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Journal of Dental Research
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Low HEMA Conjugation Induces High Autoantibody Titer in Mice

E. Sandberg1,*, G. Bergenholtz2, H. Kahu1 and U.I. Dahlgren1,3

1 Section for Oral Immunology,
2 Section for Endodontology, Faculty of Odontology, and
3 Department of Rheumatology, Faculty of Medicine, The Sahlgrenska Academy, Göteborg University, Box 450, SE 405 30 Göteborg, Sweden;


Figure 1
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  		Figure 1. IgG antibody activity to native mouse serum albumin (MSA) in sera from animals immunized with MSA carrying various numbers of 2-hydroxyethylmethacrylate (HEMA) molecules: 3, 7, 15, or 22, respectively. Displayed are also control groups immunized with unconjugated MSA or adjuvant alone. IgG antibody activities are expressed in arbitrary ELISA units, as compared with a standard included in each of the assays. Dots denote 1 animal. Horizontal bars indicate median value. The autoantibody production was significantly higher in animals immunized with MSA carrying only 3 HEMA molecules (MSAH3), compared with those animals that were immunized with the other conjugation degrees (n = 8 for all groups, except for the one given MSAH22, where n = 7). The Mann-Whitney U-test was used for statistical comparison (* = p < 0.05, ** = p < 0.01, and *** = p < 0.005).

 

Figure 2
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  		Figure 2. Total (A) and anti-MSA specific (B) antibody activity in serum from mice immunized with mouse serum albumin (MSA) carrying various numbers of 2-hydroxethylmethacrylate (HEMA) molecules: 3, 7, 15, or 22, respectively. Displayed are also control groups immunized with unconjugated MSA or adjuvant alone. Each dot represents 1 animal, and the horizontal bars indicate median values. (A) Significantly higher serum IgE concentration was found in animals immunized with MSA carrying only 3 HEMA molecules, compared with those immunized with MSA carrying 22 (n = 8 for all groups, except for the one given MSAH22, where n = 7). Statistical comparison was done by the Mann-Whitney U-test. (B) The specific IgE anti-MSA antibody activities in sera from animals treated as in (A). The antibody activities to native MSA are expressed as arbitrary ELISA units compared with a standard. Only animals immunized with MSAH3 displayed this antibody activity.

 

Figure 3
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  		Figure 3. IgG antibody activity to 2-hydroxyethylmethacrylate (HEMA) when conjugated to a carrier that was not used in the immunizations: ovalbumin (OVA). Sera were taken from animals immunized with mouse serum albumin (MSA) carrying various numbers of (HEMA) molecules: 3, 7, 15, or 22, respectively. Displayed are also control groups immunized with unconjugated MSA or adjuvant alone. IgG antibody activities are expressed in arbitrary ELISA units as compared with a standard included in each of the assays. The IgG antibody activity to OVA has been subtracted. Each dot represents 1 animal, and the horizontal bars indicate median values (n = 8 for all groups, except for the one given MSAH22, where n = 7). Antibodies that could bind to HEMA, regardless of the carrier, were mainly induced by MSA carrying 15 or more HEMA molecules. The Mann Whitney U-test was used in statistical comparison between animals immunized with MSAH3 and those immunized with MSAH22.

 

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  		Figure 4. Avidity index for antibodies induced in animals immunized with mouse serum albumin (MSA) carrying 3 or 15 2-hydroxyethylmethacrylate (HEMA) molecules, respectively. Serum was diluted, on average, 1:5000 for MSAH3 and 1:2000 for MSAH15 to yield a similar antibody activity. Wells were incubated with potassium thiocyanate (KSCN) at different concentrations, ranging from 0.1 to 3.2 M, in duplicate. Triplicate wells were incubated with PBS so that maximal activity could be assessed. The avidity is expressed as the KSCN concentration where 50% of the antibody activity is eluted compared with the activity in wells incubated with innocuous PBS. Each group consisted of 8 animals. One animal was omitted from the MSAH15 group, due to low anti-MSA antibody activity. Each dot represents 1 mouse, and the horizontal bars indicate median values. The Mann Whitney U-test was used in statistical comparison.

 

Journal of Dental Research, Vol. 84, No. 6, 537-541 (2005)
DOI: 10.1177/154405910508400610
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