Transformation of Trichoderma species with dominant selectable markers

Summary We have developed a transformation system for Trichoderma hamatum and Trichoderma harzianum Rifai, using dominant markers for selection based on the Escherichia coli hygromycin B phosphotransferase gene (hph) and the -tubulin gene (bml) from Neurospora crassa, respectively. Transformation frequencies and protoplast regeneration were low in both species. All the T. hamatum hygromycin-resistant transformants analysed were mitotically stable, in contrast to those of T. harzianum derived by benomyl resistance, in which only 50% of the transformants analysed were stable. Molecular analysis of transformants showed the integration of the transforming DNA into the genome and indicated that the number and sites of integration varied among the transformants.     

Double-masked randomized clinical trial evaluating the effect of a triclosan/copolymer dentifrice on periodontal healing after one-stage full-mouth debridement

Background: This study evaluates the effect of triclosan/copolymer dentifrice on the 6‐month clinical response of patients with generalized severe chronic periodontitis (GSCP) treated with one‐stage, full‐mouth ultrasonic debridement (FMUD). Methods: Thirty patients diagnosed with GSCP (≥8 teeth presenting probing depth [PD] ≥5 mm and bleeding on probing [BOP]) were selected and randomly allocated to a control group (n = 15) subjected to FMUD and daily use of a placebo dentifrice or to a test group (n = 15) subjected to FMUD and daily use of a triclosan/copolymer dentifrice. Patients were analyzed for the following parameters: full‐mouth plaque index (FMPI), full‐mouth BOP score (FMBS), gingival recession, PD, and clinical attachment level (CAL). Patients were evaluated at 3 and 6 months by a calibrated and masked examiner. Results: Initially, the groups presented similar periodontal conditions, with no significant differences in any of the parameters evaluated (P >0.05). In both groups, improvements in all periodontal parameters (P <0.05) were seen at the completion of the experimental period. Additionally, the test group showed lower FMPI (3 months) and FMBS (3 and 6 months) than the control group (P <0.05). Moreover, the CAL gain was significantly greater in the test group, especially at initially deep pockets (PD ≤7 mm). Whereas in the control group the CAL gain in deep pockets was 2.7 ± 0.6 mm, in the test group the CAL gain was 3.6 ± 1.4 mm (P <0.05). Conclusion: Within the limits of the present study, the use of triclosan/copolymer dentifrice promoted additional clinical benefits in the treatment of GSCP treated by one‐stage FMUD.     

Impact of resveratrol on bone repair in rats exposed to cigarette smoke inhalation: histomorphometric and bone-related gene expression analysis

This study investigated the effect of resveratrol on bone healing and its influence on the gene expression of bone-related markers in rats exposed to cigarette smoke. Two calvarial defects were created in each of 60 rats, which were assigned equally (n = 20) to three groups: (1) resveratrol (10 mg/kg) + smoke exposure (SMK + RESV); (2) placebo + smoke exposure (SMK + PLA); or (3) placebo + no smoke exposure (NS + PLA). Substances were administered daily for 30 days following surgery. Smoke inhalation was started 7 days before surgery and continued for 30 days after surgery. One defect was processed for histomorphometric analysis and the other was used for mRNA quantification of bone-related gene expression by qPCR. The remaining defect was smaller in the SMK + RESV (2.27 ± 0.61 mm, P = 0.0003) and NS + PLA (2.17 ± 0.74 mm, P = 0.0005) groups than in the SMK + PLA group (3.12 ± 0.47 mm). Higher levels of Runx2 were observed in the NS + PLA group than in the smoke exposure groups (vs. SMK + PLA, P = 0002; vs. SMK + RESV, P = 0.052); levels of Lrp-5 were also higher in the no smoke exposure group (vs. SMK + RESV, P = 0.009; vs. SMK + PLA, P = 0.003). Resveratrol therapy decreased RANKL/OPG expression when compared to placebo (SMK + RESV vs. SMK + PLA, P = 0.017). Dkk1 levels were decreased in the SMK + RESV group when compared to the SMK + PLA (P = 0.006) and NS + PLA groups (P = 0.005). In conclusion, resveratrol optimizes the repair of critical-sized bone defects, up-regulating the gene expression of important bone remodelling markers in rats exposed to cigarette smoke inhalation.     

Occurrence of peri‐implant diseases and risk indicators at the patient and implant levels: A multilevel cross‐sectional study

Background : High prevalence rates of peri‐implant diseases have been reported; however, the lack of standardization of definition criteria has lead to variations in the observed estimates. In addition, scarce data are available concerning patient and implant related factors associated to peri‐implantitis. The aim of this study was to determine the prevalence of peri‐implant diseases and their risk indicators at the patient and implant levels. Methods : One hundred forty‐seven patients with 490 dental implants were included. Dental implants were clinically and radiographically evaluated to determine their peri‐implant conditions. Patient‐related conditions and implant and prosthetic‐related factors were recorded. Multivariable Poisson regression was fitted and prevalence ratios (PR) were reported. Results : 85.3% of implants (95%CI 80.2 to 90.4) had mucositis and 9.2% (95%CI 4.7 to 13.7) had peri‐implantitis. 80.9% (95%CI 73.8 to 86.8), and 19.1% (95%CI 12.6 to 25.5) of patients had mucositis and peri‐implantitis. At the patient level, it was observed an increased probability of peri‐implantitis in individuals with pocket depths ≥6 mm (PR = 2.47) and with ≥4 implants (PR = 1.96). Smoking increased the probability of peri‐implantitis by three times (PR = 3.49). The final multilevel Poisson regression model at the implant level indicated that platform switching reduced the probability of peri‐implantitis (PR = 0.18) and implants in function for ≥5 years increased this probability (PR = 2.11). The final model including patient and implant level indicators demonstrated that higher time of function (PR = 2.76) and smoking (PR = 6.59) were associated with peri‐implantitis. C onclusion : Peri‐implant diseases are highly prevalent in the studied sample, and factors associated with the occurrence of peri‐implantitis were presence of pockets ≥6 mm, smoking, time of function, and type of platform.     

Peri-Implant Repair Using a Modified Implant Macrogeometry in Diabetic Rats: Biomechanical and Molecular Analyses of Bone-Related Markers

DM has a high prevalence worldwide and exerts a negative influence on bone repair around dental implants. Modifications of the microgeometry of implants have been related to positive results in bone repair. This study assessed, for the first time, the influence of an implant with modified macrodesign based on the presence of a healing chamber in the pattern of peri-implant repair under diabetic conditions. Thirty Wistar rats were assigned to receive one titanium implant in each tibia (Control Implant (conventional macrogeometry) or Test Implant (modified macrogeometry)) according to the following groups: Non-DM + Control Implant; Non-DM + Test Implant; DM + Control Implant; DM + Test Implant. One month from the surgeries, the implants were removed for counter-torque, and the bone tissue surrounding the implants was stored for the mRNA quantification of bone-related markers. Implants located on DM animals presented lower counter-torque values in comparison with Non-DM ones, independently of macrodesign (p < 0.05). Besides, higher biomechanical retention levels were observed in implants with modified macrogeometry than in the controls in both Non-DM and DM groups (p < 0.05). Moreover, the modified macrogeometry upregulated OPN mRNA in comparison with the control group in Non-DM and DM rats (p < 0.05). Peri-implant bone repair may profit from the use of implants with modified macrogeometry in the presence of diabetes mellitus, as they offer higher biomechanical retention and positive modulation of important bone markers in peri-implant bone tissue.     

Influence of Dyslipidemia and Diabetes Mellitus on Chronic Periodontal Disease

Background: Periodontal disease is closely related to certain systemic conditions, such as type 2 diabetes mellitus (DM2), and, as recently described, dyslipidemia, a condition with alterations in blood lipids levels. However, more than acting as disease modifiers, these conditions commonly occur as comorbidities, possibly synergically affecting periodontal tissues. The aim of the current study is to identify whether DM2 and dyslipidemia are related to the occurrence and severity of chronic periodontitis. Methods: A total of 254 individuals participated: 56 were patients with DM2, 67 had dyslipidemia, 74 had DM2 and dyslipidemia, and 57 were systemically healthy individuals. The clinical examination included a full‐mouth evaluation of periodontal probing depth, plaque score, bleeding on probing, and clinical attachment level (CAL). Blood samples were taken to assess fasting plasma glucose, low‐density lipoprotein, high‐density lipoprotein, and triglyceride levels. These parameters, as well as other medical conditions (i.e., smoking habits and body mass index), were considered in multiple regression analyses for data analyses (α = 5%). Results: Dyslipidemia was not related to periodontal disease (P >0.05). At the same time, DM2, age, and smoking showed a statistical and positive association, an increase in percentage of sites with CAL ≥3 and ≥5 mm. Regarding the percentage of sites presenting severe destruction (CAL ≥7 mm), only DM2 remained a significant risk factor (P <0.05). Conclusions: It could be concluded that dyslipidemia did not influence periodontal conditions in participants with normal health or those with DM2. However, age, smoking habits, and especially DM2 were significantly associated with loss of CAL.     

CHARACTERIZATION OF BONE CELLS OBTAINED FROM THE CALVARIA OF NEONATAL RATS (OSTEO-1) AFTER SERIAL SUBCULTURE

The objective of the present study was to characterize bone cells grown in two culture media, and to determine the effective concentration of OP-1 on the growth of osteo-1 cells. Subcultured rat bone cells (osteo-1) were grown in alpha-modified Eagle’s minimal essential medium (α-MEM) and Dulbecco’s modified Eagle’s medium (DMEM) and total protein content, alkaline phosphatase activity and the formation of mineralized nodules were evaluated after 7, 14 and 21 days. Cells were exposed to different concentrations of rhOP-1 for 1, 3, 5 and 7 days and compared with an untreated control. Osteo-1 cells presented a significant increase in alkaline phosphatase activity and calcium deposits were observed at 21 days. Cells treated with 10 and 20 ng/mL rhOP-1 for 24 h showed a significant increase in cell viability when compared to control. Osteo-1 cells cultured on DMEM demonstrated an osteoblastic phenotype as indicated by high alkaline phosphatase activity and the presence of calcified nodules. The results suggest that low concentrations of OP-1 may promote an osteogenic effect on osteo-1 cells.