DOHaD 07 20 OA 1398 - Proof - Hi

Journal of Developmental Origins of Health and Disease
Maternal consumption of ɷ3 attenuates metabolic disruption

elicited by saturated fatty acids enriched-diet in offspring
rats
Journal: Journal of Developmental Origins of Health and Disease
Manuscript ID DOHaD-07-20-OA-1398
Manuscript Type: Original Article
Date Submitted by the

18-Jul-2020
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Author:
Complete List of Authors: Alves, Debora; Universidade Federal de Pernambuco, Department of

Nutrition
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Bloise, Aline; Universidade Federal de Pernambuco, Department of

Nutrition
Mata, Laura; Universidade Federal de Pernambuco, Department of
Nutrition
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Rocha-Junior, Reginaldo; Universidade Federal de Pernambuco,

Academic Center of Vitória
Lima-Júnior, Nelson; Universidade Federal de Pernambuco, Academic
Center of Vitória
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Menezes, Luiza; Universidade Federal de Pernambuco, Academic Center

of Vitória
Silva, Elionay; Universidade Federal de Pernambuco, Center Academic of
Vitória
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de-Oliveira, Yohanna; Universidade Federal da Paraiba, Department of

Nutrition
Wanderley, Almir; Universidade Federal de Pernambuco, Department of
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Physiology and Pharmacology

Alves, José Luiz ; Universidade Federal da Paraíba, Department of
Nutrition
Nogueira, Viviane; Federal University of Pernambuco, Department of
Physical Education and Sports Science
Silva, Joao; Federal University of Pernambuco, Academic Center of
Vitória
Model or Method: Animal, Small animals < Animal
Developmental Stage, Child growth and health < Developmental Stage,

Topic: Body composition < Outcome/System, Metabolic syndrome <
Outcome/System
High fat diet (HFD) intake during gestation and lactation has been
associated with an increased risk of developing cardiometabolic disorders
in adult offspring. We investigated whether metabolic alterations
resulting from the consumption of HFD are prevented by addition of
Abstract: omega-3 (ɷ3) in the diet. Wistar rat dams were fed with Control (C:
19% of lipids and ɷ6:ɷ3=12), HF (HF: 33% lipids and ɷ6:ɷ3=21) or HF
enriched with omega-3 (HFω3: 33% lipids and ɷ6:ɷ3=9) diet during
gestation and lactation, and their offspring food consumption,
Cambridge University Press

Page 1 of 29 Journal of Developmental Origins of Health and Disease
murinometric measurements, serum levels of metabolic markers, insulin

and pyruvate sensibility tests were evaluated. The maternal HFD induced
increased body weight at birth, dyslipidemia and elevated fasting glucose
levels in the HF group. The enrichment of omega-3 in the maternal HFD
led to lower birth weight and improved lipid, glycemic and transaminase
biochemical profile of the HFω3 group until the beginning of adulthood.
However, at later adulthood of the offspring, there was no improvement
in these biochemical parameters. Our findings confirm the hypothesis
that the maternal consumption of HFD enriched with ɷ3 is able to
attenuate or prevent metabolic disruption elicited by HFD in offspring
until 90 days old, but not in the long term, as observed at 300 days old
of the offspring.
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Journal of Developmental Origins of Health and Disease Page 2 of 29
1 Maternal consumption of ɷ3 attenuates metabolic disruption elicited by saturated fatty

2 acids enriched-diet in offspring rats
3 Alves, DS1, Bloise, AMNLG1, Silva, LML1, Rocha-Junior, RL1, Lima-Júnior, NC1, Menezes, LGS1, Silva,
4 EGS1, De Oliveira, Y2, Wanderley, AG3, de-Brito-Alves, JL2, Nogueira, VO1, Costa-Silva, JH1*
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6 1Laboratory of Nutrition, Physical Activity and Phenotypic Plasticity, Department of Physical Education
7 and Sport Sciences, Universidade Federal de Pernambuco, UFPE, Vitória de Santo Antão – PE, 55608-680,
8 Brazil.
9 2 Department of Nutrition, Universidade Federal da Paraíba, UFPB, João Pessoa – PB, 58051-900, Brazil.
10 3 Department of Physiology and Pharmacology, Universidade Federal de Pernambuco, UFPE, Recife
11 - PE, 50760-901, Brazil.

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13 *Corresponding author:
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14 João Henrique Costa-Silva

15 Núcleo de Educação Física e Ciências do Esporte – CAV - UFPE
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16 Rua Alta do Reservatório, S/N, Bela Vista, Vitória de Santo Antão, PE.
17 CEP: 55608-680
18 Phone/fax: 55 81 31144101
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19 Email: joao.hcsilva@ufpe.br
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Alves, DS et al. Maternal ω3 and metabolic parameters in offspring
22 Abstract
23
24 High fat diet (HFD) intake during gestation and lactation has been associated with an increased risk of
25 developing cardiometabolic disorders in adult offspring. We investigated whether metabolic alterations
26 resulting from the consumption of HFD are prevented by addition of omega-3 (ɷ3) in the diet. Wistar rat
27 dams were fed with Control (C: 19% of lipids and ɷ6:ɷ3=12), HF (HF: 33% lipids and ɷ6:ɷ3=21) or HF
28 enriched with omega-3 (HFω3: 33% lipids and ɷ6:ɷ3=9) diet during gestation and lactation, and their
29 offspring food consumption, murinometric measurements, serum levels of metabolic markers, insulin and
30 pyruvate sensibility tests were evaluated. The maternal HFD induced increased body weight at birth,
31 dyslipidemia and elevated fasting glucose levels in the HF group. The enrichment of omega-3 in the
32 maternal HFD led to lower birth weight and improved lipid, glycemic and transaminase biochemical profile
33 of the HFω3 group until the beginning of adulthood. However, at later adulthood of the offspring, there
34 was no improvement in these biochemical parameters. Our findings confirm the hypothesis that the
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35 maternal consumption of HFD enriched with ɷ3 is able to attenuate or prevent metabolic disruption elicited
36 by HFD in offspring until 90 days old, but not in the long term, as observed at 300 days old of the offspring.
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38 Keywords: Phenotypic plasticity, high fat diet, alpha-linolenic, transmissible chronic diseases.
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40 1. Introduction
41 The advent of nutritional transition, markedly characterized by increased consumption of high

42 fat and high sugar diet associated with sedentary lifestyle, has been linked to increased prevalence of
43 noncommunicable diseases such as type 2 diabetes mellitus, obesity, arterial hypertension in worldwide (1-
44 3). According to the World Health Organization, more than 1.9 billion adults had overweight and
45 approximately 650 million were obese in the year 2016. In addition, approximately 41 million children
46 under 5 years of age and 340 million children and adolescents (5 to 19 years old) had overweight or obesity,
47 respectively (4). It is found that a significant number of obese subjects are resistant to insulin and that
48 obesity is associated with low-grade systemic inflammation (5), with the immune system related in the
49 development of insulin resistance (6).
50 The excessive consumption of HFD rich in saturated fatty acid during pregnancy and/or
51 lactation has been described as an important risk factor for development of metabolic disorders, such as
52 insulin resistance, dyslipidemias and cardiorespiratory dysfunction in offspring later in life (7). These
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53 findings suggest that nutritional strategies during pregnancy and breastfeeding periods could be considered
54 as an important intervention window to prevent and reduce the risk of maternal lipid disturbances, thus
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55 protecting the offspring against the metabolic disorders later in life.

56 The consumption of diets enriched with omega-3 during pregnancy
57 and lactation was associated with reduced adiposity, decreased adipocyte size and decreased serum leptin
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58 levels in the offspring (8). In addition to lower adiposity, omega 3 intake was also related to improved
59 insulin sensitivity, which is shown to decrease with age in the offspring of male rats (9).
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60 Therefore, the aim of this study was to investigate whether the enrichment of omega-3 in
61 maternal HFD during gestation and lactation is able to attenuate the effects of HFD consumption over
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62 murinometric measurements, food consumption, and biochemical profile in the offspring rats.
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65 2. Method
66
67 2.1 Animals and experimental groups
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69 Virgins female albino rats (n=30) from the Wistar line (Rattus norvegicus) were obtained from
70 the experimental animal room of the Academic Center of Vitória (CAV), Federal University of
71 Pernambuco, Brazil. At 85 to 90 days old (-d-old) and body weight comprised between 220 and 260 g, the
72 rats were placed for mating in the ratio of 1:3 (male:female). The determination of pregnancy was
73 performed from the observation of the presence of spermatozoa in the vaginal smear, defining the first day
74 of pregnancy. From the 1st day of gestation until the 21st day of lactation, the progenitor rats were separated
75 and placed in individual cages and randomly allocated according to their diets: C (Control, 19% lipids;
76 n=3), HF (High in saturated fatty acids content, 33% lipids; n=3) and HFω3 (High in saturated fatty acids
77 content, 33% lipids and enriched with omega-3; n=3) with chow and water ad libitum. After weaning, the
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78 male offspring were feed with standard chow for rats (Presence®, Neovia Group, São Paulo, Brazil) and
79 water ad libitum and they were separated in three groups according to their mothers’ diets: C (n=35), HF
80 (n=27) and HFω3 (n=27) groups
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81 Temperature and humidity were maintained within the range of 22 to 24 ° C and 55 to 65%
82 respectively, with 12h light and dark cycle (lights on from 06:00 to 18:00h). Offspring from each dam were
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83 reduced to 8 male rats per litter. In cases where the litter was composed of less than 8 male rats, female rats
84 were used to standardize litter size until weaning.
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86 2.2 Nutritional manipulation
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87 The C group received a standard diet in accordance with AIN-93G (10) with 19% of the energy
88 coming from the fat, 20% of the proteins and 61% of the carbohydrates, with ω6: ω3 ratio=12.66; the HF
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group received a diet rich in lipids and saturated fatty acids, which 33% of the energy coming from the fats,
90 20% of the proteins and 47% of the carbohydrates, with 6:ω3 ratio=21.22; and the HFω3 group received a
91 high fat diet enriched with omega-3, with ω6:ω3 ratio=9.45. The standard diet contained around 14.6 kJ/g
92 and the high fat diets around 18.8 kJ/g. HFDs had a higher content of saturated fatty acids and the diet
93 enriched with omega-3 contained less omega-6 (Table 1). After weaning, at 21-d-old, the animals received
94 commercial standard diet for rats (Presence®, Neovia Group, São Paulo, Brazil) ad libitum.
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96 2.3 Food consumption of dams and offspring
97 The water and food intake were evaluated on alternated days at the beginning of the clear period
98 (at 8 o'clock in the morning), by the difference between the quantity offered and the remnant of the cage.
99 Food consumption was performed throughout the gestation and lactation period (11), and the pups were
100 followed since 22 until 300-d-old. From the evaluation of dietary intake and weight gain by means of body
101 mass measurement, the food efficiency coefficient (FEC) was calculated according to the equation: FEC =
102 weight gain in the period evaluated / food consumption in the same period. The number of calories
103 consumed as well as macronutrients was obtained from the centesimal composition of the diets used.
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105 2.4 Murinometric measurements of the offspring

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107 They were measured the body weight, naso-anal length and abdominal circumference in the 1st,
108 7th, 14th, 21st, 30th, 90th and 300th-d-old. After obtaining measurements, Lee index was calculated on the 90th
109 and 300th d-old from the relation between the cubic root of the body weight and the naso-anal length of the
110 animal (12).
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112 2.5 Biochemical analysis of dams and offspring
113 Dams at 19 days of pregnancy and offspring at 22 (subsequent to weaning), 30, 90, and 300-d-old,
114 were fasted overnight and then anesthetized with Ketamine (80mg/Kg i.p) and xylazine (10mg/Kg i.p.) for
115 collection of blood samples through the rupture of the retro orbital plexus. After coagulation, the blood was
116 centrifuged at 1372 g for 10 minutes to obtain the serum, which was transferred to an Eppendorf tube and
117 stored at -20 ° until the accomplishment of the biochemical analyzes by means of the equipment Automatic
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118 Chemistry Analyzer (Pioway Medical Lab Equipment Co., Ltd.). In the dams and offspring were analyzed:
119 total proteins (TP), albumin (ALB), glycemia (GLY), triglycerides (TRI), total cholesterol (TC), alanine
120 aminotransferase (ALT), aspartate aminotransferase (AST), high-density lipoprotein cholesterol (HDL-c).
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121 Levels of low-density lipoprotein cholesterol (LDL-c) and very low-density lipoprotein cholesterol (VLDL-
122 c) were obtained by Friedwald calculations. (13).
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124 2.6 Glucose tolerance test (GTT), pyruvate tolerance test (PTT) and insulin sensitivity test (IST) of
125 the offspring
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127 At 90 and 300-d-old, the GTT was performed in rats after a 6-hour fast. For GTT, glucose load
128 20% glucose (2mg / g by weight) was administered by oral gavage. Blood samples were taken from the tail
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129 vein before glucose administration (in duplicates) and, subsequently at T15, T30, T60 and T120 minutes
130 (14).
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131 At 90 and 300-d-old, the PTT was performed after overnight, the animals were separated into
132 individual cages. Glucose measurement at time T0 was performed in duplicate, and then the sodium
133 pyruvate to 50% (2g/kg of weight ) was injected intraperitoneally, after this the blood glucose was measured
134 (in duplicate) at T15, T30, T45, T60 and T120 minutes after administration sodium pyruvate (Sigma-
135 Aldrich Brasil Ltda, P2256) (14).
136 At 90 and 300-d-old, the IST was performed after a 6-hour fast, the animals were separated into
137 individual cages. Glucose measurement at time T0 was performed in duplicate, and then the insulin (1 mU/g
138 of weight, i.v.) after this the blood glucose was measured (in duplicate) at T15, T30, T45, T60 and T120
139 minutes after administration of regular human insulin (HUMULIN R, 100UI/ml) (14).
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141 2.7 Statistical analysis
142 The results were expressed as mean ± epm. The analysis of sample normality was performed
143 using the Kolmogorov-Sminorv test. The comparison among the groups was performed using the one-way
144 ANOVA and two-way ANOVA tests (for GTT and IST), followed by the Bonferroni post-test. The
145 trapezoidal rule was used to determine the area under the curve (AUC). The data were analyzed in the

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146 Graph Pad Prism program (GraphPad Software Corporation, version 5.0, 2007). The level of significance
147 considered was p <0.05.
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149
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150 3. Results
151 3.1 Food consumption evaluations of dams (gestation and lactation) and offspring (22 to 300-d-old)
152 The food intake during pregnancy was similar among the groups (Table 3), but the consumption
153 of omega-3 was higher in the HFω3 group (p=0.0007).
154 In assessing consumption during lactation (Table 4), we observed that in the first week there
155 was no significant difference among groups regarding consumption in grams, carbohydrates and proteins.
156 The rats that received HF and HFω3 had higher consumption of lipids (p=0.005), and HFω3 consumed
157 more omega-3 (p=0.0002). In the second week of lactation, dams submitted to the HF diet had higher
158 protein intake than the control group (p=0.03). However, the centesimal composition of the diets shows
159 that the amount of protein is higher in high-fat diets due to the addition of foods that are sources of vegetable
160 protein (Table 2). In this week, there was also increase in the consumption of lipids in the HF and HFω3
161 groups compared to the C group (p=0.0004). HFω3 group also consumed more omega-3 (p=0.0002). No
162 difference was observed over food intake in grams, calories and carbohydrates among the groups. In the
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163 third week, the rats that received HF and HWω3 had higher consumption of lipids (p=0.04), and HFω3
164 group consumed more omega-3 (p=0.02). There was no difference in consumption in grams, calories and
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165 others nutrients among groups.

166 Food consumption also was verified in the offspring at 22 to 30, 30 to 60, 60 to 90, and 90 to
167 300-d-old, obtaining the FEC in the periods evaluated (Table 5). In the period from 22 to 30 days, the
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168 animals of the HF group had lower intake of grams, calories and macronutrients when compared to the
169 control group. There was no difference among the groups in the FEC value. From 30 to 60 days, no
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170 difference in consumption was observed among the groups, however the HFω3 offspring presented FEC
171 values higher than the C group (p<0.05), with no difference in HF offspring. At age 60 to 90 and 90 to 300
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172 days, no differences were found in the consumption and FEC values among the groups.
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3.2 Evaluation of murinometric measurements of the offspring
175 The HF group had higher body weight at 1-d-old when compared to the C and HFω3 (Table 6).
176 At the 7-d-old (p<0.0001) and 14-d-old (p<0.0001) the HF and HFω3 groups presented higher body weight.
177 At 21-d-old (p=0.002) the HFω3 group showed higher body weight when compared to the other groups. At
178 30, 90 and 300-d-old no difference was found among the groups.
179 Both HF and HFω3 groups presented increased abdominal circumference at the 1-d-old
180 (p=0.002) when compared to the C. At 7-d-old (p=0.005) and 14-d-old (p=0.0009) only the HF group
181 showed increased circumference. At 21-d-old no difference was found among the groups. However, at 30-
182 d-old (p=0.004) the HF group showed increased abdominal circumference when compared to the other
183 groups. At 90 and 300-d-old no difference was found among the groups.
184 The HF group showed increased naso-anal length in relation to the C and HFω3 at 1-d-old
185 (p=0.004) and 7-d-old (p<0.0001). At 14-d-old no difference was found among the groups. At 21-d-old
186 (p<0.0001) both HF and HFω3 were higher in relation to C group. However, the HF group was larger at
187 30-d-old (p<0.0001) when compared to the other groups. At 90 and 300-d-old the groups showed similar
188 body growth, and no difference was found among the groups.
189 As for Lee's index at 90 and 300-d-old no difference was found among the groups.

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191 3.3 Biochemical assessment in dams at 19 days of pregnancy
192 Serum levels of TP, ALB, GLY, TRI, CT, VLDL-c, ALT and AST were similar among group at
193 19 days of gestation (Table 7). However, dams of HF group displayed increased serum levels of TC when
194 compared with C and HFω3 dams (Table 7).
195 3.4 Biochemical assessment in male offspring
196 At 22-d-old (Table 8), no differences were found in serum levels of ALB, TP, HDL-c, LDL-c and
197 AST between the groups. In relation to GLY, only the HF group presented higher levels (p=0.02) when
198 compared to the C group. The HF group also had higher serum levels of TC (p=0.003), ALT (p=0.001) and
199 lower levels relative to the ratio AST/ALT (p=0.004) when compared to the other groups. Already the
200 HFω3 group presented lower levels of TRI (p=0.02) and VLDL-c (p=0.01) when compared to the C group.
201 At 30-d-old (Table 8), differences were found only in the serum levels of LDL-c, with the HF
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202 group showing higher levels (p=0.001) when compared to the C group.
203 At 90-d-old (Table 8), no differences were found in serum levels of ALB, TP, HDL-c, LDL-c
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204 and ALT between the groups. In relation to GLY, the HF group presented higher levels (p˂0.0001) when
205 compared to the other groups. The HF group also presented higher serum levels of TC (p=0.01), TRI
206 (p=0.02) and VLDL-c (p=0.02) when compared to the C group. Already the HFω3 group presented lower
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207 levels of AST (p=0.0007) and AST/ALT ratio (p=0.0002) when compared to the C and HF groups.
208 At 300-d-old (Table 8), no differences were found in serum levels of GLY, TP, HDL-c, LDL-c
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209 and ALT among the groups. In relation to ALB, the HF group presented higher levels (p=0.01) when
210 compared to the C group. Both HF and HFω3 groups presented higher serum levels of TC (p=0.008), TRI
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211 (p=0.0002) and VLDL-c (p=0.002) when compared to the C group. Already both HF and HFω3 groups
212 presented lower levels of AST (p=0.0001) and AST/ALT ratio (p=0.0001) when compared to the C group.
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214 3.5 GTT, PTT and IST of the offspring at 90 and 300-d-old
215 In the GTT at 90-d-old (Fig 1a) and at 300-d-old of life (Fig 1c), no difference was found in
216 glycemia levels among the groups at T0, prior to glucose administration and in the T15, T30, T60 and T120
217 minutes after glucose administration. At 300-d-old of life (Fig 1b), also no difference was found in glycemia
218 levels among the groups at T0, prior to glucose administration and in the T15, T30, T60 and T120 minutes
219 after glucose administration. In the AUC for glucose at 90-d-old (Fig 1b) and at 300-d-old of life (Fig 1d),
220 no difference was found also among the groups.
221 In the PTT at 90-d-old (Fig 2a) and at 300-d-old (Fig. 2c), no difference in glycemia was found
222 among the groups at T0, prior to pyruvate administration and in the T15, T30, T45, T60 and T120 minutes
223 after administration of pyruvate. In the AUC for glucose at 90-d-old (Fig 2b) and at 300-d-old of life (Fig
224 2d), no difference was found also among the groups.
225 Regarding the IST at 90-d-old (Fig 3a) and at 300-d-old (Fig 3c), no difference in glycemia was
226 found among the groups at T0, before administration of insulin and in the T15, T30, T45, T60 and T120
227 minutes after insulin administration. In the AUC for glucose at 90-d-old (Fig 3b) and at 300-d-old of life
228 (Fig 3d), no difference was found also among the groups.

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229 4. Discussion
230
231 Daily maternal consumption in grams did not differ among the groups in our survey, different
232 from that found by Ferro Cavalcante et al. (2013) (15) and Carvalho et al. (2013) (16). Although we did
233 not find this difference, the dams' caloric intake did not distinguish among the groups similarly to that found
234 by these authors and Khan et al. (2005) (17). Animals often show reduction in feed intake when they receive
235 a diet with higher caloric density under the ad libitum regime, as they tend to consume a constant amount
236 of energy (18). The composition of high-fat diets has lower carbohydrate content and higher lipid content
237 compared to AIN-93G, which contributed to the reduction in carbohydrate consumption in dams submitted
238 to the high-fat diet supplemented with omega 3 and higher consumption of lipids among two groups in
239 relation to the control group, according to Ferro Cavalcante et al (2013) (28) who also showed a higher
240 consumption of lipids in the last week of pregnancy to the detriment of less consumption of carbohydrates
241 in pregnant women submitted to a Westernized diet.
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242 In assessing the food consumption during pregnancy, we were able to analyze only the third
243 gestational week. Although we have not demonstrated consumption during the first and second gestational
244 weeks, it is in the last third of gestation that the mobilization of essential fatty acid stores in the maternal
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245 adipose tissue increases to the fetus, depositing in the retina and cerebral cortex and greater speed in the
246 growth rate (19, 20). There are few studies that assess the development of mothers and few reports maternal
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247 food consumption during the perinatal period.

248 We found that dietary intake of the animals that received HFD was lower from 22 to 30-d-old
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249 and, in the other periods, we did not observe a difference in dietary intake among the groups. Similar to
250 this, Pérez et al. (2015) evaluated the consumption of 45 to 60-d-old and did not find differences between
251 the animals that received HFDs during gestation, lactation and after weaning. However, Desai et al. (2014)
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252 observed higher intakes up to 18 weeks of age. Animals submitted to the HFD may present a change in the
253 signaling of satiety pathways with higher caloric intake through independent feeding even after the
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254 ingestion of breast milk (21). The HFω3 offspring presented higher FEC values between 30 and 60-d-old
255 in relation to the animals submitted to the control diet. It is known that up to 42-d-old, considered in animals
256 as early adolescence, there may be changes in body composition due to the nutritional environment, with
257 repercussions on long-term health (22). From the 60-d-old, no differences were observed between the
258 groups. Correia-Santos et al. (2014) (23) also evaluated the nutritional manipulation with HFD and HFD-
259 omega-3-rich from the linseed oil and did not verify differences as to the coefficient of feed efficiency of
260 weaning at 180-d-old.
261 In agreement with an early study, male offspring of dams fed with a HF diet during pregnancy
262 and lactation had increased body weight at 1, 7 and 14-d-old when compared to the C group (24). Although
263 male offspring of HFω3 group have had lower body weight at 1st-d-old in comparison to C and HF group,
264 on the 7th and 14th-d-old, HFω3 also displayed higher body weight when compared to the C group. In
265 addition, HFω3 group had higher body weight when compared to the other groups at 30-d-old. Taken
266 together, the findings suggest that increased maternal omega-3 consumption during pregnancy was not
267 effective to reduce body in early life of the offspring of dams feed with HF diet. This may be due the
268 overwhelming effect of the high satured fat acids present in the high fat diets, which it was reported to be

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269 associated with increased body mass in the offspring (7). Lastly, no differences in body weight were
270 observed among the groups at 30, 90 and 300-d-old. According to a systematic review carried with animal
271 studies, it is not clear whether an increased ômega-3 supply during the perinatal period influences body fat
272 mass in the offspring yet (25).
273 At early life (1 to 14-d-old), the HF group showed larger circumference than the C. It has been
274 proposed that maternal high fat diet consumption leads to increased visceral fat in offspring (26). At 21-d-
275 old, no difference was found between the groups and at the 30th-d-old, the HF group again presented larger
276 circumference when compared to the C. In all ages evaluated, the HFω3 showed similar circumference of
277 the control. It has also been shown in animals that maternal supplementation of DHA during pregnancy and
278 lactation reduces visceral and subcutaneous adipose tissue, which is associated with the risk for the
279 development of obesity (27).
280 Regarding the lipid biochemical profile evaluated, dams who consumed a diet rich in saturated
281 fatty acids had higher TC levels when compared to the C and HFW3 groups. The offspring from HF dams,
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282 at 22-d-old, also presented higher TC levels when compared to the other groups. Regarding the TRI levels,
283 the HFW3 offspring showed a reduction when compared to the C group. At 30-d-old, increase in LDL-c
284 levels was found in the group HF when compared with control group. At 90-d-old of age, offspring HF also
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285 had higher CT levels than the control group and higher levels of TRI and VLDL-c were found in these
286 animals in relation to the control, without difference when compared to the control and HFW3 groups. Our
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287 data corroborate with other studies that used HFD during pregnancy and lactation and found metabolic
288 changes such as hypertriglyceridemia, increased levels of TC, and LDL-c (28).
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289 Studies have demonstrated that omega-3 can improve lipid profile by lowering TRI levels, TC
290 and LDL-c fraction (25). Through its binding to PPARs (peroxisome proliferator-activated receptor),
291 omega-3 can regulate lipid metabolism, acting on the modulation of the expression of genes related to fatty
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292 acid oxidation. PPARs has action in regulate the expression of critical genes in lipid metabolism, fatty acid
293 oxidation, adipocyte development and lipoprotein metabolism. PPAR expression through omega-3 is
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294 accompanied by suppression of SREBP (sterol regulatory element-binding proteins), inhibiting lipogenesis
295 with reduced expression of fatty acid synthase (FAS) and acetyl CoA carboxylase (ACC), and consequently
296 promoting a decrease in serum levels of fatty acids, triacylglycerols and VLDL-c (29). It also reduces the
297 expression of enzymes involved in cholesterol synthesis and hepatic triglyceride synthesis such as
298 phosphatidic acid phosphatase and diacylglycerol acyltransferase and increased expression of lipoprotein
299 lipase to capture circulating lipoprotein triglycerides: VLDL-c and chylomicrons, can justify its lipid-
300 lowering action (30). These studies may justify the best lipid parameters observed in the HFW3 group.
301 The animals submitted to the diet rich in saturated fatty acids presented high levels of ALT,
302 whereas animals that received a high-saturated fatty acids diet enriched with omega-3 presented better
303 levels of transaminases at 22-d-old. Already at 90-d-old, the HFW3 group presented lower levels of AST
304 than the control group and HF. The consumption of diet rich in saturated fats increases the expression of
305 genes that signal adipogenic pathways in the liver (31), besides inducing and / or aggravating liver injury
306 with subsequent increase in plasma levels of transaminases (32, 33). In contrast, omega-3 ingestion reduces
307 levels of transaminases, mainly ALT in individuals with non-alcoholic fatty liver disease (34, 35).

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308 However, in our study at 300-d-old, no improvement was found in the lipid profile and in the
309 levels of liver transaminases, when compared to the HF group. A dysfunction of the adipose tissue is
310 verified with aging. The mechanisms are still not clear, but the inflammation has an important role. Low-
311 grade systemic inflammation is commonly seen in aging, with an increase in pro-inflammatory cytokines,
312 such as TNF-alpha, capable of leading to suppression of adipocyte differentiation (36, 37). A decrease is
313 found in the activity peroxisome proliferator activated-receptor γ (PPAR-γ) that is responsible for the
314 processes of differentiation and maturation of preadipocyte into an adipocyte (38, 39) and increase of
315 adipogenic suppressors (40, 41). In addition to the increase in ectopic adiposity, the impaired adipose tissue
316 function can cause an increase in systemic free fatty acids (FFA).
317 Saturated fatty acids can be considered the most harmful of FFA, being able to induce insulin
318 resistance and the development of atherosclerosis through lipid mediators such as ceramides and
319 diacylglycerols, thus causing an increased risk of metabolic diseases (42). These studies may justify the
320 overlap of the effects of SFA about enrichment of omega-3 in the lipid biochemical profile of the HFW3
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321 group at the age of 300 days, suggesting that the enrichment of omega-3 was able to attenuate the effects
322 of SFA up to 90 days , not being able to attenuate in the long term, with aging, as seen at 300-d-old of these
323 animals.
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324 Regarding fasting blood glucose levels, only the HF group presented higher values at 22-d-old
325 when compared to the control. At 90-d-old, serum fasting glucose levels were higher in the HF group when
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326 compared to the other groups. In addition to its lipid-lowering role, omega-3 intake is associated with
327 increased insulin sensitivity, reduced with age, as observed in offspring of rats (43). Studies have found in
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328 the offspring of adult rats of dams who received a diet rich in omega-3 increase in the number of pancreatic
329 islets, without altering the volume of the pancreas (44).
330 The PTT was performed to evaluate the hepatic production of fasting glucose from
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331 gluconeogenesis, since pyruvate is a common metabolite of the major precursors used in gluconeogenesis
332 (lactate, amino acids and glycerol) (45). The PTT also predicts the degree of hepatic insulin sensitivity. In
iew
333 the analyzes of the GTT, PTT and IST, no differences were found among the groups at the times evaluated
334 and in the AUC for glucose. However, the progression of diabetes mellitus goes through intermediary
335 stages, from altered fasting glycemia and glucose intolerance, and these stages are due to insulin resistance
336 associated with pancreatic β-cell dysfunction. Although we found no difference in these tests, our study
337 verified a tendency to develop diabetes mellitus in the group submitted to a diet rich in saturated fatty acids.
338 Since fasting hyperglycemia at 22 and 90-d-old in these animals may provide evidence of beta cell
339 dysfunction (46) , it is possible that in a more advanced stage impaired of tolerance to glucose, pyruvate
340 and insulin sensitivity can be found in these animals. However, at 300-d-old of these animals, we found no
341 difference among the groups in these tests, or in the biochemical parameters of fasting blood glucose,
342 suggesting that the diet rich in SFA used in this study was not able to lead to the development of type II
343 diabetes mellitus until this age evaluated.
344 In conclusion, this study demonstrated that maternal consumption of a diet rich in saturated
345 acids is associated with impaired in some murinometric and biochemical parameters. In addition, our
346 findings confirm that enrichment with omega-3 is able to attenuate some changes in the lipid profile, that
347 are present in diseases cardiovascular and metabolic, as observed in the offspring that received a diet rich

12
348 in SFA and enriched with omega-3 during pregnancy and lactation, up to 90-d-old of the offspring, but not
349 in the long term, as demonstrated at 300-d-old. More studies are needed to evaluate the mechanisms related
350 to metabolic changes elicited by the consumption of maternal HFD, as well as the role of the omega-3
351 during the perinatal period. A complete understanding of these mechanisms and the role of omega-3 may
352 lead to the development of therapeutic strategies and government policies to improve public health.
353
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354 Acknowledgements
355 We thank Danilo Augusto Ferreira Fontes for the technical support.
356
357 Financial Support
358 We thank Fundação de Amparo à Ciência e Tecnologia de Pernambuco – PE, Brazil (grant nº
359 PRONEM 0797-4.05/14; IBPG-1478-4.05/16; IBPG-0313-4.05/19) and Conselho Nacional de
360 Desenvolvimento Científico e Tecnológico - Brazil (grant nº 311386/2019-9) for financial support.
361
362 Conflicts of interest
363 None.
364
365 Ethical Standards
366 The authors assert that all procedures contributing to this work comply with the ethical standards
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367 of the relevant national guides on the care and use of laboratory animals of Conselho Nacional de Controle
368 de Experimentação Animal (CONCEA, MCTI - Brazil) and has been approved by the institutional
369 committee of the Federal University of Pernambuco, Brazil (protocol 23076.049500/2016-37).
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592

1
1 Figure captions
2
3 Figure 1. Glucose tolerance test and area under the curve for glucose at 90 and 300 days of offspring
4 of males whose dams were submitted to control, HF and HFW3 diets during gestation and lactation.
5 The rats were submitted during gestation and lactation to control diet (C, 19% lipids) or high SFA (HF,
6 33% lipids) or high content lipid in SFA and enriched with omega-3 (HFW3, 33% lipids), according to the
7 experimental group. Glucose was measured at time 0 before glucose administration (after 6 hours of fasting)
8 at time 15, 30, 60 and 120 minutes after glucose administration (panels A and C). The area under the curve
9 (AUC) for glucose were calculated by using the trapezoidal rule (panels B and D). The black line or bar
10 represents the control group, the dashed line or black bar the HF group and the gray line or bar the HFW3
11 group. The values were expressed as mean ± SEM, after the two-way ANOVA test (panels A and C) or
12 one-way ANOVA test (panels B and D) followed by Bonferroni post-test. (* p˂0.05 vs. Control, # p˂0.05
13 vs. HF; n= 10-18).
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14
15 Figure 2. Pyruvate tolerance test and area under the curve for glucose at 90 and 300 days of offspring
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18
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33% lipids) or high content lipid in SFA and enriched with omega-3 (HFW3, 33% lipids), according to the
19 experimental group. Glucose was measured at time 0 before glucose administration (after 12 hours of
20 fasting) at time 15, 30, 45, 60 and 120 minutes after pyruvate administration (panels A and C). The area
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21 under the curve (AUC) for glucose were calculated by using the trapezoidal rule (panels B and D). The
22 black line or bar represents the control group, the dashed line or black bar the HF group and the gray line
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23 or bar the HFW3 group. The values were expressed as mean ± SEM, after the two-Way ANOVA test
24 (panels A and C) or one-way ANOVA test (panels B and D) followed by Bonferroni post-test. (* p˂0.05
iew
25 vs. Control, # p˂0.05 vs. HF; n= 10-18).

26
27 Figure 3. Insulin sensitivity test and area under the curve for glucose at 90 and 300 days of offspring
30 33% lipids) or high content lipid in SFA and enriched with omega-3 (HFW3, 33% lipids), according to the
31 experimental group. Glucose was measured at time 0 before glucose administration (after 6 hours of fasting)
32 at time 15, 30, 45, 60 and 120 minutes after insulin administration (panels A and C). The area under the
33 curve (AUC) for glucose were calculated by using the trapezoidal rule (panels B and D). The black line or
34 bar represents the control group, the dashed line or black bar the HF group and the gray line or bar the
35 HFW3 group. The values were expressed as mean ± SEM, after the two-Way ANOVA test (panels A and
36 C) or one-way ANOVA test (panels B and D) followed by Bonferroni post-test. (* p˂0.05 vs. Control, #
37 p˂0.05 vs. HF; n= 10-18).

2
38 Fig. 1
A)
B) C
GTT- 90 days 90 days
HF
250 HFW3
C
25000
GTT AUC(mg/dL x 120 min)

200 HF
Glucose (mg/dL)
HFW3
20000
150
100 15000
50 10000
0 5000
T0 T15 T30 T60 T120
Time (min) 0
39
C)
D)
GTT- 300 days C
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300 days HF
250 HFW3
C
200 HF GTT AUC(mg/dL x 120 min) 20000
Glucose (mg/dL)
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HFW3
150 15000
100
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10000
50
5000
0
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T0 T15 T30 T60 T120

Time (min) 0
40
41
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42
iew
43 Fig.2
44
A)
PTT- 90 days B)
200 C
C 90 days HF
HF HFW3
20000
PTT AUC(mg/dL x 120 min)
Glucose (mg/dL)
150
HFW3
15000
100
10000
50
5000
0
T0 T15 T30 T45 T60 T120
Time (min) 0
45

3
C)
PTT- 300 days D)
C
200
C 300 days HF
HFW3
HF 20000
PTT AUC(mg/dL x 120 min)

Glucose (mg/dL)
150
HFW3
15000
100
10000
50
5000
0
T0 T15 T30 T45 T60 T120
Time (min) 0
46
47 Fig.3
48
A)
Fo
IST- 90 days B)
C
150
C 90 days HF
HFW3
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HF 10000
Glucose (mg/dL)
IST AUC(mg/dL x 120 min)
HFW3
100
8000
ee
6000
50
4000
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0 2000
T0 T15 T30 T45 T60 T120
Time (min) 0
49
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50
C)
iew
IST-300 days
D)
C
150 300 days HF
C
15000 HFW3
IST AUC(mg/dL x 120 min)
HF
Glucose (mg/dL)
100 HFW3
10000
50
5000
0
T0 T15 T30 T45 T60 T120
Time (min) 0
51
52
53

1
1 Table 1. Composition regarding the ingredients of experimental diets

2
Ingredient / quantity AIN-93 G High Fat Diet Omega-3 high fat diet
per 100g of diet with flaxseed oil 3.5%
Corn starch 39 15 15
Dextrinized corn starch 13 - -
Wheat flour - 12 12
Corn flour biscuit - 7 7
Soy flour - 6 6
Lard - 2 2
Butter - 8 8
Casein (>85%) 20 20 20
Guar gum - 0.5 0.5
Sucrose 10 18 18
Linseed oil - - 3
Soybean oil 7 7 3
Fiber (cellulose) 5 0.3 0.3
Vitamin mix 1 0.7 0.7
Mineral mix 3 2 2
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DL-methionine 0.3 0.2 0.2

Choline bitartrate 0.2 0.2 0.2
BTH 0.001 0.01 0.01
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Monosodium glutamate - 0.2 0.2

Sodium Chloride - 0.3 0.3
Total (g) 100 100 100
kJ /100g 15 19 18
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% Total fat 18 33 32
% Proteins 20 19 18
% Carbohydrates 61 46 49
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% Omega-3 0.2 0.2 0.4

% Omega-6 3 5 3
3
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The AIN-93G diet was adapted as recommended by Reeves et al., 1993; The hyperlipid diet was adapted from the study of Ferro
4 Cavalcante et al., 2013; The nutritional composition of calories and caloric percentage of fats, proteins and carbohydrates was
5 determined from the centesimal analysis of the diets carried out in the Laboratory of Bromatology of the Academic Center of Vitória
6 of the Federal University of Pernambuco.
7
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8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33

2
Table 2. Centesimal composition of nutrients from formulated experimental diets
Nutrient (g/100g) AIN-93G Hyperlipidic Hyperlipidic enriched

with omega-3
Proteins 18 22 21
Lipids 6 17 16
Carbohydrates 68 53 56
Calories 3 4 4
34 The analysis of proximate composition was carried out at the Bromatology Laboratory of the Academic Center of Vitória at the Federal
35 University of Pernambuco, according to the AOAC (1995) methodology for determining moisture, proteins, lipids and ashes. The
36 amount of carbohydrate present in the sample was obtained by difference.
37
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38 Table 3. Daily and total intake of experimental diets during the third week of gestation by dams submitted
39 to Control (C), High Fat Diet (HF) and High Fat Diet enriched with Omega-3-rich (HFω3) diets during
40 gestation.
41
42
Group
Variables
C HF HFω3
M±SEM M±SEM M±SEM
g/day 14 ± 1 13 ± 1 10 ± 0.8
Kcal/day 52 ± 5 59 ± 4 46 ± 4
Carbohydrates/day 8 ± 0.8 7 ± 0.5 5 ± 0.4*
Protein/day 2 ± 0.2 3 ± 0.2 2 ± 0.1
Lipids/day 1 ± 0.09 2 ± 0.1* 1 ± 0.1*
Omega-3/day 0.02 ± 0.001 0.02 ± 0.001 0.04 ± 0.001*
g/kg 48 ± 6 45 ± 2 34 ± 2
Kcal/kg 180 ± 23 205 ± 10 156 ± 10
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43 The pregnant rats received a control diet received with 18% lipids, a high-fat diet (HF) with 33% lipids or a high-fat diet enriched
44 with omega 3 (HFω3) with 33% lipids with 3.5% flaxseed oil, according to the experimental group, during pregnancy. Values were
45 expressed as mean ± EPM. One-Way ANOVA and Bonferroni post-test: Control [n = 3], HF [n = 3], HFω3 [n = 3]), * p˂0.05 vs.
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46 Control; # p˂0.05 vs. HF.

47
48
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iew

4
49 Table 4. Food intake in grams, calories and macronutrients and coefficient of dietary efficiency of dams
50 subjected to the Control, High Fat Diet (HF) and High Fat Diet enriched with Omega-3-rich (HFω3) during
51 lactation.
52
Group
Variables
C HF HFω3
1st week of lactation

Consumption (g) 162 ± 13 166 ± 18 181 ± 21
Calories (Kcal) 599 ± 48 753 ± 84 819 ± 95
Carbohydrates (g) 96 ± 7 88 ± 10 101 ± 11
Protein (g) 28 ± 2 37 ± 4 38 ± 4
Lipids (g) 11 ± 1 28 ± 3* 29 ± 3*
Omega-3 (g) 0.3 ± 0.01 0.3 ± 0.02 0.7 ± 0.04*
Fo
2th week of lactation

Consumption (g) 215 ± 31 265 ± 17 213 ± 14
Calories (Kcal) 798 ± 115 1201 ± 78 961 ± 66
Carbohydrates (g) 127 ± 18 141 ± 9 119 ± 8
rP
Protein (g) 38 ± 5 59 ± 4* 44 ± 3
Lipids (g) 14 ± 2 45 ± 3* 34 ± 2*
Omega-3 (g) 0.4 ± 0.03 0.5 ± 0.01 0.8 ± 0.03*
ee
3th week of lactation

Consumption (g) 281 ± 72 291 ± 31 265 ± 64
Calories (Kcal) 1089 ± 217 1319 ± 140 1198 ± 288
rR
Carbohydrates (g) 166 ± 42 155 ± 16 148 ± 35

Protein (g) 50 ± 12 65 ± 7 55 ± 13
Lipids (g) 19 ± 5 49 ± 5* 43 ± 10*
ev
Omega-3 (g) 0.5 ± 0.08 0.5 ± 0.03 1 ± 0.14*
53 The rats were submitted during gestation to a control diet (19% lipids) or with a high content of saturated fatty acids, HF, with 33%
54 lipids or with a high content of saturated fatty acids enriched with omega-3 with 3.5 % flaxseed oil, HFω3, according to the
iew
55 experimental group. Values were expressed as mean ± SEM. There was no statistical difference after the application of One-Way
56 ANOVA and Bonferroni post-test (N = 3) or Kruskal-Wallis Test and Dunns post-test when values of the variables did not present
57 normal distribution. * p˂0.05 vs. Control; # p˂0.05 vs. HF.
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73

5
74 Table 5. Food intake in grams, calories and macronutrients and coefficient of dietary efficiency of offspring
75 of rats subjected to the Control (C), High Fat Diet (HF) and High Fat Diet enriched with Omega-3-rich
76 (HFω3) after lactation.
77
Group
Period/ Nutrient
C HF HFω3
22 to 30-d-old
Consumption (g) 110 ± 1 100 ± 2* 106 ± 1
Calories (Kcal) 380 ± 4 346 ± 6* 367 ± 3
Proteins (g) 28 ± 0.3 25 ± 0.4* 27 ± 0.6
Carbohydrates (g) 60 ± 0.7 55 ± 0.9* 58 ± 0.5
Food Efficiency 0.4 ± 0.009 0.4 ± 0.01 0.4 ± 0.01
Consumption (FEC)
30 to 60-d-old
Consumption (g) 727 ± 9 735 ± 17 741 ± 7
Calories (Kcal) 2503 ± 33 2528 ± 58 2550 ± 24
Fo
Proteins (g) 184 ± 2 186 ± 4 191 ± 1

Carbohydrates (g) 397 ± 5 401 ± 9 404 ± 3
Food Efficiency 0.2 ± 0.009 0.2 ± 0.006 0.3 ± 0.009*
Consumption (FEC)
rP
60 to 90-d-old
Consumption (g) 618 ± 30 597 ± 14 615 ± 8
ee
Calories (Kcal) 2128 ± 105 2056 ± 48 2115 ± 27

Proteins (g) 157 ± 7 151 ± 3 156 ± 2
Carbohydrates (g) 337 ± 16 326 ± 7 335 ± 4
Food Efficiency 0.1 ± 0.008 0.1 ± 0.01 0.09 ± 0.007
rR
Consumption (FEC)
90 to 300-d-old
ev
Consumption (g) 10218 ± 5 8259 ± 2* 10611 ± 5#

Calories (Kcal) 35149 ± 2 28410 ± 1* 36501 ± 2
Proteins (g) 740 ± 0.3 607 ± 0.2 780 ± 0.3
Carbohydrates (g) 1609 ± 0.5 1300 ± 0.3 1671 ± 0.4
iew
Food Efficiency 0.1 ± 0.01 0.1 ± 0.01 0.1 ± 0.01

Consumption (FEC)
78 The rats were submitted during gestation to a control diet (19% lipids) or with a high content of saturated fatty acids, HF, with 33%
79 lipids or with a high content of saturated fatty acids enriched with omega-3 with 3.5 % flaxseed oil, HFω3, according to the
80 experimental group. Values were expressed as mean ± SEM. There was no statistical difference after the application of One-Way
81 ANOVA and Bonferroni post-test (N = 12-17) or Kruskal-Wallis Test and Dunns post-test when values of the variables did not present
82 normal distribution. * p˂0.05 vs. Control; # p˂0.05 vs. HF.
83

6
84 Table 6. Murinometric parameters of male offspring whose dams were submitted to Control (C), High Fat
85 Diet (HF) and High Fat Diet enriched with Omega-3-rich (HFω3) diets during gestation and lactation
86
Group
Days old / Variables
C HF HFω3
M ± SEM M ± SEM M ± SEM
1-d-old
Body Weight (g) 6 ± 0.1 7.± 0.1* 6 ± 0.1#
Abdominal circumference 4 ± 0.1 4 ± 0.1* 4 ± 0.1*
(cm)
Naso-anal-length (cm) 5 ± 0.1 5 ± 0.1* 5 ± 0.1#
7-d-old
Body Weight (g) 18 ± 0.2 20 ± 0.1* 20 ± 0.5*
Abdominal circumference 6 ± 0.1 6 ± 0.1* 6 ± 0.1
(cm)
Naso-anal-length (cm) 7 ± 0.1 7 ± 0.1* 7 ± 0.1#
Fo
14-d-old
Body Weight (g) 32 ± 0.3 34 ± 0.5* 36 ± 0.6*
Abdominal circumference 8 ± 0.1 8 ± 0.1* 8 ± 0.1
rP
(cm) 8 ± 0.1 8 ± 0.1 8 ± 0.1

Naso-anal-length (cm)
21-d-old 54 ± 0.6 54 ± 0.8 57 ± 0.7*#

ee
Body Weight (g) 10 ± 0.1 10 ± 0.1 10 ± 0.1

Abdominal circumference 10 ± 0.1 11 ± 0.1* 11 ± 0.2*
(cm)
rR
93 ± 2 97 ± 2 100 ± 2
30-d-old 12 ± 0.1 12 ± 0.2* 11 ± 0.2#
Body Weight (g) 13 ± 0.1 15 ± 0.3* 13 ± 0.1#
ev
Abdminal circumference (cm)

370 ± 6 358 ± 4.3 380 ± 7
iew
90-d-old 19 ± 0.3 18± 0.3 20 ± 0.3

Body Weight (g) 22 ± 0.5 22 ± 0.4 22 ± 0.4
Abdominal circumference 0.3± 0.007 0.3 ± 0.005 0.3 ± 0.005
(cm)
Naso-anal-lenght (cm)
Lee index (g/cm3) 472 ± 14 498 ± 16 511 ± 15
20 ± 0.5 21 ± 0.4 21 ± 0.4
300-d-old 24 ± 1 26 ± 0.3 25 ± 1.0
Body Weight (g) 0.3 ± 0.01 0.3 ± 0.005 0.3 ± 0.017
Abdominal circumference
(cm)
Naso-anal-lenght (cm)
Lee index (g/cm3)
87 The offspring rats were separated into three groups according to their dams diets during gestation and lactation: control diet, C group,
88 (19% lipids) or high fat diet, HF group, with 33% lipids, or with high fat diet omega-3-rich, HFω3 group, with 3.5% flaxseed oil.
89 Values were expressed as mean ± SEM. There was no statistical difference after the application of One-Way ANOVA and Bonferroni
90 post-test (N = 7-35). * p˂0.05 vs. C; # p˂0.05 vs. HF.
91
92
93
94
95

7
96 Table 7. Fasting biochemical profile at 19 days of gestation of the dams who were submitted to Control
97 (C), High Fat Diet (HF) and High Fat Diet enriched with Omega-3-rich (HFω3) diets
Age / Variables Group

C HF HFω3
19 days of gestation
Albumin (g/dL) 4 ± 0.1 4 ± 0.1 4 ± 0.1
Total protein (g/dL) 6 ± 0.2 6 ± 0.1 6 ± 0.1
Triglycerides (mg/dL) 145 ± 21 121 ± 17 119 ± 17
Total cholesterol 73 ± 1 90 ± 3* 34 ± 4#
(mg/dL)
HDL-c (mg/dL) 9±1 9±2 16 ± 3
LDL-c (mg/dL) 22 ± 11 56 ± 7* 23 ± 7
VLDL-c (mg/dL) 34 ± 6 24 ± 3 23 ± 3
Glucose (mg/dL) 120 ± 4 129 ± 6 109 ± 10
ALT (U/L) 49 ± 19 54 ±10 37 ± 6
AST (U/L) 95 ± 24 131 ± 8 97 ± 12
AST/ALT 2 ± 0.3 3 ± 0.5 2 ± 0.7
Fo
101
rP
post-test (n = 3-9). * p˂0.05 vs. C; # p˂0.05 vs. HF.
102
103
ee
104
105
rR
106
107
108
ev
109
110
iew
111
112
113
114
115
116
117
118
119
120
121
122
123
124
125
126

8
127 Table 8. Fasting biochemical profile at 22, 30, 90, and 300 days old (-d-old) from the offspring of males
128 whose dams were submitted to Control (C), High Fat Diet (HF) and High Fat Diet enriched with Omega-
129 3-rich (HFω3) diets during gestation and lactation.
130
Age/ Variables Group
C HF HFω3
22-d-old
Albumin (g/dL) 4 ± 0.02 4 ± 0.01 4 ± 0.02
Triglycerides (mg/dL) 138 ± 6 124 ± 6 116 ± 4*
Total cholesterol (mg/dL) 116 ± 1 126 ± 3* 117 ± 2#
HDL-c (mg/dL) 23 ± 3 28 ± 3 31 ± 4
LDL-c (mg/dL) 51 ± 7 63 ± 9 61 ± 6
VLDL-c (mg/dL) 28 ± 1 24 ± 1 23 ± 0.8*
Glucose (mg/dL) 108 ± 3 119 ± 3* 117 ± 1
ALT (U/L) 17 ± 0.7 22 ± 1* 17 ± 0.6#
AST (U/L) 152 ± 4 137 ± 3 142 ± 4
Fo
AST/ALT 8 ± 0.7 5 ± 0.8* 8 ± 0.4#
30-d-old
Albumin (g/dL) 4 ± 0.02 4 ± 0.03 4 ± 0.03
rP

Triglycerides (mg/dL) 162 ± 11 151 ± 10 183 ± 15
Total cholesterol (mg/dL) 120 ± 2 123 ± 2 125 ± 3
ee
HDL-c (mg/dL) 26 ± 1 27 ± 3 31 ± 2
LDL-c (mg/dL) 64 ± 2 78 ± 4* 60 ± 2
VLDL (mg/dL) 28 ± 2 22 ± 2 24 ± 3
Glucose (mg/dL) 90 ± 5 88 ± 3 100 ± 4
rR
ALT (U/L) 30 ± 1 34 ± 2 38 ± 2
AST (U/L) 149 ± 4 143 ± 6 141 ± 0.04
AST/ALT 5 ± 0.3 4 ± 0.6 3 ± 0.4
ev
90-d-old
Albumin (g/dL) 4 ± 0.03 4 ± 0.04 4 ± 0.04
iew

Triglycerides (mg/dL) 97 ± 1 110 ± 1* 108 ± 4
Total cholesterol (mg/dL) 80 ± 1 87 ± 1* 82 ± 1
HDL-c (mg/dL) 13 ± 0.5 12± 1 10 ± 0.7
LDL-c (mg/dL) 47 ± 1 52 ± 3 46 ± 3
VLDL (mg/dL) 19 ± 0.3 22 ± 1* 21 ± 0.8
Glucose (mg/dL) 120 ± 3 142 ± 3* 126 ± 3#
ALT (U/L) 44 ± 1 44 ±1 45 ± 1
AST (U/L) 143 ± 4 134 ± 6 115 ± 3*#
AST/ALT 3 ± 0.1 3 ± 0.1 2 ± 0.08*#
300-d-old
Albumin (g/dL) 4 ± 0.06 4 ± 0.04* 4 ± 0.04
Triglycerides (mg/dL) 59 ± 3 83 ± 4* 86 ± 5*
Total cholesterol (mg/dL) 78 ± 3 87 ± 2* 88 ± 2*
HDL-c (mg/dL) 52 ± 6 54 ± 6 55 ± 4
LDL-c (mg/dL) 25 ± 5 21 ± 4 25 ± 4
VLDL (mg/dL) 12 ± 0.7 16 ± 0.8* 16 ± 1*
Glucose (mg/dL) 126 ± 10 135 ± 7 135 ± 4
ALT (U/L) 55 ± 3 53 ± 7 62 ± 4
AST (U/L) 175 ± 7 139 ± 9* 115 ± 5*
AST/ALT 3.2 ± 0.1 2.4 ± 0.2* 1.9 ± 0.1*

9
134 post-test (N = 19-35). * p˂0.05 vs. C; # p˂0.05 vs. HF.
135
136
137
138
139
140
141
142
143
Fo
144
rP
ee
rR
ev
iew

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Journal of Developmental Origins of Health and Disease

Maternal consumption of ɷ3 attenuates metabolic disruption

Journal: Journal of Developmental Origins of Health and Disease

Manuscript Type: Original Article

Date Submitted by the

Complete List of Authors: Alves, Debora; Universidade Federal de Pernambuco, Department of

Bloise, Aline; Universidade Federal de Pernambuco, Department of

Rocha-Junior, Reginaldo; Universidade Federal de Pernambuco,

Menezes, Luiza; Universidade Federal de Pernambuco, Academic Center

de-Oliveira, Yohanna; Universidade Federal da Paraiba, Department of

Physiology and Pharmacology

Model or Method: Animal, Small animals < Animal

Developmental Stage, Child growth and health < Developmental Stage,

Cambridge University Press

murinometric measurements, serum levels of metabolic markers, insulin

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1 Maternal consumption of ɷ3 attenuates metabolic disruption elicited by saturated fatty

11 - PE, 50760-901, Brazil.

14 João Henrique Costa-Silva

Cambridge University Press

Cambridge University Press

41 The advent of nutritional transition, markedly characterized by increased consumption of high

55 protecting the offspring against the metabolic disorders later in life.

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105 2.4 Murinometric measurements of the offspring

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165 others nutrients among groups.

3.2 Evaluation of murinometric measurements of the offspring

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195 3.4 Biochemical assessment in male offspring

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247 food consumption during the perinatal period.

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Cambridge University Press

Cambridge University Press

Cambridge University Press

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400 offspring. The Journal of nutritional biochemistry. 2015;26(2):99-111.

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462 research. 2009;48(1):52-61.

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Cambridge University Press

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25 vs. Control, # p˂0.05 vs. HF; n= 10-18).

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GTT AUC(mg/dL x 120 min)

T0 T15 T30 T60 T120

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PTT AUC(mg/dL x 120 min)

IST AUC(mg/dL x 120 min)

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1 Table 1. Composition regarding the ingredients of experimental diets

DL-methionine 0.3 0.2 0.2

Monosodium glutamate - 0.2 0.2

% Omega-3 0.2 0.2 0.4

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Table 2. Centesimal composition of nutrients from formulated experimental diets

Nutrient (g/100g) AIN-93G Hyperlipidic Hyperlipidic enriched

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M±SEM M±SEM M±SEM

46 Control; # p˂0.05 vs. HF.