Inflammatory biomarkers in dentinal fluid as an approach to molecular diagnostics in pulpitis.

AIM: To explore a set of inflammatory biomarkers obtained from dentinal fluid (DF) from patients with symptomatic irreversible pulpitis (IP), reversible pulpitis (RP) and normal pulp (NP). METHODOLOGY: A cross-sectional exploratory study was performed, recruiting 64 patients on the basis of their respective pulp condition. DF samples were obtained from all patients (23, from IP patients; 20, from RP patients; and 21, from NP patients). Quantification of biomarkers was performed using a Luminex® MAGPIX platform system and multiplex assay kits. The Kruskal-Wallis test was used for comparisons with regard to pulp state. A simple logistic regression model and the odds ratio (OR) with a 95% level of confidence (P = 0.05) were used to evaluate associations between biomarker levels and pulpal diagnosis. The performance discrimination of the biomarkers was evaluated through the construction of a receiver operating characteristic (ROC) curve by calculating the area under the curve (AUC) for IP versus RP after logistic regression modelling. Youden criteria were used to establish cut-off points for biomarkers alone with AUC > 70 and P-value < 0.05, or estimated probabilities from the multivariable logistic model. RESULTS: The biomarkers that had significantly higher values in participants with IP versus RP were IL-1α, VEGF-α and FGF acid (P < 0.05). FGF acid (OR: 12.62; P = 0.0085; CI 95% 1.91-83.29) and VEGF-α (OR: 2.61; P = 0.0252; CI 95% 1.13-6.03) were associated with pulp diagnoses of IP versus RP. The AUC-ROC curve for FGF acid was 0.79. The model containing FGF acid, IL-1α, IL-6 and TIMP-1 had an AUC-ROC of 0.92 for IP versus RP with a significant difference from the FGF acid ROC curve (P = 0.0231). CONCLUSIONS: Dentinal fluid could be used to assay pulpal mediators in the molecular diagnosis of pulpitis. Despite the limitation of the clinical diagnostics used in the present study, it was possible to detect a difference between irreversible symptomatic pulpitis and reversible pulpitis associated with the following combined biomarkers: FGF acid + IL-6 + IL-1α, +TIMP-1.

The Four Pillars for Successful Regenerative Therapy in Endodontics: Stem Cells, Biomaterials, Growth Factors, and Their Synergistic Interactions.

Endodontics has made significant progress in regenerative approaches in recent years, thanks to advances in biologically based procedures or regenerative endodontic therapy (RET). In recent years, our profession has witnessed a clear conceptual shift in this therapy. RET was initially based on a blood clot induced by apical bleeding without harvesting the patient's cells or cell-free RET. Later, the RET encompassed the three principles of tissue engineering, stromal/stem cells, scaffolds, and growth factors, aiming for the regeneration of a functional dentin pulp complex. The regenerated dental pulp will recover the protective mechanisms including innate immunity, tertiary dentin formation, and pain sensitivity. This comprehensive review covers the basic knowledge and practical information for translational applications of stem cell-based RET and tissue engineering procedures for the regeneration of dental pulp. It will also provide overall information on the emerging technologies in biological and synthetic matrices, biomaterials, and signaling molecules, recent advances in stem cell therapy, and updated experimental results. This review brings useful and timely clinical evidence for practitioners to understand the challenges faced for a successful cell-based RET and the importance of preserving or reestablishing tooth vitality. The clinical translation of these current bioengineering approaches will undoubtedly be beneficial to the future practice of endodontics.

Cell-Based Regenerative Endodontics for Treatment of Periapical Lesions: A Randomized, Controlled Phase I/II Clinical Trial.

A randomized controlled phase I/II clinical trial was designed to evaluate the safety and efficacy of encapsulated human umbilical cord mesenchymal stem cells in a plasma-derived biomaterial for regenerative endodontic procedures (REPs) in mature permanent teeth with apical lesions. The trial included 36 patients with mature incisors, canines, or mandibular premolars showing pulp necrosis and apical periodontitis. Patients were randomly and equally allocated between experimental (REP) or conventional root canal treatment (ENDO) groups. On the first visit, cavity access and mechanical preparation of the root canal were performed. Calcium hydroxide medication was used, and the cavity was sealed. Three weeks later, patients were treated following their assigned protocol of ENDO or REP. Clinical follow-up examinations were performed at 6 and 12 mo. Categorical variables were evaluated by Fisher's exact test. Quantitative variables were compared using the Mann-Whitney test. The evolution over time of the percentage of perfusion units and the dimensions of lesion and cortical compromise were explored. After the 12-mo follow-up, no adverse events were reported, and the patients showed 100% clinical efficacy in both groups. Interestingly, in the REP group, the perfusion unit percentage measured by laser Doppler flowmetry revealed an increase from 60.6% to 78.1% between baseline and 12-mo follow-up. Sensitivity tests revealed an increase of the positive pulp response in the REP group at 12-mo follow-up (from 6% to 56% on the cold test, from 0% to 28% on the hot test, and from 17% to 50% on the electrical test). We present the first clinical safety and efficacy evidence of the endodontic use of allogenic umbilical cord mesenchymal stem cells encapsulated in a plasma-derived biomaterial. The innovative approach, based on biological principles that promote dentin-pulp regeneration, presents a promising alternative for the treatment of periapical pathology (ClinicalTrials.gov NCT03102879).

Effectiveness of selected premilking teat-cleaning regimes in reducing teat microbial load on commercial dairy farms.

AIMS: To determine the effectiveness of premilking teat-cleaning regimes in reducing the teat microbial load and effect on milk quality. METHODS AND RESULTS: The effectiveness of several premilking teat-cleaning regimes in reducing teat microbial load was assessed using 40 cows on each of the four commercial UK dairy farms with herringbone parlours during two sampling periods. In the first experiment, all the treatments reduced teat total viable count (TVC), but there was no significant difference between the hypochlorite wash and dry wipe, iodine dip and dry and alcohol-medicated wipe or dry wipe alone. In the second experiment, the chlorine wash and dry wipe was significantly more effective in reducing teat TVC than a water wash and dry, chlorine dip and dry or a dry wipe. There was no relationship between cleaning regime and milk TVC, Enterobacteriaceae or Escherichia coli levels. CONCLUSIONS: All of the cleaning techniques studied reduced teat microbial load, however, the chlorine wash and dry was the most effective. SIGNIFICANCE AND IMPACT OF THE STUDY: The premilking teat-cleaning techniques studied reduced the teat microbial load and therefore reduced the potential for milk contamination; however, a wash including an effective disinfectant followed by a dry wipe was the most effective.

Formulation of integrated control of Boophilus microplus in Paraguay: analysis of natural infestations.

A 3-year study into the seasonal variation in the numbers of Boophilus microplus ticks infesting three breeds of beef cattle in Paraguay was carried out to provide the basic data necessary to formulate tick control recommendations. The effect of tick burden on weight gain was also analysed. Economic parameters for tick control were calculated. Half body counts of standard female B. microplus were carried out every 2 weeks, at the same time engorged female ticks were placed in the field to observe time to oviposition, larval hatching and 50% survival time. Cattle were weighed every 2 months. Peak tick burdens were encountered in late autumn, with a winter low. Observations of the non-parasitic cycle of the tick suggested that four generations year-1 were possible. High tick burdens were well correlated to low weight gains or weight losses, depending on the breed of cattle. Compensatory weight gains were seen in all three breeds at the end of winter when tick burdens decreased. The economic threshold for tick treatment was calculated at 53 and 54 ticks per animal in 2 separate years of the study. Recommendations for tick control programmes are given.

The safety and efficacy of Australian tick-borne disease vaccine strains in cattle in Paraguay.

Glycerol preserved, frozen tick-borne disease vaccine strains developed in Australia were imported into Paraguay to test their safety in pregnant Holando heifers and their efficacy against challenge from inoculated local field strains of Babesia bigemina, B. bovis and Anaplasma marginale in Hereford X Criolla heifers. The two Babesia strains proved to be safe and the B. bovis K strain was very effective in providing immunity to a local field strain of B. bovis. The B. bigemina efficacy trial was inconclusive, possibly due to the avirulent nature of the local field strain used in challenge. The A. centrale strain did not prove to be as safe as would be desirable in safety trials, neither did it provide as good protection as the Babesia strains in the efficacy trial. It was concluded that the Babesia strains provided good protection against field challenge in Paraguay and were safe to use in highly susceptible cattle, however an alternative to A. centrale should be sought to provide protection against local strains of A. marginale.

Investigation of farmers' perspectives on the disposal of fallen livestock and animal by-products in Great Britain.

Two questionnaires were completed by a selection of farms in Great Britain during 2008 and 2009 to ascertain the role of the National Fallen Stock Company (NFSCo) in fallen livestock disposal, the current disposal methods used for fallen livestock and other animal by-products (ABPs), and factors determining use of a particular method. The results demonstrated a significant difference (P<0.001) in NFSCo membership in relation to geographical location and a significant difference (P<0.001) in disposal choice. Farmers' perceptions as to why they did or did not join NFSCo, and why they used particular disposal methods are discussed. Disposal methods of other ABPs (aborted fetuses/stillborn animals and placentas) demonstrated the illegal disposal of aborted fetuses/stillborn animals (19.5 per cent of respondents) and placentas (57.6 per cent of respondents). Overall, 13.7 per cent of respondents used a variety of illegal disposal methods for fallen livestock carcases and other ABPs.

Aislación y caracterización de células madre mesenquimales provenientes de pulpa y folículo dentario humano / Isolation and characterization of mesenchymal stem cells from human dental pulp and follicle

Recientemente se ha descubierto que diversos tejidos dentales son fuente importante de Células Madre Mesenquimales (CMM). En la cavidad oral podemos encontrar CMM en la pulpa, en el folículo dental, papila y en la encía entre otros lugares. Varios estudios avalan el extenso potencial terapéutico de las CMM en terapias de regeneración. El objetivo de este estudio es aislar, cultivar células madres mesenquimales de pulpa y folículo dental humano, caracterizar su inmunofenotipo y su potencial de diferenciación a linaje osteogénico, condrogénico y osteogénico. Se cultivaron células de pulpa y folículo dental de terceros molares de dientes permanentes jóvenes humanos. Los cultivos de CMM fueron monitoreados por microscopia óptica, las células se inmunotipificaron por citometría de flujo. Posteriormente se evaluó su capacidad de diferenciaron a los tres linajes mencionados. En estas condiciones experimentales se comprobó que las células aisladas y cultivadas de pulpa y folículo dental correspondían a células madre mesenquimales humanas, siendo éstas últimas más fáciles de obtener y proliferar. Las CMM de folículo dental poseen mayor potencial de crecimiento y capacidad de diferenciación en comparación a las CMM de pulpa dental, probablemente debido a su estado evolutivo más inmaduro.

It was recently discovered that dental tissues are important sources of mesenchymal stem cells (MSCs). In the oral cavity MSCs can be found in the pulp, dental follicle, apical papilla and gingival tissue, among others. Many studies support the therapeutic potential of MSCs in regenerative therapies. The objective of this study was to isolate and culture mesenchymal stem cells from human dental pulp and follicle, and to characterize their immunophenotype and differentiation potential to adipogenic, chondrogenic and osteogenic lineages. Oral cavity stem cells were cultured from pulp and dental follicle of wisdom teeth from young permanent teeth. Immunotypification of MSCs was performed by flow cytometry and cultures were evaluated for their ability to differentiate into the three lineages mentioned. Our results corroborate that cultured oral MSC cells isolated from pulp and dental follicle were mesenchymal in origin, being the latter more easy to obtain. Dental follicle MSCs have greater growth potential and differentiation capacity compared to dental pulp MSCs, probably due to their more immature developmental state.

Human teeth with periapical pathosis after overinstrumentation and overfilling of the root canals: a scanning electron microscopic study.

AIM: The aim of this study was to determine whether overinstrumentation followed by immediate overfilling could be a potential risk in the treatment of infected root canals. METHODOLOGY: Thirty-five human teeth with infected root canals were overinstrumented and overfilled approximately 45 min after their extraction. The experimental teeth were enlarged up to size 40 and the overinstrumentation and overfilling were checked with the aid of a magnifying glass. The specimens were fixed in glutaraldehyde plus sodium cacodylate solution and prepared for scanning electron microscope examination. RESULTS: Bacteria were detected on the flute of the files and mostly at the root apices around the main foramen, remaining firmly attached to resorptive lacunae despite the fact that the apices had undergone great changes, including fracture or zipping. A control group consisting of 10 human teeth root canals containing vital pulps were also overinstrumented and overfilled. No bacteria were detected on the flutes of the files, at the apices or on the extruded master cone overfilling these samples. CONCLUSIONS: The high percentage of bacteria adhering to the resorptive lacunae or in the flutes of files used in overinstrumented human teeth with infected root canals carry a potential risk for postoperative pain, clinical discomfort and flare-ups. The hazards observed in these circumstances do not support the one-visit treatment of teeth having acute or chronic periapical abscesses.