1. Marco Songini on behalf of the Sardinian IDDM Study Groups The Type 1 Diabetes Sardinia (Hot&Cold Spot) Project: what did we learn so far?

2. Dr. Marco Songini is the director of the Diabetes Unit at the S. Michele Hospital in Cagliari (Sardinia- Italy) He is also the vice-president of ASRIS (Association for the Study of Type 1 Diabetes in Sardinia)

3. The island of Sardina has the second highest incidence of type 1 diabetes in the world and a high prevalence of other autoimmune diseases (celiac disease, thyroid autoimmune diseases) has been reported. The type 1 diabetes prevalence has rapidly increased after the second world war. The island also offers a relatively small population with a homogenous genetic background selected by centuries of isolation.

4. Immune markes of type 1 diabetes (ICA, GADA, IA2, IAA) appear several years before the clinical onset of the disease, as was seen in family members of diabetic patients, but relatively little is know about the prevalence and prognosis of such markers in the general population. The aim of the Hot and Cold Spot Project is to investigate the prevalence of immune markers in Sardinian population and to evaluate their prognostic significance to develop a screening procedure.

5. History of the Hot&Cold Spot Project (1) Eurodiab - Ace: Sardinia is an ‘hot spot’ for Type 1 diabetes in Europe first recruitment of newborn from the general population: launch of the Sardinian Newborn IDDM study (SNI) ICA assay in 1,800 serum samples of school children from the general population enrolled for an epidemiological study of goitre prevalence in Sardinia; first milestone of the Sardinian School children IDDM Study (SSI) the Military Service approach (secular trend of type 1 diabetes among Sardinians) ending of the recruitment of the SSI (total cohort  10,000 children) 1995 1994 1993 1990 1989

6. History of the Hot&Cold Spot Project (2) stop of recruitment of SNI (total cohort  19,000 children) The Sardinian Migrants IDDM study (SMI) in the province of Pavia Coeliac Disease in the Northern Sardinia school children the Post-partum Thyroiditis and Neonatal Hypothyiroidism studies the Gestational Diabetes (GDM) Study The Environmental / Veterinarian / Ecological studies - first results 1996 1997 1998 1999HLA typing in ‘immunologically at risk’ children for type 1 diabetes

7. 0 5 10 15 20 25 30 35 40 45 Time trends in Type 1 Diabetes incidence ( 0 - 14 years) 19601965197019751980198519901994 Calendar year Incidence (per 100,000) Finland (40) Sardinia (35) Sweden (25.8) Norway (21.2) Denmark (16.4) Hungary (8.8)

8. The Sardinian Schoolchildren IDDM Study (SSI) (1) Survival Function LATENCY (months) 100806040200 Cum Survival 1,0,9,8,7,6,5 Number of Abs 3 1 0 2

9. The Sardinian Schoolchildren IDDM Study (SSI) (2) Oristano45/100,000/year Cagliari38/100,000/year Nuoro35/100,000/year Sassari30/100,000/year 0 1 2 3 4 5 6 7 8 9 10 1 Ab >= 2 Abs %

10. The Sardinian Newborn-IDDM Study (SNI) (1) Number recruited ~ 19,000 ICA tested = 15,509 (cord blood) 18 Centres from the 4 Sardinian provinces involved in the Study ICA  JDF-u 2.3% 2.3% ICA 6-19  JDF-u 1.7% 1.7% ICA  JDF-u 0.6%

11. The Sardinian Newborn-IDDM Study (SNI) (2) FOLLOW UP ICA  JDF-u GADA  IA-2icA  % prevalence 0.0 0.4 0.8 1.2 1.6 2.0 1 st YEAR 2 nd YEAR 3 rd YEAR 2,959 2,1252,1171,148399399388

12. 0 1 2 3 4 5 6 7 cordblood 1 23 6789101112131415 % prevalence age (years) ICA  JDF-u GADA  IA-2icA  P r e v a l e n c e o f i s l e t - r e l a t e d a u t o a n t i b o d i e s (ICA, GADA and IA - 2icA) according to the different age (from the S a r d i n i a n N e w b o r n and the S a r d i n i a n S c h o o l C h i l d r e n S t u d i e s)

13. The COELIAC DISEASE study (1) ~ 1,600 Sardinian school children investigated for AGA-IgG, AGA-IgA and AEA Prevalence of coeliac disease (confirmed by intestinal biopsy) 10.5/1000 children the highest reported so far in a background population

14. 79 126 11* Coeliac Disease and pre -Type 1 Diabetes in Sardinian schoolchildren (2) Islet- related Abs CD-related Abs ICA + IgG-AGA = 8 GADA + IgG-AGA = 1 GADA + IgG/IgA-AGA + AEA = 2 *

15. The MILITARY SERVICE approach Secular trend of type 1 diabetes prevalence at 19 years among male conscripts in Sardinia 0,00 1,00 2,00 3,00 4,00 5,00 6,00 7,00 1936 1938 1940 1942 1944 19461948 1950 1952 1954 1956 1958 1960 1962196419661968 1970 1972 1974 1976 1978 Birth cohorts Prevalence (per 1000)

16. The ENVIRONMENTAL and ECHOLOGICAL studies Birth seasonality Onset seasonality Temp, pop. density, urban/rural Average rainfall Time- and space-clustering overlap with: - malaria - talassemia - G-6-P-D deficiency Nitrate intake (bottle and tap waters) Cow’s and breast milk feeding YES NO YES NO

17. The GESTATIONAL DIABETES (GDM) Study ~ 100 Sardinian mothers with GDM investigated for ICA, GADA and IA-2icA 8 women resulted positive for at least 1 islet- related autoantibody at the time of OGTT After a follow up of 4 yrs, 5 became diabetic (3 insulin-dependent and 2 non insulin-dependent)

18. AUTOIMMUNE THYROIDITIS in Sardinian school children (1) ~ 8,000 Sardinian schoolchildren from the general population investigated for ATA Overall, the prevalence of ATA was 3.7% and the prevalence of a subclinical thyroiditis was about 0.9% No correlations were found between prevalence of ATA and urinary iodine excretion or prevalence of goitre 0 1 2 3 4 5 6 6-7 7-88-99-10 10-11 11-1212-13 13-14 >14 Males Females Age (years) ATA pos (%)

19. ATA and ICA in Sardinian school children (2) ICA + 325 ATA + 211 16

20. AUTOIMMUNE THYROIDITIS and PREGNANCY (3) At the time of delivery, the prevalence of ATA and ICA was 11.8 and 2.6%, respectively (0.4% with both specificities) 0 20 40 60 80 100 ORCANUSS ATA at low titers ATA at high titers 0 5 10 15 20 25 30 35 40 ICA*  20 JDFu ICA 5-19 JDFu ICA < 5 JDFu ~ 2,500 Sardinian mothers at delivery time were investigated for ATA and ICA Prevalence of ATA (%)

21. The SARDINIAN MIGRANTS study In 10 individuals the diagnosis of T1D has been confirmed giving a prevalence (4/1000) similar to that registered in the island and 3 times higher than the ones registered in Northern Italy. 3 subjects were already diabetic at the time of migration, and 7 developed the disease after the migration to Pavia. the prevalence of type 1 diabetes was assessed in ~ 2,200 born in Sardinia and migrated to Pavia 0 5 10 15 20 25 30 35 Age at onset of T1D (years) Before migration (Nr=3) After migration (Nr=7)

22. What did we learn so far? (1) Islet-related autoantibodies can appear very early in life (with particular reference to GADA) and they play a predictive role towards the future onset of type 1 diabetes. The appearance of islet-related autoantibodies progressively increases in the first years of life. The combination of more than 1 islet-related autoantibody (rather than which autoantibody) is the best predictor for the development of type 1 diabetes in the Sardinian school children population.

23. What did we learn so far? (2) Even though some epidemiological evidences suggest a role of the environment on the etiopathogenesis of type 1 diabetes, none of the variables considered so far have shown their influence; however other variables need to be further investigated. Coeliac disease shows an high prevalence among Sardinian school children and then it deserves more large investigations. It seems not to play a relevant role on the etiopathogenesis of type 1 diabetes in Sardinia instead.

24. The prevalence of thyroid-related autoantibodies seems not to be as much as high among Sardinian school children, even though they live in an Island at high risk for other autoimmune diseases. The same findings appear among the pregnant mothers, in whom the prevalence of ATA is not significantly higher than the ones registered in other matched populations. However, the possible immunosuppressive role of pregnancy on these parameters needs to be further investigated. What did we learn so far? (3)

25. Trying to further improve the prediction of type 1 diabetes in the general population by carrying out HLA genetic typing in children found ‘at immunological risk’ during the screening. What we are doing now (1) Studying the immunological and genetic markers for type 1 diabetes and other autoimmune diseases in Sardinian migrants and their relatives. Investigating other putative environmental factors which can play a role towards the etiopathogenesis of type 1 diabetes (e.g., chemicals, toxins, vaccinations, viral infections, etc.).

26. What we are doing now (2) Broadening the original investigation for type 1 diabetes on the prevalence of coeliac disease and autoimmune thyroid diseases and to study their associations. Comparing data from Sardinia and other areas (continental Italy, Finland, Sweden, Spain) by new collaborative studies. Investigating LADA within the Sardinian type 2 patients.

27. 0 25,000 50,000 75,000 100,000 125,000 150,000 175,000 200,000 225,000 250,000 0 2 4 6 8 10 12 14 16 18 20 years $ cost saved Cost of predicting T1-DM from birth and in school children background population modified from Hahl et al. Diabetologia (1998) 41:79-85 1 2 3 4

28. Benefits of a predictive screening for Type 1 diabetes in the general population Early diagnosis / Early insulin treatment (In U.S.A. about 50 deaths yearly from DKA) 1. ? Prevention and/or delay of diabetic complications 2. Prevention of the onset of type 1 diabetes in ‘at risk’ individuals (? magic bullet) 3. Identification of ‘non at-risk’ individuals (>99%) 4.

29. In spite of Gian Franco’s breakthrough twenty-five years ago with the ICA, the mystery of Type 1 diabetes still remains deeply hidden... … but we strongly believe that along with him in this beautiful Island, we have a very good chance to unravel the causes of the autoimmune diseases. ? ? ? ? ? ? ? ? ? ? The IDDM-Sardinia Study Groups ? ?