Original study - ZZI 01/2011

Comparative microarray analyses of inflamed human periimplant and periodontal tissues in vivo

The diseases periodontitis and periimplantitis were compared by means of an intermediate stage of comparing the respective disease with the expression profiles of healthy perio-dontium (neutral reference point). The two inflamed tissues (periodontal and periimplant inflamed tissue) were first compared with the healthy periodontium and in a second step the two diseases were compared with one another. The normalized gene expression values are summarized in Table 2 and assessed as increased (= I), decreased (= D) and no change (= NC). Genes with an at least two-fold higher or lower (fold change ratio +/? 2) difference in the signal intensity were determined as differently regulated. This corresponded to an approximate deviation of +/? 0.5 points in the CTs (cycle thresholds) of the real-time (RT) PCR, and this was defined as a significant change.

Results

Sample verification of the obtained mRNA

The real-time (RT) PCR performed prior to microarray analysis to determine the inflammatory marker (transcription factor) NF-?B showed a markedly increased expression profile in the chronically inflamed tissues (periodontitis and periimplantitis) compared with healthy PDL. Alongside the clinical diagnosis, this finding confirmed that the tissue samples actually consisted of chronically inflamed tissue (periodontitis or periimplantitis). The expression profile of NF-?B between the two chronically inflamed tissues was again very similar.

Results of the microarray analyses

When 22,283 human genes were tested, 2,079 genes with a different gene expression with an at least two-fold higher or lower difference (fold change ratio +/? 2) were demonstrated. The expression data of 1,093 genes were significantly up-regulated in the chronically inflamed periimplant tissue compared with chronically inflamed periodontal tissue (fold change ratio + 2) and the expression of 986 genes was significantly down-regulated (fold change ratio ? 2). Table 3 provides a summary of ECM components and their degrading enzymes on which our study focused.

Real-time (RT) PCR results

To verify the results obtained from the microarray tests, a selection of proteins was investigated using real-time (RT) PCR. This showed that cathepsins B and C (CTSB and CTSC) had increased gene expression on real-time PCR in periimplantitis compared with healthy periodontal tissue. The ratios were 25.599 and 21.873. From the matrix metalloproteinase group (MMPs) MMP-2, -8, -9 and -23B were tested and an increased expression level was found for MMP-2 and -9 in inflamed periimplant tissue. The ratio was 3.802 and 25.368 respectively. No significant changes were found in MMP-8 and MMP-23B for periimplantitis compared with periodontitis.

Out of the collagen group, collagen IV (alpha1) was tested and an increased ratio (24.489) was found.

Comparison of the microarray and real-time (RT) PCR results

Because of the greater sensitivity of the real-time (RT) PCR, gene expression differences were found with a few genes that could not be demonstrated using microarray technology (MMP9, CTSB, CTSC). Otherwise, the results of the two methods were in agreement (Table 4).

Discussion

The clinical features of chronic periimplantitis suggest a similar etiology to that of chronic periodontitis. This has also been confirmed by a series of studies [20]. The primary etiologic factor in both diseases is based on increased plaque accumulation on the tooth or implant, with resulting destructive changes in the surrounding hard and soft tissue. However, it must be borne in mind that there are two different types of tissue (tooth = desmodontium = soft tissue; implant = bone = hard tissue), which can lead to a different pathogenesis of the two diseases. However, there is as yet no clear demarcation between periimplantitis and periodontitis.

In this study 22,283 human genes in healthy tissue, chronically inflamed periodontal tissue and chronically inflamed periimplant tissue were compared by means of microarray technology. Because of the large amount of data, this study concentrated on the ECM components and their degrading enzymes, on the one hand in order to filter out significantly differently expressed genes within these two important groups and on the other hand, in order to identify possible differences between periodontitis and periimplantitis, at least at the mRNA level.

Since both the performed microarray technology and real-time PCR investigate mRNA exclusively, it must be added at this point that detection of mRNA alone does not signify the presence or secretion of the corresponding protein and thus its biological activity. Below we discuss the possible roles of the investigated genes, assuming that the amount of mRNA correlates with the amount of corresponding proteins in the periimplant tissue and in the periodontal tissue. This corresponds to current expert opinion.

Matrix metalloproteinases (MMPs)

A series of studies has confirmed the increased expression of MMP-8 in both adult and juvenile periodontitis (current nomenclature: chronic and aggressive periodontitis) [16]. In our investigations, no significant changes in MMP-8 and MMP-23B were found for periimplantitis compared with perio-dontitis. However, this does not mean that these MMPs do not play any role in periimplantitis; rather, it means that MMP-8 and MMP-23B are not expressed differently in periimplantitis compared with periodontitis. However, an increase in the MMP-2 and MMP-9 concentrations was demonstrated in periimplantitis. The correlation between the rise in MMP-2 and destruction of the periodontal structures has already been shown [28].

PAGE: 1 | 2 | 3 | 4 | 5 | 6