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Lymphland International Lymphedema Online
Lymphedema and vitamins’
The American Journal of Clinical Nutrition 26: FEBRUARY 1973, pp. 185-190. Printed in U.S.A. 185
Ethel F#{246}ldi-8#{246}rcs#{a2n4d6}k M. F#{246}ldi
In 1 97 1, the unexpected fact has been
described (1) that surgically induced acute
experimental lymphedema in the rat flourished
with the usual laboratory diet rich in
vitamins, can be treated successfully by the
administration of various vitamins, e.g., pyndoxine,
pantothenic acid, and particularly, a
highly active representative of the vitamin
P family,2 coumarin (5 , 6-benzo-a-pyron).
These data, obtained by means of plethysmographic
assessment of the volume of
lymphedema, were confirmed by histology
(2, 3).
Based on these results, the question has
to be raised whether an inadequate supplying
of the organism with vitamins would aggravate
lymphedema. After having obtained an
affirmative answer to this question, therapeutic
trials were performed in avitaminotic
lymphedematous animals.
Material and methods
Experiments were performed in 150 male Wistar
rats (body weights given in Fig. 1). The animals
were divided into three groups.
Group 1
Group 1 comprised rats fed ad libitum an artificial
diet rich in vitamins (Table 1). On the 56th
day, these animals were divided into five subgroups.
Subgroup 1.1 (n = 10). From the 56th to the
63rd day, the rats were given daily ip injections of
saline, 10 mI/kg body wt. On the 60th day, the
rats were anesthetized with nembutal (50 mg/kg
ip). Preoperative volume of the neck and head was
measured by electronic plethysmography by means
of the apparatus constructed by Bundschuh (Griesheim,
W. Germany). This was followed by a
radical cervical lymphatic blockage. From a midline
incision reaching from the mandibula to the
sternum, lymph nodes were carefully prepared and
ligated. Plethysmographic measurements were repeated
on the 63rd day.
Subgroup 1.2 (n = 10). In this subgroup, the
procedure was the same as in subgroup 1.1, with
just one difference. Instead of saline, the rats were
treated with the following vitamins in milligrams
per kilogram body weight: vitamin B1, 40; lactoflavin,
23; niamid, 160; pyridoxine, 16; pantothenic
acid, 240; biotin, 2 and cyanocobalamin, 80 pg/kg
body weight.
Subgroup 1.3 (n 10). Procedure was the same
as in subgroups 1.1 and 1.2 with one difference; i.e.,
the animals were treated with 25 mg/kg coumarin
(5 , 6-benzo-alpha-pyron).
Subgroup 1.4 (,z 10). Procedure differed from
that used in subgroups 1.1, 1.2, and 1.3 in one
respect; the rats were treated with 500 mg/kg tnhydroxy-
ethyl-rutin.
Subgroup 1.5 (n 10). In this subgroup, the
same procedure was followed as with subgroup 1.1
except that instead of a cervical lymphatic blockage,
only a sham operation was performed.
Group 2
Rats were fed a diet (ad libitum) that was poor
in B-complex vitamins (Table 1). On the 35th day,
the rats were divided into five subgroups. (This day
was chosen instead of the 56th because vitamin
B deficiency was already marked and the state of
the rats deteriorated rapidly.)
Subgroup 2.1 (n 10). From the 35th to the
42nd day, the rats were given daily ip injections of
saline. On the 39th day, they were treated as those
in subgroup 1 . 1 . Plethysmographic measurements
were repeated on the 42nd day.
Subgroup 2.2 (n 10). In this subgroup, procedure
was the same as in subgroup 2.1, with one
exception. Instead of saline, the rats were treated
with the same vitamins as those in subgroup 1.2.
Subgroup 2.3 (n 10). The rats in this group
were treated with coumarin; otherwise the procedure
was the same as in subgroup 2.1 and 2.2.
Subgroup 2.4 (n = 10). Our procedure differed
from that used in subgroups 2.1, 2.2, and 2.3 in
one respect, these rats were treated with trihydroxyethyl-
rutin.
Subgroup 2.5 (n = 10). In this subgroup, the
procedure was the same as in subgroup 2.1 except
that instead of a lymphatic blockage, only a sham
operation was performed.
Group 3
Rats in this group were fed the Sherman-LaMer-
Campbell diet (4), free of vitamin P but supplemented
with ascorbic acid (Table 1). This group
1 From the Lymphological Research Laboratory,
Schaper and Br#{252}mmer, Salzgitter-Ringelheim, West
Germany.
2 The controversial views concerning vitamin P
are handled in the Discussion.
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300. -
200.
100.
0 ‘ı 2 3 14 5 6 7 8
FIG. 1. Changes of body weight in the three groups.
186 F#{212}LDI-B#{246}RCS#{246A}KND FOLDI
E_Eı
ı ı
I weeks ı
also was fed ad libitum. On the 56th day, the rats
were divided into five subgroups.
Subgroup 3.1 (n = 10). From the 56th to the
63rd day, the rats were given saline. On the 60th
day, they were handled as those belonging to subgroup
I . 1 . Plethysmographic measurements were
repeated on the 63rd day.
Subgroup 3.2 (,i = 10). In this subgroup, procedure
was the same as in subgroup 3.1 but instead of
saline, the rats were treated with the same vitamins
as those in subgroup 1.2.
Subgroup 3.3 (n 10). These rats were given
coumanin in addition to the treatment received by
subgroups 3.1 and 3.2.
Subgroup 3.4 (,z 10). The procedure for these
rats differed from that used in subgroups 3.1, 3.2,
and 3.3 in one respect: they were treated with tnhydroxy-
ethyl-rutin.
Subgroup 3.5 (ii = 10). With this subgroup also,
a sham operation was performed instead of
lymphatic blockage. Otherwise, the procedure was
the same as with subgroup 3.1.
Within the three groups, volumes of lymphedemas
were expressed as the difference between
the postoperative (V2) and the preoperative (V1)
TABLE 1
volumes. In order to compare data of the three
groups with each other, it was necessary to eliminate
the differences existing in the V1 values. Therefore,
volumes of lymphedemas (L) were expressed
with the formula:
\.ı - \rl
L:=100ı
Statistical analysis was performed by appropriate
forms of Student’s t test.
Results
The extent of lymphedema was significantly
higher in both avitaminotic, salinetreated
subgroups (subgroup 2. 1 and 3.1)
than in subgroup 1.1 (normal diet) (Figs.
2, 3). In group 2, hair loss, rhagades, and a
markedly diminished growth rate could be
observed (Fig. 1). There were no such signs
in group 3.
A statistically significant protective effect
Composition of diets used in group I to LII
Group I (Altromin ı 1,000) Group II (Altromin C 1,000 without Group III (ShermanıLaMerıCampbelI)C
B vitamins)
Casein
Rice starch-flour
Soja oil
Cellulose powder
Minerals
Vitamins#{176}
Percent
22
64
2
4
6
2
Casein
Rice starch-flour
Soja oil
Cellulose powder
Minerals
Vitaminsb
I
Percent
22 Butter
64 Milk powder
2 Yeast powder
4 I NaCI
6
2
Percent
10
30
59
1
a Vitamins: DL-methionine; 1,200 mg; vitamin A, 15,000 lU; vitamin Dı, 500 IU; vitamin E, 150 mg;
vitamin K,, 10 mg; vitamin B1 . HCI, 20 mg; vitamin B2, 20 mg; vitamin B3. HCI, 15 mg; vitamin B12, 30 ı.
tg;
pantothenic acid, 50 mg; nicotinic acid, 50 mg; cholinchlorid, I ,000 mg; folic acid, 10 mg; biotin, 200,ug;
inositol, 100 mg; p-aminobenzoic acid, 100 mg; and ascorbic acid, 20 mg.
b Vitamins: vitamin A, 15,000 LU; vitamin Dı, 500 IU; vitamin E, 150 IU; vitamin K3, 10 mg; and
ascorbic acid, 20 mg.
The mixture was heated for 1 hr at 100 C. After cooling down, 100 mg ascorbic acid was added to
each 100 g of the dry food. Daily consumption of food/rat 30 g. Daily consumption of vitamin C/rat
- 25 mg.
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Prot.ct#{233}Cı,f ,f.ct
.5
LYMPHEDEMA AND VITAMINS 187
FIG. 4. Protective effects of various treatments.
BVK = vitamin B complex.
21i terized by a generalized increase in blood
capillary permeability. In vitamin B6 deficiency,
brought about by the administration
of desoxypyridoxine, even the permeability
20 of the blood-brain barrier is increased (6). In
group 2 of these series, a B-complex avitaminosis
has been induced; its effect on the
blood capillaries can certainly be expected
15. to be even more pronounced. With plasma
proteins leaking out of the blood capillaries,
lymphatic blockage will, of course, lead to a
more extensive lymphedema than in normal
ıo. animals with a physiological degree of protein
permeability.
On the other hand, permeability of the
Fio. 2. Extent of lymphedema (percent volume
increase after lymphatic blockage); 1) in rats fed a
normal diet (subgroup 1.1); 2) in rats fed the Sherman-
LaMer-Campbell diet (subgroup 2.1); 3) in
rats with a vitamin B complex deficiency (subgroup
3.1).
could be achieved against lymphedema in
rats fed the normal diet by coumarin and by
vitamin B complex treatment; against lymphedema
aggravated by B-avitaminosis, by FIG. 3. Lymphedema in a rat fed a normal diet
coumarin treatment, and in P-avitaminosis (subgroup 1.1; left) and in a rat in a vitamin Pby
coumarin treatment (Fig. 4). deficiency state (subgroup 3.1; right).
Discussion
After lymphatic blockage, lymph will
accumulate in the occluded lymphatics. Stagnating
lymph, however, will dilate them. The
high intralymphatic pressure will induce
“mural insufficiency” (5), i.e., induce the
diffusion of lymph through the wall of lymph
vessels, and will force open interendothelial
junctions in initial lymphatics; here too,
lymph will escape back into the tissues. At
the same time, plasma protein molecules,
steadily escaping from the blood capillaries,
will accumulate in the interstice. They are,
normally, removed by lymphatics. Lymphedema,
finally, will be the result of raised
oncotic pressure in the interstitial fluid.
It is well known that beriberi is characDownloaded
from www.ajcn.org by on May 7, 2009
188 FOLDI-BORCSOK AND FıLDI
FIG. 5. Vitamin P family.
lymph vessels may eventually be increased
by B-complex avitaminosis leading to a mural
insufficiency of lymph drainage, a hypothesis
awaiting elucidation.
Normal connective tissue cells incorporate
and digest plasma proteins; in this way, they
become colloidosmotically inactive (7, 8).
In B-complex avitaminosis quite a number
of the most important mitochondrial
enzyme systems are severely damaged. In
lymphedema of normally fed animals, the
extent of lymphedema may be limited to an
appreciable extent through this mechanism.
The possibility that plasma protein catabolism
in these connective tissue cells
could be subnormal or absent may well be a
further factor meriting additional studies.
The matter of vitamin P has been heavily
debated since 1936, when Szent-Gy#{246}rgyi (9)
described the fact that experimental scurvy
is caused not only by a deficiency in ascorbic
acid (discovered by the same author) but
also by a concomitant deficiency in vitamin
P. Based on the original studies of Szent-
Gy#{246}rgyi and on those of Javillier and Lavollay
(10), we are now aware of the fact that
vitamin P is not just one chemical compound,
as e.g., vitamin B1. It is rather similar to
vitamin K, which is not a single molecule
either. Just as various molecules possess
vitamin K activity, other chemically related
compounds possess qualitatively, but not
quantitatively, the same action on blood
capillaries; if they are entirely absent from
the food, blood capillary permeability and
fragility will increase (Fig. 5). The administration
of the diet described by Sherman,
LaMer and Campbell (4), which is free of
vitamin P and vitamin C, produces an increase
in blood capillary fragility and permeability
not only in the guinea pig, but in
the rat, which is also able to synthesize
ascorbic acid. It has been shown by Benk#{246}
et al. (11) that in rats fed this diet, cerebral
edema will appear. V#{225}rkonyi and co-workers
(12) have shown that cerebral edema is
caused by an increased permeability of the
blood-brain barrier, as indicated by the
Evans-blue fluorescence method.
In sharp contrast to Clark and MacKay
(13) who consider “many of the physiologic
and pharmacologic nonspecific effects which
they (i.e., members of the vitamin P family)
are said to exert” as an “alarm reaction,”
we share the arguments of Javillier and
Lavolley (10): “Any organic substance,
which is contained in natural food and is
responsible for the maintenance of a physiologic
equilibrium is a vitamin. The question
to be answered is, whether there exist, in the
OH
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LYMPHEDEMA AND VITAMINS 189
normal animal and human food, substances,
small amounts of which are maintaining
blood capillary fragility and permeability
at normal levels. If such is the case-and
such actually it is-these substances are
vitamin P substances . . . .“
In the present series, the extent of lymphedema
was significantly increased by feeding
the Sherman et al. diet, supplemented (in
spite of the rat’s ability to synthesize this
vitamin) with ascorbic acid. This fact can
readily be explained, as in the case of Bcomplex
avitaminosis, by an increased protein
permeability of the blood capillaries.
Again, there arises the possibility that permeability
of lymph vessels may be increased
too.
The therapeutic effect of B-complex vitamins
and of vitamin P (coumarin) treatment
in animals fed the usual laboratory diet
(group 1) enriched in various vitamins may
have the following explanation:
The treatment may 1) prevent the normal
leakage of plasma protein molecules through
blood capillaries; 2) it may cause collateral
lymphatics to open more rapidly; and 3)
it may increase the catabolism of plasma proteins
by connective tissue cells. From what is
known of the passage of protein molecules
through continuous, nonfenestrated blood
capillaries, it would seem that the first possibility
is highly unlikely. The second possibility
is unlikely too. We feel the last theory
to be the correct answer.
Two questions arise in connection with
the therapeutic trials performed in rats suffering
from B-complex avitaminosis. a) The
ineffectiveness of the vitamin B complex
therapy is a rather unexpected finding. As a
rule, the consequences of any avitaminosis
are swept away by the administration of the
missing vitamin. But, as bone destruction
brought about by mechanical strain in vitamin
D avitaminosis cannot be cured by the
administration of vitamin D, so we assume
that vascular and/or cellular lesions brought
about by B avitaminosis of long duration are,
in combination with lymphatic blockage, too
serious as to be influenced by the vitamin
therapy in the manner and the dosage employed
in our experiments. b) The effectiveness
of vitamin P (only the protective effect
of coumarin was biologically and statistically
significant) therapy in lymphedema of rats
suffering of B-complex avitaminosis is a surprising
finding, but an analogous result has
already been described by F#{246}ldiand Zolt#{225}n
(14). The authors induced a combined pyridoxine-
pantothenic acid deficiency in the rat
by the administration of the two specific
antivitamins, deoxypyndoxine and omegamethylpantothenate.
Specific deterioriation
of the function of the central nervous system
could be effectively prevented by coumarin.
To quote another well-known fact, consequences
of A-avitaminosis can be treated
effectively by the administration of vitamin
C. The most effective therapeutic agent
against lymphedema aggravated by the administration
of the Sherman-LaMer-Campbell
diet was coumarin.
We are, of course, aware of the fact that
these results can be interpreted by the
assumption of some unidentified pharmacological
effect of the vitamins used in our
experiments. Dosage-response curves, and
autoradiographic histology could bring the
definite answer to this question. The fact that
various vitamins exert a therapeutic effect in
lymphedema and that a deficiency in various
vitamins aggravate it cannot be regarded as
an argument in favor of a specific interrelationship
between vitamins and lymphedema.
Axeirod ( 1 5) has described quite similar
relationships between vitamins and immunologic
responses. Coumarin has been found
to be effective in the treatment of some types
of lymphedema in the human being too (J.
S. Calnan and J. J. Pflug, personal cornmunication).
Based on these studies, we propose the
hypothesis that in the etiopathornechanism
of some of those gigantic lymphedemas leading
to massive destruction in tropical countries,
dietetic factors may play an aggravating
role but, we are, of course, aware of the
fact that the present work was perfornied in
the rat and, at the present state of our
knowledge, there is no evidence that the
described mechanism exists in man.
Summary
Both B-complex avitaminosis and P avitaminosis
aggravate lymphedema in the rat.
A member of the vitamin P family, coumarin
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190 F#{212}LDI-BORCSOK AND FOLD!
(devoid of any antithrombotic activity)
proved to be a highly active therapeutic agent
against both forms of avitaminosis-aggravated
lymphedemas. ı
References
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