Desensitising Celiac Disease Patients With the Human Hookworm

Biopsies were fixed in neutral buffered formalin, processed and carefully orientated and embedded in paraffin wax. Sections (3 µm) were stained with H&E. Slides from both time-points were de-identified, shuffled and graded by Dr John Croese after which results from poorly orientated slides were verified by Dr Andrew Clouston. The Vh:Cd ratios were measured on 5 randomly selected well-orientated sites. The null hypothesis is that hookworm infection will not protect against mucosal damage following 12-week exposure to gluten in celiac disease.

Biopsies were fixed in neutral buffered formalin, processed and carefully orientated and embedded in paraffin wax. Sections (3 µm) were stained with anti-CD3. All slides were de-identified and graded by Dr John Croese. The IEL percentages were measured on 2 or more randomly selected well-orientated villi. The null hypothesis is that hookworm infection will not protect against mucosal IEL influx following 12-week exposure to gluten in celiac disease.

The Marsh score is a defined but qualitative assessment assigned a value to allow for comparison. The scores were evaluated by consensus between the primary (chief) investigator and the study pathologist. The Marsh score was graded 0, 1, 2, 3A (assigned-4), 3B (-5) and 3C (-6); rage 1-6 with normal=0 and severe inflammation=6. Because the scoring is vulnerable to artefact, only a 2-point shift was regarded as a significant intra-individual change. The scores were graded after week-36 on biopsies de-identified shuffled. An upward shift was interpreted to reflect a significant worsening of gluten-associated inflammation. The comparison reported evaluated changes from baseline (week-24) to post-low-dose gluten challenge (week-24; GC-1g). The objective for using the Marsh score was to identify individuals who might have experienced a severe worsening in pathology due to GC-1g that might not be reflected in the Vh:Cd group analysis.

The trial was extended with pre-trial and mid-trial anti-tTG antibody levels used to compare with the post-trial levels. Anti-tTG is a serological measure of tissue transglutaminase-2 antibodies. In active celiac disease, levels are increased. In treated disease, levels are low (normal cut-off was <15 IU/mL). A significant increase compared to baseline in tTG can be expected 2 weeks after consuming 3g of gluten daily for 2 weeks in people with celiac disease who have been maintaining a gluten-free diet, but who are not taking other treatment.