Inhibition of Neurogenic InØammation by the Amazonian
Herbal Medicine Sangre de Grado
Mark J. S. Miller, Nathalie Vergnolle,
Webb McKnight, Rabi A. Musah,
Cathy A. Davison, Ann Marie Trentacosti,
Jane H. Thompson, Manuel Sandoval,
and John L. Wallace
———————————————-
Department of Pediatrics, and Center for Cardiovascular Sciences, Albany Medical College, Albany, New York, New York, U.S.A.;Department of Pharmacology and Therapeutics, University of Calgary, Calgary, Alberta, Canada; Department of Chemistry, State University of New York at Albany, and Rainforest Phytoceuticals, LLC, Delmar, New York, New York, U.S.A.This study was designed to determine if the Amazonian medicinal sangre de grado, confers bene- Æt by suppressing the activation of sensory afferent nerves. Methods: (i) vasorelaxation of rat mesenteric arteries in response to calcitonin gene-related pep-tide; (ii) rat paw edema in response to protease-activating peptide receptor 2±activating peptide; (iii) rat paw hyperalgesia in response to low-dose pro-tease-activating peptide receptor 2±activating peptide or prostaglandin E; (iv) gastric hyperemia in response luminal capsaicin; (v) a clinical trial of a sangre de grado balm in pest control workers. The parent botanical was fractionated for evaluation of potential active components. In preconstricted rat mesenteric arteries, highly diluted sangre de grado (1:10,000) caused a shift to the right of the calcitonin gene-related peptide dose±response curve (p < 0.01). Paw edema in response to protease-activating pep-tide receptor 2±activating peptide (500mg) was reduced by as single topical administration sangre de grado balm (1% concentration, p < 0.01) for at least 6 h. Hyperalgesia induced by either low-dose pro-tease-activating peptide receptor 2±activating peptide(50mg) or prostaglandin E2 was prevented by sangre de grado balm. A fraction possessing analgesic and capsaicin antagonistic properties was isolated and high-performance liquid chromatography and gas chromatography±mass spectrometry analysis indica-
ted that it was a proanthocyandin oligomer. In pest control workers, sangre de grado balm (Zangrado) was preferred over placebo, for the relief of itching,
pain, discomfort, edema, and redness in response to wasps, Ære ants, mosquitoes, bees, cuts, abrasions, and plant reactions. Subjects reported relief within minutes. We conclude that sangre de grado is a potent inhibitor of sensory afferent nerve mechan-isms and supports its ethnomedical use for disorders characterized by neurogenic infammation. Keywords: capsaicin/Croton lechleri/itch, pain/sensory afferent nerves. J Invest Dermatol 117:725±730, 2001 Sangre de grado (SdG), also known as Sangre de drago or Dragon’s blood, is a viscous, red tree sap that is used extensively by indigenous cultures of the Amazon River basin for its remarkable healing properties (Duke and Vasquez, 1994; Jones, 1995; Phillipson, 1995). Derived from several Croton species (C. dracanoides, C. palanostigma, C. lechleri), SdG is common throughout Amazonia, with the highest quality originating in the upper jungle of Peru and Ecuador. The tree is fast growing, reaching heights of 30±45 ft (10±15 m) in 3 y. Whereas the sap can be harvested like rubber, trees that are repeatedly tapped are prone to fungal infections, thereby diminishing productivity. Current experimental farming techniques focus on growing and felling the trees in a 2±3 y cycle; at this time a single tree will produce approximately 1.5 l of sap. Mechanistically, little is known about SdG given the region and cultures from which it originates. Applied to the skin for abrasions, cuts, scratches, blisters, bites, and stings, SdG forms a long-standing barrier possibly due to its ability to coprecipitate with proteins or other matrix elements (Uchida et al , 1990). In so doing it is claimed to foster accelerated wound healing and does so with reduced pain, infammation, and scarring (Vaisberg et al , 1989; Porras-Reyes et al, 1993; Chen et al , 1994). SdG’s applications in Amazonia are not limited to cutaneous disorders. SdG is also taken orally, in dilute form, for severe gastrointestinal distress, e.g., gastritis, gastric ulcer, intestinal infections, and inØammation (Maxwell, 1990; Duke and Vasquez, 1994; Jones, 1995). In addition, because SdG has hemostatic properties it is sometimes used to arrest excessive bleeding associated with childbirth (this hemostatic effect is also useful for severe cuts and lacerations). As this information is largely anecdotal, the purpose of the present investigation was to evaluate the mechanisms underlying the proposed efÆcacy of SdG in a scientiÆcally rigorous manner.
———————————–
MATERIALS AND METHODS
Vasorelaxation of rat mesenteric arteries Mesenteric arteries were
removed from anesthetized 3 mo old female Fisher 344 rats (National
Manuscript received December 19, 1999; revised May 7, 2001; accepted
for publication May 22, 2001. Reprint requests to: Dr. Mark J.S. Miller, Department Pediatrics, MC-8, Albany Medical College, 47 New Scotland Ave., Albany, NY 12208, U.S.A. Email: [email protected]
Abbreviations: protease activated receptor-2 activating peptide,
PAR-2 AP; calcitonin gene-related peptide, CGRP; sangre de grado,
SdG; vanilloid receptor, VR. 0022-202X/01/$15.00¥ Copyright
2001 by The Society for Investigative Dermatology, Inc. 725 Institutes of Aging) for the study of diameter changes, as previously described by us (White
et al, 1996). The small intestine was excised immediately after sacriÆce of the rat and placed in ice-cold bicarbonate buffer solution of the following composition (in mm): NaCl, 130; KCl, 4.7; MgSO4 ¥7H2O, 1.17; KH2 PO4, 1.18; NaHCO3, 14.9; dextrose, 5.5,NaCa2 EDTA, 0.03; CaCl2 ¥2H2O, 1.6. Resistance-size arteries (3rd±4th order branches of the superior mesenteric artery; approximately 250mm outer diameter) were cleaned of adherent fat and connective tissue and cannulated at each end with glass microcannulae in an arteriograph chamber. These arteries are located at the margin of the intestine, just before the arteries branch and actually enter the intestinal wall. The arteries were pressurized and maintained at 60 mmHg with a servo control device. Warm buffer (37∞C) bubbled with a mixture of 95% O2, 5% CO2 was circulated through the arteriograph chamber at a rate of
20 ml per min. All drugs and mediators were added to the superfusion solution. The arteriograph was placed on the stage of an inverted microscope, and the image of the artery projected on to a video monitor. The image of the
artery was continuously scanned and measurements of the outer diameter
stored in a computer Æle for analysis. Arteries were allowed to equilibrate
at 60 mmHg, in warmed, circulating buffer for a minimum of 45 min
prior to the initiation of the experimental protocol. Arteries were constricted with an EC 80 concentration of phenylephrine (the concentration needed to produce 80% of the maximal contractile response), and calcitonin gene-related peptide (CGRP) was added to the superfusate bath at increasing concentrations until a full relaxation response was achieved. The superfusate was washed, re-equilibrated and pheny-lephrine was administered to raise vascular tone. SdG (1:10,000 dilution)was added to the bath and the dose±response curve to CGRP repeated. In a separate study, the CGRP alone administration was repeated without coadministration of SdG. At the conclusion of the experiment, the bath was rinsed, the vessel preconstricted and the vasodilator response to acetylcholine evaluated as an example of a sensory afferent independent vasodilator. Relaxations of arterial segments to CGRP are expressed as a percentage of the phenylephrine-constricted diameter. Data were analyzed using a nonlinear regression of sigmoidal dose±response curves (using GraphPad Prism 2.01) from which an EC50 value (concentration of CGRP that elicited 50% of the maximum response), maximum response and slope. The negative logEC50 values (pD2) in the absence and presence of SdG (1:10 000 dilution) were compared using a paired test (Statistica, Statsoft). Rat paw edema Sprague±Dawley rats (250 g) were anesthetized with sodium pentobarbitol. Twenty minutes prior to the intradermal injection of protease activated receptor-2 activating peptide (PAR2-AP) (SLIGRL-NH2, 500mg) into the rat footpad, rats received either 40 mg of placebobalm or balm containing SdG (1%, Zangrado Bug Bite Balm, Rainforest Phytoceuticals, LLC, Delmar, NY, http://www.amazonmedicines.com) opically to the footpad. This dose of PAR2-AP was chosen as based on our previous experience that it elicits paw edema through neurogenic mechanisms (Steinhoff et al , 2000). Basal paw volume was measured before PAR 2 -AP administration, and every subsequent hour for a total of 6 h, as previously described (Vergnolle et al , 1999; Steinhoff
et al, 2000). Paw volume was measured using a hydroplethysmometer bath (Ugo Basile, Italy) by an individual unaware of the treatment group. Rat paw hyperalgesia Induced by PAR2-AP Rats were treated in a manner similar to that described above for paw edema studies except that the dose of PAR2-AP was reduced to 50mg (intradermal injection) to avoid complications
associated with edema formation. Rats were divided into two treatment
groups: (1) control, who received the placebo balm (devoid of SdG), and
(2) Zangrado balm (1% SdG). Both were delivered topically (40 mg)
15 min before intradermal administration of PAR2 -AP. Paw withdrawal
latency time to a thermal stimulus, as determined in a Hargreave’s
apparatus, was used as the index of pain sensitivity. This involves the
measurement of the time it takes for a rat to move (withdraw) its foot
away from a thermal stimulus (Vergnolle et al , 1999). Withdrawal times
were determined in each group prior to PAR2-AP administration (basal),
and then 30 and 60 min after PAR2 -AP administration. A reduction in
latency withdrawal time is used as an index of hyperalgesia.
Induced by prostaglandin (PG) E2 Similarly to the experiments where
hyperalgesia was induced by PAR2 -AP, paw hyperalgesia was also
induced by PGE2(0.3mg), administered by intradermal injection. Basal
withdrawal times were established and either placebo or Zangrado balm
containing 1% SdG were applied topically to the paw. Withdrawal times
were determined every 15 min over the course of 1 h, at which time
the PGE2 hyperalgesia response had returned to baseline.
Rat gastric hyperemia in response to capsaicin
As previously
outlined (Gronbech and Lacy, 1996), gastric blood Øow was measured by
a laser Doppler Øow meter placed on the gastric luminal surface of
anesthetized, laparotomized rats. This involves the use of a plexiglass
chamber used for isolating the gastric mucosal surface whereas the
stomach remains fully innervated and with its blood supply intact. The
luminal surface is continually bathed in a buffered salt solution to which
capsaicin (320
m
m), alone or with pretreatment with SdG (1% solution)
or SdG fractions. Hyperemia is expressed as a percentage change from
baseline.
SdG fractionation and structural analysis
The SdG latex (30 ml)
was mixed with 270 ml of acetate buffer (1 mm sodium acetate, 1 mm
acetic acid, pH 4), vortexed and centrifuged at 1200
3
g
for 3 min After
removal of the supernatant the precipitant was then mixed with
methanol (30 ml), vortexed, and centrifuged at 1200
3
g
for 3 min. The
supernatant was applied to a C18 sephadex cartridge. Eluants were
collected with various water±methanol (0, 20, 40, 60, 80, and 100%)
washes and dried for analysis of biologic properties. Similarly, the super-
natant (20
m
l) was also subjected to high pressure liquid chromatography
analysis. After Æltration in a 20
m
m Ælter, the supernatant components
were separated using a Varian Dynamax high pressure liquid chromato-
graphy system equipped with two Model SD-200 pumps, and a model
330 diode array detector, and a microsorb-MV C-18 reverse phase
column (25
3
0.46 cm) at a Øow rate of 1 ml per min. The mobile
phase was a gradient of 15±45% solvent A for 30 min, followed by an
increase to 88% solvent A, where solvent A is methanol, and solvent B is
0.1% acetic acid. Fractions were collected for biologic assay as well as
structural identiÆcation of constituents by gas chromatography±mass
spectroscopy and
1
H nuclear magnetic resonance.
Clinical evaluation in pest control workers
Terminex pest control
workers were invited to participate in a placebo-controlled trial. Either a
balm containing SdG (1%, Zangrado Bug Bite Balm, Rainforest
Phytoceuticals, LLC; http://www.amazonmedicines.com) or placebo
(balm base without the SdG) was applied to various skin conditions
encountered over a 3 mo period at the discretion of the participants.
The balms were coded and their nature was not disclosed to the
participants during the study. Because of their occupation, these pest
control workers incur various cutaneous afØictions. Participants were also
asked to keep a log of the type of afØictions, associated symptoms, time
to achieve relief, complications or side-effects, and whether reapplication
was necessary. A total of 11 participants were enrolled but only 10
participated in the study. The time frame was open-ended, but was
concluded after 3 mo because the spring infestations had yielded a
reasonable number of “events”. Because of the random nature of these
afØictions in a “real world” situation, participants were asked to apply
either balm at their discretion. If for example, more than one bite
occurred at any one time then participants were asked to apply both
balms on different locations in order to facilitate comparison. Some
events by their nature were isolated, however, and so balms were applied
in series as opposed to in parallel. At the conclusion of the trial logged
information was decoded and analyzed. No workers were encouraged to
be attacked by insects, rather all “events” occurred as a hazard of their
occupation.
RESULTS
CGRP vasorelaxation
Rat mesenteric arteries, preconstricted
with phenylephrine, relaxed in response to addition of CGRP to
the superfusate (pD2 = 8.84
6
0.08, n = 5). Addition of SdG to
the superfusate at a dilution of 1:10 000 resulted in a signiÆcant shift
of the dose±response curve to the right (pD2 = 8.08
6
0.1; n = 5,
p < 0.01), as indicated in
Fig 1
. Vasorelaxant responses to acetyl-
choline were unaltered by SdG (data not shown). Repetitive dose±
response curves to CGRP in the absence of SdG were identical.
Rat paw edema
Edema induced by intradermal injection of
PAR
2
-AP is the result of sensory afferent nerve activation, as this
response can be abolished by neonatal capsaicin treatment (sensory
afferent ablation), and neurokinin receptor antagonism (Vergnolle
et al
, 1999; Steinhoff
et al
, 2000). To address the potential effects of
SdG, we applied a balm containing 1% SdG to the rat paw 20 min
prior to PAR
2
-AP injection, with edema monitored for a
subsequent 6 h (
Fig 2
, n = 8 in each group). In both groups
PAR
2
-AP caused a marked increase in paw volume by 1 h,
indicative of edema development. Paw volume then slowly
726
MILLER
ET AL
THE JOURNAL OF INVESTIGATIVE DERMATOLOGY
declined over the next 5 h in the placebo group. In SdG-treated
paws, the decline in paw volume was more dramatic and sustained
despite its single topical application, resulting in approximately 50%
less edema (p < 0.01,
Fig 2
).
Rat paw hyperalgesia
Intradermal injection of PAR
2
-AP
resulted in a decrease in the latency withdrawal period to a heat
source (
Fig 3
). This is indicative of a hyperalgesic state.
Pretreatment with topical SdG (1%, Zangrado Bug Bite Balm)
prevented the induction of hyperalgesia, with the latency
withdrawal time remaining at its baseline level despite PAR
2
-AP
administration (
Fig 3
). Zangrado balm did not affect withdrawal
latency time in rats that did not receive PAR
2
-AP (data not shown)
indicating that it was not acting as an anesthetic. Hyperalgesia was
also induced by intradermal PGE
2
, which is thought to induce an
increased sensitivity to pain perception by raising the resting
potential of sensory afferent nerve Æbers. In these experiments
intradermal PGE
2
resulted in a signiÆcant reduction in paw
withdrawal time (
Fig 4
) and this effect was blocked by a single
topical administration of SdG balm.
Potential thermal analgesic bioactivity of fractionated SdG was
assessed using PAR
2
-AP as the algesic agent. Two fractions of SdG
were ineffective (fractions 2 and 3), with paw withdrawal times
being indistinguishable from vehicle-treated animals (
Fig 5
). In
contrast, fraction 5 was fully effective in preventing the hyperalgesic
response to intradermal PAR
2
-AP, comparable with that observed
with the parent botanical SdG (
Fig 3
).
Capsaicin-induced hyperemia
Gastric hyperemia in response
to luminal capsaicin (320
m
m) was evident in control animals, with
an average increase in basal blood Øow of 60% (
Fig 6
). In rats
treated with either SdG or its derivative, fraction 5, this capsaicin-
induced hyperemia response was largely abolished (p < 0.01). The
concentration chosen for assessment, 1%, was based on the
concentration that was effective topically in rat edema and
analgesic assays as well as the clinical evaluation in pest control
workers.
Structural determination of active chemical con-
stituents
Analysis of fraction 5, a fraction that shared the same
biologic properties of the parent botanical, indicated that it was a
pure compound with a molecular weight of 930, and likely to
be an oligomer of proanthocyandins. Proanthocyandins are the
predominant chemical component of SdG but yet, to date,
proanthocyandins have not been reported to possess sensory
afferent nerve suppressant activity. It should be noted that
fraction 3, which was ineffective as an analgesic agent (
Fig 5
),
possessed the same molecular weight and proanthocyandin
Figure 2. SdG and PAR-
2
AP induced Edema Formation.
PAR-
2
AP used was SLIGRL-NH2. It was injected into the rat paw at a
dose of 500
m
g. Either the placebo balm (
closed circles
,
solid lines
) or the
Zangrado Bug Bite Balm (
open circles
,
broken lines
) were applied topically
(40 mg) to the paw 20 min prior to injection of the SLIGRL-NH
2
. Paw
volume was measured using a hydroplethysmometer, by an individual
unaware of the treatments the rats had received. Results depict the
mean
6
SEM for eight rats in each group. The asterisk indicates a
signiÆcant difference (p < 0.05) between placebo and Zangrado balm
groups as determined by
ANOVA
and
post hoc
analysis with Student±
Newman±Keuls t test.
Figure 3. Hyperalgesia induced by PAR-
2
AP and the effects of
SdG.
Intradermal injection of PAR-
2
AP (50
m
g) resulted in a state of
hyperalgesia as indicated by a reduction in latency withdrawal time from
baseline in rats treated with placebo balm (
*
p < 0.05 compared with
basal values, n = 5,
ANOVA
and
post hoc
analysis with Dunnett’s multiple
comparison test). Zangrado balm (containing 1% SdG, n = 7) completely
prevented the induction of hyperalgesia, with withdrawal latency being
unaltered from baseline. Results are depicted as mean
6
SEM.
Figure 1. SdG and CGRP-induced vascular relaxation.
Dose±
response curves to CGRP in rat mesenteric arteries preconstricted with
phenylephrine (PE). Data are expressed as a percentage of the PE tone.
Vascular preparations were made from Æve animals in each group.
Sequential dose±response curves to CGRP were superimposable (
closed
circles
, data not shown). After establishing the response to CGRP alone
SdG (1:10,000 dilution) was introduced into the superfusate, and the
response to CGRP repeated. SdG caused a signiÆcant shift of the dose±
response curve to the right (
open circles
,
broken lines
, p < 0.05). Results
are depicted as mean
6
SEM for Æve animals in each group.
VOL. 117, NO. 3 SEPTEMBER 2001
INHIBITION OF NEUROGENIC INFLAMMATION
727
oligomer base as fraction 5. In other words, it appears that only
certain isomers of these proanthocyandin oligomers possess the
desired activity, with some proanthocyandin isomers (fraction 3)
having no analgesic activity, whereas others (fraction 5) display
both analgesic and capsaicin antagonistic properties. At this time
further comparative analysis of how the spatial arrangement of these
complex oligomers is not available. Given the complex nature of
the components of this botanical it should be noted that chemical
constituents other than proanthocyandins may be active.
Clinical evaluation of a SdG-based balm
An evaluation of a
commercially available SdG balm (Zangrado Bug Bite Balm) for
relief of the symptoms of insect bites, stings, and other skin
conditions was done in Terminex pest control workers in New
Orleans, LA. These workers are prone to these hazards as part of
their occupation. Participants compared the effects of a placebo
balm with a balm containing 1% SdG over a 3 mo period. As noted
in
Table I
, the primary afØiction during this period was Ære ant
bites, with all participants suffering at least one attack. Fire ants are
endemic throughout the south-eastern United States, and they
elicit an initial painful bite followed by an itching response that can
persist for weeks. Because of the uncontrolled environment in
which this study was performed every participant did not
experience the same number or type of encounters. Some
individuals encountered the same afØiction multiple times (for
example Ære ants).
Figure 5. Effects of SdG fractions on the hyperalgesia induced by
PAR
2
-AP.
Intradermal injection of PAR
2
-AP (50
m
g) resulted in the
induction of thermal hyperalgesia as indicated by a reduction in latency
withdrawal time from baseline in rats treated topically with the vehicle
balm. Balm containing 1% of fractions 2, 3, or 5 isolated from the parent
botanical, SdG, were evaluated at 30 and 60 min after PAR
2
-AP
administration. Fraction 5 completely prevented the induction of
hyperalgesia, with withdrawal latency being indistinguishable from
baseline. In contrast, fractions 2 and 3 were ineffective analgesic agents,
producing similar results to the vehicle control. Results are depicted as
mean
6
SEM, n = 5 (
*
p < 0.05 compared with basal values, n = 5,
ANOVA
and
post hoc
analysis with Dunnett’s multiple comparison test).
Figure 6. Effects of SdG and fraction 5 on the gastric hyperemia
induced by luminal capsaicin.
Gastric mucosal blood Øow was
measured by a laser Doppler Øow meter in rats. Luminal exposure to
capsaicin (320
m
m) greatly increased mucosal blood Øow (expressed as a
percentage change from baseline, mean
6
SEM). in rats pretreated with
luminal SdG or fraction 5 (both 1% solutions), the hyperemic response
to capsaicin was largely absent (n = 5,
*
p < 0.01
vs
vehicle).
Table I. AfØictions experienced by pest control workers
a
AfØiction
Individuals reporting
a speciÆc afØiction
Total no. of
individuals enrolled
Bee
1
10
Wasps
6
10
Mosquito
1
10
Fire ant
10
10
Other ants
6
10
Plant reactions
1
10
Cut
5
10
Abrasion
1
10
a
The number of individuals reporting an individual afØiction is noted. Not all
participants encountered the same proÆle of afØictions, as to be expected, as these
events were random and reØect the occupational hazard. It is important to note
that an individual participant may have experienced the same type of afØiction
multiple times, but it is only noted here once. This table only describes the proÆle
of events that were recorded over a 3 mo period, and not the number of events
that any single participant encountered.
Figure 4. Hyperalgesia induced by PGE
2
and the effects of SdG.
Intradermal injection of PGE
2
(0.3
m
g) resulted in a state of hyperalgesia
as indicated by a reduction in latency withdrawal time from baseline in
rats treated with placebo balm (
*
p < 0.05 compared with basal values,
n = 5,
ANOVA
and
post hoc
analysis with Dunnett’s multiple comparison
test). Zangrado balm (containing 1% SdG, n = 5) completely prevented
the induction of hyperalgesia, with withdrawal latency being unaltered
from baseline. Results are depicted as mean
6
SEM.
728
MILLER
ET AL
THE JOURNAL OF INVESTIGATIVE DERMATOLOGY
The symptoms associated with these encounters are recorded in
Table II
. Itching (10 of 10 participants) was the predominant
symptom, reØecting the preponderance of Ære ant bites among the
workers. The placebo balm offered relief in only two cases, whereas
the SdG relieved symptoms in all individuals, for all applications.
The SdG balm was preferred over placebo balm by all participants
and relief was reported to occur in less than 2 min, on average. The
diversity of conditions in which the SdG offered symptomatic relief
was broad (
Table I
). That combined with the speed by which it
achieved relief, is highly suggestive that beneÆt was related to a
sensory afferent-based mechanism of action.
DISCUSSION
Despite the widespread use of SdG in Amazonia as an analgesic,
antidiarrheal, and wound-healing agent few in the Western world
are aware of its existence and little is known about how it achieves
these therapeutic beneÆts. We postulated that these beneÆts may
result from a suppression of sensory afferent nerve activation and
the present results support this conclusion. This hypothesis was
generated from experience and the knowledge that sensory afferent
nerves serve as broad-based sentinels in the skin, gut, and lung, and
that the rapidity by which SdG relieved pain and itching was
consistent with a neurogenic mechanism. In addition, the
serendipitous personal observation by an author that SdG relieved
the symptoms of cutaneous capsaicin (sensations associated with an
overly spicy meal) focused our attention to sensory afferent nerve
mechanisms.
, 1996; Wallengren, 1997; Quinlan
et al
,
1998; Scholzen
et al
, 1999). Potentially any skin condition
characterized by itching, pain, edema, redness, and discomfort
may receive beneÆt from an agent that inhibits sensory nerve
afferent activity.
Ethnomedical applications for SdG are not conÆned to the skin
(Maxwell, 1990). The oral intake of SdG for diarrhea and intestinal
distress may well be due to its ability to modify sensory afferent
mechanisms in the gut (Miller
et al
, 2000). Sensory afferent nerves
are major components of the enteric nervous system, and
contribute not only to secretion, but also to cramping, discomfort,
and pain perception. One may speculate that SdG could also
provide therapeutic beneÆt for various pulmonary disorders that
involve sensory afferent nerves. These include airway hyperrespon-
siveness ± cough, asthma (Reidel
et al
, 1997; Choi and Kwon,
1998; Tohda
et al
, 1998), and viral infections (Piedimonte
et al
,
1990, 1997; Yamawaki
et al
, 1995), provided an adequate delivery
system could be designed. In addition, sensory afferent nerves and
their neuropeptides have also implicated in the pathogenesis of
arthritis (McDougall
et al
, 1999; Vilensky and Cook, 1998)
We conclude that SdG offers a valuable tool for determining the
role of sensory afferent nerves in inØammation. Considering the
recent advances provided by studies of the VR1 knock-out mice,
demonstrating signiÆcant thermal analgesia resulting from the
failure to activate sensory afferent nerves (Caterina
et al
, 2000;
Davis
et al
, 2000), any botanical that suppresses sensory nerve
activity warrants a detailed evaluation of its therapeutic potential.
Indeed, SdG’s actions as an analgesic and anti-inØammatory agent
may provide a new and valuable therapeutic approach to a variety
of inØammatory disorders and serves to highlight the valuable
medicinal resources that are still contained within the rain forests of
Amazonia.
REFERENCES
Ansel JC, Kaynard AH, Armstrong CA, Olerud J, Bunnett N, Payan D:
Skin±nervous system interactions.
J Invest Dermatol
106:198±204, 1996
Caterina MJ, LefØer A, Malmberg AB,
et al
: Impaired nociception and pain sensation
in mice lacking the capsaicin receptor.
Science
288:306±313, 2000
Chen ZP, Cai Y, Phillipson JD: Studies on the anti-tumour, anti-bacterial, and
wound-healing properties of dragon’s blood.
Planta Med
60:541±545, 1994
Choi DC, Kwon OJ: Neuropeptides and asthma.
Curr Opin Pulm Med
4:16±24, 1998
Davis JB, Gray J, Gunthorpe MJ,
et al
: Vanilloid receptor-1 is essential for
inØammatory thermal hyperalgesia.
Nature
405:183±187, 2000
Duke J, Vasquez R:
Amazonian Ethnobotanical Dictionary
. Boca Raton, FL: CRC
Press, Inc., 1994
Gabriel SE, Davenport SE, Steagall RJ, Viaml V, Carlson T, Rozhon RJ: A plant-
derived inhibitor of cAMP-mediated Øuid and chloride secretion.
Am J Physiol
276:G58±G63, 1999
Goebeler M, Henseleit U, Roth J, Sorg C: Substance P and calcitonin gene-related
peptide modulate leukocyte inÆltration to mouse skin during allergic contact
dermatitis.
Arch Dermatol Res
286:341±346, 1994
Gronbech JE, Lacy ER: Role of sensory afferent neurons in hypertonic damage and
restitution of the rat gastric mucosa.
Gastroenterology
111:1474±1483, 1996
Jones K:
Cat’s Claw. Healing Vine of Peru
. Seattle: Sylvan Press, 1995
Maxwell N:
Witch Doctor’s Apprentice, Hunting for Medicinal Plants in the Amazon
, 3rd
edn. New York, NY: Citadel Press, 1990
McDougall JJ, Ferrell WR, Bray RC: Neurogenic origin of articular hyperemia in
early degenerative joint disease.
Am J Physiol
276:R745±R752, 1999
Miller MJS, McNaughton WK, Zhang X-J,
et al
: Treatment of gastric ulcers and
diarrhea with the Amazonian herbal medicine sangre de grado.
Am J Physiol
279:G192±G200, 2000
Phillipson JD: A matter of some sensitivity.
Phytochemistry
38:1319±1343, 1995
Piedimonte G, McDonald DM, Nadel JA: Glucocorticoids inhibit neurogenic plasma
extravasation and prevent virus-potentiated extravasation in the rat trachea.
J
Clin Invest
86:1409±1415, 1990
Piedimonte G, Pickles RJ, Lehmann JR, McCarthy D, Costa DL, Boucher RC:
Replication-deÆcient adenoviral vector for gene transfer potentiates airway
neurogenic inØammation.
Am J Respir Cell Mol Biol
16:250±258, 1997
Pieters L, De Bruyne T, Claeys M, Vlietinick A: Isolation of a dihydrobenzofuran
lignan from South American dragon’s blood (
Croton spp
) as an inhibitor of cell
proliferation.
J Nat Prod
56:899±906, 1993
Porras-Reyes RH, Lewis WH, Roman J, Simchowitz L, Mustoe TA: Enhancement
of wound healing by the alkaloid taspine deÆning mechanisms of action.
Proc
Soc Exp Biol Med
203:18±25, 1993
Quinlan KL, Song I-S, Bunnett NW,
et al
: Neuropeptide regulation of human
dermal microvascular endothelial cell ICAM-1 expression and function.
Am J
Physiol
275:C1580±C1590, 1998
Rao GSR, Gerhat MA, Lee 3d RT, Mitscher LA, Drake S: Antimicrobial agents
from higher plants: Dragon’s blood resin.
J Nat Prod
45:646±648, 1982
Reidel F, Benden C, Philippou S, Streckert HJ, Marek W: Role of sensory
neuropeptides in PIV-3-infection-induced airway hyperresponsivess in guinea
pigs.
Respiration
64:211±219, 1997
Scholzen TE, Brzoska T, Kalden DH, O’Reilly F, Armstrong CA, Luger TA, Ansel
JC: Effect of ultraviolet light on the release of neuropeptides and
neuroendocrine hormones in the skin: mediators of photodermatitis and
cutaneous inØammation.
J Invest Dermatol Symp Proc
4:55±60, 1999
Steinhoff M, Vergnolle N, Young SH,
et al
: Agonists of proteinase-activated
receptor-2 induce inØammation by a neurogenic mechanism.
Nature Med
6:151±158, 2000
Sterner O, Szallasi A: Novel natural vanilloid receptor agonists: new therapeutic
targets for drug development.
Trends Pharmacol Sci
20:459±465, 1999
Tohda Y, Nanbu Y, Tanaka A, Kubo H, Fukuoka M, Nakajima S: Role of substance
P in increased airway hypersensitivity following induced stress in guinea pig
asthma model.
J Invest Allergol Clin Immunol
8:340±345, 1998
Ubillas R, Jolad SD, Bruening RC,
et al
: SP-303, an antiviral oligomeric
proanthocyanidin from the latex of
Croton lechleri
(Sangre de Grado).
Phytomed
1:77±106, 1994
Uchida S, Ohata H, Niwa M, Mori A, Nonaka G, Nishioka I, Ozaki M:
Prolongation of life span of stroke-prone spontaneously hypertensive rats
(SHRSP) ingesting persimmon tannin.
Chem Pharm Bull
38:1049±1052,
1990
Vaisberg AJ, Milla M, del Carmen Planas M,
et al
: Taspine is the cicatrizant principle
in sangre de grado extracted from
Croton lecheleri
.
Plant Med
55:140±143, 1989
Vanner S, MacNaughton WK: Capsaicin-sensitive afferent nerves activate
submucosal secretomotor neurons in guinea pig ileum.
Am J Physiol
269:G203±G209, 1995
Vergnolle N: Review article: proteinase-activated receptors ± novel signals for
gastrointestinal pathophysiology.
Aliment Pharmacol Ther
14:257±266, 2000
Vergnolle N, Hollenberg MD, Sharkey KA, Wallace JL: Characterization of the
inØammatory response to proteinase-activated receptor-2 (PAR-2)-activating
peptides in the rat paw.
Br J Pharmacol
127:1083±1090, 1999
Vilensky JA, Cook JA: Neurogenic acceleration of osteoarthritis.
Curr Opin
Rheumatol
10:251±255, 1998
Wallengren J: Vasoactive peptides and skin.
J Invest Dermatol Symp Proceedings
2:49±55, 1997
Wallengren J, Ha
kanson R: Effects of substance P, neurokinin A and calcitonin gene-
related peptide in human skin and their involvement in sensory nerve-mediated
responses.
Eur J Pharmacol
143:263±273, 1987
Wallengren J, Ha
kanson R: Effects of capsaicin, bradykinin and prostaglandins in the
human skin.
Br J Dermatol
126:111±117, 1992
White RM, Rivera CO, Davison CB: Differential contribution of endothelium
function to vascular reactivity in conduit and resistance arteries from DOC-salt
hypertensive rats.
Hypertension
27:1245±1253, 1996
Yamawaki I, Geppetti P, Bertrand C, Chan B, Massion P, Piedimonte G, Nadel JA:
Viral infection potentiates the increase in airway blood Øow produced by
substance P.
J Appl Physiol
79:398±404