Heterozygous activating mutations in the KCNJ11 gene are a common cause of permanent neonatal diabetes (PNDM) (1,2). In contrast to the autoimmune type 1 diabetes, patients with KCNJ11 mutations do not have serological markers of autoimmune β-cell destruction at disease onset (1,35). In such patients, hyperglycemia does not result from insulin-secreting cell destruction but rather from impaired insulin secretion. In addition, the majority of cases can be corrected with sulfonylurea therapy (2,6). Here, we report that carriers of the KCNJ11 mutation that are immunonegative at onset may show presence of islet antibodies in the further course of the disease.

We sought to evaluate the clinical and genetic characteristics of patients with neonatal diabetes in Poland, using the Nationwide Registry, which was established in 2005 (7,8). Automatic sequencing of the KCNJ11 gene allowed identification of 15 patients with heterozygous mutations that cause neonatal diabetes. Sera from 11 carriers of the KCNJ11 gene mutation were available for the present study. None of the patients had a family history of type 1 diabetes. Informed consent was obtained from all subjects or their parents. The study was conducted in accordance with the Declaration of Helsinki (revised in 2000) and accepted by the local ethics committee in Lodz, Poland.

β-Cell antibody analysis

At diabetes onset.

Available data on islet antibody measurement at the onset of diabetes were limited. Five of our patients were diagnosed with diabetes before islet antibodies were implemented for clinical use. The rest of the obtained results come from several local laboratories. None of the tested patients had islet antibodies.

At study entry.

Autoantibody analysis was performed in the Immunopathology Laboratory at the Department of Pediatrics, Medical University of Lodz, Poland, which is a reference laboratory for islet antibodies measurement and is a regular participant of international proficiency testing programs. The methods of antibody measurements were verified in the reference laboratory at the Barbara Davis Center for Childhood Diabetes in Denver, Colorado, with an inconsistency of 3% (courtesy of Prof. George Eisenbarth; see supplemental Figs. 1 and 2 for more information [available in an online appendix at http://dx.doi.org/10.2337/dc06-2440]).

Nonparametric statistics using the Mann-Whitney U test were applied to assess the differences between groups.

Of 11 patients tested for the presence of islet antibodies, 5 were positive (Table 1). Among patients with >10 years’ duration of neonatal diabetes, more than one-half showed the presence of at least one islet antibody. The relationship between disease duration and the occurrence of autoantibodies is shown in supplemental Fig. 2. Of five sera collected from the R201H mutation carriers, two (in subjects aged 13 and 50 years) were positive for islet antibodies. Moreover, two subjects with phenotypically more severe mutation, V59M, were negative for tested autoantibodies (Table 1). Thus, it is likely that the type of mutation is not related to the presence of autoantibodies. Despite detectable markers of islet-specific autoimmune process, all five patients responded well to sulfonylurea treatment, which indicates that a sufficient number of β-cells remained to maintain glucose homeostasis.

Humoral markers of autoimmune type 1 diabetes are absent at onset of PNDM. This finding is consistent with all published studies on neonatal diabetes, due to mutation in the KCNJ11 gene (1,35). In our patients’ records, antibody measurement at disease onset were available for only five subjects, and, systematically, all patients were negative for β-cell–specific autoantibodies.

Our results demonstrate that immunonegative-at-onset carriers of the KCNJ11 mutation may show seroconversion with long duration of the disease. It is known that apoptosis in β-cells can be elicited by various stimuli, including the perturbation of the metabolic and signal pathways (912). Gain-of-function mutations in Kir6.2 severely disturb metabolism of pancreatic β-cells, possibly promoting increased cell turnover. Recurrent exposure of tissue-specific antigens could lead to primary sensitization of immune cells. It appears that in this subgroup of patients, an autoimmune process may occur as a secondary effect to severe cell dysfunction, which results from KCNJ11 mutation.

All studies published to date that tested behavior of cells in the presence of mutated Kir6.2 focused on short-term effects. For instance, Lin et al. (13) did not observe a proapoptotic effect of mutated Kir6.2 in the insulin-secreting cell line INS-1 during their up to 1 week of experiments. Among patients with the KCNJ11 mutation, seroconversion occurs after at least a 10-year duration of PNDM. Therefore, it would be interesting to check the long-term effect of mutated Kir6.2 on the survival of insulin-secreting cells.

It has been reported that not all patients with PNDM caused by mutated Kir6.2 can transfer from insulin to sulfonylurea therapy (2). In some cases, it can be explained by the severity of mutation, as was demonstrated by in vitro studies (14). Interestingly, two diabetic mothers of two children also affected who responded well to sulfonylureas were resistant to this therapy. This observation shows that mutation type is not the only limiting factor in successful transfer to sulfonylureas and that diabetes duration may have an additional impact. Presence of islet antibodies proves that autoimmune response can be triggered in patients with mutated Kir6.2 protein. Ongoing processes may be responsible for the destruction of pancreatic islets, which in turn preclude treatment with sulfonylureas. Conversely, it is still speculative whether the autoimmune process could constitute an explanation for an observed lack of response to sulfonylurea therapy in some individuals with a long duration of PNDM.

Moreover, our finding is of clinical importance regarding type 1 diabetes diagnosis qualification versus type 2 or gestational diabetes. The presence of islet antibodies in patients with long duration of neonatal diabetes exemplifies that immune reaction against β-cells is not exclusively observed in type 1 diabetes. Therefore, diabetes diagnosis at age <6 months and lack of antibodies at the onset (but not in the later course of the disease) should constitute inclusion criteria for genetic evaluation of PNDM.

Table 1—

Detailed characteristics of the study group with autoantibody measurements

Diabetes duration (years)Age at onset (months)MutationSexAntibody at onsetIslet cell antibodies (N<5JDF)GAD anitbody (N<9.1 IU/ml)Insulin-associated protein 2 antibody (N<20 IU/ml)
Pol2 13 R201H 9.4 159.5 
Pol9 20 V59M NA 0.0 0.0 
Pol11 22 K170N NA 10 4.0 67.8 
Pol13 35 R201H NA 1.8 16.4 
Pol14 21 R201H NA 1.9 17.3 
Pol16 V59M 0.0 0.0 
Pol19 50 R201H NA 40 1.1 20.8 
Pol23 H46L 6.3 10.3 
Pol31 11 E229K 10 3.6 7.6 
Pol32 19 G53D 0.0 59.0 
Pol33 11 R201H 0.2 0.0 
Diabetes duration (years)Age at onset (months)MutationSexAntibody at onsetIslet cell antibodies (N<5JDF)GAD anitbody (N<9.1 IU/ml)Insulin-associated protein 2 antibody (N<20 IU/ml)
Pol2 13 R201H 9.4 159.5 
Pol9 20 V59M NA 0.0 0.0 
Pol11 22 K170N NA 10 4.0 67.8 
Pol13 35 R201H NA 1.8 16.4 
Pol14 21 R201H NA 1.9 17.3 
Pol16 V59M 0.0 0.0 
Pol19 50 R201H NA 40 1.1 20.8 
Pol23 H46L 6.3 10.3 
Pol31 11 E229K 10 3.6 7.6 
Pol32 19 G53D 0.0 59.0 
Pol33 11 R201H 0.2 0.0 

F, female; M, male; N, normal value.

This study was supported by the Polish Ministry of Education and Science (Grants 2P0E13629 and 2P05E05129).

We thank Dr. Liping Yu from the Barbara Davis Center in Denver, Colorado, for islet antibodies evaluation and Prof. George Eisenbarth for critical discussion.

1.
Gloyn AL, Pearson ER, Antcliff JF, Proks P, Bruining GJ, Slingerland AS, Howard N, Srinivasan S, Silva JM, Molnes J, Edghill EL, Frayling TM, Temple IK, Mackay D, Shield JP, Sumnik Z, van Rhijn A, Wales JK, Clark P, Gorman S, Aisenberg J, Ellard S, Njolstad PR, Ashcroft FM, Hattersley AT: Activating mutations in the gene encoding the ATP-sensitive potassium-channel subunit Kir6.2 and permanent neonatal diabetes.
N Engl J Med
350
:
1838
–1849,
2004
2.
Pearson ER, Flechtner I, Njølstad PR, Malecki MT, Flanagan SE, Larkin B, Ashcroft FM, Klimes I, Codner E, Iotova V, Slingerland AS, Shield J, Robert JJ, Holst JJ, Clark PM, Ellard S, Søvik O, Polak M: Hattersley AT, the Neonatal Diabetes International Collaborative Group: Switching from insulin to oral sulfonylureas in patients with diabetes due to Kir6.2 mutations.
N Engl J Med
355
:
467
–477,
2006
3.
Vaxillaire M, Populaire C, Busiah K, Cave H, Gloyn AL, Hattersley AT, Czernichow P, Froguel P, Polak M: Kir6.2 mutations are a common cause of permanent neonatal diabetes in large cohort of french patients.
Diabetes
53
:
2719
–2722,
2004
4.
Sagen JV, Raeder H, Hathout E, Shehadeh N, Gudmundsson K, Baevre H, Abuelo D, Phornphutkul C, Molnes J, Bell GI, Gloyn AL, Hattersley AT, Molven A, Sovik O, Njolstad PR: Permanent neonatal diabetes due to mutations in KCNJ11 encoding Kir6.2: patient characteristics and initial response to sulfonylurea therapy.
Diabetes
53
:
2713
–2718,
2004
5.
Massa O, Iafusco D, D'Amato E, Gloyn AL, Hattersley AT, Pasquino B, Tonini G, Dammacco F, Zanette G, Meschi F, Porzio O, Bottazzo G, Crino A, Lorini R, Cerutti F, Vanelli M, Barbetti F, Early Onset Diabetes Study Group of the Italian Society of Pediatric Endocrinology and Diabetology: KCNJ11 activating mutations in Italian patients with permanent neonatal diabetes.
Hum Mutat
25
:
22
–27,
2005
6.
Tonini G, Bizzarri C, Bonfanti R, Vanelli M, Cerutti F, Faleschini E, Meschi F, Prisco F, Ciacco E, Cappa M, Torelli C, Cauvin V, Tumini S, Iafusco D, Barbetti F, Early Onset Diabetes Study Group of the Italian Society of Paediatric Endocrinology and Diabetology: Sulfonylurea treatment outweighs insulin therapy in short-term metabolic control of patients with permanent neonatal diabetes mellitus due to activating mutations of the KCNJ11 (KIR6.2) gene.
Diabetologia
49
:
2210
–2213,
2006
7.
Skupien J, Malecki MT, Mlynarski W, Klupa T, Wanic K, Gach A, Solecka I, Sieradzki J: Assessment of insulin sensitivity in adult subjects with permanent neonatal diabetes mellitus due to mutations in the KCNJ11 gene encoding Kir6.2.
Rev Diabetic Stud
3
:
17
–20,
2006
8.
Malecki MT, Skupien J, Klupa T, Wanic K, Mlynarski W, Gach A, Solecka I, Sieradzki J: Transfer to sulphonylurea therapy of adult subjects with permanent neonatal diabetes due to KCNJ11 activating mutations: evidence for improvement in insulin sensitivity.
Diabetes Care
30
:
147
–149,
2007
9.
Horwitz MS, Ilic A, Fine C, Rodriguez E, Sarvetnick N: Presented antigen from damaged pancreatic beta cells activates autoreactive T cells in virus-mediated autoimmune diabetes.
J Clin Invest
109
:
79
–87,
2002
10.
Faideau B, Larger E, Lepault F, Carel JC, Boitard C: Role of β-cells in type 1 diabetes pathogenesis
Diabetes
54(Suppl. 2)
:
S87
–S96,
2005
11.
Lipson KL, Fonseca SG, Urano F: Endoplasmic reticulum stress-induced apoptosis and auto-immunity in diabetes.
Curr Mol Med
6
:
71
–77,
2006
12.
Lee SC, Pervaiz S: Apoptosis in the pathophysiology of diabetes mellitus.
Int J Biochem Cell Biol
39
:
497
–504,
2007
13.
Lin CW, Lin YW, Yan FF, Casey J, Kochhar M, Pratt EB, Shyng SL: Kir6.2 mutations associated with neonatal diabetes reduce expression of ATP-sensitive K+ channels: implications in disease mechanism and sulfonylurea therapy.
Diabetes
55
:
1738
–1746,
2006
14.
Koster JC, Remedi MS, Dao C, Nichols CG: ATP and sulfonylurea sensitivity of mutant ATP-sensitive K+ channels in neonatal diabetes: implications for pharmacogenomic therapy
Diabetes
54
:
2645
–2654,
2005

Published ahead of print at http://care.diabetesjournals.org on 2 May 2007. DOI: 10.2337/dc06-2440.

Additional information for this article can be found in an online appendix at http://dx.doi.org/10.2337/dc06-2440.

A table elsewhere in this issue shows conventional and Système International (SI) units and conversion factors for many substances.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C Section 1734 solely to indicate this fact.

Supplementary data