Hepatic enzymes and immuno inflammatory response to BIO-C® Temp bioceramic intracanal medication implanted into the subcutaneous tissue of rats

Camila Soares Lopes1, Mateus Machado Delfino1, Mário Tanomaru-Filho1, Estela Sasso-Cerri2, Juliane Maria Guerreiro-Tanomaru1 & Paulo Sérgio Cerri2

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Objective:
To evaluate the biocompatibility and hepatotoxicity of a new intracanal medication, BIO-C® Temp (BIO), compared to those of the gold standard calcium hydroxide-based intracanal medication (Calen; CAL).
Clinical implications: The biocompatibility of a material means the ability to be in contact with biological tissues and not cause damage (and this is often associated with postoperative pain) and hepatotoxicity means the damage caused by the material to the liver. The results of this study are important as they demonstrate the biocompatibility of BIO-C® Temp and the absence of hepatotoxicity in the material.

Results:

Morphological findings, capsule thickness and numerical density of the number of inflammation cells
‍At 7 days, all groups' capsules contained inflammatory cells, primarily macrophages and lymphocytes, as well as a few fibroblasts (Fig. 3A-C). However, there were significant differences between the groups (p 0.0001); the lowest levels of inflammatory cells were found in GC capsules, while the highest levels were found in CAL specimens (Fig. 3A-M). The number of inflammatory cells in BIO and GC decreased significantly over time (p 0.0001). Although the number of inflammatory cells in CAL specimens decreased significantly from 7 to 15 days (p 0.0001), no significant difference (p = 0.61) was found between the 15- and 30-day periods. At 60 days, the number of inflammatory cells in all groups was lower (p 0.0001) than at other time points (Fig. 3M).
FIGURE 3(A-L) Light micrographs displaying the capsules' interiors. Inflammatory cells (arrows), fibroblasts (Fb), collagen fibers (CF), and particles of material (arrowheads) are seen in the capsules. The control group's capsules at 15, 30, and 60 days exhibit a gradual increase in collagen fibers (CF) and fibroblasts (Fb) but few inflammatory cells (arrows). BV: blood vessels; ELE. Bars: 18 μm. (M) The graph displays the numbers representing the numerical density of inflammatory cells within the capsules, expressed as the mean ± standard deviation. In each period, the comparison between the groups is indicated by superscript letters; different letters = significant difference. The superscript numbers indicate the analysis of each group over time; different numbers = significant difference.
Tukey's test (p ≤ 0.05).
Immunohistochemical detection of IL-6
According to Fig. 5M, the lowest values of cells immunolabeled with IL-10 were found in all groups at 7 days. From 7 to 60 days, IL-10 immunoexpression increased significantly in all groups (p < 0.0001). At 7 days, no significant difference was detected between the groups (p > 0.05). At 15, 30, and 60 days, the number of IL-10 immunolabeled cells was significantly lower in the CG specimens than in the BIO and CAL groups (p < 0.0001). On the 15th day, the highest values were found in the BIO specimens, while at 30 and 60 days, the highest values were observed in the CAL samples (p < 0.0001).
FIGURE 5Light micrographs displaying isolated capsule sections from sections subjected to hematoxylin counterstaining and immunohistochemistry to detect IL-10 (brown/yellow color). Few immunolabeled cells are visible in the capsules of all groups, particularly at day seven (A–C). In all groups, obvious immunolabeling is present in mast cells (MC). Arrows, inflammatory cells; BV, blood vessels; material particles, arrowheads.
Bars: 18 μm. (M) The graph displays the numbers for the numerical density of IL-10-immunolabeled cells in the capsules, expressed as the mean ± standard deviation. In each period, the comparison between the groups is indicated by superscript letters; different letters = significant difference. The superscript numbers indicate the analysis of each group over time; different numbers = significant difference. Tukey's test (p ≤ 0.05).
Birefringent collagen content in capsules
Quantitative analysis (Fig. 6G) showed that the amount of birefringent collagen was similar between the groups (p ˃ 0.05) at 7 and 15 days. The amount of birefringent collagen was significantly higher in the BIO specimens than in the CAL specimens at 60 days (p < 0.0001). The analysis also revealed a significant increase for collagen in the BIO group at 60 days (p < 0.0001).
FIGURE 6Light micrographs showing portions of the capsule from sections subjected to picrosirius-red and analyzed under polarized illumination. At 7 days, fine birefringent collagen fibers (in red-orange) are dispersed in the capsules. At 60 days, the capsules show thick bundles of birefringent collagen; a marked birefringence is observed in the capsules of the BIO (D) and CG (F) groups compared to the CAL (E) group. Bars: 18 μm. (G) The graph shows the values (expressed as mean ± standard deviation) of the amount of birefringent collagen (in percentage) in the capsules. In each period, the comparison between the groups is indicated by superscript letters; different letters = significant difference. The superscript numbers indicate the analysis of each group over time; different numbers = significant difference. Tukey's test (p ≤ 0.05).
Conclusion: Compared to CAL specimens, BIO specimens showed immunoexpression of the pro-inflammatory cytokine IL-6 and a lower quantity of inflammatory cells throughout all periods. The reduction in these parameters was accompanied by a significant increase in collagen content and in the immunoexpression of IL-10, a cytokine involved in tissue repair, over time. Our findings indicate that Bio-C Temp is biocompatible and has no hepatotoxicity effect.