Influence of a nucleotide oligomerization domain 1 (NOD1) polymorphism and NOD2 mutant alleles on Crohn's disease phenotype

Table 1 Primers for NOD2 and NOD1 genotyping

	Forward	Reverse	Size (bp)
NOD2
R702W	5´-AGATCACAGCAGCCTTCCTG-3´	5´-CACGCTCTTGGCCTCACC-3´	185
G908R	5´-CTCTTTTGGCCTTTTCAGATTCTG-3´	5´-CAGCTCCTCCCTCTTCACCT-3´	163
L1007finsC NOD1	5´-GGCAGAAGCCCTCCTGCAGGGCC-3´	5´-CCTCAAAATTCTGCCATTCC-3´	151
ND1+32656	5´-TGACTGTGTGTGACTCTCTCTGC-3´	5´-TGGTGAAAGCTCTCCACTATCTC-3´	250

Table 2 Genotype at NOD2 polymorphisms in CD cases and healthy controls

Mutation	Group	Genotype count, n (%)			OR (95% CI)1	P2
		WT/WT	Heterozygous	Homozygous
R702W	CD	75 (77.32)	21 (21.65)	1 (1.03)	7.04 (1.58-31.30)	0.004
	Controls	48 (96.00)	2 (4.00)	0
G908R	CD	92 (94.85)	5 (5.15)	0		0.166
	Controls	50 (100)	0	0
L1007finsC	CD	89 (91.75)	8 (8.25)	0	4.40 (0.53-36.25)	0.167
	Controls	49 (98.00)	1 (2.00)	0

¹ORs and probability values for disease status associated with carriage of at least 1 mutant allele (heterozygous, compound heterozygous and homozygous were grouped together). ²P-values were calculated with the Fisher's Exact test when comparing controls and CD patients.

Table 3 Distribution of NOD2 mutations in CD patients and controls

NOD2 genotype	CD Patients,n (%)	Controls,n (%)	OR (95% CI)1	P2
At least one variant3	30 (30.92)	3 (6)	7.01 (2.02-24.30)	< 0.001
Heterozygous	24 (24.74)	3 (6)	5.61 (1.59-19.0)	0.003
Compound heterozygous4	5 (5.15)	0
Homozygous	1 (1.03)	0

¹ORs and probability values for disease status associated with NOD2 genotype. ²P-values are calculated with the Fisher's Exact test when comparing controls and CD patients.

³At least one variant was considered if any subject had at least one copy of the variant allele.

⁴Compound heterozygous was defined as the presence of two different variants.

Table 4 Genotype frequencies of ND₁+32656 in CD patients and controls

Group	NOD2	n1	NOD1 genotype, n (%)
			*1*1	*1*2	*2*2
CD		97	58 (59.79)2	36 (37.11)	3 (3.09)
	WT/WT	67	40 (59.70)	25 (37.31)	2 (2.98)
	Mutant3	30	18 (60.00)	11 (36.66)	1 (3.33)
Controls		50	26 (52.00)	17 (34.00)	7 (14.00)
	WT/WT	47	23 (48.93)	17 (36.17)	7 (14.89)
	Mutant	3	3 (100)	0	0

CD: Crohn's disease; WT: Wildtype. ¹Number of CD patients or controls;

²Results are expressed as number of CD patients (% of CD patients);

³NOD2 mutant was considered any subject that inherited at least one copy of the variant allele.

Table 5 Clinical characteristics of CD patients according to NOD2 genotype

Clinical features	n1	NOD2 genotype, n (%)					P2OR (95% CI)
		WT/WT	R702W/WT	G908R/WT	L1007fsinsC/WT	Heter.compound & homozygous
Age at diagnosis
< 40 yr (A1 + A2)	80	54 (55.67)3	13 (13.40)	3 (3.09)	4 (4.12)	6 (6.18)	0.57 1.56 (0.46-5.27)
> 40 yr (A3)	17	13 (13.40)	3 (3.09)	0	1 (1.03)	0
Location
Ileal (L1)	34	21 (21.65)	8 (8.25)	1 (1.03)	2 (2.06)	2 (2.06)	0.26 1.67 (0.69-4.06)
Colonic (L2)	23	20 (20.62)	0	1 (1.03)	1 (1.03)	1 (1.03)	0.04 0.26 (0.07-0.96)
Ileocolonic (L3)	40	26 (26.80)	8 (8.25)	1 (1.03)	2 (2.06)	3 (3.09)	0.5 1.38 (0.57-3.29)
Behavior
Non-stricturing, non-penetrating (B1)	37	24 (24.74)	8 (8.25)	3 (3.09)	2 (2.06)	0	0.5 1.37 (0.56-3.29)
Stricturing (B2)	41	34 (35.05)	5 (5.15)	0	0	2 (2.06)	0.01 0.29 (0.11-0.78)
Penetrating (B3)	19	9 (9.28)	3 (3.09)	0	3 (3.09)	4 (4.12)	0.02 3.22 (1.14-9.06)

CD: Crohn's disease; WT: Wildtype. ¹Number of CD patients in each subgroup; ²P-values, odds ratios and confidence intervals refer to the comparison of presence versus absence of the at least one mutant NOD2 allele;

³Results are expressed as number of CD patients (% of CD patients).

Table 6 Clinical characteristics of NOD2 WT/WT CD patients according to NOD1 genotype

Clinical features	n1	NOD2 WT/WT
		NOD1:	*1*1	*1*2	*2*2
Age at diagnosis
< 40 yr (A1 + A2)	54		62.962	35.18	1.85
> 40 yr (A3)	13		46.15	46.15	7.69
Location
Ileal (L1)	21		57.14	38.09	4.76
Colonic (L2)	20		70	25	5
Ileocolonic (L3)	26		53.84	46.15	0
Behavior
Non-stricturing,	24		62.5	33.33	4.16
non-penetrating (B1)
Stricturing (B2)	34		61.76	35.29	2.94
Penetrating (B3)	9		44.44	55.55	0
Controls	47		48.93	36.17	14.89

WT: Wildtype. ¹Number of CD patients or controls in each subgroup;

²Values are expressed as the percent of patients in each clinical subgroup.