Perry R. Klokkevold

The effect of chitosan (poly-N-acetyl glucosamine) on lingual hemostasis in heparinized rabbits☆

PURPOSE The purpose of this study was to evaluate the effect of chitosan on lingual hemostasis in rabbits whose coagulation pathway had been impaired by administration of intravenous heparin. MATERIALS AND METHODS Bleeding times were measured for bilateral (15 mm x 2 mm) tongue incisions in 10 New Zealand white rabbits. Using a randomized, blinded experimental design, one incision in each animal was treated with chitosan, and the other was treated with the control vehicle without chitosan. Activated coagulation times and extraoral bleeding times were measured for each animal before, during, and after heparinization. RESULTS Intravenous infusion of heparin more than tripled the mean activated coagulation time and increased mean systemic bleeding time by 40%. In this heparinized animal model, lingual incisions receiving the experimental substance showed a 43% improvement in bleeding time as compared with lingual incisions receiving the control solution (P< or =.001). Chitosan treatment brought bleeding time of the lingual incision for heparinized animals within the normal range. Scanning electron microscopic evaluation of the incisions treated with chitosan showed an altered red blood cell morphology and an unusual affinity between erythrocytes. CONCLUSIONS Topical application of chitosan to lingual incisions effectively decreased intraoral bleeding time in a therapeutically anticoagulated (heparinized) rabbit model. Chitosan facilitated lingual hemostasis, possibly through interaction with erythrocytes, linking them together to establish a cellular clot or hemostatic plug.

Tissue-engineered growth of bone by marrow cell transplantation using porous calcium metaphosphate matrices

In this study we investigated not only osteoblastic cell proliferation and differentiation on the surface of calcium metaphosphate (CMP) matrices in vitro but also bone formation by ectopic implantation of these cell-matrix constructs in athymic mice in vivo. Interconnected porous CMP matrices with pores 200 microm in size were prepared to use as scaffolds for rat-marrow stromal-cell attachment. Cell-matrix constructs were cultured in vitro, and cell proliferation and ALPase activities were monitored for 56 days. In addition to their being cultured in vitro, cell-matrix constructs were implanted into subcutaneous sites of athymic mice. In vitro these porous CMP matrices supported the proliferation of osteoblastic cells as well as their differentiation, as indicated by high ALPase activity. In vivo the transplanted marrow cells gave rise to bone tissues in the pores of the CMP matrices. A small amount of woven bone formation was detected first at 4 weeks; osteogenesis progressed vigorously with time, and thick lamellar bones that had been remodeled were observed at 12 weeks. These findings demonstrate the potential for using a porous CMP matrix as a biodegradable scaffold ex vivo along with attached marrow-derived mesenchymal cells for transplantation into a site for bone regeneration in vivo.

Effect of chitosan on lingual hemostasis in rabbits.

Bleeding times were measured for bilateral (15 mm x 2 mm) tongue incisions in 14 New Zealand white rabbits. Using a randomized, blinded experimental design, one incision in each animal was treated with chitosan and the other was treated with control vehicle without chitosan. Extraoral bleeding and coagulation times were also measured for each animal preoperatively, postoperatively, and prior to killing to verify normal bleeding parameters and to evaluate possible systemic effects associated with topical application. Comparison of lingual incisions receiving the experimental substance versus those receiving control solution showed enhanced hemostasis manifested by a 32% (P less than .05) decrease in bleeding time.

Effect of chitosan on lingual hemostasis in rabbits with platelet dysfunction induced by epoprostenol

Chitosan, a complex carbohydrate derivative of shellfish exoskeleton, is shown to enhance lingual hemostasis in rabbits treated with a known antagonist of platelet function, epoprostenol (prostacyclin or PGI2). Bleeding times were measured for bilateral (15 mm x 2 mm) tongue incisions in 10 New Zealand white rabbits. Using a randomized, blinded experimental design, one incision in each animal was treated with chitosan and the other was treated with control vehicle without chitosan. Extraoral bleeding and coagulation times were measured for each animal before, during, and after infusion of epoprostenol. Continuous infusion of epoprostenol increased mean systemic bleeding time 95%. In this platelet dysfunction animal model, lingual incisions receiving the experimental substance showed a 56% improvement in bleeding time in comparison with lingual incisions receiving control solution (P = .003).

Optical protein sensor for detecting cancer markers in saliva

A surface immobilized optical protein sensor has been utilized to detect Interleukin-8 (IL-8) protein, an oral cancer marker, and can reach limit of detection (LOD) at 1.1 pM in buffer without using enzymatic amplification. Only after applying enzymatic amplification to increase the signal level by a few orders of magnitude, ELISA can reach the LOD of 1 pM level. We then develop the confocal optics based sensor for further reducing the optical noise and can extend the LOD of the surface immobilized optical protein sensor two orders in magnitude. These improvements have allowed us to detect IL-8 protein at 4.0 fM in buffer. In addition, these sensitive LODs were achieved without the use of enzymatic signal amplification, such that the simplified protocol can further facilitate the development of point-of-care devices. The ultra sensitive optical protein sensor presented in this paper has a wide number of applications in disease diagnoses. Measurements for detecting biomarkers in clinical sample are much more challenging than the measurements in buffer, due to high background noise contributed by large collections of non-target molecules. We used clinical saliva samples to validate the functionality of the optical protein sensor. Clinical detection of disease-specific biomarkers in saliva offers a non-invasive, alternative approach to using blood or urine. Currently, the main challenge of using saliva as a diagnostic fluid is its inherently low concentration of biomarkers. We compare the measurements of 40 saliva samples; half from oral cancer patients and half from a control group. The data measured by the optical protein sensor is compared with the traditional Enzyme-Linked Immunosorbant Assay (ELISA) values to validate the accuracy of our system. These positive results enable us to proceed to using confocal optical protein sensor to detect other biomarkers, which have much lower concentrations.

Anionic carbohydrate-containing chitosan scaffolds for bone regeneration

Scaffolds derived from naturally occurring polysaccharides have attracted significant interest in bone tissue engineering due to their excellent biocompatibility and hydrophilic nature favorable for cell attachment. In this study, we developed composite chitosan (CH) scaffolds containing anionic carbohydrate, such as chondroitin 4-sulfate (CS) or alginate (AG), with biomimetic apatite layer on their surfaces, and investigate their capacity to deliver progenitor cells (bone marrow stromal cells, BMSC) and model proteins with net-positive (histone) and net-negative charge (bovine serum albumin, BSA). The incorporation of CS or AG in CH scaffolds increased compressive modulus of the scaffolds and enhanced apatite formation. Initial burst release of histone was significantly higher than that of BSA from CH scaffold, while the addition of CS or AG in the scaffolds significantly reduced the initial burst release of histone, indicating strong electrostatic interaction between histone and negatively charged CS or AG. The apatite layer created on scaffold surfaces significantly reduced the initial burst release of both BSA and histone. Furthermore, apatite-coated scaffolds enhanced spreading, proliferation, and osteogenic differentiation of BMSC seeded on the scaffolds compared to non-coated scaffolds as assessed by live/dead and alamarBlue assays, scanning electron microscopy (SEM), alkaline phosphatase (ALP) activity, and Picrosirius red staining. This study suggests that apatite-coated CH/CS composite scaffolds have the potential as a promising osteogenic system for bone tissue engineering applications.

Dynamic Changes in the Subgingival Microbiome and Their Potential for Diagnosis and Prognosis of Periodontitis

ABSTRACT The human microbiome influences and reflects the health or disease state of the host. Periodontitis, a disease affecting about half of American adults, is associated with alterations in the subgingival microbiome of individual tooth sites. Although it can be treated, the disease can reoccur and may progress without symptoms. Without prognostic markers, follow-up examinations are required to assess reoccurrence and disease progression and to determine the need for additional treatments. To better identify and predict the disease progression, we aim to determine whether the subgingival microbiome can serve as a diagnosis and prognosis indicator. Using metagenomic shotgun sequencing, we characterized the dynamic changes in the subgingival microbiome in periodontitis patients before and after treatment at the same tooth sites. At the taxonomic composition level, the periodontitis-associated microorganisms were significantly shifted from highly correlated in the diseased state to poorly correlated after treatment, suggesting that coordinated interactions among the pathogenic microorganisms are essential to disease pathogenesis. At the functional level, we identified disease-associated pathways that were significantly altered in relative abundance in the two states. Furthermore, using the subgingival microbiome profile, we were able to classify the samples to their clinical states with an accuracy of 81.1%. Follow-up clinical examination of the sampled sites supported the predictive power of the microbiome profile on disease progression. Our study revealed the dynamic changes in the subgingival microbiome contributing to periodontitis and suggested potential clinical applications of monitoring the subgingival microbiome as an indicator in disease diagnosis and prognosis. IMPORTANCE Periodontitis is a common oral disease. Although it can be treated, the disease may reoccur without obvious symptoms. Current clinical examination parameters are useful in disease diagnosis but cannot adequately predict the outcome of individual tooth sites after treatment. A link between the subgingival microbiota and periodontitis was identified previously; however, it remains to be investigated whether the microbiome can serve as a diagnostic and prognostic indicator. In this study, for the first time, we characterized the subgingival microbiome of individual tooth sites before and after treatment using a large-scale metagenomic analysis. Our longitudinal study revealed changes in the microbiota in taxonomic composition, cooccurrence of subgingival microorganisms, and functional composition. Using the microbiome profiles, we were able to classify the clinical states of subgingival plaque samples with a high accuracy. Follow-up clinical examination of sampled sites indicates that the subgingival microbiome profile shows promise for the development of diagnostic and prognostic tools. Periodontitis is a common oral disease. Although it can be treated, the disease may reoccur without obvious symptoms. Current clinical examination parameters are useful in disease diagnosis but cannot adequately predict the outcome of individual tooth sites after treatment. A link between the subgingival microbiota and periodontitis was identified previously; however, it remains to be investigated whether the microbiome can serve as a diagnostic and prognostic indicator. In this study, for the first time, we characterized the subgingival microbiome of individual tooth sites before and after treatment using a large-scale metagenomic analysis. Our longitudinal study revealed changes in the microbiota in taxonomic composition, cooccurrence of subgingival microorganisms, and functional composition. Using the microbiome profiles, we were able to classify the clinical states of subgingival plaque samples with a high accuracy. Follow-up clinical examination of sampled sites indicates that the subgingival microbiome profile shows promise for the development of diagnostic and prognostic tools.

Periodontal regeneration - furcation defects: a consensus report from the AAP Regeneration Workshop.

BACKGROUND Treatment of furcation defects is a core component of periodontal therapy. The goal of this consensus report is to critically appraise the evidence and to subsequently present interpretive conclusions regarding the effectiveness of regenerative therapy for the treatment of furcation defects and recommendations for future research in this area. METHODS A systematic review was conducted before the consensus meeting. This review aims to evaluate and present the available evidence regarding the effectiveness of different regenerative approaches for the treatment of furcation defects in specific clinical scenarios compared with conventional surgical therapy. During the meeting, the outcomes of the systematic review, as well as other pertinent sources of evidence, were discussed by a committee of nine members. The consensus group members submitted additional material for consideration by the group in advance and at the time of the meeting. The group agreed on a comprehensive summary of the evidence and also formulated recommendations for the treatment of furcation defects via regenerative therapies and the conduction of future studies. RESULTS Histologic proof of periodontal regeneration after the application of a combined regenerative therapy for the treatment of maxillary facial, mesial, distal, and mandibular facial or lingual Class II furcation defects has been demonstrated in several studies. Evidence of histologic periodontal regeneration in mandibular Class III defects is limited to one case report. Favorable outcomes after regenerative therapy for maxillary Class III furcation defects are limited to clinical case reports. In Class I furcation defects, regenerative therapy may be beneficial in certain clinical scenarios, although generally Class I furcation defects may be treated predictably with non-regenerative therapies. There is a paucity of data regarding quantifiable patient-reported outcomes after surgical treatment of furcation defects. CONCLUSIONS Based on the available evidence, it was concluded that regenerative therapy is a viable option to achieve predictable outcomes for the treatment of furcation defects in certain clinical scenarios. Future research should test the efficacy of novel regenerative approaches that have the potential to enhance the effectiveness of therapy in clinical scenarios associated historically with less predictable outcomes. Additionally, future studies should place emphasis on histologic demonstration of periodontal regeneration in humans and also include validated patient-reported outcomes. CLINICAL RECOMMENDATIONS Based on the prevailing evidence, the following clinical recommendations could be offered. 1) Periodontal regeneration has been established as a viable therapeutic option for the treatment of various furcation defects, among which Class II defects represent a highly predictable scenario. Hence, regenerative periodontal therapy should be considered before resective therapy or extraction; 2) The application of a combined therapeutic approach (i.e., barrier, bone replacement graft with or without biologics) appears to offer an advantage over monotherapeutic algorithms; 3) To achieve predictable regenerative outcomes in the treatment of furcation defects, adverse systemic and local factors should be evaluated and controlled when possible; 4) Stringent postoperative care and subsequent supportive periodontal therapy are essential to achieve sustainable long-term regenerative outcomes.

Mental nerve neuropathy: A symptom of Waldenström's macroglobulinemia

The uncontrolled production of immunoglobulins in Waldenströms macroglobulinemia, a neoplastic plasma cell disorder, results in hyperviscosity of blood and multisystem organ derangement. Previously reported orofacial manifestations have been limited to osteolytic lesions of the jaw, ulcerations of the mucosa and tongue, gingival bleeding, and enlargement of salivary and lacrimal glands. Mental nerve anesthesia resulting from IgM deposition on the myelin sheath is a previously unreported early warning sign of advancing disease. Its occurrence heralds the need for aggressive medical management.

Effect of Implant Diameter and Ridge Dimension on Stress Distribution in Mandibular First Molar Sites—A Photoelastic Study

The long-term clinical success of a dental implant is dependent upon maintaining sufficient osseointegration to resist forces of occlusion. The purpose of this study was to investigate the effect of implant diameter on stress distribution around screw-type dental implants in mandibular first molar sites using photoelastic models. The design included models with different buccal-lingual dimension. Twelve composite photoelastic models were assembled using 2 different resins to simulate trabecular and cortical bone. Half of the models were fabricated with average dimensions for ridge width and the other half with narrower buccal-lingual dimensions. One internal connection implant (13 mm length) with either a standard (4 mm), wide (5 mm), or narrow (3.3 mm) diameter was embedded in the first molar position of each photoelastic model. Half the implants were tapered and the other half were straight. Full gold crowns in the shape of a mandibular first molar were fabricated and attached to the implants. Vertical and angled loads of 15 and 30 pounds were applied to specific points on the crown. Wide-diameter implants produced the least stress in all ridges while narrow-diameter implants generated the highest stress, especially in narrow ridges. It may be that the volume and quality of bone surrounding implants influences stress distribution with a greater ratio of cortical to trabecular bone, thus providing better support. Models with wide-diameter implants loaded axially had a more symmetrical stress distribution compared to standard and narrow diameter implants. A more asymmetrical stress pattern developed along the entire implant length with angled loads. Implant diameter and ridge width had considerable influence on stress distribution. Narrow-diameter implants produced more stress than wide diameter implants in all conditions tested.