Expression levels of seven candidate genes in human peripheral blood mononuclear cells and their association with preeclampsia

Figures & data

Table 1. General characteristics and risks factors for PE of the study population.

Characteristic	Controls (n = 69)	Mild PE (n = 60)	Severe PE (n = 48)	p Value
Maternal age, median (range)	26 (17–36)	27 (15–40)	27.5 (17–40)	0.061
Number of pregnancies, mean (range)	2 (1–6)	2 (1–9)	2 (1–5)	0.769
Family history of PE, n (%)	3 (4.35)	3 (5.0)	5 (10.42)	0.364
Family history of mellitus diabetes  type 2, n (%)	27 (39.13)	27 (45.0)	22 (45.83)	0.713
Primipaternity, n (%)	21 (30.43)	21 (35.0)	12 (25.0)	0.533
Use of barrier contraceptive  method, n (%)	25 (36.23)	19 (31.7)	13 (27.1)	0.578
Nulliparous, n (%)	25 (36.23)	22 (36.7)	19 (39.6)	0.663

Table 2. Endogenous gene selection.

	Study group
Endogenous gene	Controls Ct mean (±SD)	Severe PE Ct mean (±SD)	p Value
18S Ribosomal RNA	8.250 (±0.067)	9.860 (±0.147)	<0.001*
GAPDH	26.123 (±0.756)	25.757 (±0.736)	0.581
HPRT1	26.751 (±1.665)	26.017 (±1.098)	0.307

*Statistical difference.

Table 3. Selection of the quantification method.

Gene	Study group	Endogenous gene	ΔCt	Slope absolute value
VEGF-A	Controls	HPRT1	2.69	0.3067
		18S ribosomal	21.45	0.1466
		GAPDH	4.89	0.5065
	Severe PE	HPRT1	2.79	0.1844
		18S ribosomal	21.14	0.3733
		GAPDH	5.14	0.4754
TGF-β1	Controls	HPRT1	5.89	0.2322
		18S ribosomal	12.93	0.1184
		GAPDH	3.75	0.3781
	Severe PE	HPRT1	6.02	0.2412
		18S ribosomal	12.38	0.3170
		GAPDH	3.79	0.4091
HMOX-1	Controls	HPRT1	1.30	0.2151
		18S ribosomal	17.52	0.1013
		GAPDH	0.84	0.3610
	Severe PE	HPRT1	1.33	0.2426
		18S ribosomal	17.01	0.3184
		GAPDH	0.90	0.4105

Figure 1. Distribution of gene expression levels in the PE cases. Selected genes (VEGF-A, TGF-β1, HMOX-1 and SOD-1, respectively) were analyzed by real-time PCR. Total PBMC RNA isolated from normal pregnant women (n = 69) and women with PE (n = 108) were analyzed. Gene expression was calculated using the relative standard curve method and was expressed as fold-change relative to the average of expression in the controls.

Table 4. Gene expression level of the study genes.

		PE severity expression level, median (range)			PE onset expression level, median (range)
Gene	All PE cases Mean (±SD) n = 108	Mild PE n = 60	Severe PE n = 48	p Value†	Early PE n = 40	Late PE n = 68	p Value†
HMOX-1	−0.6 (1.9)	−1.0 (2.6 to −4.1)	−1.1 (2.0 to −12.0)	0.736	−1.1 (2.6 to −12.0)	−1.0 (2.0 to −4.0)	0.640
SOD-1	0.3 (1.6)	1.0 (4.4 to −3.6)	1.1 (5.5 to −2.4)	0.278	0.5 (4.4 to −3.6)	1.1 (5.5 to −2.4)	0.248
VEGF-A	−1.6 (1.8)	−1.9 (7.8 to −4.9)	−1.8 (1.8 to −4.8)	0.514	−1.7 (7.8 to −4.9)	−1.9 (1.8 to −4.4)	0.163
TGF-β1*	−10.2 (13.1)	−9.8 (4.5 to −34.8)	−8.4 (35.5 to −37.6)	0.650	−7.7 (4.5 to −31.6)	−10.2 (35.5 to −37.6)	0.649

†p Value obtained from the gene expression level comparison between subgroups (mild versus severe PE, or early versus late PE, respectively) by Mann-Whitney U statistic.

*One patient was excluded from TGF-β1 analysis because the cDNA sample was insufficient for qPCR amplification.

Table 5. Analysis of PE cases with an expression fold change of 20% for the evaluated genes and their association with severity and/or onset of disease.

		Severity (n = 108)			Onset (n = 108)
Gene	Expression status‡	Mild PE	Severe PE	p Value¶	Early PE	Late PE	p Value¶
HMOX-1	Over	12	10		6	16
	Under	26	24	0.842	20	30	0.300
	Total†	38	34		26	46
SOD-1	Over	20	22		14	28
	Under	20	11	0.152	15	16	0.194
	Total§	40	33		29	44
VEGF-A	Over	4	5		6	2
	Under	48	39	0.728	29	59	0.048*
	Total±	52	44		35	61
TGF-β1	Over	9	16		8	17
	Under	50	32	0.028*	31	51	0.598
	Total††	59	48		39	68

‡Fold change of 20%.

†Thirty-six, §thirty-five and ±twelve cases, respectively, did not show differences in their expression levels respect to controls and they were excluded from the analysis.

††One patient was excluded from TGF-β1 test because the cDNA sample was insufficient for qPCR amplification of this gene.

¶p Value obtained by Chi square statistic.

*Statistical difference.