[Effects of Bruton's tyrosine kinase on the proliferation and differentiation of osteoclasts].

OBJECTIVE: To observe the effect of Bruton's tyrosine kinase (BTK) on the proliferation and differentiation of osteoclasts and to explore the mechanism of BTK on bone destruction in periapical periodontitis. METHODS: After RAW264.7 cells induced with 100 ng·L⁻¹ receptor activator for nuclear factor-κB ligand (RANKL) for 5 days, osteoclast induction was confirmed by light microscopy, tartrate-resistant acid phosphatase (TRAP) staining, and quantitative real-time PCR (RT-qPCR). Then, BTK-small interfering RNA (BTK-siRNA) was transfected into cells induced for 5 days. After 24 h, the expression of TRAP mRNA was measured using RT-qPCR, and the proliferation and differentiation of osteoclasts were detected using CCK-8 and TRAP activity assay. Statistical analysis was performed. RESULTS: After RAW264.7 was induced with RANKL for 5 days, a large number of round, ellipse, irregularly protuberant, and TRAP-positive macrophages were observed under light microscopy. The expression of TRAP mRNA significantly reduced after 24 h of BTK-siRNA transfection (P<0.05). The detection of CCK-8 and TRAP activities showed that the proliferation and differentiation of osteoclasts significantly decreased (P<0.05). CONCLUSIONS: Silencing of BTK can inhibit the proliferation and differentiation of osteoclasts. BTK can be used as a new target for the inhibition of osteoclasts.

[Analysis of relationship between the detection of Enterococcus faecalis in post-treatment endodontic disease and clinical symptom or sign].

OBJECTIVE: To detect the Enterococcus faecalis (E. faecalis) in post-treatment endodontic disease, and to analyze the relationship between the occurrence of E. faecalis and clinical symptom. METHODS: 108 teeth which need root canal retreatment were collected, and the clinical symptoms and physical signs were recorded. Bacterium samples from root canal were taken, and genome DNA from bacterial samples were extracted. The occurrence of E. faecalis by means of the polymerase chain reaction was investigated. RESULTS: The detection rate of E. faecalis in cases of root canal retreatment was 47.2%, while in cases with symptoms or signs, or cases with both symptoms and signs, the root canal E. faecalis detection rates were 52.6%, 57.9%, 62.5%. The detection rates of E. faecalis between cases with clinical symptom and without clinical symptom demonstrated statistical significance (P < 0.05). The detection rates between cases with both clinical symptom and manifestly aneretic root and cases without clinical symptom and manifestly aneretic root had statistical significance (P < 0.05). In the group of clinical symptom, the detection rate of E. faecalis in cases with biting pain was 66.7%, clearly higher than those without biting pain (P < 0.05). CONCLUSION: The occurrence of E. faecalis in cases of root canal retreatment correlates with clinical symptoms.

[Use of atomic force microscopy to observe the surface and the dynamic forming process of biofilms of Enterococcus faecalis].

OBJECTIVE: To observe the surface of Enterococcus faecalis and the dynamic forming process of those biofilms using atomic force microscopy (AFM) in air condition. METHODS: The surface of Enterococcus faecalis which were dried in air were observed with AFM. We used the cellulose nitrate film to construct the Enterococcus faecalis biofilms model in vitro, and then placed the biofilms under AFM to observe the surface changes of biofilms' development. RESULTS: The cell surfaces of strain Enterococcus faecalis were not regular because of the presence of the amorphous substance on the colony surface, which congregated globular, fibrous structure. Gradually determined that at 6 h the initial biofilm formed and at 24 h the biofilms maintained the steady-state. AFM height images showed topographical changes due to biofilms' development, which were used to characterize several aspects of the bacterial surface, such as the presence of extracellular polymeric substance, and the biofilms' development stage. CONCLUSION: Application of AFM in physiological conditions could be useful in observing Enterococcus faecalis surface ultrastructure and dynamic process of biofilms formation.

[Effect of Enterococcus faecalis on apoptosis rate and the release of matrix metalloproteinase-8 of polymorphonuclear leukocytes].

OBJECTIVE: To evaluate the release of matrix metalloproteinase-8 (MMP-8) and apoptosis rate of polymorphonuclear leukocytes (PMNs) after PMNs was triggered by Enterococcus faecalis (E. faecalis) in vitro. METHODS: The activated E. faecalis suspension was prepared and added to PMNs suspension as experiment group. As a positive control, phorbol myristate acetate (PMA) was used. As negative control, PMNs suspension was incubated with PBS. The release of MMP-8 was measured at 0, 20, 60, 120 min by ELISA method. E. faecalis lysate acted on PMNs as experiment group, PMNs suspension was incubated with PBS as negative control, samples in two groups were incubated at 37 degrees C for 2, 5, 10, 15 h. The apoptosis rate of PMNs was tested by Flow Cytometry. RESULTS: At 0 min, there was no significant difference of MMP-8 release in the experiment group and positive control (P>0.01); whereas at 60, 120 min, E. faecalis induced a significant lower MMP-8 release compared with the positive control (P<0.01). The apoptosis rate of PMNs in both groups increased along with time, and apoptotic rate in experiment group was higher than that in the control group at 2, 5, 10, 15 h (P<0.01). CONCLUSION: After E. faecalis act on PMNs, no significant release of MMP-8 from PMNs was observed. E. faecalis don't induce PMNs apoptosis delay.

[Effect of different decalcification condition on tooth-clearing technique].

OBJECTIVE: To evaluate shrinkage range of cleared teeth caused by nitric acid with different temperature and concentration. METHODS: 48 human teeth were root canal-prepared and filled, then randomly and averagely divided into six groups on the basis of temperature and density of nitric acid and the condition of whether or not added the oscillate. Group A was 20 degrees C with 6% nitric acid, group B was 20 degrees C with 6% nitric acid and oscillate, group C was 20 degrees C with 3% nitric acid, group D was 20 degrees C with 3% nitric acid and oscillate, group E was 30 degrees C with 6% nitric acid and oscillate, group F was 30 degrees C with 3% nitric acid and oscillate. After achieving the standard of the decalcification, all the specimens were gradually dehydrated, and then cleared and conserved using methyl salicylate. Time-consumed and shrinkage range of all the specimens were recorded and analyzed. RESULTS: The time of decalcification in group E was the fastest, then was group F, group B. Group C was the last one. The anastole of the specimens was group E > group B > group A, group F > group D > group C, group B > group D, group E > group D, there was significant difference (P < 0.05). Group C had significant difference with other groups (P < 0.05). The anastole rate of the specimens had no significant difference between group A and group B, group C and group D, group B and group F, group D and group F. CONCLUSION: In 20 degrees C, 3% nitric acid with oscillate to carry out the decalcification can use less time and get less anastole. The result of the tooth-clearing technique is the best.