Validation of a DNA methylation reference panel for the estimation of nucleated cells types in cord blood

Figures & data

Table 1. Sample characteristics for the Gen3G birth cohort used for validation.

Sample Characteristic (n = 154)	Mean (SD) or n (%)
Maternal Characteristics
Nulliparous	75 (48.7%)
Age (years)	28.2 (4.1)
Child Characteristics
Birth weight (grams)	3,400 (436)
Gestational age at birth (weeks)	39.4 (1.2)
Female	85 (55.2%)
European descent	152 (99%)

Figure 1. Scatter plots and linear regression line for the comparison of the measured (y-axis) and estimated (x-axis) nucleated cell type composition from the cord blood reference panel.

Table 2. Measured and predicted proportions of nucleated cell types using the cord blood reference panel and the adult blood reference panel in the Gen3G birth cohort (n = 154).

	Direct measurements	Cord blood reference panel				Adult blood reference panel
Nucleated Cell Type	Mean (SD)	Mean (SD)	r	R^2	rMSE	Mean (SD)	r	R^2	rMSE
nRBCs	5.4% (3.6)	12.2% (3.9)	0.92	0.85	1.4%	NA	NA	NA	NA
Lymphocytes	31.3% (7.2)	46.4% (9.6)	0.77	0.58	4.6%	45.1% (9.0)	0.69	0.47	5.2%
Granulocytes	52.9% (7.8)	39.2% (10.4)	0.72	0.52	5.4%	49.3% (8.0)	0.68	0.46	5.7%
Monocytes	10.5% (2.3)	9.6% (2.5)	0.51	0.25	2.0%	11.8% (2.6)	0.44	0.19	2.1%

NA: not available.

r: correlation coefficient.

R²: coefficient of determination.

rMSE: root mean square error for the residuals from the linear regression model.

Figure 2. Distributions for the nucleated cell types directly measured in cord blood (red) and the estimates obtained from the cord blood reference panel using the raw DNA methylation data and 5 different normalization methods.

Table 3. Evaluation of normalization methods for the prediction of nucleated cell types in cord blood using the cord blood reference panel, compared to the complete blood count measured cell types in the Gen3G birth cohort (n = 154).

	Raw data			Illumina (Studio)			Noob (Default)			FunNorm			Quantile			SWAN
Cell types	r	R^2	rMSE	r	R^2	rMSE	r	R^2	rMSE	r	R^2	rMSE	r	R^2	rMSE	r	R^2	rMSE
nRBCs	0.92	0.84	1.4%	0.93	0.87	1.3%	0.92	0.85	1.4%	0.92	0.85	1.3%	0.86	0.73	1.9%	0.92	0.84	1.4%
Lymphocytes	0.76	0.58	4.7%	0.76	0.58	4.7%	0.77	0.58	4.6%	0.77	0.59	4.6%	0.75	0.55	4.8%	0.76	0.58	4.7%
Granulocytes	0.73	0.53	5.3%	0.72	0.52	5.4%	0.72	0.52	5.4%	0.72	0.52	5.4%	0.71	0.50	5.5%	0.73	0.52	5.4%
Monocytes	0.52	0.27	2.0%	0.54	0.29	2.0%	0.51	0.25	2.0%	0.53	0.27	2.0%	0.53	0.28	1.9%	0.54	0.29	2.0%

r: correlation coefficient.

R²: coefficient of determination.

rMSE: root mean square error for the residuals from the linear regression model.

Figure 3. Quantile-quantile plot of the log₁₀ (P values) for the epigenome-wide association of gestational age at birth among models adjusted for cell type composition using the adult reference methylation panel, the direct clinically measured cell types, and the cord blood reference methylation panel.