Celiac disease
DQ2 or DQ8 HLA variants are necessary for onset and development
of celiac disease
Typical symptoms of CD:
 diarrhea
 abdominal distention
 delayed growth
High risk for the disease:
 first degree of patients with CD (up to 12-15%)
 others disorders: -type 1 diabetes
-Down syndrome
-deficit IgA
MALATTIA CELIACA
GEMELLI MONOZIGOTI
GEMELLI DIZIGOTI
80 %
20 %
HLA TYPING MAY BE USEFUL
a negative test for HLA DQ2-DQ8 has a high negative predictive
value wich may have a role in those with borderline histology or
serology
a positive family history is another indication where HLA typing
may be useful. Those who are HLA compatible for CD need follow up
also if are initially negative, whereas those are HLA DQ2 and DQ8
negative may be discharged.
wheat
GLUTEN
-gliadins
-gliadins
-gliadins
Glutenins
+
α
γ
ω
RUN
Gliadin structure
α-gliadin
290 aa
C:
N
/translation="MVRVPVPQLQPQNPSQQQPQEQVPLVQQQQFPGQQQPFPPQQPY
PQPQPFPSQQPYLQLQPFPQPQLPYPQPQLPYPQPQLPYPQPQPFRPQQPYPQSQPQY
SQPQQPISQQQQQQQQQQQQKQQQQQQQQILQQILQQQLIPCRDVVLQQHSIAYGSSQ
VLQQSTYQLVQQLCCQQLWQIPEQSRCQAIHNVVHAIILHQQQQQQQQQQQQPLSQVS
FQQPQQQYPSGQGSFQPSQQNPQAQGSVQPQQLPQFEEIRNLALETLPAMCNVYIPPY
CTIAPVGIFGTNYR
- gliadin
product="alpha-gliadin"
ORIGIN
1
61
121
181
241
301
361
421
481
541
601
661
721
781
841
atggttagag
gagcaagttc
caacagccat
ccatttccgc
ctaccatatc
cagtattcgc
caacaaaaac
ctaattccat
gttttgcaac
atccccgagc
caacagcaac
cagcctcaac
caggcccagg
gcgctagaga
ccagttggca
ttccagtgcc
cattggtaca
atccgcagcc
agccgcaact
cgcagccgca
aaccacaaca
aacaacaaca
gcagggatgt
aaagtactta
agtcgcggtg
aacaacaaca
aacaatatcc
gctctgtcca
cgctacctgc
tcttcggtac
acaattgcag
acaacaacaa
gcaaccattt
accatatccg
accatttcga
accaatttcg
acaacaacaa
tgtattgcaa
ccagctggtg
ccaagccatc
acaacaacaa
atcaggccag
gcctcaacaa
aatgtgcaat
taactatcga
ccacaaaatc
tttccagggc
ccatcacaac
cagccgcaac
ccacaacaac
cagcagcagc
cagatccttc
caacacagca
caacaattgt
cacaatgttg
caacaaccgt
ggctccttcc
ctgccccagt
gtctatatcc
tga
catctcagca
agcaacaacc
aaccatatct
taccatatcc
catatccaca
agcagcagca
aacaaatttt
tagcgtatgg
gttgtcagca
ttcatgctat
tgagccaggt
agccatctca
ttgaggaaat
ctccatattg
acaaccacaa
atttccacca
gcagctgcaa
gcagccgcaa
atcgcaacca
acaacaacaa
gcaacaacaa
aagctcacaa
gctgtggcag
tattctgcat
ctccttccaa
gcaaaaccca
aaggaaccta
caccattgct
33-mer large peptide
LQLQPFPQPQLPYPQPQLPYPQPQLPYPQPQPF
1
LQLQPFPQPQLPYPQPQLPYPQPQLPYPQPQPF
1
3
2
N
- gliadin
The trimolecular complex
Peptide
 1-domain
1-domain
HLA
 2-domain
2-domain
Ig homology
folds
intestinal lumen
toxic gluten (33 mer)
lamina propria
MICA (MHC class I- related chain A)
NKG2D
intestinal lumen
toxic gluten
Il-15
O
C
H2N
NH2
O
O-
C
CH2
CH2
CH2
CH2
C
C
H
Gln
COOH
H2N
H
Glu
COOH
Linfocita T4 helper maturo
Secrezione di
Lymphocytes B
Linfocita T4
helper
Secrezione di IL1 da parte del macrofago
IL2
Proliferazione dei linfociti B attivati
plasmacellule
SEROLOGICAL TESTS
Tissue transglutaminase antibody [ELISA]
Endomysial antibody [Indirect immunofluorescence]
Antibodies against deamidated gliadin [ELISA]
Schermo televisivo
Endoscopio
Telecamera
Duodenal biopsy is necessary alwais?
The advent of highly sensitive and specific serological markers has led
to some protagonist proposing that celiac disase can be diagnosed without
the need for a biopsy.
However, this is an area of controversy.
Sequence
Specific Primers
SSP
Match esatto → Amplificazione
(Allele specifico)
Mismatch → Nessuna Amplificazione
(Allele non specifico)
GENOTYPES
DQ2 → DQA1*0501 - DQB1*0201
DQ8 → DQA1*03 - DQB1*0302
Chromosome 6
Genomic sequence: 6,295 bases
Aminoacids: 232
HLA SYSTEM
Class II
DP
DQ
Class III
Class I
DR
B C
E
B1 A1
tel
cent
C4A
TNXA
TNXB
C4B
gene espresso
RP1
gene non espresso
RP2
CYP21P
CYP21
30 kb
A
DQA1 gene map
5’
3’
IVS1 (4kb)
1
2
3
4
chain 
chain 
1
1
NH2
N2H
2
COOH
2
COOH
There are 27 DQA1 nucleotide sequences currently reported in the HLA sequence database
http://www.ebi.ac.uk/imgt/hla
Specific PCR amplifications
group 1 → DQA1*01
group 2 → DQA1*02, *04, *05
group 3 → DQA1*03
Sequencing-based typing (SBT)
Chromosome 6
DQB1 gene
Genomic sequence: 7,347 bases
Aminoacids: 261
Chr6
Classe II
DP
DQ
Classe III
Classe I
DR
B C
E
B1 A1
tel
cent
C4A
gene espresso
gene non espresso
RP1
TNXA
RP2
CYP21P
TNXB
C4B
CYP21
30 kb
A
DQB1 gene
Seq. Leader
1-domain
5
5’
1
1,5kb
2- domain
transmembrane and tail
95
3’
2,5kb
2
chain 
3
chain 
1
1
NH2
N2H
2
COOH
2
COOH
4
5
NEW THERAPEUTIC APPROACH
111
endopeptidase
Flavobacterium meningosepticum
pH
pH
pH
pepsin
C:
Bacterial prolyl-endopeptidase from Flavobacterium meningosepticum removes gluten toxicity by cleaving it into small fragments
Gli enzimi prolyl oligopeptidase hanno l’ottimo di pH tra 7 e 8 e così non possono funzionare a pH acido dello stomaco; inoltre esse sono efficientemente distrutte
dalla pepsina secreta dallo stomaco. Queste proprietà implicano che la supplementazione orale con prolil oligopeptidasi non è ancora sufficiente a degradare il glutine
prima che arrivi nella parte prossimale del duodeno.
Aspergillus niger prolyl-endopeptidase
pH
pH
compatible
Resistant
to degradation
pH
by pepsin
C:
The prolyl-endoprotease from Aspergillus niger (AN-PEP) is a member of the serine peptidase family, and this degrades gluten peptides
rapidly. This AN-PEP is capable of accelerating the degradation of gluten in a gastrointestinal model that closely mimics in-vivo
digestion. The pH optimum of the enzyme is compatible with that found in the stomach, and the enzyme is resistant to degradation by
pepsin.
Grazie per l’attenzione
Piero Sammarco