Relationship between methylation and colonic inflammation in inflammatory bowel disease

Table 1 Patient characteristics n (%)

Patient characteristics	Value
Number of patients	38
Patients with ulcerative colitis (UC)/Crohn’s colitis (CC)	29 (76)/9 (24)
Men	23 (58)
Age (yr): mean ± SD	55.24 ± 14.11
Disease duration (yr): mean ± SD	17.82 ± 8.29
Number of colonoscopies	62
Number of colonoscopies in UC/CC	49 (79)/13 (21)
Disease extension1
Left colitis (UC) or segmentary colitis (CC)	20 (33)
Extensive colitis (UC and CC)	42 (68)
Medication at endoscopy1
Oral 5-ASA	40 (65)
IMM (AZA/6MP)	18 (29)/3 (5)
TNF inhibitor (IFX/ADA)	2 (3)/2 (3)
CRP (mg/dL): mean ± SD	8.25 ± 16.56
Fecal calprotectin (μg/g): mean ± SD	306.5 ± 690.1
Reason for colonoscopy
Endoscopic activity assessment	5 (8)
Dysplasia surveillance	57 (92)
Clinical activity
Active disease	5 (8)
Symptomatic remission	57 (92)
Endoscopic activity
Endoscopic remission	24 (39)
Endoscopic activity	38 (61)
Histological activity and presence of dysplasia
Dysplasia	8 (13)
Acute histological activity	23 (37)
Chronic histological activity	24 (39)
No histological activity	7 (11)
Methylation status
SLIT2 (patients with any methylated sample)	40 (65)
TGFB2 (patients with any methylated sample)	23 (42)

¹As therapy regimens overlapped, the total is 104.7%. ASA: Aminosalicylic acid; CS: Corticosteroids; IMM: Immunomodulators; AZA: Azathioprine; 6MP: 6 mercaptopurine; MTX: Methotrexate; TNF: Tumor necrosis factor; IFX: Infliximab; ADA: Adalimumab.

Table 2 Relationship between methylation status and disease activity n (%)

Relationship	SLIT2 methylation (yes/no)	TFGB2 methylation (yes/no)
Endoscopic activity1
Activity	38/31 (55)	22/41 (35)
Remission	28/129 (18)	18/127 (12)
P value	< 0.001	< 0.001
Histological activity
Acute	26/20 (57)	7/29 (20)
Chronic	25/80 (24)	23/82 (22)
No activity	10/55 (15)	5/52 (9)
P value	< 0.001	NS
Clinical activity2
Active	5/0 (100)	2/2 (50)
Remission	35/22 (61)	21/30 (41)
P value	0.151	NS
Biological activity
Active (CRP ≥ 5 mg/L + FC ≥ 250 μg/g)	7/0 (100)	5/2 (71)
Remission (CRP < 5 mg/L + FC < 250 μg/g)	12/7 (63)	4/13 (24)
P value	0.134	0.061

¹Endoscopic activity was defined as Mayo > 0 for ulcerative colitis (UC) or Crohn’s disease (CD) activity index of severity (CDEIS) > 0 for CD. Endoscopic remission was defined as Mayo = 0 for UC or CDEIS = 0 for CD;

²Clinical activity, for UC patients symptomatic remission was defined as a Mayo stool frequency subscore of 0 or 1 and a Mayo rectal bleeding subscore of 0; and for CD patients as CD activity index < 150. CRP: C-reactive protein; FC: Faecal calprotectin; NS: Not significant.

Table 3 Methylation status and disease activity according to disease location

Variable (% samples)	Distal colon	Proximal colon	Pvalue
Endoscopic activity1	48/101 (48)	21/121 (17)	< 0.001
Active histological activity2	30/97 (31)	16/115 (13)	0.004
Methylation of SLIT2	43/101 (43)	23/121 (19)	< 0.001
Methylation of TGFB2	24/90 (27)	16/117 (14)	0.022

¹Endoscopic activity was defined as Mayo > 0 for ulcerative colitis or Crohn’s disease activity index of severity > 0 for Crohn’s disease;

²Active histological activity was defined as the presence of neutrophils in conjunction with epithelial cell damage.

Table 4 Histological activity and disease location as predictors of SLIT2 and TGFB2 methylation status

Multivariate analysis	Distal location	Acute histological activity1
SLIT2
Coefficient (β)	0.95	1.45
OR (95%CI)	2.57 (1.34-4.99)	4.25 (2.07-8.72)
P value	0.005	< 0.001
TGFB2
Coefficient (β)	0.95	0.002
OR (95%CI)	2.59 (1.21-5.54)	1.01 (0.39-2.57)
P value	0.014	0.996

¹Acute histological activity was defined as the presence of neutrophils in conjunction with epithelial cell damage.

Table 5 Changes in inflammatory status and SLIT2 methylation from colonoscopy 1 to colonoscopy 2

Changes in methylation of SLIT2	Changes in endoscopic inflammation
		Stable (n = 50)		Change (n = 16)
		ER→ER	Active→active	ER→active	Active→ER	Total
Stable	UM → UM	25	6	2	1	34
(n = 44)	M → M	2	5	0	3	10
Change	UM → M	2	2	7	0	11
(n = 22)	M → UM	7	1	3	0	11
Total		36	14	12	4	66

ER: Endoscopic remission; Active: Endoscopic activity; UM: Unmethylated; M: Methylated.