676
Views & Citations10
Likes & Shares
Introduction: In recent years, the use of food supplements like co-adjuvants in the
treatment of various diseases has increased; being one of the most studied and
consumed the algae Spirulina maxima,
which possesses important demonstrated biological effects including antioxidant
and anti-inflammatory activities.
Objective: This study sought to determine if there is a direct link between these
two pharmacological mechanisms in models of acute and chronic inflammation in vivo using a trademark of Spirulina
powder.
Methods: The acute
anti-inflammatory effect of Spirulina was evaluated in the model of sub-plantar
edema induced by carrageenan and its antioxidant activity was determined in a
model of experimental arthritis induced with complete Freund’s adjuvant.
Results: For acute
inflammation, Spirulina at 400 mg/kg generated an anti-inflammatory effect
(51%) similar to indomethacin (58%) in fifth hour, whereas in the chronic
model, also dose of 400 mg/kg demonstrated an anti-inflammatory effect on days
14 and 21 of the study (51% and 38%), close to phenylbutazone effect in the
same days (53% and 52%), as well this dose showed an increase in body weight
gain during the experiment compared to CFA control group. Finally, 400 mg/kg
dose of Spirulina decreased protein carbonyl content (69%) and lipid
peroxidation (81%) in edema tissue compared to arthritic mice without
treatment. And for antioxidant enzymes activities this dose decrease SOD
activity (42.98%) and increase CAT (3216%) and GSH-Px (130%) activities when
compared to CFA control group.
Conclusion: Thus, it was demonstrated that Spirulina at 400 mg/kg dose generated an
anti-edematous effect associated to antioxidant activity in vivo since it decreases oxidative damage over biomolecules of
sub-plantar edema tissue during chronic inflammation, as well increase and
regulate the antioxidant enzymatic endogenous response.
Keywords: Spirulina
maxima, Acute inflammation, Chronic inflammation, Oxidative stress
Abbreviations: 2,4-DNPH: 2,4-Dinitrophenylhidrazine; ANOVA: Analysis of Variance; BIRMEX: Biological Laboratories Reagents México; BW: Body Weight; CAT: Catalase Enzyme; CFA: Complete Freund’s; COX-2: Cyclooxygenase 2; EtOAc: Ethyl Acetate; EtOH: Ethanol; GSH: Reduced Glutathione; GSH-Px: Glutathione Peroxidase; HCl: Hydrochloric Acid; IBD: Inflammatory Bowel Disease; i.g.: intragastric; IMC: Indomethacin; iNOS: induced Nitric Oxide Synthase; MDA: Malondialdehyde; MEC: Molar Extinction Coefficient; OS: Oxidative Stress; PBS: Phosphate Buffer Solution; PBZ: Phenilbutazone; PGE2: Prostaglandin E2; RM: Repeated Measures; ROS: Reactive Oxygen Species; s.c.: subcutaneous; SEM: Standard Error of the Mean; Sm: Spirulina maxima; SNK: Student Newman Keuls; SOD: Superoxide Dismutase Enzyme; TBA: Thiobarbituric Acid; TBARS: Thiobarbituric Acid Reactive Substances; TCA: Trichloroacetic Acid; TP: Total Proteins; UV/Vis: Ultraviolet/Visible
INTRODUCTION
Inflammation is a beneficial body reaction
caused by a stimuli [1-3], this could be external (induced by a biological or
physical damage) or internal (auto-immune pathologies), with the objective of
identify and isolate the damaged area, eliminate the causal agent and finally
reset the homeostasis in the affected zone. This immune process has two
well-known phases: acute and chronic, both characterized by vasodilatation,
macrophages migration, reactive oxygen species (ROS) production and edema, also
it exhibits two pharmacological pathways: the cellular and the oxidative one,
in this last one is where the respiratory burst takes place for the generation
of ROS and other free radicals with high reactivity [2-6]. Nowadays, the use of
food supplements as co-adjuvants in the treatment of diseases has increased,
due to allopathic medicine failure [7-11]. Algae from Spirulina genus have been
one of the most studied as a source of active compounds [12-15], which did not
show toxicity [16-18]. Also its properties have been proven through scientific
studies: anti-viral [19], hypoglycemic [20], hypolipidemic [21], anti-cancer
[22], anti-anemic [23], and anti-inflammatory [24,25]. Previous work
demonstrated that laboratory cultivated S.
maxima protects against oxidative damage in joint tissue as well as serum
during arthritis in rats [26].
The aim of this work was to demonstrate the
ability of Spirulina maxima to
generate an anti-inflammatory effect right on the paw edema tissue, through its
antioxidant activity in vivo over
oxidative stress associated to Complete Freund’s Adjuvant (CFA)-induce
arthritis in mice. Also its beneficial effect during acute inflammation was
evaluated.
METHODS
Reagents and
chemicals
All chemicals used in this study were analytical grade and were purchased from Sigma-Aldrich (St. Louis, MO, E.U.A.). λ-Carrageenan (No. CAT 9064-57-7), Complete Freund’s Adjuvant (No.CAT F5881, 1 mg/mL), Bradford reagent (No. CAT B6916), trichloroacetic acid (No. CAT T6399), thiobarbituric acid (No. CAT S564508), hydrochloric acid (No. CAT H1758), 2,4-dinitrophenylhidrazine (No. CAT D199303), ethyl acetate (No. CAT 270989), ethylic alcohol (No. CAT E7023), guanidine (No. CAT PH016683), epinephrine (No. CAT 1236970), superoxide dismutase (No. CAT S5395), acetic acid (No. CAT 1005706), hydrogen peroxide (No. CAT 216763), glutathione reductase (No. CAT G9297), HEPES (No. CAT H3375), sodium azide (No. CAT 769320), and NADPH (No. CAT N5130).
Biological material
Lyophilized powder of Spirulina maxima (Sm) was obtained from Solarium Biotechnology
Company, S.A., Santiago, Chile. According to the company procedures complete S. maxima biomass was cultivated and put
in distilled water and then subjected to defrost and freeze cycles, then
stirred to obtain an aqueous extract with all the cellular components in
suspension and finally put in a freeze dryer (Quanta-S Model) coupled to a
vacuum system (Leybold Rotary Vane Model) to lyophilize the water at a
temperature of -50°C and obtain the final dried sterile powder.
Laboratory animals
Male CD1 mice (weighing 20 ± 5 g) were
acquired from BIRMEX (Biological Laboratories Reagents México, D.F.), with a 7
day conditioning period, 12 h light-dark cycles, at a temperature of 25 ± 2°C,
with 55-80% humidity, food (Lab Rodent Chow) and water ad libitum. The experimental protocol was carried out according to
the National Commission of Scientific Investigation and Bioethics and the
Postgraduate and Research Secretariat of the National Polytechnic Institute
(Project no. 20121433) and following
all the guidelines mentioned in the Mexican Official Norm (NOM-062-ZOO-1999) in
relation to the handling and use of laboratory animals.
Inflammation murine
models in vivo:
Carrageenan-induced
edema in mice: This model was performed as described by
Dominguez-Ortiz et al. [27] and Garcia-Rodriguez et al. [28]. All experimental
groups were formed randomly with CD1 male mice (n=8). Treated groups received
by intragastric (i.g.) route indomethacin (IMC) (10 mg/kg) and Sm (400 and 800
mg/kg), 1 h prior to the subcutaneous (s.c.) injection of carrageenan (20 µl,
2% dissolved in sterile isotonic saline solution), prepared at the moment. The
reference drug and S. maxima were
solubilized in Tween80:water (1:9) and carrageenan control received only
vehicle. The percentage of inhibition was calculated by comparing the
measurement of the paw edema at different times (1, 2, 3, 5 and 7 h) (Et) using
a digital micrometer (Mitutoyo model 293-831) and value of time zero (baseline)
(E0). Results were analyzed with the formula described by Olajide et
al. [29]:
% Inhibition = ((Et-Eo)
carrageenan control - (Et-Eo) treated / (Et-Eo)
carrageenan control) × 100
CFA-induced
experimental arthritis in mice: This model was
carried out according to Rasool et al. [25]. All experimental groups control
and treated (n=8) were injected s.c. with 25 µl of CFA in the hind paw in day
zero (Eo). Treatment groups were administered by i.g. route with PBZ
(100 mg/kg) and Sm (400 and 800 mg/kg) daily from day 7 to 21, because physical
clinical signs of experimental arthritis begin to be observed on day 7. All
samples were solubilized in Tween 80:water (1:9) and group of arthritic animals
without treatment only received vehicle. Development of paw edema were measured
at different times (1, 4, 7, 14 and 21 days) (Et) using a digital
micrometer (Mitutoyo
model 293-831) and the value of day
zero (baseline) (Eo). Body weight (BW) gain was also measured the
same days. Percent inhibition of edema in each group was calculated from 7 to
21 day comparing with CFA group without treatment as follows:
%
inhibition = ((Et-Eo) CFA group – (Et-Eo)
Treated group / (Et-Eo) CFA group) × 100
Oxidative stress (OS) measurement: For OS evaluation, animals with CFA-induced
experimental arthritis, were euthanized on the day 21 of the experiment, later
sub-plantar edema tissue was obtained and placed in an ice bath, after this 500
mg were homogenized in 2 ml of cold phosphate buffer solution (PBS, pH 7.3),
from which appropriated aliquots were taken for the determination of the
concentration of carbonyl proteins and the lipid peroxidation rate. 1 ml of the
remaining homogenate was centrifuged (12500 rpm for 15 min at 4°C) and 25 µl of
the supernatant was taken for the quantification of total proteins (TP) by the
Bradford’s technique [30] and the rest of the supernatant was used for the
antioxidant enzymes assays. All the OS measures were realized in an ultraviolet-visible (UV-VIS) spectrophotometer (Shimadzu Model
UV-1700 Double Beam Scanning).
Lipid peroxidation: Evaluation
of lipid peroxidation process was performed by the modified method of Buëge and
Aust [31]. To an aliquot of 500 ml of homogenate were added 1 ml of
thiobarbituric acid reactive substances (TBARS) reagent (trichloroacetic acid
(TCA 16%), thiobarbituric acid (TBA 0.5%), hydrochloric acid (HCl 0.3 N), mixed
by vortexing and then incubated under boiling conditions (at 92°C for 15 min).
Later the samples were placed in ice bath for 10 min and then centrifuged (4000
rpm for 10 min at 4°C). Finally, the absorbance of the supernatant was measured
at 535 nm against a TBARS reagent blank (without homogenate). The results were
expressed as moles of malondialdehyde (MDA)/mg of protein/g of tissue, using
the molecular extinction coefficient (MEC) of 1.56 × 105 M-1cm-1.
Carbonyl proteins: This assay was performed
according to the method described by Parvez and Raisuddin [32], in which to a
300 ml of homogenate, 300 ml of 20% TCA were added, and centrifuged (11500 rpm for 5 min at
4°C). The supernatant was discarded and 150 ml of 2,4-dinitrophenylhidrazine (2,4-DNPH) (10 mM in HCl 2 M) were
added to the precipitated and reconstituted by vortexing for 5 min and subsequently
incubated in a water bath for 1 h at 37°C. After that the samples were
centrifuged (11000 rpm for 10 min at 4°C), the supernatant was discarded and
the precipitated was washed with 1 ml of a solution of ethyl acetate:ethanol
(EtOH-AcOEt, 1:1) and vortexed for 5 min, repeating this step for three times.
The final precipitated was dissolved in 1 ml of guanidine (6 M in 0.2 mM PBS,
pH 2.3) and vortexed for 5 min, then the samples were incubated at 37°C for 40
min and after centrifuged (5500 rpm for 5 min at 4°C). Finally, the supernatant
was measured at 360 nm. As reagent blank 1 ml of HCl 2 M was used. The results
were expressed as moles of reactive carbonyls CO●/mg of protein/g of
tissue, according the MEC 21000 M-1cm-1.
Antioxidant enzymes: Superoxide dismutase
(SOD) was determined as described by Misra and Fridovich [33], measuring
absorbance at 30 s and at 5 min and the difference between these two values was
used to calculate adrenochrome concentration with MEC of 4020 M−1cm−1
at 480 nm. Data was extrapolated in a calibration curve (y=−0.004x + 0.0518; R2=0.9536)
and results were expressed as SOD International Units (IU)/g tissue. Catalase
(CAT) activity was evaluated as described by Radi et al. [34], measuring
absorbance at time zero and at 1 min and difference was employed to calculate
hydrogen peroxide (H2O2) denaturalization with MEC of
0.043 mM−1cm−1 at 240 nm; results were expressed as mmol
H2O2 consumed/min/g tissue. According to this method, one
CAT IU/g tissue represents 1 μmol of H2O2 consumed/min/g
tissue. Glutathione peroxidase (GSH-Px) activity was determined according to
Plagia and Valentine [35], measuring absorbance at time zero and at 1 min with
MEC of 6.2 mM−1cm−1 for 360 nm; results were expressed as
mmol NADPH consumed/min/g tissue.
STATISTICAL ANALYSIS
SigmaPlot ver.
12.0 statistical software (2011-2012) was utilized for analysis of results.
Data is presented as standard error (±) of the mean (SEM). Data from
sub-plantar edema diameter in both models of inflammation (acute and chronic)
and body weight gain during experimental arthritis were analyzed with a
repeated measures (RM) two-way analysis of variance (ANOVA), while results of
oxidative stress parameters were analyzed with one-way ANOVA. The difference
between means was determined by the Student Newman Keuls (SNK) post hoc test.
Results considered statistically significant were those that showed a value p<0.05, according to the confidence
interval destined for biological tests (95%).
RESULTS AND DISCUSSION
Acute inflammation
For acute
inflammation model, we found that IMC generated a statistical decrease (p<0.05), over sub-plantar edema
formation in the first 2 hours after the inoculation of the carrageenan (37 and
41%) compared to carrageenan control at the same hours (0.42 ± 0.02 and 0.44 ±
0.04 mm, respectively), while Sm at 400 and 800 mg/kg caused an inhibition of
23 and 37%, respectively in the second hour compared to those mice injected
with carrageenan and administered only with the vehicle (0.44 ± 0.04 mm) (Figure 1). Furthermore, it was
observed that Sm generated significant edema inhibition at both doses (400 and
800 mg/kg) of 51 and 24%, respectively, at the fifth hour of measure, being the
first dose similar in inhibition to that shown by IMC (58%) (p<0.05), when compared the three
groups to carrageenan control group values at the same hour (0.67 ± 0.03 mm).
In this model of acute inflammation this stage (5 h), is characterized by the
release of free radicals by the immune cells and leukocyte infiltration (Figure 1).
This effect
described above could respond to the suppression of pro-inflammatory cytokines
or vasodilators like histamine, which are known to be generated and released at
this time due to the pharmacokinetic of this acute inflammation model. S. maxima effect on interleukins was
demonstrated by Romay et al. [36] and Gemma et al. [37], in which Spirulina decreased the concentration of tumor
necrosis factor-alpha (TNF-α) and tumor necrosis factor-beta (TNF-β), key
mediators to initiate the inflammatory process.
The previously
described effect may be due to the antioxidant activity of the components of Spirulina maxima, because it is known
that their inactive ROS, being these chemo-attractants during the immune
process and reduce in this way the leukocyte migration and metabolic activity
of neutrophils [38,39]. Another possible mechanism through which this alga
inhibited the edema growth, would be by the suppressive effects of its
constituents like the phycocyanin over the expression of induced nitric oxide
synthase (iNOS) and cyclooxygenase 2 (COX-2), both important enzymes in the
development of the edema generation [40-43] and thus the synthesis of
prostaglandin E2 (PGE2)[44]. To elucidate these important pharmacological steps
further experiments need to be done, such as in vivo evaluation of myeloperoxidase in this acute inflammation
model or the in vitro determination
of COX-2 and iNOS activities, as well nitric oxide concentration.
Chronic inflammation
Although there
was no group of healthy mice as a negative control in various publications
where work has been done with experimental arthritis induced with CFA, it has
been shown that this adjuvant undoubtedly generates a measurable and visible
severe inflammatory condition compared to healthy animals, both in rats and
mice, and more than our vehicle group it was our CFA control without treatment
to compare the anti-edematous and antioxidant effects [26]. During the
development of this experiment, PBZ-treated group showed a statistical increase
in BW at day 14 (263.23%) and 21 (170.31%), compared to arthritic mice without
treatment in the same days (0.94 ± 0.39 and 1.28 ± 0.56 g, respectively).
Furthermore, animals administered with Sm at 400 mg/kg exhibited a significant
increase of BW on days 14 (138.30%) and 21 (329.69%) when compared to
CFA-induced control group that only was administered with vehicle, while the
higher tested dose of Sm (800 mg/kg), did not show a positive effect in the
gain of BW during the chronic inflammation model (Table 1).
In the model of
chronic inflammation induced by CFA, one of the general indicators that there
is in fact an immune process is the loss of BW due to the hyperalgesia
generated by pro-inflammatory cytokines [26,39]. The weight loss can be
explained in two ways, due to pain produced during chronic inflammation,
provoking the immobility of the animals and leading to the alteration of normal
feeding habits [45] or by bad absorption syndromes of nutrients in the
intestine[46]. In this study, the treatment with Spirulina maxima at lower dose was able to maintain BW and although
this alga has not been used on inflammatory
bowel disease (IBD) models, the same protective effect was found in
isolated natural products such as curcumin, which generated the inhibition of
TNF-β, IL-1β and IL-6, decreased the immune response and favored both
absorption and transport of nutrients [47] and it is known that Spirulina maxima has molecules that
inhibit this cytokines.
For paw edema
formation induced by CFA, PBZ-treated group showed a significant decrease in
size on day 14 and 21 (53.14 and 52.18%, respectively) (p<0.05), compared to CFA un-treated group (1.59 ± 0.10 and 1.27
± 0.09 mm, respectively). Animals that were administered with the 400 mg/kg
dose of Sm, showed a significant greater inhibition at day 14 (51.22%) than in
day 21 (37.97%) when compared to CFA control group; however, higher evaluated
dose of Sm (800 mg/kg) only generated a statically moderate anti-inflammatory
effect in both days (37.92 and 26.21%, respectively), than the dose of 400
mg/kg (Figure 2).
The inhibitory
effect generated by Sm on the edema development, is probably, among other
possible pharmacological mechanisms, due to the suppression of ROS during the
oxidative phase of chronic inflammation [39]. During an immune process there
exist two pathways in which it begins and is exacerbated, one that depends on
enzymes like COX-2 and another that is oxidative, the last one is related to
the generation of free radicals. Today is known that OS is involved in the
development of various human immune pathologies related to both acute and
chronic inflammation [6].
OS microenvironment in edema tissue
The tests of OS
in edema tissue of arthritic animals showed that CFA control group exhibited
high levels of oxidized proteins (21.84 ± 1.87 μm CO●/g tissue), and
an increased lipid peroxidation (2.24 ± 0.43 μm MDA/g tissue), while
PBZ-treated group exhibited a statistical decrease in the concentration of
carbonyl proteins (41.70%) and lipid peroxidation (71.87%) (p<0.05). Furthermore, Sm administered
groups at both doses 400 and 800 mg/kg had a statistical decrease in the
concentration of oxidized proteins (69.49 and 59.29%, respectively), while for
lipid peroxidation Sm treated arthritic animals exhibited a significant
decrease in a non-dose dependent effect as well as in the oxidation of
proteins, in which 400 mg/kg dose caused a greater decrease (80.87%) while
higher tested dose 800 mg/kg generated a lesser antioxidant effect protecting
lipid from oxidative damage (49.13%), when both groups were compared to CFA
un-treated group of arthritic mice (Table
2).
In this context
food supplements like Spirulina maxima has
a large number of components with antioxidant activity, such as phycobiliproteins and unsaturated fatty acids like the
gamma linolenic acid and these ones also have a suppressing effect over
pro-inflammatory cytokines, such as TNF-α, IL-1β, IL-6 and IL-12 [12,40,48]. By
inhibiting these chemical mediators of the inflammation, the effect could be at
different levels of the immune process, such as the generation of free radicals
or the differentiation and maturation of T cells, that are characteristic of a
chronic inflammatory process [46,49].
An increase of
LPO and PCC indicates that there is an important and directly relation between
the experimental monoarthritis onset and the OS, in which this last one helps
in the maintenance and exacerbation of the chronic immune process [50]. During
the development of this research work, arthritic animals treated with Spirulina maxima, had decreased their
oxidized proteins levels, as also the rate of lipid peroxidation. This can be
explained by the fact that this alga contains several bioactive compounds,
including C-phycocyanin, which possess great antioxidant activity reported by
Romay et al. [51] and confirmed by Riss et al. [52]. Other substances with high
antioxidant activity are beta carotene and gamma linolenic acid [41,42].
For the
antioxidant response, those mice that were from the CFA control group exhibited
an antioxidant enzymatic activity for SOD of 2257.05 ± 1.96 IU/g tissue, CAT
60.01 ± 0.03 UI/g tissue and GSH-Px of 3.09 ± 0.20 µmol NADPH/g tissue, while
PBZ-treated arthritic mice showed for SOD activity an increase of 80.53%
(p<0.01), for CAT enzyme a statistical decrease of 66.62% (p<0.05) and for GSH-Px enzyme a
significant decrease of 68.28% compared to CFA control group (Table 3). On the other hand, those
arthritic mice treated with Spirulina
maxima generated a regulation on the antioxidant enzymatic response in a no
dose-dependent manner, where dose of 400 mg/kg showed a statistical decrease in
SOD activity of 42.98% (p<0.05), however for CAT and GSH-Px activities it
generated a significant increase of 3216% and 130%, respectively compared to
CFA control group. Arthritic mice group treated with S. maxima at dose of 800 mg/kg showed a lower beneficial effect and
different behavior compared to the previous described dose over the antioxidant
response in sub-plantar edema tissue with an increase for SOD enzyme activity
(89.27%), and a decrease for CAT and GSH-Px activities of 49.94% and 35.28%,
respectively when compared to CFA control group (Table 3).
This behavior
is commonly known due to the existence of link between oxidative stress and
inflammatory processes, where these last ones have an oxidative pathway for the
pathogen elimination as well for the signaling during immune response, this
occurs when the host organism could not regulate the inflammatory response so
this triggers a chain of cellular and immune reactions which establish an
oxidative stress and makes the pass from acute to chronic inflammation [53]. It
has been proven that an increase in ROS amounts at articular level stimulates a
greater expression of TNF-α in blood, favoring in this way the propagation of
the inflammatory chronic response due to this interleukin acts as a
chemo-activator of T lymphocytes and activated macrophages [54]. Also it has
been proven that the antioxidant enzymatic defenses are compromised during a
chronic inflammatory process such as arthritis, where levels of reduced
glutathione (GSH) are below the normal values generated in articular tissue and
also in systemic level [55].
In the
articular tissue of arthritic mice treated with Spirulina maxima we found a significant decrease so much of
oxidized proteins as well lipid peroxidation, and generated also an increase of
both enzymes CAT and GSH-Px at dose of 400 mg/kg, while for SOD activity a
decreased was observed, these results match to those published by Kuriakose and
Kurup in 2011 [56], where a treatment with S.
laxissima stimulates a greater antioxidant enzymatic response in which the
only activity of SOD could not be consider as beneficial to the host organism
since it only reduces the reactivity of the anion superoxide for a molecule of
hydrogen peroxide (which is still cytotoxic), so the coupled system of SOD-CAT
or SOD-GSH-Px is needed.
This mechanism
could be explained due to this observed antioxidant effect is generated by the
cellular components of these cyanobacteria such as gamma linoleic acid which
has demonstrated biological activities like antioxidant, anti-inflammatory and
anti-proliferative [57,58]. These cyanobacteria also possesses an important
content of amino acids like cysteine which is a precursor molecule of the GSH,
which increase the levels of this compound in the host body, as well as
glutathione reductase enzyme [59].
In the other
hand, it’s well-known that S. maxima
has also a large number of essential minerals needed as enzymatic co-factors
such as selenium [60]. Another important component of these cyanobacteria is
the ferredoxin which are molecules in charge of reduce NADP to NADPH, important
co-factor in the regulation of the organism’s oxide-reduction environment [61].
The antioxidant
activity of S. maxima has been widely
studied and supported in diverse pre-clinical experimental animal models of
oxidative stress where the beneficial effect of these cyanobacteria was
reported previously, for example Canchihuaman et al. [62] in a lead-induced
liver oxidative stress model, S. maxima
treatment decrease lipid peroxidation and favored the antioxidant enzymatic
activity in liver tissue.
For everything
described previously, S. maxima
capacity to generate an anti-inflammatory effect and antioxidant activity is
due to the ability of its components to inhibits the pro-inflammatory cytokine
TNF-α, which favored the immune response and also increases transcription of
oxidative stress during inflammation via the NF-kB pathway, as well the
antioxidant effect of this cyanobacteria is due to its molecular and cellular
components such as polyunsaturated fatty acids and antioxidant enzymes like
glutathione reductase [54,63].
Finally through
all the experiments 400 mg/kg was the best dose, which generated both
anti-edematous effect in both acute and chronic models, increase BW gain in
arthritic mice, and reducing oxidative damage. In previous works same dose of Spirulina fusiformis, showed beneficial
effects in various chronic inflammation models induced by zymosan [64],
collagen [39] and CFA-induced experimental arthritis in rats[26,65].
This beneficial
effect at dose of 400 mg/kg is due may be to saturation on gastric (20%) and
intestinal (80%) absorption of Spirulina constituents
such as proteins and poly-unsaturated acids which increase their
bio-availability when this alga is orally consumed, nonetheless in previous
published works this kind of biological effect is well-known as hormesis, where
a bi-phasic dose-response is observed due to the phenomena of saturation in the
absorption site, for this reason lower doses allow the perfect balance between
absorption and blood concentration of the evaluated compounds, allowing their
distribution in the body and increasing their bioavailability [66], however
pharmacokinetics of Spirulina remain unknown and need further studies[63].
CONCLUSION
In conclusion
due to what we have previously described and found in this research work, we
demonstrated that, in fact exists a direct relationship between the antioxidant
activity of Spirulina maxima and its
anti-inflammatory properties in vivo
on a located zone in which develops a chronic immune process, exhibiting a
higher anti-inflammatory effect at dose of 400 mg/kg as well as a greater
ability to regulate the oxidative stress develop within these immune processes.
Thus Spirulina maxima could works as
a co-adjuvant in the treatment of chronic degenerative diseases related to
acute and chronic inflammation by inhibiting the oxidative damage over
biomolecules and regulating the antioxidant enzymatic endogenous response in
compromised tissue.
CONFLICT OF INTERESTS
The authors
declare that there is no conflict of interest regarding the publication of this
paper.
1.
Chen L, Deng H, Cui H, Fang J, Zuo Z, et al. (2018)
Inflammatory responses and inflammation-associated diseases in organs.
Oncotarget 9: 7204-7218.
2.
Rathes A, Geng S, Lee C, Li L (2018) Cellular and
molecular mechanisms involved in the resolution of innate leukocyte
inflammation. J Leuko Biol 104: 535-541.
3.
Kim B, Lee JH, Seo MJ, Eom SH, Kim W (2016) Linarin
down-regulates phagocytosis, proinflammatory cytokine production and activation
marker expression in RAW264.7 macrophages. Food Sci Biotechnol 25: 1437-1442.
4.
Parhiz H, Roohbakhsh A, Soltani F, Rezaee R (2015)
Antioxidant and anti-inflammatory properties of the citrus flavonoids
hesperidin and hesperetin: An updated review of their molecular mechanisms and
experimental models. Phytother Res 29: 323-331.
5.
Zhu GF, Guo HJ, Huang Y, Wu CT, Zhang XF (2015)
Eriodictyol, a plant flavonoid, attenuates LPS-induced acute lung injury
through its anti-oxidative and anti-inflammatory activity. Exp Ther Med 10:
2259.
6.
Chen X, Zhang S, Xuan Z, Ge D, Chen X, et al. (2017)
The phenolic fraction of Mentha haplocalyx
and its constituent linarin ameliorate inflammatory response through
inactivation of NF-κB and MAPKs in lipopolysaccharide-induced RAW264.7 cells.
Molecules 22: 811.
7.
Heo SJ, Hwang JY, Choi JI, Han JS, Kim HJ, et al.
(2009) Diphlorethohydroxycarmalol isolated from Ishige okamurae, a brown algae, a potent [alpha]-glucosidase and
[alpha]-amylase inhibitor, alleviates post-prandial hyperglycemia in diabetic
mice. Eur J Pharmacol 615: 252-256.
8.
Prasad S, Phromnoi K, Yadav VR, Chaturvedi MM,
Aggarwal BB (2010) Targeting inflammatory pathways by flavonoids for prevention
and treatment of cancer. Planta Med 76: 1044-1063.
9.
Aggarwal BB (2010) Targeting inflammation-induced
obesity and metabolic diseases by curcumin and other nutraceuticals. Annu Rev
Nutr 30: 173-199.
10.
Kim S, Wijesekara I (2010) Development and
biological activities of marine-derived bioactive peptides: A review. J Funct
Foods 2: 1-9.
11.
Pangestuti R, Kim SK (2011). Neuro-protective
effects of marine algae. Mar Drugs 9: 803-818.
12.
Belay A (2002) The potential application of
Spirulina (Arthrospira) as a nutritional and therapeutic supplement in health
management. JANA 5: 27-48.
13.
Sánchez N, Bu M, León N, Pérez-Saad H (2002)
Fundamentos de una posible acción beneficiosa de la Spirulina platensis en las neuropatías periféricas. Rev Cubana
Plant Med 7: 146-150.
14.
Celekli A, Yavuzatmaca M (2009) Predictive modeling
of biomass production by Spirulina
platensis as function of nitrate and NaCl concentrations. Bioresour Technol
100: 1847-1851.
15.
Ponce-Canchihuamán J, Pérez-Méndez O,
Hernández-Muñoz R, Torres-Durán P, Juárez-Oropeza M (2010) Protective effects
of Spirulina maxima on hyperlipidemia
and oxidative-stress induced by lead acetate in the liver and kidney. Lipids
Health Dis 9: 35-41.
16.
Sánchez M, Bernal-Castillo J, Rozo C, Rodríguez I
(2003) Spirulina (Arthrospira): An edible microorganism: A review. Univer
Scient 8: 7-20.
17.
Gutiérrez-Salmeán G, Fabila-Castillo L,
Chamorro-Cevallos GA (2015) Nutritional and toxicological aspects of Spirulina
(Arthrospira). Nutr Hosp 32: 34-40.
18.
Chamorro G, Salazar M, Favila L, Bourges H (1996).
Farmacología y toxicología del alga Spirulina. Rev Invest Clín 48: 389-399.
19.
Mader J, Gallo A, Schommartz T, Handke W, Nagel CH,
et al. (2016) Calcium spirulan derived from Spirulina
platensis inhibits herpes simplex virus 1 attachment to human keratinocytes
and protects against Herpes labialis.
J Allergy Clin Immunol 137: 197-203.
20.
Setyaningsih I, Bintang M, Madina N (2015)
Potentially anti-hyperglycemic from biomass and phycocyanin of Spirulina fusiformis Voronikhin by in vivo test. Procedia Chem 14: 211-215.
21.
Hernández Lepe MA, Wall-Medrano A, Juárez-Oropeza
MA, Ramos-Jiménez A, Hernández-Torres RP (2015). Spirulina and its
hypolipidemic and antioxidant effects in humans: A systematic review. Nutr Hosp
32: 494-500.
22.
Czerwonka A, Kaławaj K, Sławińska-Brych A, Lemieszek
MK, Bartnik M, et al. (2018). Anticancer effects of the water extract of a
commercial Spirulina (Arthrospira) platensis product on the human lung
cancer A549 cell line. Biomed Pharmacother 106: 292-302.
23.
Selmi C, Leung PS, Fischer L, German B, Yang CY, et
al. (2011) The effects of Spirulina on anemia and immune function in senior
citizens. Cell Mol Immunol 8: 248-254.
24.
Pham TX, Lee JY (2016) Anti-inflammatory effect of Spirulina platensis in macrophages is
beneficial for adipocyte differentiation and maturation by inhibiting nuclear
factor-κB pathway in 3T3-L1 adipocytes. J Med Food 19: 535-524.
25.
Rasool M, Sabina EP, Lavanya B (2006)
Anti-inflammatory effect of Spirulina fusiformis
on adjuvant-induced arthritis in mice. Biol Pharm Bull 29: 2483-2487.
26.
Gutiérrez-Rebolledo GA, Galar-Martínez M,
García-Rodríguez RV, Chamorro-Cevallos GA, Hernández-Reyes AG, et al. (2015).
Antioxidant effect of Spirulina (Arthrospira)
maxima on chronic inflammation
induced by Freund’s complete adjuvant in rats. J Med Food 18: 865-871.
27.
Domínguez-Ortiz MA, Muñoz-Muñiz OD, García-Rodríguez
RV, Vázquez-Hernández M, Gallegos-Estudillo J, et al. (2010) Anti-oxidant and
anti-inflammatory activity of Moussonia deppeana (Schldl. and Cham) Hanst. Bol
Latinoam Caribe Plantas Med Aromát 9: 13-19.
28.
García-Rodríguez RV, Gutiérrez-Rebolledo GA,
Méndez-Bolaina E, Sánchez-Medina A, Maldonado-Saavedra O, et al. (2014) Cnidoscolus chayamansa McVaugh, an
important anti-oxidant, anti-inflammatory and cardio-protective plant used in
Mexico. J Ethnopharmacol 151: 937-943.
29.
Olajide OA, Makinde JM, Awe SO (1999) Effects of the
aqueous extract of Bridelia ferruginea
stem bark on carrageenan-induced edema and granuloma tissue formation in rats
and mice. J Ethnopharmacol 66: 113-117.
30.
Bradford M (1976) A rapid and sensitive method for
the quantization of microgram quantities of protein utilizing the principle of
protein dye binding. Anal Biochem 72: 248-254.
31.
Buege J, Aust S (1978). Microsomal lipid
peroxidation. Methods Enzymol 52: 302-310.
32.
Parvez S, Raisuddin S (2005) Protein carbonyls:
novel biomarkers of exposure to oxidative stress-inducing pesticides in
freshwater fish Channa punctata
(Bloch). Environ Toxicol Pharmacol 20: 112-117.
33.
Misra HP, Fridovich I (1972) The role of superoxide
anion in the autoxidation of epinephrine and a simple assay for superoxide
dismutase. J Biol Chem 247: 3170-3175.
34.
Radi F, Turrens JF, Chang LY, Bush KM, Crapo JD, et
al. (1991) Detection of catalase in rat heart mitochondria. J Biol Chem 266:
22028-22034.
35.
Plagia DE, Valentine WN (1967) Studies on the
quantitative and qualitative characterization of erythrocyte glutathione
peroxidase. J Lab Clin Med 70: 158-169.
36.
Romay C, Delgado R, Remirez D, González R, Rojas A
(2001). Effects of phycocyanin extract on tumor necrosis factor-alpha and
nitrite levels in serum of mice treated with endotoxin. Arzneimittelforschung
51: 733-736.
37.
Gemma C, Mesches MH, Sepesi B, Choo K, Holmes DB, et
al. (2002). Diets enriched in foods with high antioxidant activity reverse
age-induced decreases in cerebellar beta-adrenergic function and increases in
pro-inflammatory cytokines. J Neurosci 22: 6114-6120.
38.
Deng R, Chow TJ (2010) Hypolipidemic, antioxidant
and anti-inflammatory activities of microalgae Spirulina. Cardiovasc Ther 28:
e33-e45.
39.
Kumar N, Kumar P, Singh S (2010) Immunomodulatory
effect of dietary Spirulina platensis
in type II collagen induced arthritis in mice. RJPBCS 1: 877-885.
40.
Remirez D, Fernández V, Tapia G, González R, Videla
LA (2002) Influence of C-phycocyanin on hepatocellular parameters related to
liver oxidative stress and Kupffer cell functioning. Inflamm Res 51: 351-356.
41.
Bai SK, Lee SJ, Na HJ (2005) β-Carotene inhibits
inflammatory gene expression in lipopolysaccharide-stimulated macrophages by
suppressing redox-based NF-kappa-β activation. Exp Mol Med 37: 323-334.
42.
Katsuura S, Imamura T, Bando N, Yamanishi R (2009)
β-carotene and beta-cryptoxanthin but not lutein, evoke redox and immune changes
in RAW264 murine macrophages. Mol Nutr Food Res 53: 1396-1405.
43.
Patel A, Mishra S, Ghosh PK (2006) Antioxidant
potential of C-phycocyanin isolated from cyanobacterial species Lyngbya, Phormidium and Spirulina
spp. Indian J Biochem Biophys 43: 25-31.
44.
Inglis JJ, Nissim A, Lees DM, Hunt SP, Chernajovsky
Y, et al. (2005) The differential contribution of tumor necrosis factor to
thermal and mechanical hyperalgesia during chronic inflammation. Arthritis Res
Ther 7: R807-816.
45.
Franch A, Castellote C, Castell M (1994) Blood
lymphocyte subsets in rats with adjuvant arthritis. Ann Rheum Dis 53: 461-466.
46.
Rasool M, Sabina EP (2009) Appraisal of
immunomodulatory potential of Spirulina
fusiformis and in vivo and in vitro study. J Nat Med 63: 169-175.
47.
Ali T, Yun L, Rubin DT (2012) Risk of post-operative
complications associated with anti-TNF therapy in inflammatory bowel disease.
World J Gastroenterol 18: 197-204.
48.
Romay CH, González R, Ledón N, Remirez D, Rimbau V
(2003) C-Phycocyanin: A biliprotein with antioxidant, anti-inflammatory and
neuroprotective effects. Curr Protein Pept Sci 4: 207-216.
49.
Ravi M, Lata De S, Azharuddin S, Solomon P (2010)
The beneficial effects of Spirulina focus on its immunomodulatory and
antioxidant properties. Nutr Diet Suppl 2: 73-83.
50.
Kumar VL, Roy S (2007) Calotropis procera latex extract affords protection against
inflammation and oxidative stress in Freund’s complete adjuvant-induced
monoarthritis in rats. Mediators Inflamm 2007: 1-7.
51.
Romay C, Armesto J, Remirez D, González R, Ledon N,
et al. (1998) Antioxidant and anti-inflammatory properties of C-phycocyanin
from blue-green algae. Inflamm Res 47: 36-41.
52.
Riss J, D´ecordé K, Sutra T, Delage M, Baccou JC, et
al. (2007) Phycobiliprotein C-phycocyanin from Spirulina platensis is powerfully responsible for reducing
oxidative stress and NADPH oxidase expression induced by an atherogenic diet in
hamsters. J Agric Food Chem 55: 7962-7967.
53.
Lonkar P, Dedon PC (2011) Reactive species and DNA
damage in chronic inflammation: Reconciling chemical mechanisms and biological
fates. Int J Cancer 128: 1999-2009.
54.
Filippin LI, Vercelino R, Marroni NP, Xavier RM
(2008) Redox signaling and the inflammatory response in rheumatoid arthritis.
Clin Exp Immunol 152: 415-422.
55.
Siems W, Bresgen N, Brenke R, Siems R, Kitzing M, et
al. (2010) Pain and mobility improvement and MDA plasma levels in degenerative
osteoarthritis, low back pain and rheumatoid arthritis after infrared
A-irradiation. Acta Biochem Pol 57: 313-319.
56.
Kuriakose GC, Kurup MG (2011) Antioxidant and
anti-hepatotoxic effect of Spirulina
laxissima against carbon tetrachloride induced hepatotoxicity in rats. Food
Funct 2: 190-196.
57.
Olveira G, Olveira C, Acosta E, Espildora F,
Garrido-Sanchez L, et al. (2010) Fatty acid supplementation improves
respiratory, inflammatory and nutritional parameters in adults with cystic
fibrosis. Arch Bronconeumol 46: 70-77.
58.
Olendzki BC, Leung K, Van Buskirk S, Reed G, Zurier
RB (2011) Treatment of rheumatoid arthritis with marine and botanical oils:
Influence on serum lipids. Evid Based Complement Altern Med 2011: Article ID
827286.
59.
Gershwin ME, Belay A (2007) Spirulina in human
nutrition and health. 5th Edn. Editorial CRC Press Taylor and
Francis Group: New York, United States of America, pp: 159-312.
60.
Cases J, Vacchina V, Napolitano A, Caporiccio B
(2001) Selenium from selenium-rich Spirulina is less bioavailable than selenium
from sodium selenite and selenomethionine in selenium-deficient rats. J Nutr
131: 2343-2350.
61.
Fukuyama K, Ueki N, Nakamura H, Tsukihara T,
Matsubara H (1995) Tertiary structure of [2Fe-2S] ferredoxin from Spirulina platensis refined at 2.5 •ð
resolution: Structural comparisons of plant-type ferredoxins and an
electrostatic potential analysis. Biochemistry 117: 1017-1023.
62.
Canchihuamán JC, Méndez OP, Muñoz RH, Durán PV,
Oropeza MA (2010) Protective effects of Spirulina
maxima on hyperlipidemia and oxidative-stress induced by lead acetate in
the liver and kidney. Lipids Health Dis 9: 35-43.
63.
Deng R, Chow TJ (2010) Hypolipidemic, anti-oxidant
and anti-inflammatory activities of microalgae Spirulina. Cardiol Ther 28:
e33-e45.
64.
Remirez D, González R, Merino N, Rodríguez S,
Ancheta O (2002) Inhibitory effects of Spirulina in zymosan-induced arthritis
in mice. Mediators Inflamm 11: 75-79.
65.
Ali EAI, Barakat BM, Hassan R (2015) Anti-oxidant
and angiostatic effect of Spirulina
platensis suspension in complete Freund’s adjuvant-induced arthritis in
rats. PLoS One 10: e0121523.
66.
Gálvez I, Torres-Piles S, Ortega-Rincón E (2018)
Balneotherapy, immune system and stress response: A hormetic strategy? Int J
Mol Sci 19: 1687.
QUICK LINKS
- SUBMIT MANUSCRIPT
- RECOMMEND THE JOURNAL
-
SUBSCRIBE FOR ALERTS
RELATED JOURNALS
- Journal of Cardiology and Diagnostics Research (ISSN:2639-4634)
- International Journal of Anaesthesia and Research (ISSN:2641-399X)
- Journal of Clinical Trials and Research (ISSN:2637-7373)
- Journal of Alcoholism Clinical Research
- International Journal of Clinical Case Studies and Reports (ISSN:2641-5771)
- Journal of Renal Transplantation Science (ISSN:2640-0847)
- Ophthalmology Clinics and Research (ISSN:2638-115X)