Erythrocyte folate concentrations, CpG methylation at genomically imprinted domains, and birth weight in a multiethnic newborn cohort

Figures & data

Table 1. Characteristics of 496 study participants by maternal race/ethnicity

	Non-Hispanic Black (n = 189)	Non-Hispanic White (n = 133)	Hispanic (n = 154)	Others (n = 20)	p-values
Characteristic of study participants	#(%)	#(%)	#(%)	#(%)
Maternal age 18–20 y 21–25 y 25–30 y 31–35 y 36+years	33 (17.5) 73 (38.6) 42 (22.2) 21 (11.1) 20 (10.6)	3 (2.3) 19 (14.3) 45 (33.8) 54 (40.6) 12 (9.0)	16 (10.4) 46 (29.9) 42 (27.3) 35 (22.7) 15 (9.7)	2 (10.0) 3 (15.0) 6 (30.0) 5 (25.0) 4 (20.0)	< 0.0001
Maternal Marital Status Not partnered (never married/ divorced) Living with partner (cohabit/married) Other Missing	106 (56.1) 71 (37.6) 7 (3.7) 5 (2.6)	12 (9.0) 120 (90.2) 1 (0.8) 0 (0.0)	37 (24.0) 114 (74.0) 3 (2.0) 0 (0.0)	4 (20.0) 16 (80.0) 0 (0.0) 0 (0.0)	< 0..0001*
Maternal Education < High School High School - < college College graduate Graduate/professional school Missing	5 (2.7) 99 (52.7) 69 (36.7) 15 (7.9) 1 (0.5)	3 (2.3) 19 (14.3) 55 (41.4) 55 (41.4) 1 (0.8)	62 (40.3) 75 (48.7) 10 (6.5) 7 (4.5) 0 (0.0)	2 (10.0) 7 (35.0) 4 (20.0) 7 (35.0) 0 (0.0)	< 0.0001
Parity None One Two Three or more	50 (26.5) 44 (23.2) 34 (18.0) 61 (32.3)	43 (32.3) 44 (33.1) 25 (18.8) 21 (15.8)	33 (21.4) 32 (20.8) 36 (23.4) 53 (34.4)	6 (30.0) 6 (30.0) 3 (15.0) 5 (25.0)	0.02
Maternal BMI pre-pregnancy < 25 25- < 30 30- < 35 35- < 40 40+ Missing	60 (31.6) 50 (26.5) 33 (17.5) 21 (11.1) 19 (10.1) 6 (3.2)	75 (56.4) 26 (19.6) 22 (16.5) 4 (3.0) 4 (3.0) 2 (1.5%	37 (24.0) 62 (40.3) 21 (13.6) 10 (6.5) 5 (3.3) 19 (12.3)	12 (60.0) 6 (30.0) 1 (5.0) 0 (0.00) 0 (0.0) 1 (5.0)	< 0.0001
Mean BMI (sd)	29.4 (8.2)	25.2 (5.8)	27.5 (5.5)	23.4 (3.2)	< 0.0001
Maternal cigarette smoking   Ever smoked   Quit sometime during pregnancy   Never   Missing	16 (8.5) 48 (25.4) 125 (66.1) 0 (0.0)	29 (68.2) 18 (13.5) 85 (63.9) 1 (0.8)	8 (5.2) 6 (3.9) 139 (90.2) 1 (0.7)	4 (20.0) 2 (10.0) 14 (70.0) 0 (0.0)	< 0.0001
Gestational age a delivery > 37 wk < = 37weeks	157 (83.1) 32 (16.9)	116 (87.2) 17 (12.8)	135 (87.7) 19 (12.3)	16 (80.0) 4 (20.0)	0.5
Sex of offspring Male Female	93 (49.2) 96 (50.8)	75 (56.4) 58 (43.6)	72 (46.8) 82 (53.2)	9 (45.0) 11 (55.0)	0.4
Prenatal physical activity No Yes	130 (68.8) 59 (31.2)	119 (89.5) 14 (10.5)	114 (74.0) 40 (26.0)	17 (85.0) 3 (15.0)	0.0002
Gestational/Type II diabetes mellitus Yes No	14 (7.4) 174 (92.1)	5 (3.8) 128 (96.2)	6 (4.5) 146 (95.5)	0 20 (100)	0.001*
Missing	1 (0.5)	0	2 (1.3)	0
Birth weight (mean, sd)	3164.0 (535.1)	3452.9 (548.7)	3342.1 (498.2)	3126.1 (512.2)	< 0.0001
Maternal folate levels (µg/L) in all	194.6 (83.2)	247.6 (66.6)	222.3 (64.1)	204.4 (83.6)	< 0.0001
1st trimester blood draw (n = 326)	191.3 (79.9)	249.3 (66.1)	215.5 (61.2)	214.7 (55.1)	< 0.0001
2nd trimester blood draw (n = 160)	198.2 (92.9)	243.8 (69.5)	228.9 (69.4)	175.5 (119.7)	0.02
3rd trimester blood draw (n = 6)	208.3 (57.4)	188.0 (NA)	229.0 (35.2)	283.5 (NA)	0.4

* P values based on Fisher’s exact test. White pregnant mothers are older, more likely to be married, have a higher educational attainment, have a lower pre-pregnancy BMI > 30 or diabetes . NA-no variability.

Table 2. Regression coefficients for the association between maternal folate and offspring birth weight

	Erythrocyte folate concentrations
Characteristic	Regression coefficient (SE) p-value
Intercept	3044.68 (179.08)
Maternal folate concentrations       1st Quartile	Referents
2nd Quartile	143.18 (63.24) 0.024
3rd Quartile	117.32 (63.99) 0.067
4th Quartile	133.88 (65.20) 0.041
Male sex	140.82 (43.55) 0.001
Gestational age < 37 wk	-620.70 (61.84) 0.000
Race/ethnicity       Non-Hispanic Black	-243.63 (60.96) 0.000
Hispanic	-121.74 (62.43) 0.052
Other	-213.85 (117.48) 0.069
Non-Hispanic White	Referents
Pre-pregnancy BMI	11.88 (3.40) 0.001

After adjusting for maternal race/ethnicity, offspring sex, cigarette smoking, gestational age at delivery, gestational age at blood draw, physical activity, maternal pre-pregnancy body mass index (BMI) and delivery route, we note a relationship between maternal folate concentrations and birth weight consistent with a threshold effect.

Figure 1. Maternal folate and birth weight by pre-pregnancy BMI < 30 and BMI ≥ 30Kg/m². Adjusted for race/ethnicity, gestational age, cigarette smoking, gestational age at delivery, pre-pregnancy BMI and physical activity during pregnancy. *p-value < 0.01, ** p-value < 0.001

Table 3. Regression coefficients and standard errors for associations between maternal folate levels in quartiles and DNA methylation fractions at 9 DMRs regulating imprinted genes

	All		BMI < 30		BMI ≥ 30
DMR	Mean, β, SE	p-value	β, SE	p-value	β, SE	p-value
MEG3 (mean, se)	67.2% (0.64)
Quartile 2	-1.16 (0.56)	0.041	-0.77 (0.64)	0.237	-2.02 (1.17)	0.092
Quartile 3	-1.93 (0.57)	0.001	-1.47 (0.69)	0.037	-2.72 (1.05)	0.013
Quartile 4	-2.02 (0.58)	0.001	-1.68 (0.68)	0.017	-2.47 (1.16)	0.039
NNAT (mean, se)	56.7% (0.84)
Quartile 2	-0.83 (0.71)	0.241	-0.85 (0.84)	0.314	-1.02 (1.32)	0.441
Quartile 3	-1.34 (0.73)	0.068	-1.90 (0.92)	0.039	-0.21 (1.21)	0.866
Quartile 4	-1.15 (0.74)	0.119	-2.00 (0.91)	0.028	1.16 (1.27)	0.361
PEG10/SGCE (mean, se)	44.8% (0.37)
Quartile 2	-0.22 (0.32)	0.489	-0.08 (0.34)	0.820	-0.83 (0.61)	0.174
Quartile 3	-0.44 (0.32)	0.178	0.23 (0.35)	0.518	-2.08 (0.56)	0.000
Quartile 4	-0.14 (0.33)	0.661	-0.11 (0.36)	0.769	-1.12 (0.58)	0.054
MEG3-IG (mean, se)	49.9% (0.69)
Quartile 2	-0.54 (0.60)	0.372	0.04 (0.71)	0.953	-2.36 (1.19)	0.048
Quartile 3	-0.67 (0.60)	0.263	-0.43 (0.75)	0.566	-1.31 (1.04)	0.206
Quartile 4	-0.68 (0.61)	0.269	-0.50 (0.75)	0.504	-0.92 (1.16)	0.426
PLAGL1 (mean, se)	50.0% (0.46)
Quartile 2	0.20 (0.39)	0.607	0.62 (0.50)	0.209	-0.44 (0.69)	0.521
Quartile 3	-1.01 (0.40)	0.011	0.01 (0.53)	0.988	-1.86 (0.62)	0.003
Quartile 4	-0.08 (0.41)	0.851	0.55 (0.53)	0.305	-1.04 (0.64)	0.105
PEG3 (mean, se)	36.0% (0.26)
Quartile 2	0.43 (0.22)	0.031	-0.12 (0.28)	0.679	1.66 (0.38)	0.000
Quartile 3	0.22 (0.23)	0.294	-0.24 (0.30)	0.428	1.01 (0.34)	0.003
Quartile 4	0.06 (0.23)	0.602	-0.28 (0.29)	0.339	0.63 (0.37)	0.089
PEG1/MEST (mean, se)	43.7% (0.48)
Quartile 2	-0.57 (0.42)	0.177	-0.13 (0.55)	0.816	-1.51 (0.60)	0.013
Quartile 3	-0.09 (0.43)	0.835	0.40 (0.57)	0.483	-1.38 (0.58)	0.017
Quartile 4	0.39 (0.44)	0.367	0.56 (0.58)	0.333	0.01 (0.60)	0.984
H19 (mean, se)	46.0% (0.37)
Quartile 2	-0.03 (0.32)	0.920	0.37 (0.35)	0.287	-2.17 (0.74)	0.003
Quartile 3	-0.20 (0.33)	0.545	0.04 (0.37)	0.908	-0.99 (0.67)	0.138
Quartile 4	0.09 (0.33)	0.777	0.26 (0.37)	0.484	-0.77 (0.71)	0.278
IGF2 (mean, se)	51.0% (0.49)
Quartile 2	-1.22 (0.42)	0.004	-1.17 (0.50)	0.020	-1.39 (0.80)	0.083
Quartile 3	-0.47 (0.43)	0.271	-1.06 (0.52)	0.041	0.64 (0.74)	0.388
Quartile 4	-0.04 (0.43)	0.929	0.07 (0.52)	0.886	-0.43 (0.78)	0.586

Relationships adjusted for maternal race/ethnicity, cigarette smoking, delivery route, gestational age at blood draw, gestational age and delivery, physical activity, pre-pregnancy BMI. Referents are infants born to women with folate levels in the lowest quartiles. Maternal folate concentrations are associated with DNA methylation at all DMRs for either all subjects, those born to obese or non-obese women-associations are nonlinear.

Table 4. Regression coefficients for associations between offspring DMR methylation at the 9 regions and birth weight

DMR	β SE	p-value
*MEG3	-10.92 (4.12)	0.008
NNAT	-7.57 (4.04)	0.062
*PEG10/SGCE	18.11 (5.77)	0.002
MEG3-IG	0.42 (9.13)	0.963
*PLAGL1	12.33 (3.78)	0.001
PEG3	1.36 (8.97)	0.879
PEG1/MEST	-5.60 (5.68)	0.325
*H19	20.25 (6.76)	0.003
IGF2	6.10 (5.71)	0.286

Referents are infants with methylation levels in the fourth quartile. Models are adjusted for maternal race, sex, cigarette smoking, gestational age at delivery, gestational age at blood draw, physical activity, pre-pregnancy BMI, and delivery route. *Significant associations are noted between birth weight and methylation fractions at DMRs regulating SGCE, and PLAGL1, and one of two DMRs regulating the IGF2/H19 and DLK1/MEG3 imprinted domains.