Similarity on neural stem cells and brain tumor stem cells in transgenic brain tumor mouse models

Although it is believed that glioma is derived from brain tumor stem cells, the source and molecular signal pathways of these cells are still unclear. In this study, we used stable doxycycline-inducible transgenic mouse brain tumor models (c-myc⁺/SV40Tag⁺/Tet-on⁺) to explore the malignant trans-formation potential of neural stem cells by observing the differences of neural stem cells and brain tumor stem cells in the tumor models. Results showed that chromosome instability occurred in brain tumor stem cells. The numbers of cytolysosomes and autophagosomes in brain tumor stem cells and induced neural stem cells were lower and the proliferative activity was obviously stronger than that in normal neural stem cells. Normal neural stem cells could differentiate into glial fibrillary acidic protein-positive and microtubule associated protein-2-positive cells, which were also negative for nestin. However, glial fibrillary acidic protein/nestin, microtubule associated protein-2/nestin, and glial fibrillary acidic protein/microtubule associated protein-2 double-positive cells were found in induced neural stem cells and brain tumor stem cells. Results indicate that induced neural stem cells are similar to brain tumor stem cells, and are possibly the source of brain tumor stem cells.     

Efficacy of locally-delivered doxycycline microspheres in chronic localized periodontitis and on Porphyromonas gingivalis

Aim: This study aimed to assess the efficacy and effect of locally‐delivered doxycycline microspheres with scaling and root planing in periodontal pocket therapy and on Porphyromonas gingivalis, respectively. Methods: Twenty sites with a probing pocket depth of 4–6 mm were divided into two groups: a control group consisting of scaling and root planing, with one application of doxycycline microspheres only at baseline, and a test group consisting of scaling and root planing, with an application of doxycycline microspheres at baseline and 1 and 3 months. Clinical readings included the plaque index, gingival index, probing pocket depth, and relative attachment level. Rapid polymerase chain reaction method was used for the detection of P. gingivalis. Results: A statistically‐significant reduction in probing pocket depth and attachment gain was found in both groups; the test group showed a significant reduction in probing pocket depth and attachment gain compared with the control at 3 and 6 months. P. gingivalis cell count in the test group was significantly reduced at all the time periods, except from 1 to 3 months. Conclusion: Local drug delivery of doxycycline microspheres significantly improved the treatment outcomes in periodontal pocket therapy and reduced P. gingivalis in the periodontal pocket.     

Concentration of 3 tetracyclines in plasma, gingival crevice fluid and saliva

BACKGROUND: Systemically-administered tetracyclines have been used widely for treatment of periodontal diseases with little understanding of their delivery characteristics to periodontal tissues. This study was designed to measure concentrations of 3 tetracyclines in gingival crevice fluid (GCF), plasma and saliva of following systemic administration. METHOD: The concentration of tetracycline (TC), minocycline (MN) and doxycycline (DX) was measured in gingival crevice fluid (GCF), plasma and saliva of 20 subjects following single sequential standard oral systemic doses. Gingival crevice fluid concentration was measured at 4 sites (2 shallow and 2 deep) before administration, and at 1 h and 2 h following administration. Plasma and saliva concentrations were measured from in samples at the same time points. No antibacterial activity was detected before administration. The highest concentrations were measured 2 h after administration. RESULTS: The average concentrations at 2 h were highest in plasma (TC = 1.02, MN=2.18, DX=2.35 microg/ml). Intermediate concentrations were measured in GCF (TC=0.61, MN= 1.49, DX= 1.65 microg/ml). Saliva concentrations (TC=0.09 MN=0.31, DX=0.47 microg/ml) were the lowest of the 3 fluids monitored. Data are presented indicating that the average GCF concentration of systemically administered tetracyclines is less than the that of plasma concentration. The concentration of tetracyclines in GCF was strongly associated with plasma concentration, indicating a primary role of drug absorption in the delivery of these systemically administered antibiotics to the site of action in periodontal therapy. The average GCF concentration in individuals varied widely (between 0 and 8 microg/ml) with approximately 50% of samples not achieving levels of 1 microg/ml. CONCLUSION: These observations suggest that poor absorption of orally-administered tetracyclines in many individuals may account for much of the variability in clinical response to antibiotics observed in practice.     

Therapy with adjunctive doxycycline local delivery in patients with type 1 diabetes mellitus and periodontitis

OBJECTIVES: The purpose of this study was to evaluate the effect of subgingival administration of doxycycline as an adjunct to periodontal therapy in type 1 diabetes mellitus (DM) patients. MATERIAL AND METHODS: Twenty-two paired periodontal defects > or =5.0 mm were treated in 11 patients (35-55 years old). After initial therapy the sites were randomly assigned into test (scaling and root planing+subgingival administration of 10% doxycycline hyclate gel) or control (scaling and root planing+subgingival placebo gel) groups. The clinical parameters of clinical attachment level (CAL), probing depth (PD) and gingival margin level (GML) for recession determination were assessed at baseline, after 6 weeks, and 6, 9 and 12 months, using a computerized probe. Data were statistically evaluated using Duncan and F tests. RESULTS: Between study group comparisons indicated PD reduction and CAL gain were greater in the test group than in the control group at 6 weeks and 6, 9 and 12 months but only statistically significant at 12 months (p<0.05). Within study group comparisons indicated statistically significant differences were found for CAL and PD values favouring the adjunctive doxycycline group from baseline to 6 weeks and 6, 9 and 12 months (p<0.05). CONCLUSIONS: These findings suggest that subgingivally delivered doxycycline hyclate produces additional favorable clinical results to periodontal therapy in type 1 DM patients.     

α1-Proteinase inhibitor in gingival crevicular fluid of humans with adult periodontitis: serpinolytic inhibition by doxycycline

The serum protein, α₁-proteinase inhibitor (α₁PI), defends the host against serine proteinases, e.g. PMN elastase. Using a rabbit anti-serum against human α₁-PI, this protein in GCF was quantified from a standard curve constructed from dot-blot analysis and characterized by Western blot. GCF was collected on filter paper strips from healthy (H), gingivitis (G) and adult periodontitis (AP) patients, then extracted with Tris/NaCl/CaCl₂ buffer, pH 7.6. α₁-PI concentration increased with G and was highest in AP subjects. H sites only showed intact α₁-PI (52 kDa); no degradation fragments (48 kDa) were detected. In G and AP subjects, α₁-PI degradation fragments were seen in 17% and 71% of GCF samples, respectively. Both collagenase and α₁-Pl-degrading activities in GCF increased with severity of inflammation (GCF flow). Moreover, the α₁-PI degrading (or serpinolytic) activity was characterized as a matrix metalloproteinase, probably collagenase, based on its in vitro response to a panel of different proteinase inhibitors including doxycycline. We propose: (1) that collagenase promotes periodontal breakdown not only by degrading collagen, but also by depleting α₁-PI regulation of elastase and other serine-proteinases, thereby favoring a broader attack on extracellular matrix (ECM) constituents, and (2) based on a recent longitudinal double-blind study using the techniques described above for α₁-PI analysis, that low-dose doxycycline administration to humans with adult periodontitis can inhibit this broad cascade of ECM degradation.     

Effects of scaling and root planing and sub-antimicrobial dose doxycycline on oral and systemic biomarkers of disease in patients with both chronic periodontitis and coronary artery disease

OBJECTIVES: This study evaluated the effects of scaling and root planing (SRP) +/- sub-antimicrobial dose doxycycline (SDD) on gingival crevicular fluid (GCF) levels of matrix metalloproteinase (MMP) -1, -8, -13 and on serum levels of high-sensitivity C-reactive protein (HsCRP) and lipid fractions in patients with both chronic periodontitis (CP) and coronary artery disease (CAD). MATERIAL AND METHODS: Thirty-six patients were randomly distributed into two groups (Placebo or SDD; 6 weeks) and both received two regimens of SRP. At baseline and 6 weeks, GCF and blood were collected and clinical indices were recorded. MMPs, HsCRP and lipid fractions were assayed. RESULTS: There were statistically significant improvements for all clinical parameters, GCF volumes, GCF MMPs and serum levels of HsCRP, apolipoprotein-A (APO-A), high-density lipoprotein (HDL) and lipoprotein-a between pre- and post-treatment in both groups. Between groups, there were statistically significant greater improvements in pocket depth (PD), gingival index (GI), APO-A and HDL, favouring the group receiving SDD adjunctive to SRP (p < 0.05). CONCLUSION: Greater improvement was detected for PD and GI, and for serum levels of APO-A and HDL cholesterol when using SRP+SDD compared with SRP+placebo in this study. An investigation with larger numbers of patients and a longer duration of drug treatment is needed to confirm these preliminary findings.     

Subantimicrobial dose doxycycline effects on alveolar bone loss in post-menopausal women

AIM: Determine the efficacy of 2-year continuous subantimicrobial dose doxycycline (SDD; 20 mg bid) on alveolar bone in post-menopausal osteopenic, oestrogen-deficient women undergoing periodontal maintenance in a 2-year double-blind, placebo-controlled, randomized clinical trial. MATERIAL AND METHODS: One-hundred and twenty-eight subjects randomized to SDD or placebo (n=64 each). Posterior vertical bite wings taken at baseline, 1 and 2 years for alveolar bone density (ABD), using radiographic absorptiometry (RA) and computer-assisted densitometric image analysis (CADIA), and alveolar bone height (ABH). Statistical analyses utilized generalized estimating equations; primary analyses were intent to treat (ITT). Results are presented as SDD versus placebo. RESULTS: Under ITT, there was no statistically significant effect of SDD on ABD loss (RA: p=0.8; CADIA: p=0.2) or ABH loss (p=0.2). Most sites (81-95%) were inactive. For subgroup analyses, mean CADIA was higher with SDD for non-smokers (p=0.05) and baseline probing depths > or =5 mm (p=0.003). SDD was associated with 29% lower odds of more progressive ABH loss in women >5 years post-menopausal (p=0.05) and 36% lower among protocol-adherent subjects (p=0.03). CONCLUSIONS: In post-menopausal osteopenic women with periodontitis, SDD did not differ overall from placebo. Based on exploratory subgroup analyses, additional research is needed to determine the usefulness of SDD in non-smokers, subjects >5 years post-menopausal and in deeper pockets. Protocol registered at (ClinicalTrials.gov). Identifier: NCT00066027.     

Inflammatory response to chlorhexidine, minocycline HCl and doxycycline HCl in an in vivo mouse model

Aim: To examine the effect of locally delivered antimicrobial drugs on the inflammatory response in an in vivo mouse chamber model.
Material and Methods: Two weeks following chamber implantation, 24 BALB/c mice, in the experimental group, were given an intra-chamber challenge of heat-killed Porphyromonas gingivalis , followed immediately by injection of the specific antimicrobial drug: 2000  μ g/ml chlorhexidine (CHX); 1500  μ g/ml minocycline HCl;and 1500  μ g/ml doxycycline HCl (concentrations achieved in the periodontal pocket with commercial controlled-release delivery systems). A second group of 24 animals received only the antimicrobial treatment without P. gingivalis challenge. Intra-chamber exudates were sampled at 2 and 24 h following the challenge, and leucocytes, TNF α , IFN γ and IL-10 were evaluated.
Results: At 2 h, minocycline HCl induced high levels of IL-10, TNF α and IFN γ , while CHX reduced the levels of TNF α and IFN γ . By 24 h, these responses were attenuated. Following bacterial challenge, the antibacterial agents attenuated the inflammatory process, each in its own fashion.
Conclusions: Antibacterial agents applied locally have the ability to induce an inflammatory response. They also modify the inflammatory response to P. gingivalis independent of their antimicrobial effect. CHX and doxycycline HCl appear to have the most marked anti-inflammatory effect.     

The effect of topical doxycycline usage on gingival crevicular fluid MMP-8 levels of chronic and aggressive periodontitis patients: a pilot study

Abstract:  The aim of this study was to evaluate the efficacy of topical subgingival application of doxycycline hyclate (DH) gel adjunctive to non-surgical periodontal therapy on gingival crevicular fluid (GCF) matrix metalloproteinase (MMP)-8 levels in chronic and aggressive periodontitis patients. Forty teeth of 10 chronic periodontitis patients and 32 teeth of eight aggressive periodontitis patients were screened for 6 months. Scaling and root planing (SRP) was applied to the control sites and DH gel adjunctive to SRP was applied to the test sites of each patient simultaneously. GCF MMP-8 levels were analysed at baseline, 7 days; and at 1, 3 and 6 months by Sandwich Elisa Method. At 1, 3 and 6 months, probing depth ( P  < 0.0051) and plaque scores and bleeding on probing values ( P  = 0.000) significantly decreased in each group when compared with the baseline, but there was no statistically significant difference between the test and control sites. GCF MMP-8 levels reduced presenting statistically significant differences on 7 days, 1, 3 and 6 months in four of the groups ( P  < 0.05); however, intergroup differences were not statistically significant. Developing functional and immunological-based chair-side MMP tests might serve as useful adjunctive diagnostic tools when monitoring the effects of DH gel application.     

Susceptibility of Porphyromonas gingivalis in biofilms to amoxicillin,doxycycline and metronidazole

The susceptibility of Porphyromonas gingivalis to amoxicillin, doxycycline and metronidazole was determined by a standardized method taking into account the biofilm mode of growth of subgingival bacteria. The minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of 48-h biofilms of P. gingivalis established on membrane filters in a Modified Robbins Device were determined by agar dilution. The results were compared to (i) conventional MIC determinations, (ii) the susceptibility of planktonic cultures with cell numbers equal to those of the biofilms, and (iii) results for detached biofilm cells. The MICs of the biofilms of the six reference strains and clinical isolates containing 107-8 cells/filter were much higher than the conventional MIC values. However, the MIC of planktonic cultures of equal cell numbers also increased, indicating that an inoculum effect is part of the explanation of the increased resistance of biofilms. Still, the MBCs of biofilms were 2-8 times, and those for doxycycline up to 64 times, greater than the MBC values for planktonic cultures.