ANCA and ASCA Profiles in a Moroccan Population With Inflammatory Bowel Diseases

Abstract

Introduction

Diagnosing inflammatory bowel disease (IBD) is hindered by the invasive procedures required for accurate classification as Crohn’s disease (CD) or ulcerative colitis (UC). As alternatives, non-invasive tests using anti-Saccharomyces cerevisiae antibodies (ASCA) and anti-neutrophil cytoplasmic antibodies (ANCA) have gained significance. This study evaluated ANCA and ASCA antibody frequencies in IBD and their role in disease characterization in a Moroccan population.

Methods

Conducted at Marrakech’s Mohammed VI University Hospital from 2014 to 2018, this cross-sectional study included patients with suggestive symptoms or confirmed IBD diagnosis based on clinical, endoscopic, and histological criteria. Immunological investigations detected p-ANCA, c-ANCA, and ASCA using immunofluorescence and immunodot assays.

Results

Among 60 participants (mean age: 33.1 ± 11.75 years), the 20–30-year age group was most affected (31.67%). CD, UC, and indeterminate colitis (IC) were diagnosed in 46.67%, 45%, and 8.33% patients, respectively. Gastrointestinal symptoms were prevalent (98.3%), with ANCA+/ASCA-profile in 41% of UC patients versus 11% in CD, and ANCA-/ASCA + profile exclusive to CD (50%). ANCA positivity was significantly associated with UC, rectal syndrome, and inflammatory syndrome, whereas ASCA positivity was significantly associated with CD and König’s syndrome (p < 0.05).

Conclusion

This study highlighted demographic and phenotypic particularities of IBD in a Moroccan population. Non-invasive tests using ASCA and ANCA antibodies offer valuable alternatives to invasive procedures, facilitating personalized management strategies. Variations in ANCA and ASCA profiles provide insights into disease characterization and inform tailored treatment approaches.

1. Introduction

Inflammatory bowel diseases (IBD), including Crohn’s disease (CD) and ulcerative colitis (UC), pose significant challenges in diagnosis and management. Despite recognized factors such as smoking and appendectomy influencing IBD risk and progression, the mechanisms underlying these associations remain elusive.1–5 Current diagnostic approaches rely on a combination of clinical, morphological, and histological criteria to accurately classify CD and patients with UC.6 However, the invasive nature of these diagnostic procedures underscores the need for non-invasive alternatives.

Among the myriad serum antibodies (Ab) proposed as diagnostic markers for CD or UC, anti-Saccharomyces cerevisiae antibodies (ASCA) and antineutrophil cytoplasmic antibodies (ANCA) have emerged as prominent candidates.7 These antibodies have potential not only for distinguishing CD from UC but also for refining diagnosis, characterizing disease phenotypes, and predicting disease course.8

Despite the existence of numerous antibody markers, the rationale for focusing on ANCA and ASCA lies in their widespread use and established relevance in clinical studies.9 By examining the ANCA/ASCA profile in patients with IBD, this study aims to fill a crucial gap in the literature by elucidating their frequency and clinical associations within a Moroccan population. This investigation seeks to provide insights into the diagnostic, prognostic, and therapeutic implications of ANCA and ASCA in IBD, thereby contributing to enhanced patient care and management strategies.

2. Methods

2.1. Study design

This descriptive and analytical cross-sectional study was designed to investigate the frequency of anti-neutrophil cytoplasm antibodies (ANCA) and anti-S. cerevisiae antibodies (ASCA) in patients with inflammatory bowel disease (IBD).

2.2. Study setting

The study was conducted at the immunology laboratory and gastroenterology department of Marrakech’s Mohammed VI University Hospital in Morocco, spanning the period from 2014 to 2018.

2.3. Study participants

Patients exhibiting suggestive clinical symptoms or those with confirmed diagnosis of IBD based on clinical, endoscopic, and histological criteria were enrolled in the study. Patients were admitted to the immunology laboratory for ANCA and ASCA antibody testing.

2.4. Outcome measures

The primary outcome of the study was to establish correlations between immunological results (ANCA and ASCA antibody status) and the clinical and paraclinical characteristics of patients with IBD.

2.5. Study instruments

Clinical data, including demographic characteristics and clinical manifestations, were collected from medical records. Immunological investigations were conducted to detect ANCA and ASCA antibodies using specific assays.

2.6. Immunological investigation

ANCA detection involved indirect immunofluorescence techniques (ANCA ethanol and formaldehyde, Bio-Rad), with a threshold of 1:20. Positive results were further analyzed for anti-myeloperoxidase (MPO) and anti-proteinase 3 (PR3) antigenic specificity using an immunodot assay (BlueDot ANCA, Aesku), with a threshold of 10 index. ASCA detection was performed using an immunodot assay (BlueDot ASCA IGG/IGA, Aesku), with a threshold set at 10 index.

2.7. Data collection

Clinical data were extracted from medical records, encompassing demographic characteristics and clinical manifestations. Symptoms and diagnostic criteria were assessed at the time of initial diagnosis or during the disease.

2.8. Sample size determination

A sample size of 60 patients was calculated for this study on the basis of statistical considerations.

2.9. Statistical analysis

Data analysis involved descriptive and inferential statistics, including Chi-square and cross-tabulation, conducted using the Statistical Package for Social Sciences (SPSS) version 23. A p-value of <0.05 was considered statistically significant.

Ethical approval

Data collection adhered to global ethical guidelines, ensuring confidentiality and protection of patient data. Ethical approval was obtained before the start of the study.

3. Results

The study included 60 patients, with a mean age of 33.1 ± 11.75 years (range: 16–64 years), showing a slight female predominance (sex-ratio M/F = 0.87). The most affected age groups were 20–30 years (31.67%, n = 19) and 30–40 years (30%, n = 18) (Table 1). Among these patients, 28 (46.67%), 27 (45%), and 5 (8.33%) were diagnosed with CD, ulcerative colitis (UC), and indeterminate colitis (IC), respectively.

Table 1.

Distribution of patients according to demographic and clinical characteristics.

Category of IBD	CD n = 28 (46.7%)	UC n = 27 (45%)	IC n = 5 (8.3%)	Total
Demographic characteristics
Mean age in years [ranges]	31.8 [16–52]	35.1 [18–64]	28 [18–38]	33.1 [6–64]
Male-to-female ratio	1.15	0.8	0.25	0.87
Clinical manifestations	n (%)	n (%)	n (%)	n (%)
General signs	25 (89.3)	18 (66.7)	5 (100)	48 (80)
Physical decline	21 (75)	11 (40.7)	3 (60)	35 (58.3)
Fever	3 (10.7)	1 (3.7)	0	4 (6.7)
Edema	1 (3.6)	0	0	1 (1.7)
Skin pallor	13 (46.4)	14 (52)	3 (60)	30 (50)
Digestive signs	27 (96.4)	27 (100)	5 (100)	59 (98.3)
Chronic diarrhea	26 (92.8)	24 (88.8)	3 (60)	53 (88.3)
Abdominal pain	20 (71.4)	14 (50)	4 (80)	38 (63.3)
Rectal syndrome	10 (35.7)	21 (77.7)	1 (20)	32 (53.3)
Koenig’s syndrome	12 (42.8)	2 (7.4)	1 (20)	15 (25)
Rectal bleeding	1 (3.6)	12 (44.4)	2 (40)	15 (25)
Anoperineal lesions	14 (50)	4 (14.8)	4 (80)	22 (36.7)
Extra-digestive signs	14 (50)	8 (29.6)	2 (40)	24 (40)
Arthritis	13 (46.4)	8 (29.6)	2 (40)	23 (38.3)
Erythema	1 (3.6)	3 (11.1)	1 (20)	5 (8.3)
Eye redness	4 (14.3)	0	0	4 (6.7)
Biological manifestations
Anemia (%)	11 (39.3)	13 (48.1)	2 (40)	26 (43.3)
Inflammatory syndrome (%)	17 (60.7)	17 (63)	4 (80)	38 (63.3)
Localization
Ileal (%)	3 (5)	0	0	3 (5)
Ileoceacal (%)	14 (23.3)	0	0	14 (23.3)
Colitis (%)	3 (5)	0	0	3 (5)
Left colitis (%)	0	5 (8.3)	0	5 (8.3)
Proctosigmoiditis (%)	1 (1.7)	2 (3.3)	0	3 (5)
Proctitis (%)	2 (3.3)	10 (16.7)	0	12 (20)
Pancolitis (%)	3 (5)	11 (18.3)	0	14 (23.3)

Gastrointestinal symptoms were predominant, observed in 98.3% of cases, followed by general symptoms (80%), joint signs (38.3%), ocular signs (66.7%), and cutaneous signs (5%) (Table 1).

3.1. ANCA and ASCA frequencies

Of the patients, 33.3% (n = 20) tested positive for ANCA, predominantly exhibiting a perinuclear pattern (60%, n = 12), and less frequently a cytoplasmic pattern (10%, n = 2). Among ANCA-positive cases, 20% (n = 4) had PR3-ANCA, and 15% (n = 3) had both MPO- and PR3-ANCA. ASCA was positive in 31.7% (n = 19) of patients, with predominance of the IgA (36.8%) and IgG isotypes (31.6%). ANCA positivity was significantly associated with UC (p = 0.0003), rectal syndrome (p = 0.018), and inflammatory syndrome (p = 0.02), whereas ASCA positivity was significantly associated with CD (p = 0.0002) and König’s syndrome (p = 0.037) (Table 2).

Table 2.

ANCA subtypes and ASCA isotypes according to the clinical category of IBD.

IBD categories	CD n = 28 (100%)	UC n = 27 (100%)	IC n = 5 (100%)
ANCA-positive	5 (17.8)	15 (55.6)	0
p-ANCA	1 (3.6)	11 (40.7)	0
c-ANCA	0	2 (7.4)	0
MPO specificity	0	0	0
PR3 specificity	0	4 (14.8)	0
Both MPO and PR3	1 (3.6)	2 (7.4)	0
Undetermined specificities	3 (10.7)	2 (7.4)	0
ASCA-positive	16 (57.1)	3 (11.1)	0
ASCA-IgA	6 (21.4)	1 (3.7)	0
ASCA-IgG	5 (17.8)	1 (3.7)	0
Undetermined specificities	5 (17.8)	1 (3.7)	0

4. ASCA and ANCA profiles

Regarding disease subtypes, the ANCA-positive/ASCA-negative profile was more prevalent in UC patients (41%) than in CD patients (11%), whereas the ANCA-negative/ASCA-positive profile was exclusively found in CD patients (50%). ANCA/ASCA double negative status was observed in all IC patients (Table 3).

Table 3.

IBD patients characteristics clustered according to ANCA and ASCA results.

	ANCA-positive		ASCA-positive
	n = 20 (33.3%)	P value	n = 19 (31.7%)	P value
Demographic characteristics
Male-to-female ratio	0.66	0.630	1.11	0.528
Clinical manifestations
Digestive signs
Abdominal pain	10 (26.3)	0.691	11 (28.9)	0.426
Rectal syndrome	15 (44.1)	0.018	8 (23.5)	0.121
Koenig’s syndrome	4 (26.7)	0.630	8 (53.3)	0.037
Rectal bleeding	6 (40)	0.423	3 (20)	0.329
Anoperineal lesions	8 (22.2)	0.738	17 (47.2)	0.727
Extra-digestive signs	10 (29.4)	0.517	11 (32.4)	0.473
Biological manifestations
Anemia	9 (34.6)	0.667	6 (23)	0.251
Inflammatory syndrome	8 (21)	0.020	11 (28.9)	0.551

Information regarding the confirmation of IBD diagnosis and the duration of the disease at study entry, as well as treatment regimens and the mean time to symptom control, were not specifically addressed in this study and would require additional investigation or data collection beyond the scope of this cross-sectional analysis.

5. Discussion and conclusion

Our study observed a noteworthy prevalence of inflammatory bowel diseases (IBD), predominantly Crohn’s disease (CD), and ulcerative colitis (UC) within our patient cohort. This observation aligns with the existing literature, which commonly reports a higher incidence of CD than that of UC.10–12 However, it is essential to note that few studies have shown a predominance of UC, highlighting the variability in disease distribution across different populations.13

The mean age of our patients (33.1 years) corresponds with that of previous studies indicating that IBD primarily affects young adults without a significant gender predilection.14,15 Manifestations of IBD, such as gastrointestinal symptoms and extra-digestive manifestations, were consistent with the literature, with diarrhea and abdominal pain being the most prevalent complaints.16–18

Studies have consistently demonstrated a high frequency of ASCA and ANCA antibodies in CD and UC, respectively, suggesting their potential diagnostic utility.19,20 Our findings support this finding, indicating a statistically significant association between ANCA/ASCA profiles and CD/UC diagnosis. Specifically, the ASCA-positive/pANCA-negative and pANCA-positive/ASCA-negative profiles were highly indicative of CD and UC, respectively.21

In our study, ANCA positivity was more common in patients with UC, whereas ASCA positivity was predominantly observed in patients with CD, consistent with the literature.22–25 These antibody profiles aid in disease differentiation and classification. However, it is crucial to recognize that although statistically significant associations were observed, the sensitivity and specificity of these antibody markers, which are the gold standard for IBD diagnosis, were not evaluated in this study, which warrants further investigation.

The association between sociodemographic factors and ANCA/ASCA positivity was not statistically significant in our study, which is consistent with previous findings.13,16,24,26 However, we observed a statistically significant association between ANCA/ASCA positivity and specific clinical manifestations, such as diarrhea and disease severity, which contradicts some previous studies.10,13,24 These discrepancies underscore the need for further research to elucidate the clinical implications of ANCA and ASCA in IBD.

6. Clinical value of ANCA and ASCA in IBD

In total, the clinical relevance of ANCA and ASCA in IBD lies in the following facts:

• - Joint research of ANCA and ASCA is useful to optimize the diagnosis of CD and UC, where the clinic is especially atypical28;

• - The existence of an ANCA-negative/ASCA-positive profile favors the diagnosis of CD, and conversely, an ANCA-positive/ASCA-negative profile favors the diagnosis of UC.28

• - These autoantibodies should minimize invasive investigations and are particularly useful in cases where endoscopic procedures are difficult or contraindicated.27

• - They can help detect certain severe forms with a more aggressive course of the disease, whether for CD or UC.29

However, the emergence of new biomarkers, such as calprotectin and lactoferrin, may impact the diagnostic utility of ANCA and ASCA in IBD diagnosis.28

In conclusion, our study provides valuable insights into the prevalence and clinical implications of ANCA and ASCA antibodies in IBD in the Moroccan population. Although our findings support the diagnostic value of these antibodies, further research with larger sample sizes and diverse populations is warranted to validate our results and better define the ANCA/ASCA profile in the entire IBD population. In addition, future studies should assess the sensitivity and specificity of these antibody markers in relation to the gold standard for IBD diagnosis to strengthen their clinical utility.

Data availability statement

The data used to support the findings of this study have been included in the manuscript.