon. A large artery
coming from the capsule of the nixie penetrates and branches into the
cortex of the organ (arrowheads at the top of the figure). The LN hilus
is indicated by arrows. x30.
Acknowledgements
The authors thank Dr. Artur P. Aguas for help with the writing of
the manuscript. Mr. Alfredo Ribeiro (Depart. Experimental Surgery.
ICBAS) for surgical help and Mr. Jose' Aurelio for the photographic
work. During the course of this research work. A.C'.S. was a recipient
of a fellowship front the Calouste Gulbenkian Foundation (Lisbon.
Portugal). This research work was supported by a research grant front
the Junta Nacional de Investí garfio Científica e Tecnológica (Portu­
guese Research Council).
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