The Center for Immunology and Transplantation
University of Florida, Gainesville, July 5th, 2005
type 1 diabetes and related ADs in
Sardinia
Marco Songini, Director
Diabetes Unit - Azienda Ospedaliera “G.Brotzu”
www.aob.it, marcosongini@aob.it
Cagliari – Sardinia, Italy
Cagliari
• Gene and diseases in the founder
population of Sardinia
Sardinia and the genetic map of Europe
According to Cavalli-Sforza et al. 1994
Different colours indicate genetic distances
Genes
Isolation
Genetic drift
Consanguinity
Stable population
• Uniformly
distributedposition
• Insularity
and geographical
Paleolitic: 700-1800
inhabitants
•3rd
Frequencies
unique
for Sardinia
About
300,000 inhabitants
fromKinship
800 b.C.
Phoenicians
and
• Physical
geography
(14.000 b.C.; about
750 b.C. to 17th century • 1880-1884 = 12.26%
century
Carthaginians
RH-; MNS-M;
DIA2, HLA B18
•
Low
population
density
generations ago)
• No real colonization in terms of
238etb.C.
Romans
•Cavalli-Sforza
1945 = 8.21%
al. (1994)
• Small number ofagent
founders
of demographic development
A.D.
Vandals
• Today =456
very
rare
large-scale
admixture between
A.D. Eastern Roman
• Same ancestral• No
gene
pool
Moroni et al534
(1972)
Language
populations
empire (Byzantium)
• Skewing effect
of
selection from costal to internal
regions(the most archaic
1258 A.D. Pisans
Life expectancy
Romance
• Considerable
drift
• The
invaders
not inclined
1323 A.D. Catalan-Aragoneses
The oldest man
in the
world were
neo-latin
language
spoken so far)
towards
permanent inhabitation
of 1479mobility
A.D. Spanish
lives currently
in
Sardinia
Matrimonial
• Early
Latinization
the island
1720 A.D. Italians – Piedmont
at
birth
1year
(average
distances
of birth place of
• Isolated
location
• Little genetic
flow from invaders
(Savoys)
the
spouses)
Stable
population
•1930-32
60.5 to innovation
•54.9
Resistance
• 1850
= 13.3 Km
Struggle
for survival
i.e. not to
•1960-62
69.4
71.8 mostly
Any
lexical innovation
regards some
Southern and Northern areas
• 1911-1949
become
extinct = 20 Km
•1970-72
72.9
73.9
24,000
sq. KmPaulis
(1995)
• 1950-1965
= 39.9 Km
• Famine
and malnutrition
•1989-93
73.9
73.6
popul. 1.654.470
• Nowadays
> 40 < 50
Km
• Endemic
illness (Malaria
and
(National Institute of Statistics)
Lat. 39°-41° N
other major
infections)
Gatti (1990)
Long.8°-10° E
GENETIC DISTANCE AMONG POPULATIONS
DAN
SVE
LAP
NOR
FINLANDIA
1.40
IRI
DUT
SCO
FMG
GER
1.14
AUS
SWI
0.88
CZE
MAR
PUG
0.62
TOS
FRE
HUN
CAL
0.36
0.10
UMB
PIE
CAM
SPA
SIC
-0.16
BAS
EMI
POR
LIG
-0.42
VEN
-0.68
1.42
1.16
0.90
0.64
0.38
0.12
-0.14
-0.40
-0.66
GRE
-0.92
-1.18
-1.44
-1.70
-1.96
-0.94 SARDEGNA
-1.20
LOM
LAZ
Neighbor-joining tree of 16 European and North-African populations using DRB1-DQA1DQB1 haplotype frequencies
(Courtesy of F. Cucca)
CRE
ITA
TUR
SPA
MOR
ALG
TUN
GB
NOR
US WHI TE
ROM
CZE
FRA
SARD
BULG
GRE
ALG, Algerians; BULG, Bulgarians; CRE, Cretan; CZE, Czech; FRA, French; GB, Britons; GRE, Greeks; ITA, Italians; MOR, Moroccan;
NOR, Norwegians; ROM, Romanians; SARD, Sardinians; SPA, Spanish; TUN, Tunic: TUR, Turks; US-WHITE, American Whites.
Neighbor-joining tree of 7 Sardinian sub-populations
and continental Italians using DRB1-DQA1-DQB1 haplotype
frequencies (Courtesy of F. Cucca)
ITALY
LANUSEI CAGLIARI
CARBONIA
TEMPIO
ORISTANO SASSARI
SORGONO
1
2
3
4
5
6
7
Cagliari
Carbonia
Lanusei
Oristano
Sassari
Sorgono
Tempio
Multi-dimensional scaling analysis carried out
on the DRB1-DQA1-DQB1 haplotype frequencies
(Courtesy of F. Cucca)
1.5
SEN
1
Second dimension
GRE
0.6
BULG
SARD
KOGI
MEX-AM
CRE
0.2
- 0.2
ROM
ITA
TUN
BRA BLACK
MOR
TUR
FRA
FIN
USA WHITE
ALG
UK
CZE
SPA
SAHA
SWE
NOR
USA BLACK
POLY
KAY
CHI
HONG KONG
- 0.6
JAP
S AFR
-1
- 1.5
-1
- 0.5
0
0.5
1
First dimension
1.5
2
2.5
3
0101-0101-0501
0102-0101-0501
0103-0101-0501
0301-0501-0201
0302-0401-0402
0401-0301-0301
0401-0301-0302
0402-0301-0301
0402-0301-0302
0403-0301-0302
0403-0301-0304
0403-0301-0305
0404-0301-0301
0404-0301-0302
0404-0301-0401
0405-0301-0201
0405-0301-0302
0405-0301-0401
0405-0501-0301
0406-0301-0302
0407-0301-0301
0407-0301-0302
0407-0301-0304
0408-0301-0301
0408-0301-0302
0409-0301-0401
0701-0201-0201
0701-0201-0303
0701-0301-0201
0801-0301-0302
0801-0401-0402
0802-0401-0402
0803-0103-0601
0803-0601-0301
0804-0401-0402
0804-0501-0301
0901-0301-0201
0901-0301-0303
1001-0101-0501
1101-0102-0602
1101-0501-0301
1102-0501-0301
1103-0501-0301
1104-0501-0301
1201-0101-0501
1201-0501-0301
1301-0103-0502
1301-0103-0603
1302-0102-0501
1302-0102-0604
1301-0102-0605
1303-0201-0201
1303-0501-0301
1305-0201-0201
1401-0101-0501
1401-0101-0503
1402-0501-0301
1403-0501-0301
1501-0102-0501
1501-0102-0602
1502-0101-0501
1502-0103-0601
1503-0101-0501
1503-0102-0602
1601-0102-0501
1601-0102-0502
1601-0102-0603
1602-0102-0502
1602-0501-0301
haplotype frequency (%)
25
20
15
10
5
25
20
15
10
5
25
20
15
10
5
25
20
15
10
5
25
20
15
10
5
0
DRB1* DQA1 DQB1* haplotypes in 5 ethnic groups
Chinese
Blacks
Latin American
Caucasians
Sardinians
30% of Sardinians
Lampis, Cucca et al. 2001
Genetic clusters for T1D in Sardinians
Genetic tree for the 21
linguistic domains in Sardinia
for 12 polymorphisms
Cappello et al. 1996
Genetic tree of the Sardinian
and Italian populations for HLA
Vona et al. 1997
Genetic diseases
G6PD deficiency (favism). Prevalence (%) among males
Silvetti et al. 1969
Genetic diseases
Beta thalassemia in Sardinia
Epidemiology of beta-thalassemia (95% beta39=founder effect):
• 13% beta-thalassemia carrier rate
• 1:70 couple is at risk for
• 1:250 newborns would be affected but they are reduced to
1:4,000 thanks to prenatal diagnosis
2%
1%
2%
95%
beta° 39
frame shift 76
Rosatelli et al. 1992; Cao et al. 1996
frameshift 6
Others
Genetic diseases
Wilson’s Disease
Incidence:
Sardinia
1-2:10,000
Other countries
1-2:100,000
Heterozygous in Sardinia: 3.4%
Heterozygous in other countries: 1.1%
Loudianos et al 1999
Genetic diseases
Wilson’s Disease
mutations in Sardinian population (founder effect)
6 mutations
15%
others
(>200)
-441del15
5 others
85%
40%
60%
Loudianos et al. 1999
• Other diabetes-related autoimmune
diseases in Sardinia
Other autoimmune diseases
APECED (APS1)
Autoimmune Polyendocrinopathy Candidiasis Ectodermal Dystrophy
Autosomal recessive disorder
Incidence:
Finns
1:25,000
Iranian Jews
1:9,000
Sardinians
1:14,400
Genetic:
Mutation of AIRE (autoimmune regulator) gene: R139X (18/20
indipendent mutations found are R139X= founder effect)
Heterozygous in Sardinia for R139X: 1.7%
Rosatelli et al. 1998
Other autoimmune diseases
Celiac disease (CD)* in background population
Population
Number
Prev.
North Sardinia
Schoolchildren
1,607
1.06%
(0.6-1.6)
Italy - Marche
Schoolchildren
3,351
0.33%
(0.1-0.5)
Catassi et al. 1994
1.1%
(0.4-2.3)
Saukkonen et al 2001
(0.3-1.5)
Kolho et al. 1998
Finnish sibs
of T1D (mean
550
(95% C.I. )
Ref.
Meloni et al. 1999
age 12yrs)
Finnish adults
1,070
0.8%
* biopsy proven
Other autoimmune diseases
CD* among T1D patients
Population
Number
Prev.
(95% C.I. )
NorhtSardinian
Schoolchildren
650
4.1%
(2.7-6.0)
Frongia et al. (unpubl.)
T1D Italian
Schoolchildren
4,514
498
2.7%
3.2%
(2.2-3.1)
(1.8-5.2)
Pocecco et al. 1989
Sardinian adults
T1D patients
394
3.8%
(2.1-6.2)
Songini et al. 1998
Italian adults
T1D patients
383
820
2.6%
7.0%
(1.3-4.7)
(5.4-9.0)
* biopsy proven
Ref.
Barera et al. 1991
Sategna-Guidetti et al. 1994
De Vitis et al. 1996
Other autoimmune diseases
Myasthenia gravis (MG) in background adult populations
Population
Number
Prev*.
Incid*.
North Sardinia
270.000
11.1
0.8
Aiello et al.1997
Whole Sardinia
Denmark
1.5mil
2.8mil
4.5
7.8
0.25
0.5
Giagheddu et al. 1995
9
0.24
Whole Norway
4.5mil
Italian adults BO 815.000
FE 381.118
4.9
9.7
* x100.000
Ref.
Christensen et al. 1993
Storm-Mathisen et al. 1984
D’Alessandro et al. 1991
Other autoimmune diseases
Thyroid autoantibodies (TA) in background population
Population
Number
Prev.
Sardinian
Schoolchildren
8,484
3.7%
(3.3-4.1)
Mariotti et al. 2000
Italian
Schoolchildren
419
142
2.4%
4.3%
(1.1-4.3)
(1.6-9.0)
Aghini et al. 1999
Sardinian mothers
at delivery
2,500
11.8%
(10.5-13.1)
Olivieri et al. 2000
Mothers at
delivery (Japan, USA)
4,022
552
5.5%
19.6%
(95% C.I. ) Ref.
Fenzi et al. 1986
(4.8-6.2) Sakaihara et al. 2000
(18.1-21.1) Stagnaro et al.1990
Other autoimmune diseases
Thyroid autoantibodies among T1D patients
Population
Number
Prev. (95% C.I. )
Ref.
T1D Sardinian
Schoolchildren
650
17.6%
Ricciardi et al.
T1D Italy/Europe
Schoolchildren
212
495
16.5%
22.0%
(11.5-21.5)
(18.4-25.7)
T1D Sardinian
adults patients
399
28.3%
(23.9-22.6)
T1D European
adults patients
258
111
19.8%
27.9%
(14.9-24.6)
(19.6-36.3)
(14.6-20.5)
(unpubl.)
Lorini et al. 1996
Holl et al. 1999
Songini et al. 1998
Mangendre et al. 2000
Fernandez-Castaner et al,
1999
Sardinia: Giants’ tomb
The new concept of T1D
as a cluster of autoimmune-related
diseases:
the example of MS and T1D in Sardinia
70
93
280
75
96
165
140
55
100
25
40
65
53
17
90
120
50
110
40
50
80
35
60
13
43
85
60
60
70
150
40
80
26
120
30
30
10
90
Prevalence of MS x 100.000 in Europe (MS pts= 2.5 mil ww)
J Cl Nurology and Neurosurgery 104 (002) 182-91
Other autoimmune diseases
Multiple Sclerosis (MS) and T1D
Population
Number
MS (Prev)
(95% C.I. )
Reference
Sardinians
270,000
0.16 %
(0.14-0.17)
Granieri et al. 2000
(NW,all ages)
0.05 %
Italians (all ages)
Sardinian T1D
adults (15-50 yrs)
Sardinian T1D
424
650
Granieri et al. 1997
0.47%
(0.05-1.69)
Songini et al. (unpubl)
0.77%
(0.25-1.79)
Chessa et al. (unpubl)
schoolchildren
Population
Sardinian
Number
T1D (Prev)
(95% C.I. )
Reference
1,053
2.40%
(1.54-3.49)
Marrosu et al. 2000
6,255
0.37%
(0.22-0.52)
Muntoni et al. 1988
MS pts (9-65 yrs)
Sardinian pop
(20-59yr)
Other autoimmune diseases
Genetic association of MS with:
In Northern Europeans: DRB1*1501-DQB1*0602 (DR2)*
*Protective for T1D then T1D and MS
occur together very rarely
In Sardinians:
DRB1*0301-DQB1*0201 (DR3)*
DRB1*0405-DQB1*0301 (DR4)*
*Also predisposing to T1D, DR2 is very
rare then T1D and MS may coexist
Marrosu et al. 2000
Multiple Sclerosis (MS) and T1D in
Sardinia
Population
Number
MS (Prev)
(95% C.I. )
Reference
Sardinians
270,000
0.16 %
(0.14-0.17)
Granieri et al. 2000
(NW,all ages)
0.05 %
Italians (all ages)
Sardinian T1D
adults (15-50 yrs)
Sardinian T1D
424
650
Granieri et al. 1997
0.47%
(0.05-1.69)
Songini et al. (unpubl)
0.77%
(0.25-1.79)
Chessa et al. (unpubl)
schoolchildren
Population
Sardinian
Number
T1D (Prev)
(95% C.I. )
Reference
1,053
2.40%
(1.54-3.49)
Marrosu et al. 2000
6,255
0.37%
(0.22-0.52)
Muntoni et al. 1988
MS pts (9-65 yrs)
Sardinian pop
(20-59yr)
Genetic association of MS with T1D
In Northern Europeans: DRB1*1501-DQB1*0602 (DR2)*
*Protective for T1D then T1D and MS
occur together very rarely
In Sardinians:
DRB1*0301-DQB1*0201 (DR3)*
DRB1*0405-DQB1*0301 (DR4)*
*Also predisposing to T1D, DR2 is very
rare then T1D and MS may coexist
Marrosu et al. 2000
MS with T1D:
together at last (Allegheny County, US)
T1D
Siblings†
Parents†
Characteristics
Males
(n=148)
Females
(n=149)
Males
(n=143)
Females
(n=187)
Males
(n=84)
Females
(n=140)
Age (yrs)
41.4
42.6
42.4
42.3
69.0
67.2
Duration T1D (yrs)
35.2
35.4
----
----
----
----
Observed MS prev.
0%
2.0%
0%
0.5%
0%
0%
Expected MS prev.
n/a
0.1%
n/a
0.1%
n/a
n/a
Relative increased
risk of MS
n/a
20fold*
n/a
5 fold
n/a
n/a
Dorman JS, Songini M et al. Diabetes Care,november 2003
*p<0.01
† unaffected with T1D
n/a non applicable
Mediterranean sea
A
SASSARI
Corse (F)
20%
Tuscany
15%
NUORO
Gallura
20%
2
CAGLIARI
1
4
3
22
7 9
6
5
8
11
10
12
13
14
15
Barbagia
50%
16
17 18
19
20
MS prevalence
gradient
Gallurese-speaking
communes
23
21
B
The two Sardinian provinces under
study: Sassari and Nuoro; the southwestern to north-eastern gradient of MS
prevalence in the province of Sassari
(Pugliatti et al., 2002b), correspond to
significant differences of genetic and
linguistic domains (prevalence is
expressed as number of cases per
100,000 population for each commune
of the province: black communes >150,
grey communes >130 and <150, blank
communes < 130 MS cases). B.
Polymorphisms of mitochondrial genes
in Barbagia (central, “archaic” Sardinia),
Gallura (norther Sardinia), Corse
(insular France) and Tuscany (mainland
Italy) (Morelli et al., 2000) and
subdivision of the island in 23
linguistically and genetically domains
(Cappello et al., 1996);
Figure 5.
MS prevalence gradient
(total population)
<130
140-150
>150
Figure 6.
MS prevalence gradient
(0-15 years population)
<190
190-200
>200
A
rate per 100,000
6
Nuoro
Sassari
Ferrara
4
2
0
1965-74
1975-84
1985-93
Figure 4.
A. Temporal trends of MS incidence rates in the province of Nuoro, Sassari and
Ferrara, in the period 1965 to 1993.
Mean population (x 1000)
Figure 1
12
12
10
10
8
8
6
6
4
4
2
2
191216
194246
194751
195256
195761
196266
Five-years periods
196771
197276
197781
MS incidence rate (100,000)
1) Average annual incidence rates
(per 100,000 per year, right Y axis)
of MS in Macomer, Sardinia, from
1912 through 1981 (5-years
periods). Left Y axis expresses the
population growth (per 1,000).
Adapted from reference [12].
25
20
15
10
5
0
Caucasians
MS and T1DM
Lampis,R et al. 2001
Protetctive
T1DM
1501-0102-0602
25
20
15
10
5
1502-0101-0501
1502-0103-0601
1503-0101-0501
1503-0102-0602
1601-0102-0501
1601-0102-0502
1601-0102-0603
1602-0102-0502
1602-0501-0301
25
20
15
10
5
0406-0301-0302
0407-0301-0301
0407-0301-0302
0407-0301-0304
0408-0301-0301
0408-03010409-03010302
0701-0201-0201
0401
0701-0201-0303
0701-0301-0201
0801-0301-0302
0801-0401-0402
0802-0401-0402
0803-0103-0601
0803-0601-0301
0804-0401-0402
0804-0501-0301
0901-0301-0201
0901-0301-0303
1001-0101-0501
1101-0102-0602
1101-0501-0301
1102-0501-0301
1103-0501-0301
1104-0501-0301
1201-0101-0501
1201-0501-0301
1301-0103-0502
1301-0103-0603
1302-0102-0501
1302-0102-0604
1301-0102-0605
1303-0201-0201
1303-0501-0301
1305-0201-0201
1401-0101-0501
1401-0101-0503
1402-0501-0301
1403-0501-0301
1501-0102-0501
20
15
10
5
0405-0501-0301
25
20
15
10
5
0302-0401-0402
0401-0301-0301
0401-0301-0302
0402-0301-0301
0402-0301-0302
0403-0301-0302
0403-0301-0304
0403-0301-0305
0404-0301-0301
0404-0301-0302
0404-0301-0401
0405-0301-0201
0405-0301-0302
0405-0301-0401
0301-0501-0201
0101-0101-0501
0102-0101-0501
0103-0101-0501
haplotype frequency (%)
25
DRB1* DQA1 DQB1* haplotypes in 5 ethnic groups
Chinese
Blacks
Latin American
MS
MS
Sardinians
HLA allele
T1DM effect
MS effect
DRB1*1501 (DR2) –
DQA1 0102- DQB1 0602
Dominant
protection
Predisposition
in Caucasians
DRB1* 0301(DR3) –
DQA1 0501 - DQB1 0201-
Predisposition
Predisposition
in Sardinians
other loci outside the MHC region may be
shared between MS and T1DM and contribute
to the susceptibility to both the diseases.
The strong effect of having multiple MS
affected relatives suggests that
genetic variability in risk for diabetes
is linked to genetic loading for MS,
which is substantially increased in
these individuals.
These findings suggest that loci or genes
predisposing to familial MS aggregation
also contain an enrichment of diabetes
predisposing genes.
Taken together, these data suggest:
that common genes, either within the HLA
region or outside it, influences susceptibility to
both MS and T1DM
that genetic structure of Sardinians might
contain a particular combination of etiologic or a
lack of protective genes for these diseases.
Thus, Sardinians appears to be a very suitable
population for studying what genes contribute
to trigger and sustain autoimmunity in humans.
Conclusions…
To sum up (1)
• T1D has started to rise around the middle of the 20th
century and keeps increasing ww in younger ages mainly in
low risk countries (catch-up phenomenon)
• Whether it’s an earlier presentation or a true increase it’s
not clear yet
• Sardinia is confirmed to be a hot spot for clinical T1D and
islet-related autoimmunity, as for some other autoimmunerelated diseases (CD, MS, not TD)
• An increasing T1D incidence in the Island has been found
rising dramatically since the years 60’ as in other countries
• No defined geographical clustering for T1D was found
across the island
To sum up (2)
• The combination of more than 1 islet-related autoantibody
(rather than which) is the best predictor for the
development of T1D in the Sardinian background
population (SC)
• None of the classical ecological variables considered so far
has shown any firm influence towards the etiopathogenesis
of T1D in Sardinia; however other variables need to be
further investigated
• Migrants data seems to support a different role of genetic
vs environmental determinants
• There’s still a long way in front of us to possibly prevent
type 1 diabetes…
What we are doing now with the
help of so many (1)
• Trying to further improve the prediction of T1D in the
general population by studying the immunological and
genetic markers for T1D and other autoimmune
diseases in Sardinian population and migrants and their
relatives
• Investigating other putative environmental factors which
can play a role towards the etiopathogenesis of T1D
(e.g., chemicals, toxins, vaccinations, viral infections,
etc.)
What we are doing now... (2)
• Broadening the original investigation for T1D on the
prevalence of others autoimmune diseases and to
study their associations
• Comparing data from Sardinia and other areas by new
collaborative studies(RIDI,TRIGR,ENDIT, SARDINIAN
DIABFIN)
• Sensitizing Sardinian Health Autorities & population
towards primary prevention of T1D and autoimmune
diseases …..and by this way...
• improving the current care of diabetes in Sardinia!
Acknoledgements
Anthropology:
G. Vona (Sardinia-Italy)
Other autoimmune diseases:
GF Bottazzo and coworkers (Rome-Italy)
S. Mariotti and coworkers (Cagliari-Italy)
T. Meloni and coworkers (Sardinia-Italy)
M. Marrosu and coworkers (Sardinia-Italy)
Epidemiology:
The Sardinian Epidemiology Study Group
Eurodiab TIGER Study Group
IDA Study Group
RIDI Study Group
Z. Laron, I. Askenazi (Israel)
L. Bernardinelli and C. Pascutto (Pavia-Italy)
P. Contu, L. Minerba (Cagliari-Italy)
Genetic:
F. Cucca and coworkers (Sardinia-Italy)
E.A.M. Gale and coworkers (UK)
I. Loudianas (Sardinia-Italy)
Acknoledgements
Echological studies:
G.F. Bottazzo (Rome-Italy)
M. Fadda (Sardinia-Italy)
B. Elliot (New Zeland)
P. Pozzilli (Rome-Italy)
A. Contu, M. Carlini (Sardinia-Italy)
Prediction Study: G.F. Bottazzo & M. Locatelli (Rome- Italy)
S. Carta, M. Sorcini, A. Olivieri (Rome-Italy)
GF Bottazzo, A. Loviselli and the Sardinian SchooIchildren Study
Group (Sardinia-Italy)
GF Bottazzo,R. Cirillo and the Sardinian Newborn Study Group
(Sardinia-Italy)
A. Dolei, G. Delitala (Sardinia-Italy)
M.T. Tenconi, G. Devoti (Pavia-Italy)
The League of Sardinian Migrant Associations
ASRIS (Association for Study and Research of IDDM in Sardinia):
M. Porceddu (secretary)
Tina Carboni
President: G.F. Bottazzo
VicePresident: M.Songini
C. Putzu, A. Casu
R. Cavallo
R.A.S. (Regione Autonoma della Sardegna)
Italian Ministry of Healh, ADCT
GPs&Pediatricians from Sardinia….and many more!
European Union
National Institute of Health
My Diabetes staff thanks you