Consequences of cathepsin C inactivation for membrane exposure of proteinase 3, the target antigen in autoimmune vasculitis.

Membrane-bound proteinase 3 (PR3m) is the main target antigen of anti-neutrophil cytoplasmic autoantibodies (ANCA) in granulomatosis with polyangiitis, a systemic small-vessel vasculitis. Binding of ANCA to PR3m triggers neutrophil activation with the secretion of enzymatically active PR3 and related neutrophil serine proteases, thereby contributing to vascular damage. PR3 and related proteases are activated from pro-forms by the lysosomal cysteine protease cathepsin C (CatC) during neutrophil maturation. We hypothesized that pharmacological inhibition of CatC provides an effective measure to reduce PR3m and therefore has implications as a novel therapeutic approach in granulomatosis with polyangiitis. We first studied neutrophilic PR3 from 24 patients with Papillon-Lefèvre syndrome (PLS), a genetic form of CatC deficiency. PLS neutrophil lysates showed a largely reduced but still detectable (0.5-4%) PR3 activity when compared with healthy control cells. Despite extremely low levels of cellular PR3, the amount of constitutive PR3m expressed on the surface of quiescent neutrophils and the typical bimodal membrane distribution pattern were similar to what was observed in healthy neutrophils. However, following cell activation, there was no significant increase in the total amount of PR3m on PLS neutrophils, whereas the total amount of PR3m on healthy neutrophils was significantly increased. We then explored the effect of pharmacological CatC inhibition on PR3 stability in normal neutrophils using a potent cell-permeable CatC inhibitor and a CD34+ hematopoietic stem cell model. Human CD34+ hematopoietic stem cells were treated with the inhibitor during neutrophil differentiation over 10 days. We observed strong reductions in PR3m, cellular PR3 protein, and proteolytic PR3 activity, whereas neutrophil differentiation was not compromised.

Analysis of urinary cathepsin C for diagnosing Papillon-Lefèvre syndrome.

Papillon-Lefèvre syndrome (PLS) (OMIM: 245000) is a rare disease characterized by severe periodontitis and palmoplantar keratoderma. It is caused by mutations in both alleles of the cathepsin C (CatC) gene CTSC that completely abrogate the proteolytic activity of this cysteine proteinase. Most often, a genetic analysis to enable early and rapid diagnosis of PLS is unaffordable or unavailable. In this study, we tested the hypothesis that active CatC is constitutively excreted and can be easily traced in the urine of normal subjects. If this is true, determining its absence in the urine of patients would be an early, simple, reliable, low-cost and easy diagnostic technique. All 75 urine samples from healthy control subjects (aged 3 months to 80 years) contained proteolytically active CatC and its proform, as revealed by kinetic analysis and immunochemical detection. Of the urine samples of 31 patients with a PLS phenotype, 29 contained neither proteolytically active CatC nor the CatC antigen, so that the PLS diagnosis was confirmed. CatC was detected in the urine of the other two patients, and genetic analysis revealed no loss-of-function mutation in CTSC, indicating that they suffer from a PLS-like condition but not from PLS. Screening for the absence of urinary CatC activity soon after birth and early treatment before the onset of PLS manifestations will help to prevent aggressive periodontitis and loss of many teeth, and should considerably improve the quality of life of PLS patients.

In vivo evaluation of bioactive glass-based coatings on dental implants in a dog implantation model.

OBJECTIVES: Although titanium is commonly used as a favorable bone implant material due to its mechanical properties, its bioactive and osteoconductive capacity is relatively low. Calcium phosphate ceramics, predominantly hydroxyapatite (HA), have been frequently used for coating purposes to improve the bioactive properties. In view of the suggested osteopromotive capacity of bioactive glasses (BGs), this study aimed to evaluate the effect of BG incorporation into HA coatings on implant performance in terms of bone contact and bone area. MATERIALS AND METHODS: A total of 48 screw-type titanium implants with magnetron sputter coatings containing different ratios of HA and BG (HA, HABGLow, and HABGHigh; n = 8) were placed into the mandible of 16 Beagle dogs. After 4 and 12 weeks, their performance was evaluated histologically and histomorphometrically. Peri-implant bone area percentage (BA%) was determined in three zones (inner, 0-500 µm; middle, 500-1000 µm; and outer, 1000-1500 µm). Additionally, bone-to-implant contact (BIC%) and first bone-implant contact (1st BIC) were assessed for each sample. RESULTS: After 4 weeks, bone-to-implant contact for the HA- and HABGLow-coated groups was significantly higher (P < 0.05) than for the HABGHigh coatings. Mean values for overall BA% showed comparable values for both the HABGLow (58.3%)- and HABGHigh (56.3%)-coated groups. Data suggest that the relative BA around the HA-coated implants (67.8%) was higher, although this was only significant compared to the HABGHigh group. After 12 weeks, all three groups showed similar bone-to-implant contact and no differences in BA were found. CONCLUSIONS: The incorporation of BG into HA sputter coatings did not enhance the performance of a dental implant in implantations sites with good bone quality and quantity. On the contrary, coatings containing high concentrations of BG resulted in inferior performance during the early postimplantation healing phase.

Resolution, sensitivity, and in vivo application of high-resolution computed tomography for titanium-coated polymethyl methacrylate (PMMA) dental implants.

OBJECTIVES: The aims of this study were (i) to determine the spatial resolution and sensitivity of micro- versus nano-computed tomography (CT) techniques and (ii) to validate micro- versus nano-CT in a dog dental implant model, comparative to histological analysis. MATERIAL AND METHODS: To determine spatial resolution and sensitivity, standardized reference samples containing standardized nano- and microspheres were prepared in polymer and ceramic matrices. Thereafter, 10 titanium-coated polymer dental implants (3.2 mm in Ø by 4 mm in length) were placed in the mandible of Beagle dogs. Both micro- and nano-CT, as well as histological analyses, were performed. RESULTS: The reference samples confirmed the high resolution of the nano-CT system, which was capable of revealing sub-micron structures embedded in radiodense matrices. The dog implantation study and subsequent statistical analysis showed equal values for bone area and bone-implant contact measurements between micro-CT and histology. However, because of the limited sample size and field of view, nano-CT was not rendering reliable data representative of the entire bone-implant specimen. CONCLUSIONS: Micro-CT analysis is an efficient tool to quantitate bone healing parameters at the bone-implant interface, especially when using titanium-coated PMMA implants. Nano-CT is not suitable for such quantification, but reveals complementary morphological information rivaling histology, yet with the advantage of a 3D visualization.

Oral rehabilitation of a case of Papillon-Lefevre syndrome with dental implants.

Papillon-Lefevre syndrome (PLS) is a rare autosomal recessive disorder characterized by diffuse palmoplantar keratoderma and precocious aggressively progressing periodontitis, leading to the premature loss of deciduous and permanent teeth. Full mouth rehabilitation with implant supported fixed prosthesis provides numerous advantages over the conventional denture in terms of esthetics, function, longevity of the prosthesis, and patient satisfaction. We report a case of PLS in a 19-year-old female who underwent extraction of all the teeth followed by full mouth rehabilitation with implant supported fixed prostheses. A one-year follow up of the case showed functionally and esthetically stable dental implant with no signs of infection or bone loss.

Study of the osseointegration of dental implants placed with an adapted surgical technique.

OBJECTIVE: To study the osseointegration of dental implants placed with a modified surgical technique in Beagle dogs and to compare it with the conventional method. MATERIALS AND METHODS: Dental implants were placed bilaterally in the mandible of Beagle dogs using the press-fit as well as undersized implant bed preparation technique. Micro computer tomography (micro-CT) and histometric methods were used to analyze the bone implant contact and bone volume (BV) around the implants. RESULTS: The bone-to-implant contact percentage (BIC: expressed as %), first BIC (1st BIC: expressed in mm), sulcus depth (SD: expressed in mm) and connective tissue thickness (CT: expressed in mm) were analyzed for both groups. The BIC percentage was significantly higher for the undersized installed implants (P=0.0118). Also, a significant difference existed between the undersized and press-fit installed implants for the first screw thread showing bone contact (P=0.0145). There were no significant differences in mucosal response (SD and CT) for both installation procedures. Also, no significant difference was found in the BV, as measured using micro-CT, between the implants placed with an undersized technique (59.3 ± 4.6) compared with the press-fit implants (56.6 ± 4.3). CONCLUSION: From the observations of the study, it can be concluded that an undersized implant bed can enhance the implant-bone response.