Study of the Correlation between the Gingival Immunologic Defense Index and Parameters Associated with Dental Caries

 
Maria Cristina Monteiro de SOUZA-GUGELMIN1
Izabel Yoko ITO1
Geraldo MAIA CAMPOS2
 
1Faculdade de Ciências Farmacêuticas de Ribeirão Preto,
2Faculdade de Odontologia de Ribeirão Preto,
Universidade de São Paulo, Ribeirão Preto, SP, Brasil


Braz Dent J (1996) 7(2): 91-95 ISSN 0103-6440

| Introduction | Material and Methods | Results and Discussion | References |


The objective of the present study was to determine the possible existence of a correlation between the gingival immunologic defense index (GIDI) and parameters associated with dental caries such as number of teeth with cavities, number of colony forming units (CFU) of streptococci of the mutans group, decay missing filling deciduous teeth (dmft), and decay missing filling permanent teeth (DMFT). Since no correlation was detected with the above parameters, we conclude that the simple presence of caries or of CFU is not sufficient to stimulate IgA production and/or secretion. For this to occur, an unfavorable action of these parameters on gingival health is needed, with the production of gingival inflammation.


Key Words: gingival immunologic defense index, IgA, number of CFU of streptococci of the mutans group.


Introduction

In addition to periodontal disease, another oral disease - caries - has been related to salivary immunoglobulin levels, as also are DMF/dmf and number of colony forming units (CFU) of streptococci of the mutans group. In a study of unstimulated total saliva from 63 children aged 6 to 13 years, Riviere and Papagiannoulis (1987) noted that 29 presented dental caries and 34 did not. They investigated salivary IgA, IgG and IgM levels related to the streptococcus strains because, in their opinion, if salivary anti-S. mutans antibodies effectively acted on resistance to caries, then children with higher levels of these antibodies should have a smaller number of caries. However, no significant difference was detected with respect to salivary IgA levels. Camling et al. (1987) noted that individuals with active caries and/or with high decay missing filling surface (DMFS) indices manifested a tendency to present lower IgA levels in total saliva than individuals with a low caries experience. IgA levels were mostly higher in the group with a small number of S. mutans (<5 x 104). Camling and Köhler (1987), looking for a correlation between salivary S. mutans and IgA levels of mothers and their children, observed that IgA levels did not differ significantly between children whose mothers had received preventive treatment against caries and children of untreated mothers, although children of untreated mothers presented higher S. mutans levels. Reinholdt and Kilian (1987) demonstrated that bacterial IgA-proteases that cleave sIgA in the fold region can interfere with the mechanism of action of this immunoglobulin, a mechanism that impairs the adhesion of oral streptococci to hydroxyapatite and to cell surfaces. Some investigators explain the effects mediated by IgA-protease by a reduction in the ability of Fab-alpha fragments separated by cleavage to bind to the antigen. Reinholdt and Kilian (1987), however, showed that, after exposure of specific sIgA to IgA-protease, the streptococci are covered with Fab-alpha fragments. Gråhn et al. (1988) reported a total absence of correlation between salivary IgA level and presence of caries, as well as the absence of any correspondence between salivary S. mutans levels and salivary anti-S. mutans antibodies. The objective of the present study was to determine the possible existence of a correlation between the gingival immunologic defense index (GIDI) and caries, looking for a correlation between this new index and the number of teeth with caries and also with the number of CFU of the mutans group.


Material and Methods

The study was conducted on 135 healthy children aged 5 to 13 years and residing in Araraquara, State of São Paulo, from whom unstimulated total saliva samples were obtained. The dmft/DMFT indices and the number of CFU of the mutans group were also determined. Saliva samples were collected into 20 x 150 mm sterile tubes; after collection, part of the saliva was transferred to 12 x 75 mm sterile tubes and frozen for later measurement of salivary IgA content by single radial immunodiffusion as described in the first part of this research (Souza-Gugelmin et al., 1993). The portion of saliva destined to the counts of CFU of the mutans group was left in the 20 x 150 mm tubes and cooled on dry ice in a styrofoam box for transportation to Ribeirão Preto, where it was processed by the serial solution method. The method was used to determine the number of CFU of the mutans group by first diluting 0.1 ml saliva in 10.0 ml sterile phosphate buffered saline (PBS). This provided the 10-2 dilution from which the 10-3 dilution was obtained by transferring 1.0 ml to a second tube containing 9.0 ml PBS. Similarly, the successive transfer of 1.0 ml of a freshly prepared dilution to the next tube containing 9.0 ml PBS provided the subsequent dilutions, 10-4 and 10-5. The different saliva dilutions were inoculated into SB20 medium by the glass rod technique (Davey and Rogers, 1984). This medium was chosen because it is selective for the mutans group. Inoculation was carried out in duplicate and the plates were incubated in anaerobiosis jars containing 10% CO2 at 37ºC for 48 hours and at room temperature for an additional 24 hours. CFU of streptococci of the mutans groups were then counted using a stereoscopic microscope. After recording the counts of the 4 duplicate dilutions for each sample, the number of CFU of the mutans group was calculated as follows: the number of CFU of the mutans group for each dilution was multiplied by 2 and the number of zeros corresponding to the absolute value of its exponent plus 1 was added. The number of CFU for each dilution was summed and divided by the number of plates counted to obtain the mean for the first series. The same procedure was applied to the plates of the 2nd series (duplicates), and the final number of CFU of the mutans group present in 1.0 ml saliva was determined by the mean calculated for the two series. Since these values were very high their logarithms were used in the subsequent statistical calculations in order to reduce data variability. In cases in which the number of CFU in the first dilution was zero, the use of the logarithm was inviable since the logarithm of zero is infinity. To overcome this drawback, we assumed 2000 to be the minimum number of colonies that would correspond to the finding of a single colony in the lowest dilution. This strategy permitted the lowest logarithm to be 3.30103. The DMFT/dmft indices were determined by the same examiner immediately after saliva collection. The GIDI was calculated as described in the second part of this study (Souza-Gugelmin et al., 1995), i.e., by dividing the IgA level measured in saliva (mg/100 ml saliva) by the number of inflamed gingival surfaces (NIGS) present.


Results and Discussion

A total of 135 saliva samples were obtained from children aged 5 to 13 years. IgA concentration was measured in the samples by the single radial immunodiffusion technique as described previously (Souza-Gugelmin et al., 1993) and the GIDI value was then calculated. This index evaluates the immunologic potential of the gingiva with respect to IgA, as demonstrated in the second part of this study (Souza-Gugelmin et al., 1995). Table 1 lists the results obtained for the different parameters studied in relation to the different class intervals of GIDI. Table 1 is simply illustrative since in the regression and correlation tests performed in the present study we used the 135 data pairs related to each parameter. Thus, the table is simply a summary of the general table of the original data. In the present study we tested the possibility of a correlation between GIDI and parameters such as CFU of streptococci of the mutans group and number of teeth with caries (lesions present). We first tested the correlation between IgA concentrations in saliva and dmft/DMFT indices and number of CFU of streptococci of the mutans group for all 135 children. The statistical tests performed revealed a complete absence of correlation between these parameters and IgA level, with a correlation coefficient of 0.2191, i.e., corresponding to a 5.88% probability for the hypothesis of equality. When correlation with dmft was tested, the coefficient obtained was 0.0107, i.e., corresponding to a 90.66% probability for the hypothesis of equality. The correlation coefficient with respect to DMFT was 0.0992, i.e., corresponding to a 25.28% probability for the hypothesis of equality. The correlation coefficient with respect to number of CFU of the mutans group was 0.0992, i.e., corresponding to a probability of 25.28% for the hypothesis of equality. The lack of statistical significance for the calculated r values for the dmft and DMFT indices led us to conclude that there is no relationship between salivary IgA levels and these conventional indices. Indeed, an objective evaluation of the results leads to the following question: what is the role of teeth lost or exfoliated in the past or of the number of teeth with previous fillings in the production of IgA, which is always a current response to an inflammatory reaction? On this basis, we decided to use only the number of teeth with caries, i.e., with current lesions, a fact that, in addition to solving the above problem, also offered a standardization of the parameter, since most of the children sampled were in the phase of mixed dentition, with the consequent need to utilize the dmft and DMFT, while exfoliated teeth are known not to be representative of active caries. However, the present comparison between number of teeth with caries with GIDI rather than with dmft or DMFT, and not simply with IgA level continued to show a lack of significant correlation between them since the regression and correlation tests performed between GIDI and number of teeth with caries yielded r = -0.0133 (non-significant) with a p(HO) = 87.86%. The same lack of significance was obtained when the correlation between GIDI and number of CFU of streptococci of the mutans group was tested, with r = 0.0370 and p(HO) = 67.60%. These findings indicate that the simple presence of caries or of CFU of streptococci of the mutans group in the oral cavity is not sufficient to stimulate IgA production and/or secretion. For this to occur, the caries should be biologically active, i.e., they should be acting unfavorably on gingival health, provoking gingival inflammation. Only then will an antigenic stimulus occur for the production and secretion of IgA at levels sufficient to bring about a possible reversal of the clinical picture. These tests simply confirmed the impression we already had that the influence of caries, of bacterial colonies and even of poorly executed fillings on GIDI would be simply indirect and would only manifest when these elements act as direct agents of gingival inflammation due to excessive plaque accumulation and to the action of irregular tooth borders (caused by caries or by defective filling borders).


References

Camling E, Köhler B: Infection with the bacterium Streptococcus and salivary IgA antibodies in mothers and their children. Arch Oral Biol 32: 817-823, 1987

Camling E, Gahnberg L, Krasse B: The relationship between IgA antibodies to Streptococcus mutans antigens in human saliva and breast milk and the numbers of indigenous oral Streptococcus mutans. Arch Oral Biol 32: 21-25, 1987

Davey AL, Rogers AH: Multiple types of the bacterium Streptococcus mutans in the human mouth and their intra-family transmission. Arch Oral Biol 29: 453-460, 1984

Gråhn E, Tenovuo J, Lehtonen O-P, Eerola E, Vilja P: Antimicrobial systems of human whole saliva in relation to dental caries, cariogenic bacteria, and gingival inflammation in young adults. Acta Odont Scand 46: 67-74, 1988

Reinholdt J, Kilian M: Interference of IgA protease with the effect of secretory IgA on adherence of oral streptococci to saliva-coated hydroxyapatite. J Dent Res 66: 492-497, 1987

Riviere GR, Papagiannoulis L: Antibodies to indigenous and loboratory strain of Streptococcus mutans in saliva from children with dental caries and from caries-free children. Pediat Dent 216-220, 1987

Souza-Gugelmin MCM, Ito IY, Maia Campos G: Study of the correlation between salivary IgA and gingival inflammation in children. Bras Dent J 4: 91-96, 1993

Souza-Gugelmin MCM, Ito IY, Maia Campos G: Creation of a new index to evaluate the immunological potential of the gingiva and the possible risk for periodontal disease, the Gingival Immunologic Defense Index (GIDI). Braz Dent J 6: 95-102, 1995


Correspondence: Maria Cristina Monteiro de Souza-Gugelmin, Faculdade de Ciências Farmacêuticas de Ribeirão Preto, USP, 14040-903 Ribeirão Preto, SP, Brasil.


Accepted April 8, 1996
Electronic publication: February, 1997


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