Effect of Fish or Soybean Oil-Rich Diets on Bradykinin,Kallikrein, Nitric Oxide, Leptin, Corticosteroneand Macrophages in Carrageenan Stimulated Rats

Marta Wohlers,1 Roberta Araujo Navarro Xavier,1 Lila Missae Oyama,1

Eliane Beraldi Ribeiro,1 Claudia Maria Oller do Nascimento,1

Dulce Elena Casarini,2 and Vera Lucia Flor Silveira1,3

Abstract—We have previously demonstrated that both nY3 and nY6 polyunsaturated fatty acids

(PUFA)-rich diets decrease the acute inflammatory response partially explained by the high cor-

ticosterone basal levels. The present study aimed to determine the effect of hyperlipidic diets

(PUFA nY3 or nY6) on phagocytosis, hydrogen peroxide (H2O2) and nitric oxide (NO) release by

macrophages, bradykinin (BK) and NO release in the paw inflammatory perfusate and Kallikrein

(KK), corticosterone and leptin blood levels. Hyperlipidic diets decreased H2O2 release from

macrophages stimulated by carrageenan or phorbol-miristate-acetate (PMA), NO release from

macrophage stimulated by carrageenan, BK and NO release in the edema perfusate, KK plasma

levels and the increase of serum leptin after carrageenan stimulus. These data show that both fish

and soybean oil-rich diets promote similar alterations on inflammatory mediators of carrageenan

edema and a causal association with the anti-inflammatory effect of these diets.

KEY WORDS: polyunsaturated fatty acids; inflammation; corticosterone; bradykinin; nitric oxide; hydrogenperoxide; leptin.

INTRODUCTION

It has been demonstrated that dietary lipids are

incorporated into phospholipids cell membranes chang-

ing its fluidity, activity and release of intracellular

mediators [1, 2]. Many studies have shown that

alterations in the type and quantity of polyunsaturated

fatty acids (PUFA) from diet could modify immuno-

logical and inflammatory responses [3, 4].

Although the exact mechanisms responsible for

these effects are not fully elucidated, there have been

recent reports showing that PUFAs such as DHA

(docosahexaenoic acid, 22:6, nY3) and AA (arachidonic

acid, 20:4, nY6) induce the production of anti-inflam-

matory mediators, named docosatrienes, protectins,

lipoxins, and resolvins [5, 6]. Those eicosanoids have

been shown to play an important role during the

resolution phase of the inflammatory process and could

help explain the anti-inflammation induced by diets

enriched with those PUFAs [7].

Additionally, clinical studies have shown beneficial

effects of dietary supplementation with fish oil (rich in

nY3 PUFA), under chronic inflammatory conditions [8,

9] and these effects have been attributed to a decrease in

the generation of inflammatory mediators derived from

the AA, with the production of less potent inflammatory

mediators, after nY3 PUFA-rich diets treatment [9Y11].

0360-3997/06/0200-0081/0 # 2006 Springer Science+Business Media, Inc.

Inflammation, Vol. 29, Nos. 2Y3, April 2005 (# 2006)

DOI: 10.1007/s10753-006-9002-2

81

1 Physiology Department, Federal University of Sao PauloVEPM,

Sao Paulo, Brazil.2 Medicine Department, Federal University of Sao PauloVEPM, Sao

Paulo, Brazil.3 To whom correspondence should be addressed at Departamento de

Fisiologia, Disciplina de Fisiologia da Nutricao, Universidade

Federal de Sao PauloVEscola Paulista de Medicina, Rua Botucatu,

862-2- andar, Vila ClementinoVCEP, Sao Paulo 04023-060, Brazil.

E-mail: vera@ecb.epm.br

However, we previously observed that both fish

and soybean oil-rich diets were able to attenuate the

acute inflammatory process, which could be partially

attributed to increased corticosterone levels [12].

The inflammatory process is a response of the body

to injury, aimed at attacking the agent while limiting

and/or repairing tissue damage, thus helping the

reestablishment of homeostasis. The acute inflammatory

response is characterized by the development of

erythema, pain, and edema, as a result the local release

of inflammatory mediators, such as BK, NO, catechol-

amines, eicosanoids, and others [13, 14]. Macrophages

have an essential role on the inflammatory process and

dietary manipulations can change their fatty acids

composition, leading to alterations in the citotoxic

activity of these cells, such as phagocytosis and

production of reactive oxygen species [15].

The purpose of the present study was to examine

the effect of the chronic consumption of diets rich in

nY6 or nY3 PUFAs, on aspects of the acute inflamma-

tion process. For this, we evaluated macrophages

functionality and local as well as plasma release of

carrageenan edema mediators, as BK, KK, NO, and

glucocorticoids. Secondly, we aimed at verifying the

effect of these diets on plasma leptin, a hormone

released mainly from adipose tissue and involved in

the inflammatory process [16]. The association between

those mediators and the anti-inflammatory effect of the

lipid diets was examined.

MATERIAL AND METHODS

Animals and Treatment

The Committee on Experimental Research Ethics

from Federal University of Sao Paulo approved all

procedures involving animals.

Male Wistar rats (28Y30 days) weighing 65Y70 g

were supplied by the animal care facility of the Federal

University of Sao Paulo. They were housed under

controlled conditions of lighting (12:12 h lightYdark

cycle with lights on at 7:00 A.M.) and temperature (24 T1-C). During seven weeks, the rats had free access to

water and one of three types of diets: standard balanced

chow with 4% fat and 20% protein (Nuvilab) (control

group), nY6 PUFA-rich-diet prepared by adding 15% of

soybean oil to control diet (soybean group), or nY3

PUFA-rich diet prepared by adding 15% of fish oil

(Sigma) to the control diet (fish group). Twelve percent

of casein was added to the lipid diets to achieve 20% of

protein. Caloric density of diets was determined using an

adiabatic calorimeter IKA-C400. They were 17.4 KJ/g

for the standard chow and 20.5 KJ/g for both PUFA-rich

diets. Once prepared, the diets were kept frozen until

usage. The rats were provided with a fresh food cup

every day and a 24-h food intake measurement was

performed once a week. The fatty acids composition of

the diets was analyzed at the Nutrition Institute of

Federal University of Rio de Janeiro (Table 1).

Carrageenan-Induced Paw Edema

The animals were anesthetized with sodium pen-

tobarbital (40 mg/kg, i.p.). Anesthetic supplementation

(20 mg/Kg) was provided as required to keep a light

anesthesia level during the whole experiment. Carra-

geenan (0.1 mg) was injected in the right subplantar

region. Some animals were decapitated after 1, 2, 3, or

4 h later and trunk blood collected. Plasma corticoste-

rone levels were determined by fluorometry [17]. Serum

leptin was determined in the 1- and 3-h samples by

radioimmunoassay (Linco Research, Inc, USA).

In other carrageenan-stimulated animals, the tail

vein was punctured both 1 and 2 h after carrageenan for

Table 1. Fatty Acids Composition (g/100 g of diet) of Control, Fish,

and Soybean Diets

Fatty acid Control Fish Soybean

14:0 0.061 1.01 0.04

16:0 0.66 3.33 1.80

18:0 0.100 0.55 0.50

14:1 0.048 0.12 0.09

16:1 0.004 1.21 0.015

17:1 Y 0.10 Y18:1 nj9 0.80 2.02 3.43

24:1 nj9 0.002 0.05 Y18:2 nj6 2.11 1.71 8.08

18:3 nj6 0.008 0.015 0.04

18:3 nj3 0.13 0.09 0.81

20:3 nj3 Y 0.07 Y20:4 nj6 Y Y Y22:4 nj6 Y 0.06 Y22:5 nj3 Y 0.26 YEPA 0.002 1.44 0.011

DHA 0.01 1.58 0.004

Saturated 0.83 4.90 2.34

Monounsaturated 0.85 3.51 3.54

Polyunsaturated 2.26 5.22 8.94

nj6 Polyunsaturated 2.12 1.78 8.12

nj3 Polyunsaturated 0.14 3.44 0.82

Polyunsaturated: saturated 2.72 1.06 3.82

nj6 : nj3 15.14 0.52 9.90

82 Wohlers, Xavier, Oyama, Ribeiro, Nascimento, Casarini, and Silveira

determination of plasma kallikrein (KK) activity. This

was performed by measuring the production of p-nitro

anile (pNA) from the hydrolysis of the synthetic

substrate, S2302 [18].

For determination of local levels of nitric oxide

(NO) and bradykinin (BK), a coaxial perfusion of the

hind paw was performed (modified from [19]). Briefly,

the paw was cannulated before carrageenan injection

and perfused at 53 ml/min, with perfusion fluid (158

mM NaCl, 5.65 mM CaCl, 0.54 mM CaCl2, 2.90 mM

MgCl2, 178 mM NaHCO3, and 1.0 g/l glucose, main-

tained in bath at 36.5-C). Two one-hour paw perfusate

samples were collected, from 1 to 2 h and from 2 to 3

h after carrageenan, into ice-cold tubes. The samples

were centrifuged at 2.500 rpm, at 4-C, for 15 min and

the supernatant used for NO quantification. For BK

determination, the perfusate was collected into 80%

ethanol, centrifuged, and the supernatant evaporated in

gaseous nitrogen. The samples were stored at Y70-C.

NO levels were evaluated by nitrite (NO2Y) production,

using Griess reagent [20]. BK levels were determined

by reverse-phase HPLC [21].

The volume of the perfused paw was determined

by plethysmography (H. Basile, Italia), both before and

3 h after carrageenan administration.

Carrageenan-Induced Peritoneal Inflammation

The anesthetized animals received an intraperito-

neal injection of carrageenan (20 mg). Three hours

later, the animals were decapitated and the peritoneal

exudate was collected by lavage with PBS [22]. After

centrifugation, the supernatant was used for determina-

tion of NO level, by the method described above [20].

The in vitro release of NO and H2O2 from the

exudate macrophages was also determined. For this,

1 � 106 macrophages from the precipitate were

Fig. 1. Determination of phagocytosis (in vitro zymozan stimulated),

H2O2 and NO (basal, stimulated in vitro by PMA or in vivo by 20 mg

of carrageenan peritoneal) in macrophages of rats from control ( ),

soybean ( ), and fish ( ) groups. Each column represents the

mean T s.e.m. from 9 to 17 animals. (a) Different ( p < 0.05) from

control group, (b) from fish group, and (c) from respective basal.

Fig. 2. NO concentration (mM/100 ml) in peritoneal exudate of rats from

control ( ), soybean ( ) and fish ( ) groups, 3 h after peritoneal

carrageenan stimulus. Each column represents the mean T s.e.m. from 9 to

10 animals. (a) Different ( p < 0.05) from control group.

83Fish/Soybean Oil-Rich Diets Effect on BK, KK, NO, Leptin, Corticosterone and Macrophages in Rats

incubated for 1 h, at 37-C, in PBS containing 5 mM

glucose, 2 mM glutamine, 2% BSA, and 1.3 mM

calcium (incubation solution). The medium was

harvested and NO was measured as described above

[20]. H2O2 in the medium was determined by the

horseradish peroxidase-induced oxidation of phenol red

[23, 24]. Results were normalized for cells total protein

content [25].

For these assays, cell viability was examined by

Trypan blue exclusion (>96%) and cell identity exam-

ined by differential count, confirming that at least 92%

of the peritoneal exudate cells were macrophages.

In vitro Macrophage Stimulation

This procedure was performed in incubated macro-

phages harvested by peritoneal lavage of rats not

stimulated in vivo. The in vitro stimulation consisted

of adding 10 ng of PMA (phorbol myristate acetate) to

the incubation solution, for the 1-h incubation period

described above. NO and H2O2 release were determined

in the medium. Additionally, basal release of NO and

H2O2 was determined by omitting PMA from the

incubation medium of the macrophages from these

non-stimulated rats.

Macrophage Phagocytosis

Phagocytosis was addressed in non-stimulated

macrophages, by adding 1 � 107 particles of opsonized

zymozan to the incubation solution. One hour later, the

percentage of cells presenting at least three zymozan

particles was determined in a counting chamber [26].

Statistical Analysis

Data are expressed as means T S.E.M. Statistical

analysis was performed by KruskallYWallis ANOVA

followed by Dunn test, for the comparisons among the

three diet groups (control, soybean, and fish). The

MannYWhitney test was used for comparisons inside

the same group. Significance level was set at p < 0.05.

RESULTS

Food intake of the fish and soybean groups was

lower than that of the control-diet group (soybean:

25.6% and fish: 29.7%) throughout the seven weeks of

the study. The energy intake was also lower although

not significantly and body weight did not significantly

differ among the groups (data not shown).

Macrophage functionality, as evaluated by both the

phagocytosis of opsonised zymozan particles and the

basal H2O2 production, was not significantly affected by

the consumption of either the soybean or the fish diet. On

the other hand, H2O2 produced by macrophages stimu-

Fig. 3. Paw volume, 3 h after subplantar carrageenan stimulus in

control ( ), soybean ( ) and fish ( ) groups, submitted to

coaxial perfusion. Each column represents the mean T s.e.m. from 8 to

9 animals. (a) Different ( p < 0.05) from control group.

Table 2. Concentration of NO (mM/100 ml) and BK (nmol/ml) on the Paw Edema Perfusate, Collected from First to Second and from Second to

Third Hour, after Subplantar Carrageenan Stimulus, in Rats from Control, Soybean and Fish Groups

NO BK

First to second hour Second to third hour First to second hour Second to third hour

Control 2.25 T 0.68 0.94 T 0.34 58.80 T 3.76 65.93 T 6.77

Soybean 0.77 T 0.35a 0.21 T 0.11 24.14 T 2.87a 21.07 T 7.56a

Fish 0.84 T 0.22 0.09 T 0.05ab 30.68 T 3.38a 29.09 T 6.60a

Data are shown as mean T s.e.m. The number of rats in each group varied between 6 and 11. a Different (p < 0.05) from respective control group andb from respective first to second hour period.

84 Wohlers, Xavier, Oyama, Ribeiro, Nascimento, Casarini, and Silveira

lated either in vitro with PMA or in vivo by the

peritoneal injection of carrageenan, was significantly

decreased in both the lipid groups ( p = 0.0005 and

0.0002, respectively). In fact, after either in vitro or invivo stimulation, only macrophages from the control

group showed a significant increase in the H2O2 produc-

tion, in relation to the respective basal level ( p = 0.02

and p = 0.0005, respectively). The production of NO was

not affected by diet (basal) or in vitro stimulation with

PMA. The in vivo carrageenan stimulus induced incre-

ment in the NO production by peritoneal macrophages

from the control and fish groups ( p = 0.01 for both

groups) (Fig. 1).

Figure 2 shows the levels of NO in the peritoneal

exudate after in vivo stimulation with carrageenan. It

can be seen that NO concentration was lower in both

the soybean and the fish group than in the control group

( p = 0.003).

The paw edema measured 3 h after the subplantar

injection of carrageenan was lower in both soybean and

fish groups than in the control one ( p = 0.0003) (Fig. 3).

NO levels in the paw edema perfusate was also

decreased in the lipid groups in comparison to the

control group. In the soybean group, the decrease was

significant at the first to second-hour period ( p = 0.04)

and in the fish group at the second to third-hour period

( p = 0.007) after carrageenan administration. BK levels

were lower in the perfusate of both hyperlipidic groups

then that of the control rats, in both 1-h collection

periods ( p = 0.0001 and p = 0.001 at the first and

second period, respectively) (Table 2).

The amidolitic activity of plasma KK was also

diminished in the hyperlipidic groups, 1 h ( p = 0.00003)

and 2 h ( p = 0.001) after the carrageenan stimulus

(Fig. 4).

Plasma corticosterone levels did not differ among

the groups during edema development. However, the

basal levels were higher in the lipid groups (Table 3).

The lipid groups showed hyperleptinemia 1 h ( p =

0.003) and 3 h ( p = 0.0004) after the carrageenan

stimulus when compared to the control group. At the

third hour, the leptin levels were higher than at the first

Table 3. Corticosterone Plasma Concentration (mg/100 ml) from 1 to 4 h after Subplantar Carrageenan Stimulus in Rats from Control, Soybean and

Fish Groups

Basal* 1 h* 2 h 3 h 4 h*

Control 11.93 T 0.67 30.72 T 0.75b 31.12 T 1.66b 27.44 T 1.47b 29.16 T 1.46b

Soybean 21.32 T 0.72a 31.35 T 1.06b 29.68 T 1.64b 26.93 T 1.87b 33.74 T 1.49b

Fish 19.45 T 0.53a 32.98 T 0.75b 33.47 T 0.60b 30.88 T 0.89b 29.56 T 2.07b

Data are shown as mean T s.e.m. a Significantly different ( p < 0.05) from respective control group and b from respective basal group. The number of rats

in each group varied between 9 and 15. * Ours previous data from [27], to evaluate the corticosterone release during the carrageenan edema development.

Fig. 4. Amidolitic activity of plasma kalikrein (KK), 1 and 2 hours after

subplantar carrageenan stimulus in control ( ), soybean ( ) and fish

( ) groups. Each column represents the mean T s.e.m from 10 to 12

animals. (a) Different ( p < 0.05) from control group.

85Fish/Soybean Oil-Rich Diets Effect on BK, KK, NO, Leptin, Corticosterone and Macrophages in Rats

hour, in the three groups (control: p = 0.0002; soybean:

p = 0.00002; fish: p = 0.0001) (Table 4).

DISCUSSION

In the present work, we reproduced previous data

from ours and other laboratories, showing that the diets

enriched with either soybean or fish oil decreased food

intake [27Y29]. The reported increased stimulation of

cholecystokinin (CCK) secretion by high-lipid diets may

have contributed to this effect, since CCK is a hormone

involved in the satiety process [30, 31]. The energy

intake, although did not statistically differ among the

groups, tended towards reduction in both the soybean

(Y14%) and the fish group (Y17%) throughout the seven

weeks of the study. Body weight gain was similar among

the groups. This finding has been reported previously

indicating a higher food efficiency for the lipid groups

due the lower energetic cost of lipid deposition from

dietary fat than from dietary carbohydrate [28].

No significant diet effect was evident on macro-

phage phagocytosis, as evaluated 1 h after the addition

of zymozan to the isolated macrophages. However, a

trend towards reduction was seen in both the soybean

(Y17%) and the fish group (Y35%). A decrease in

macrophage phagocytosis has been reported after the

ingestion of nY6 PUFA-rich diet in comparison to the

ingestion of saturated fatty acids-rich diet [32]. In other

reports, however, the consumption of fish oil failed to

change macrophage phagocytic function [15].

The H2O2 production, stimulated by either carra-

geenan or PMA, was decreased in the macrophages

from both hyperlipidic groups, when compared to the

control group. These results are similar to other [33] in

neutrophils from rats fed a soybean-rich diet. H2O2 is an

important citotoxic agent [34] and its lower release

could contribute to the anti-inflammatory effect of the

hyperlipidic diets, as observed previously [12, 27] and

reproduced in the present study.

Some reports have shown decreased activity of

superoxide-dismutase (SOD), the enzyme catalyzing

H2O2 formation, in plasma and in different tissues of

animals treated with nY3 or nY6 PUFA-rich diets [35, 36].

It is possible that a lower SOD activity could be

associated to the low H2O2 production observed in our

study. However, there are controversial information about

the influence of lipid diets on H2O2 production and

antioxidant enzymes activity. Some studies failed to

observe changes on SOD activity in cells obtained from

animals fed with nY3 or nY6 PUFA-rich diets [33, 37]

while others found increase in H2O2 production by

macrophages from rats treated with nY3 PUFA-rich diet

[38]. These different observations could be attributed to

different cell types and quantities of different fatty acids

on the high-lipid diets used. Additionally, the reduction of

H2O2 production, observed in the present study, could

also be attributed to the high basal levels of corticoste-

rone, found in the lipid groups [12], since this hormone

can reduce H2O2 production and activity of antioxidant

enzymes, such as SOD, in macrophages in culture [39].

NO release was decreased in both the peritoneal

exudate and edema perfusate of hyperlipidic groups

stimulated in vivo with carrageenan. NO plays a role in

the carrageenan edema formation [40], promoting vaso-

dilatation and vasopermeability [41]. Thus, its lower

release in the lipid groups likely contributed to the anti-

inflammatory effect of these diets. The observed low BK

levels in the edema perfusate may have influenced NO

release, since it has been demonstrated that BK promotes

NO synthetase (NOs) activation [40]. Additionally, some

studies have shown that both nY3 PUFA [42] and nY6

PUFA-rich diets reduced NOs enzyme [43]. Thus, a low

NOs activity may also have been present in our animals.

This is a possibility to be investigated, since the lipid

groups showed, besides reduced BK release, increased

levels of corticosterone [27], which has been shown to

inhibit NOs [44].

The in vivo results were not equally reproduced in

vitro procedure. The NO production by peritoneal

macrophages from fish group, stimulated in vivo with

carrageenan, was not decreased. This disagreement

could be occurred due the absence Bin vitro^ from

plasma factors, whose interaction must be important for

the in vivo responses. After the PMA stimulus, no

differences were observed in NO production by macro-

phages from all groups, in relation to their respective

baseline. It is possible that the concentration of PMA

and/or the 1-h incubation period were not sufficient to

NO stimulation.

Table 4. Serum Leptin (mg/dl), 1 and 3 h after Subplantar Carra-

geenan Stimulus in Rats from Control, Soybean and Fish Groups

1 h 3 h

Control 1.97 T 0.23 7.82 T 1.34 b

Soybean 5.13 T 0.69 a 15.43 T 1.36 a b

Fish 7.23 T 1.19 a 17.54 T 1.14 a b

Data are shown as mean T s.e.m. a Significantly different (p < 0.05)

from respective control group and b from respective 1 h group. The

number of rats in each group varied between 9 and 14.

86 Wohlers, Xavier, Oyama, Ribeiro, Nascimento, Casarini, and Silveira

BK is an important mediator of the carrageenan

edema [13] that induces hyperalgesia, vasodilatation,

and vasopermeability. We found BK levels to be

diminished in the edema perfusate from the lipid

groups, what is probably related to the reduced kalikrein

levels we found in these animals. At the best of our

knowledge, this is the first demonstration of a causal

association between BK release and the anti-inflamma-

tory effect of fish or soybean oil-rich diets. Since BK is

a product of cininogen cleavage by KK and corticoste-

rone has been shown to decrease cininogen production

[45] the lower BK release could be related to a

reduction on cininogen levels.

It is important to note that other anti-inflammatory

mediators, such as docosatrienes, protectins, and resol-

vins may also be involved, as their production was

reportedly induced by DHA and AA [5]. Additionally,

in view of the reported stimulation of PGE2 release by

BK [46, 47], it is possible that the anti-inflammation

observed in the lipid diets may have relied, at least in

part, on a reduced production of this pro-inflammatory

prostaglandin [48]. Such an effect would be particularly

relevant in the nY6 group, since this diet has been

demonstrated to induce high PGE2 levels [49].

Leptin, a hormone with a role in appetite and energy

expenditure regulation [50], has also been involved in

the inflammatory process, being its production in-

creased, as part of the acute phase response [16, 51, 52].

Leptin levels were higher in the lipid groups, when

compared to control group, probably due to the higher

adiposity induced by the long-term consumption of these

hyperlipidic diets [53]. Additionally, higher corticosterone

basal levels in the lipid groups may have contributed to

the increased leptin, since leptin synthesis has been shown

to be dependent on the presence of glucocorticoids [16].

Increased leptin gene expression has been shown to

follow the intake of hyperlipidic diets, regardless the diet

was rich in nY6 PUFA or saturated fatty acids [54].

Contrarily, a decrease in leptin plasma levels and gene

expression has been reported in rats fed nY3 PUFA, when

compared to rats fed a saturated fatty acids-rich diet [55].

Leptin has been shown to exert a protective effect in

models of gastric injury [56] and to inhibit neutrophils

infiltration and migration in experimental colitis [52, 57].

In contrast, it was suggested a pro-inflammatory role of

leptin in the colitis model [58]. Additionally, leptin-

deficiency has been associated with low in vivo expression

of pro-inflammatory citokines in mice, and leptin addition

to their cultured macrophages increased the expression of

these citokines [59].

In the present study, the higher levels of leptin in the

lipid groups, 1 and 3 h after carrageenan, could be taken as

indicating an anti-inflammatory role of the hormone, since

these groups showed a lower edema in response to

carrageenan. However, the percentage increase of plasma

leptin in the same interval was of 297% in the control

group, while it was of 201 and 143% in the nY6 and nY3

groups, respectively. It is thus possible that the lower

increments, rather than the higher absolute levels, were

relevant, and this assumption would lead to the conclu-

sion of leptin as having a pro-inflammatory action.

Diet enrichment with nY6 PUFA has been associ-

ated with increased levels of inflammatory markers

while dietary supplementation with nY3 PUFA has been

associated with anti-inflammatory effects [60]. The

production of different eicosanoids derived from nY3 or

nY6 PUFA is one of the mechanisms hypothesized to

explain these differences. Indeed, a recent workshop on

the essentiality of these fatty acids advised an increase

of nY3 and a reduction of nY6 PUFA in the diet, to

minimize the adverse effects of AA and its metabolites

[60]. However, there is no clear evidence of the nY6

PUFA prejudicial effects [61]. In contrast, many studies

have been demonstrated beneficial effects of these fatty

acids [62, 63]. It has recently been shown that the

excess of nY6 PUFA failed to antagonize nY3 PUFA

effects and that the association of these two types of

fatty acids lowered the concentration of inflammatory

cytokines, in chronic inflammatory disease [64].

Our previous [12] and present results showed that

nY6 and nY3 PUFA-rich diets had a similar anti-

inflammatory effect and we were presently able to

correlate this effect with similar changes of inflammatory

mediators. The doubt remains as on the real beneficial

effects of reducing the acute inflammation, since the

inflammatory process is a physiological response to

tissue injury, aimed at reestablishing homeostasis.

ACKNOWLEDGMENTS

The authors would like to thank to FAPESP, CAPES

and CNPq for the financial support.

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89Fish/Soybean Oil-Rich Diets Effect on BK, KK, NO, Leptin, Corticosterone and Macrophages in Rats