Effect of the Japanese diet during pregnancy and lactation or post-weaning on the risk of metabolic syndrome in offspring

Figures & data

Table 1. Menu card of 1975 Japanese diet.

	Day 1	Day 2	Day 3	Day 4	Day 5	Day 6	Day 7
Break-fast	Rice	Raisin Bread	Rice	Toast	Rice	Toast	Rice
	Salt-Grilled Salmon	Omelet	Hard-Cured Horse Mackerel	Bacon and Eggs	Rolled Egg	Boiled Egg	Sake-Steamed Short-Necked Clams and Cabbage
	Natto (Fermented Soybeans)	Sausage and Cabbage Saute	Stewed Short-Necked Clams with Japanese Mustard Spinach	Yogurt with fruit	Natto (Fermented Soybeans)	Tuna Salad with Broccoli	Natto (Fermented Soybeans)
	Chinese Cabbage and Bean Sprout Miso Soup	Fruit	Scarlet Runner Beans Boiled in Sugar and Soy Sauce		Cabbage and Deep-Fried Tofu Miso Soup	Fruit	Tofu and Deep-Fried Tofu Miso Soup
		Milk	Miso Soup with Eggplant		Fruit	Milk
Lunch	Udon Noodles with Deep-Fried	Fried Rice	Japanese-Style Chow Mein	Sweet Potato Rice	Chicken and Egg on Rice	Rice	Sandwich
	Fruit	Seaweed Soup	Fruits Mitsumame	Simmered Freeze-Dried Tofu	Rad and White Salad	Sauteed Eggplant with Mince	Consomme
				Miso Soup with Pork and Vegetables	Food Boiled in Soy Sauce	Simmered Hijiki Seaweed	Fruit
Dinner	Rice	Rice	Rice	Rice	Rice	Rice	Rice
	Braised Meat and Vegetables	Chicken and Vegetables Chikuzen Style	Cream Stew	Simmered Mackerel in Miso	Marinated Horse Mackerel, Nanban-Style	Simmered Flatfish	Sliced Raw Fish
	Mozuku Seaweed Vinger	Chilled Tofu	Pickled Chinese Cabbage and Dried Shrimp Dipped in Sauce	Cooked Beans with Various Vegetables	Tofu Dengaku	Simmered Okara Tofu Lees	Simmered Satsuma Age (Deep-Fried Minced Fish and Vegetables) and Chinese Cabbage
	Clear Cabbage and Egg Soup	Spinach and Deep-Fried Tofu Miso Soup	Marinated Cucumber and Hijiki Seaweed	Clear Chinese Cabbage and Wakame Seaweed Soup	Clear Squash and Japanese Mustard Spinach Soup	Taro and White Radish Miso Soup	Tofu and Vegetables in a Sesame Sauce

Figure 1. Study protocol. Dams were given the Control diet or the Japanese diet during pregnancy and lactation and offspring from each group was divided into two groups given the Control diet or 1975 Japanese diet after weaning at 3 weeks.

Table 2. Primer pairs used for the quantitative RT-PCR analysis.

NM_031884	Abcg5	Forward	AGGGCCTCACATCAACAGAG
		Reverse	GCTGACGCTGTAGGACACAT
NM_153151	Acat3	Forward	CCAGTGGTCATCGTCTCGG
		Reverse	GGACAGGGCACCATTGAAGG
NM_133904	Acc	Forward	CGCTCACCAACAGTAAGGTGG
		Reverse	GCTTGGCAGGGAGTTCCTC
NM_015729	Acox1	Forward	TCCAGACTTCCAACATGAGGA
		Reverse	CTGGGCGTAGGTGCCAATTA
NM_007393	β-actin	Forward	GGCTGTATTCCCCTCCATCG
		Reverse	CCAGTTGGTAACAATGCCATGT
NM_009948	Cpt1	Forward	ATTCTGTGCGGCCCTTATTGGAT
		Reverse	TTTGCCTGGGATGCGTGTAGTGT
NM_009949	Cpt2	Forward	TGTCTTCCAAGCACTTCTGG
		Reverse	TGGATAGGCTGCAATGTCTC
NM_007824	Cyp7a1	Forward	CAAGTGTCCCCCTCTAGA
		Reverse	ACTCAATATCATGTAGTGGTGGCAAA
NM_007988	Fasn	Forward	CCTGGATAGCATTCCGAACCTG
		Reverse	TTCACAGCCTGGGGTCATCTTTGC
NM_008062	G6pdx	Forward	TGGGTCCACCACTGCCACTTTTG
		Reverse	ATTGGGCTGCACACGGATGACCA
AK079302	Hmgcar	Forward	AGCTTGCCCGAATTGTATGTG
		Reverse	TCTGTTGTGAACCATGTGACTTC
NM_013839	Lxra	Forward	CTCAATGCCTGATGTTTCTCCT
		Reverse	TCCAACCCTATCCCTAAAGCAA
M29546	Me	Forward	CCTCACCACTCGTGAGGTCAT
		Reverse	CGAAACGCCTCGAATGGT
NM_011144	Pparα	Forward	AACATCGAGTGTCGAATATGTGG
		Reverse	AGCCGAATAGTTCGCCGAAAG
NM_009270	Sqle	Forward	ATTGGCTCAGGCCTTGTATG
		Reverse	ATTGAAAGCAACCCAACAGG
NM_011480	Srebp1c	Forward	GGAGACATCGCAAACAAGC
		Reverse	TGAGGTTCCAAAGCAGACTG

Notes: Abcg5, ATP-binding cassette, sub-family G (WHITE), member 5; Acat3,acetyl-Coenzyme A acetyltransferase 3;Acc,acetyl-Coenzyme A carboxylase beta; Acox1,acyl-Coenzyme A oxidase 1, palmitoyl;β-actin, actin, beta, cytoplasmic; Cpt1,Carnitine palmitoyl transferase 1; Cpt2,carnitine palmitoyltransferase 2; Cyp7a1,cytochrome P450, family 7, subfamily a, polypeptide 1; Fasn,fatty acid synthase; G6pdx,glucose-6-phosphate dehydrogenase X-linked; Hmgcar,3-hydroxy-3-methylglutaryl-Coenzyme A reductase; Lxra,nuclear receptor subfamily 1, group H, member 3; Me,malic enzyme; Pparα,peroxisome proliferator activated receptor alpha; Sqle,squalene epoxidase;Srebp1c,sterol regulatory element binding protein 1 c.

Table 3. Nutritional compositions of test diets.

Test diets		Control diet	1975 Japanese diet
Protein	(g/100 g)	16.6	17.1
Fat	(g/100 g)	4.3	7.2
Carbohydrate	(g/100 g)	71.4	67.3
Energy	(kcal/100 g)	391	402

Table 4. Growth parameters.

	CO-CO	CO-75	75-CO	75–75	Interaction
Initial body weight (g)	11.7 ± 0.2	11.6 ± 0.2	12.1 ± 0.8	12.6 ± 0.7
Final body weight (g)	36.7 ± 0.8	33.7 ± 0.6	35.7 ± 1.3	35.6 ± 0.9
Body weight gain (g)	25.0 ± 1.0	22.0 ± 0.5	20.0 ± 3.3	23.1 ± 1.4
Food intake (g/day)	4.15 ± 0.14	3.99 ± 0.31	3.88 ± 0.21	4.12 ± 0.24
Enegry intake (kcal/day)	16.2 ± 0.6	16.0 ± 1.3	15.2 ± 0.8	16.6 ± 1.0
Tissue weight (g/100 g body weight)
Brain	1.36 ± 0.03	1.44 ± 0.02	1.43 ± 0.03	1.36 ± 0.04
Liver	4.09 ± 0.06	4.26 ± 0.08	4.10 ± 0.11	4.22 ± 0.10
Pancreas	0.85 ± 0.04	0.89 ± 0.05	0.90 ± 0.04	0.90 ± 0.08
Kidney	1.69 ± 0.07^ab	1.78 ± 0.06^b	1.47 ± 0.05 ^a	1.66 ± 0.04^ab	P, O
White adipose tissue
Mesenteric	0.75 ± 0.08^b	0.36 ± 0.05^a	0.75 ± 0.14^b	0.56 ± 0.09^ab	O
Perirenal	0.78 ± 0.12^b	0.35 ± 0.05^a	0.64 ± 0.07^ab	0.53 ± 0.09^ab	O
Epididymal	1.53 ± 0.20^b	0.79 ± 0.10^a	1.64 ± 0.16^b	1.43 ± 0.20^ab	P, O
Total	3.06 ± 0.34^b	1.51 ± 0.18^a	3.03 ± 0.33^b	2.52 ± 0.35^ab	O

Notes: Values are means ± SE, n = 10. Because there were two factors of maternal diet and post-weaning diet Statistical analysis was performed using two-way ANOVA and for items with significant differences one-way ANOVA was performed and followed by Tukey-Kramer’s test. Different superscript letters indicate significantly different means at p < 0.05. maternal diet effect, Post-weaning diet effect and interaction were described at the right side of the table : P, maternal diet effect; O, post-weaning diet effect; P × O, interaction.

Figure 2. Effects of the Japanese diet intake from pregnancy and lactation on adipocytes and liver of offspring at 7 weeks. (a) Effects of the Japanese diet intake from pregnancy and lactation on adipocytes of offspring at 7 weeks. Photographs were hematoxylin-eosin staining of epididymal adipose tissue sections from representative mice of each group (scale bar = 100 μm). (b) Bar graph of the adipocytes size ratio of the CO-CO, CO-JD, JD-CO and JD-JD groups. Statistical analysis was performed using one-way ANOVA and two-way ANOVA followed by Tukey-Kramer’s test. Different superscript letters indicate significantly different means at p < 0.05. Maternal diet effect, Post-weaning diet effect and interaction were described at the top of the graph: P, Maternal diet effect; O, post-weaning diet effect; P × O, interaction. (c) Effects of the Japanese diet intake from pregnancy and lactation on liver of offspring at 7 weeks. Photographs were hematoxylin-eosin staining of liver sections from representative mice of each group (scale bar = 100 μm).

Table 5. Biochemical parameters.

	CO-CO	CO-75	75-CO	75–75	Interaction
Serum
TG (mmol/L)	1.22 ± 0.17^b	0.64 ± 0.05^a	1.33 ± 0.18^b	1.01 ± 0.11^ab	P, O
TC (mmol/L)	2.83 ± 0.25^ab	2.34 ± 0.15^a	2.60 ± 0.15^ab	3.07 ± 0.20^b	P, P×O
PL (mmol/L)	13.7 ± 1.0^ab	11.1 ± 0.7^a	13.4 ± 0.8^ab	14.2 ± 0.9^b	P×O
Glucose (mmol/L)	6.85 ± 0.44	8.49 ± 0.34	7.98 ± 1.10	7.63 ± 0.31
Insulin (nmol/L)	0.06 ± 0.01	0.09 ± 0.02	0.06 ± 0.02	0.14 ± 0.04	O
Albumin (g/L)	29.8 ± 1.0	28.8 ± 1.0	29.1 ± 0.8	28.9 ± 1.2
Liver
TG (μmol/g)	39.5 ± 4.6^bc	19.4 ± 1.9 ^a	47.4 ± 9.3 ^c	27.0 ± 3.3^ab	O
TC (μmol/g)	44.8 ± 4.5^ab	53.6 ± 5.5^b	35.5 ± 3.5 ^a	46.6 ± 3.5^ab	O
PL (μmol/g)	28.6 ± 3.6	30.6 ± 1.4	29.0 ± 2.5	31.5 ± 1.0

Notes: Values are means ± SE, n = 10. Because there were two factors of maternal diet and post-weaning diet Statistical analysis was performed using two-way ANOVA and for items with significant differences one-way ANOVA was performed and followed by Tukey-Kramer’s test. Different superscript letters indicate significantly different means at p < 0.05. maternal diet effect, Post-weaning diet effect and interaction were described at the right side of the table : P, maternal diet effect; O, post-weaning diet effect; P × O, interaction.

Table 6. mRNA expression levels for lipid metabolism-related genes in liver.

	CO-CO	CO-75	75-CO	75–75	Function	Interaction
(Ratio)
Acc	1.00 ± 0.17^b	0.32 ± 0.04^a	0.99 ± 0.21^b	0.38 ± 0.03^a	Fatty acid synthesis	O
Fasn	1.00 ± 0.06	1.14 ± 0.15	1.26 ± 0.11	1.40 ± 0.12		P
G6pdx	1.00 ± 0.06	1.05 ± 0.08	1.32 ± 0.12	1.33 ± 0.11		P
Me	1.00 ± 0.16	0.94 ± 0.08	0.55 ± 0.17	0.62 ± 0.22		P
Srebp1c	1.00 ± 0.15	1.71 ± 0.17	0.99 ± 0.27	1.67 ± 0.35		O
Aco	1.00 ± 0.15^ab	0.51 ± 0.07^a	1.25 ± 0.24^b	0.69 ± 0.07^ab	Fatty acid β oxidation	O
Cpt1	1.00 ± 0.17^ab	0.47 ± 0.03^a	1.29 ± 0.23^b	0.71 ± 0.13^ab		O
Cpt2	1.00 ± 0.10^ab	0.77 ± 0.03^a	1.06 ± 0.08^b	0.92 ± 0.06^ab		O
Pparα	1.00 ± 0.10^b	0.66 ± 0.05^a	1.03 ± 0.07^b	0.83 ± 0.10^ab		O
Hmgcoar	1.00 ± 0.04^a	1.16 ± 0.06^ab	1.21 ± 0.11^ab	1.48 ± 0.09^b	Cholesterol biosynthesis	P
Sqle	1.00 ± 0.20^ab	0.28 ± 0.03^a	3.02 ± 0.50^b	0.60 ± 0.09^a		P, O
Abcg5	1.00 ± 0.07^ab	1.50 ± 0.13^c	0.80 ± 0.01^a	1.34 ± 0.13^bc	Cholesterol catabolism	O
Acat3	1.00 ± 0.06	0.81 ± 0.05	0.91 ± 0.07	0.89 ± 0.05
Cyp7a1	1.00 ± 0.31	0.50 ± 0.09	0.70± 0.13	0.75 ± 0.15
Lxra	1.00 ± 0.04^b	0.79 ± 0.04^a	0.88 ± 0.05^ab	0.88 ± 0.05^ab		O, P×O

Notes: Values are means ± SE, n = 10. Because there were two factors of maternal diet and post-weaning diet, Statistical analysis was performed using two-way ANOVA and for items with significant differences one-way ANOVA was performed and followed by Tukey-Kramer’s test.

Different superscript letters indicate significantly different means at p < 0.05. maternal diet effect, Post-weaning diet effect and interaction were described at the right side of the table: P, maternal diet effect; O, post-weaning diet effect; P × O, interaction.

Table 7. Growth parameters of 18 days offspring.

	CO (n = 6)	75 (n = 8)
Body weight (g)	7.16 ± 0.55	8.57 ± 0.35
Tissue weight (g/100 g body weight)
Liver	3.73 ± 0.09	3.79 ± 0.13
Total white adipose tissue	0.28 ± 0.05	0.41 ± 0.05

Notes: Values are means ± SE.

* Different superscript letters indicate significantly different means at p < 0.05.

Table 8. Biochemical parameters of 18 days offspring.

	CO (n = 6)	75 (n = 8)
Serum
TG (mmol/L)	1.15 ± 0.15	1.00 ± 0.13
TC (mmol/L)	3.55 ± 0.25	3.26 ± 0.10
PL (mmol/L)	17.3 ± 0.5	16.0 ± 0.5
Liver
TG (μmol/g)	44.6 ± 2.4	35.8 ± 1.6*
TC (μmol/g)	15.4 ± 0.4	16.9 ± 0.9
PL (μmol/g)	30.4 ± 0.8	31.7 ± 0.7

Notes: Values are means ± SE.

* Different superscript letters indicate significantly different means at p < 0.05.

Table 9. mRNA expression levels for lipid metabolism-related genes in liver of 18 days offspring.

	CO (n = 6)	75 (n = 8)	Function
Acc	1.00 ± 0.10	0.46 ± 0.06*	Fatty acid synthesis
Fas	1.00 ± 0.10	0.50 ± 0.05*
G6pdx	1.00 ± 0.11	0.73 ± 0.10
Me	1.00 ± 0.15	1.02 ± 0.18
Srebp1c	1.00 ± 0.16	0.57 ± 0.01
Acox1	1.00 ± 0.07	1.03 ± 0.07	Fatty acid β oxidation
Cpt1	1.00 ± 0.10	1.00 ± 0.16
Cpt2	1.00 ± 0.03	0.99 ± 0.07
Ppara	1.00 ± 0.08	1.00 ± 0.11
Hmgcoar	1.00 ± 0.10	0.60 ± 0.09*	Cholesterol biosynthesis
Sqle	1.00 ± 0.19	0.27 ± 0.06*
Abcg5	1.00 ± 0.14	1.19 ± 0.15	Cholesterol catabolism
Acat3	1.00 ± 0.08	1.08 ± 0.16
Cyp7a1	1.00 ± 0.12	1.32 ± 0.44
Lxra	1.00 ± 0.09	0.46 ± 0.16*

Notes: Values are means ± SE

* Different superscript letters indicate significantly different means at p < 0.05.