Correspondence

Lack of Immunity to Bovine Serum Albumin in Insulin-Dependent Diabetes Mellitus

N Engl J Med 1994; 330:1616-1617June 2, 1994

Article

To the Editor:

Atkinson and colleagues (Dec. 16 issue)1 report a lack of B-cell and T-cell immunity to bovine serum albumin (BSA) in children with insulin-dependent diabetes mellitus (IDDM). Their results are in contrast with several epidemiologic, ecologic, time-series, and experimental studies2,3.

Exposure during the first months of life to infant formulas based on cow's milk carries a significantly elevated risk of IDDM (mean odds ratio in case-control studies, 1.6; 95 percent confidence interval, 1.2 to 2.2)3. We reported elevated levels of IgG anti-BSA antibody in Finnish children with diabetes of recent onset,4 an observation confirmed in a recent large study of French children (unpublished data).

The low titers of BSA antibodies in the patients with IDDM studied by Atkinson et al.1 are surprising. The values were twice as high in the study of French children and four times higher in the diabetic children in Finland4. BSA antibodies resemble anti-insulin autoantibodies in that they are difficult to detect in standard immunoassays; technical aspects of the assay may contribute to the discrepancy. In addition, Atkinson et al. used a BSA-specific mouse monoclonal antibody to standardize their assay, which is unlikely to recognize epitopes related to diabetes in humans. Also, the secondary antihuman antibodies used contained BSA as a stabilizer, which competes for anti-BSA and reduces the sensitivity of the assay.

We found that T cells from 88 percent of 43 diabetic Canadian children were sensitized to BSA, ABBOS, and the p69 islet-cell protein (mean [±SD] stimulation index, 4.8 ±0.7)5. The absence of any T-cell responses to BSA in the study by Atkinson et al. raises the question of problems with culture conditions. The excellent, high-insulin culture medium used has a short shelf life, and we have found it necessary to prepare fresh stocks frequently.

The findings of Atkinson et al. emphasize the need for collaborative efforts to standardize assays. Their negative results signal the need for caution but do not disprove positive results from an increasing number of studies of BSA immunity in patients with IDDM.

Hans-Michael Dosch, M.D.
Hospital for Sick Children, Toronto, ON M5G 1X8, Canada

Jukka Karjalainen, M.D.
University of Oulu, SF-90220 Oulu, Finland

John VanderMeulen, M.D., Ph.D.
McMaster University, Hamilton, ON L8N 3Z5, Canada

5 References

1. 1

   Atkinson MA, Bowman MA, Kao K-J, et al. Lack of immune responsiveness to bovine serum albumin in insulin-dependent diabetes. N Engl J Med 1993;329:1853-1858
   Full Text | Web of Science | Medline

2. 2

   Akerblom HK, Savilahti E, Saukkonen TT, et al. The case for elimination of cow's milk in early infancy in the prevention of Type 1 diabetes: the Finnish experience. Diabetes Metab Rev (in press).
   

3. 3

   Gerstein HC. Cow's milk exposure and type I diabetes mellitus: a critical overview of the clinical literature. Diabetes Care 1994;17:13-19
   CrossRef | Web of Science | Medline

4. 4

   Karjalainen J, Martin JM, Knip M, et al. A bovine albumin peptide as a possible trigger of insulin-dependent diabetes mellitus. N Engl J Med 1992;327:302-307
   Full Text | Web of Science | Medline

5. 5

   Karjalainen J, Cheung R, Miyazaki I, Gaedigk R, Hui MF, Dosch H-M. BSA/p69 specific T cells in newly diabetic children. In: Pilarski L, ed. Proceedings of the Eighth Meeting of the Canadian Society for Immunology, Ste. Adele, Quebec, 1994:1.10. abstract.
   

Author/Editor Response

The authors reply:

To the Editor: We find the epidemiologic association between the consumption of cow's milk and IDDM intriguing1. We sought to confirm an immunologic basis for the association,2 but found none.

We believe the statements by Dosch et al. about our ability to detect BSA antibodies are unfounded and provide insufficient explanation of the differences between their results2 and ours. First, although the fluorescence activity of our positive serum samples was not as high as that reported by Dosch and colleagues, the titers of anti-BSA antibodies in these samples were not necessarily low, as their letter suggests. Antibody titer and mean intensity of fluorescence are different measurements, and the determination of titer requires an assay of serially diluted serum samples that was not reported in our study1.

Second, we used the BSA-specific monoclonal antibody to ensure that BSA was covalently linked to the polystyrene beads and not to standardize immunologic detection by human IgG. Our assay did in fact detect anti-BSA antibodies, since the antibody reactions could be inhibited by exogenous BSA. The antibodies, however, were not specific for IDDM, being detected in both high- and low-risk nondiabetic relatives of probands with IDDM, as well as patients with other autoimmune disorders.

Third, we disagree with the statement that standard immunoassays do not detect insulin autoantibodies. Finally, the secondary antihuman antibodies we used contained ovalbumin as a stabilizer, not BSA.

With respect to the detection of T-cell responses to BSA, we used only freshly prepared medium. Furthermore, the immunologic association between anti-islet-cell p69 autoantibodies3 and anti-BSA immunity is increasingly unclear, because detection of the former is not inhibited by the addition of cow's milk3. In addition, since epidemiologic data suggest that breast-feeding is associated with a decreased incidence of IDDM, it may be that this practice protects against the development of IDDM rather than that IDDM is induced by the ingestion of cow's milk.

The association of environmental factors with the pathogenesis of IDDM remains the subject of controversy. Further epidemiologic, immunologic, and genetic information is required to identify the cause of this disease.

Mark A. Atkinson, Ph.D.
K.J. Kao, M.D., Ph.D.
Noel K. Maclaren, M.D.
University of Florida, Gainesville, FL 32610

3 References

1. 1

   Maclaren N, Atkinson M. Is insulin-dependent diabetes mellitus environmentally induced? N Engl J Med 1992;327:348-349
   Full Text | Web of Science | Medline

2. 2

   Karjalainen J, Martin JM, Knip M, et al. A bovine albumin peptide as a possible trigger of insulin-dependent diabetes mellitus. N Engl J Med 1992;327:302-307
   Full Text | Web of Science | Medline

3. 3

   Pietropaolo M, Castano L, Babu S, et al. Islet cell autoantigen 69 kD (ICA69): molecular cloning and characterization of a novel diabetes-associated autoantigen. J Clin Invest 1993;92:359-371
   CrossRef | Web of Science | Medline

Citing Articles (2)

Citing Articles

1. 1

   T Saukkonen, E Savilahti, M Landin-Olsson, G Dahlquist. (1995) IgA bovine serum albumin antibodies are increased in newly diagnosed patients with insulin-dependent diabetes mellitus, but the increase is not an independent risk factor for diabetes. Acta Paediatrica 84:11, 1258-1261
   CrossRef

2. 2

   P. Pigny, G. Mortreux, A. Racadot, C. Stuckens, A. Boersma. (1995) Humoral immune response to bovine serum albumin in new onset and established insulin-dependent diabetes mellitus. Acta Diabetologica 32:2, 135-136
   CrossRef