[Show abstract][Hide abstract] ABSTRACT: With recent progress in the analysis of the salivary proteome, the number of salivary proteins identified has increased dramatically. However, the physiological functions of many of the newly discovered proteins remain unclear. Closely related to the study of a protein's function is the identification of its interaction partners. Although in saliva some proteins may act primarily as single monomeric units, a significant percentage of all salivary proteins, if not the majority, appear to act in complexes with partners to execute their diverse functions. Coimmunoprecipitation (Co-IP) and pull-down assays were used to identify the heterotypic complexes between histatin 5, a potent natural antifungal protein, and other salivary proteins in saliva. Classical protein-protein interaction methods in combination with high-throughput mass spectrometric techniques were carried out. Co-IP using protein G magnetic Sepharose TM beads suspension was able to capture salivary complexes formed between histatin 5 and its salivary protein partners. Pull-down assay was used to confirm histatin 5 protein partners. A total of 52 different proteins were identified to interact with histatin 5. The present study used proteomic approaches in conjunction with classical biochemical methods to investigate protein-protein interaction in human saliva. Our study demonstrated that when histatin 5 is complexed with salivary amylase, one of the 52 proteins identified as a histatin 5 partner, the antifungal activity of histatin 5 is reduced. We expected that our proteomic approach could serve as a basis for future studies on the mechanism and structural-characterization of those salivary protein interactions to understand their clinical significance.
PLoS ONE 11/2015; 10(11):e0142517. DOI:10.1371/journal.pone.0142517 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Candida albicans is the most pathogenic fungal species, commonly colonizing on human mucosal surfaces. As a polymorphic species, C. albicans is capable of switching between yeast and hyphal forms, causing an array of mucosal and disseminated infections with high mortality. While the yeast form is most commonly associated with systemic disease, the hyphae are more adept at adhering to and penetrating host tissue and are therefore frequently observed in mucosal fungal infections, most commonly oral candidiasis. The formation of a saliva-derived protein pellicle on the mucosa surface can provide protection against C. albicans on oral epithelial cells, and narrow information is available on the mucosal pellicle composition. Histatins are one of the most abundant salivary proteins and presents antifungal and antibacterial activities against many species of the oral microbiota, however, its presence has never been studied in oral mucosa pellicle. The objective of this study was to evaluate the potential of histatin 5 to protect the Human Oral Epithelium against C. albicans adhesion. Human Oral Epithelial Tissues (HOET) were incubated with PBS containing histatin 5 for 2 h, followed by incubation with C. albicans for 1 h at 37◦C. The tissues were then washed several times in PBS, transferred to fresh RPMI and incubated for 16 h at 37◦C at 5% CO2. HOET were then prepared for histopathological analysis using light microscopy. In addition, the TUNEL assay was employed to evaluate the apoptosis of epithelial cells using fluorescent microscopy. HOET pre-incubated with histatin 5 showed a lower rate of C. albicans growth and cell apoptosis when compared to the control groups (HOET alone and HOET incubated with C. albicans). The data suggest that the coating with histatin 5 is able to reduce C. albicans colonization on epithelial cell surfaces and also protect the basal cell layers from undergoing apoptosis.
Frontiers in Microbiology 08/2015; 6. DOI:10.3389/fmicb.2015.00885 · 3.99 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: OBJECTIVES:To compare the effects of stannous (Sn) and fluoride (F) ions and their combination on acquired enamel pellicle (AEP) protein composition (proteome experiment), and protection against dental erosion (functional experiment).
METHODS: In the proteome experiment, bovine enamel specimens were incubated in whole saliva supernatant for 24h for AEP formation. They were randomly assigned to 4 groups (n=10), according to the rinse treatment: Sn (800ppm/6.7mM, SnCl2), F (225ppm/13mM, NaF), Sn and F combination (Sn+F) and deionized water (DIW, negative control). The specimens were immersed 3× in the test rinses for 2min, 2h apart. Pellicles were collected, digested, and analyzed for protein content using liquid chromatography electrospray ionization tandem mass spectrometry. In the functional experiment, bovine enamel specimens (n=10) were similarly treated for pellicle formation. Then, they were subjected to a five-day erosion cycling model, consisting of 5min erosive challenges (15.6 mM citric acid, pH 2.6, 6×/d) and 2min treatment with the rinses containing Sn, F or Sn+F (3×/d). Between the treatments, all specimens were incubated in whole saliva supernatant. Surface loss was determined by profilometry.
RESULTS: Our proteome approach on bovine enamel identified 72 proteins that were common to all groups. AEP of enamel treated with Sn+F demonstrated higher abundance for most of the identified proteins than the other groups. The functional experiment showed reduction of enamel surface loss for Sn+F (89%), Sn (67%) and F (42%) compared to DIW (all significantly different, p<0.05).
CONCLUSION: This study highlighted that anti-erosion rinses (e.g. Sn+F) can modify quantitatively and qualitatively the AEP formed on bovine enamel. Moreover, our study demonstrated a combinatory effect that amplified the anti-erosive protection on tooth surface.
PLoS ONE 06/2015; 10(6). DOI:10.1371/journal.pone.0128196 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Low back pain is the most common musculoskeletal problem and the single most common cause of disability, often attributed to degeneration of the intervertebral disc. Lack of effective treatment is directly related to our limited understanding of the pathways responsible for maintaining disc health. While transcriptional analysis has permitted initial insights into the biology of the intervertebral disc, complete proteomic characterization is required. We therefore employed liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) protein/peptide separation and mass spectrometric analyses to characterize the protein content of intervertebral discs from skeletally mature wild-type mice. A total of 1360 proteins were identified and categorized using PANTHER. Identified proteins were primarily intracellular/plasma membrane (35%), organelle (30%), macromolecular complex (10%), extracellular region (9%). Molecular function categorization resulted in three distinct categories: catalytic activity (33%), binding (molecule interactions) (29%), and structural activity (13%). To validate our list, we confirmed the presence of 14 of 20 previously identified IVD-associated markers, including matrix proteins, transcriptional regulators, and secreted proteins. Immunohistochemical analysis confirmed distinct localization patterns of select protein with the intervertebral disc. Characterization of the protein composition of healthy intervertebral disc tissue is an important first step in identifying cellular processes and pathways disrupted during aging or disease progression.
PLoS ONE 02/2015; 10(2):e0117807. DOI:10.1371/journal.pone.0117807 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Genetic factors influence the effects of fluoride (F) on amelogenesis and bone homeostasis but the underlying molecular mechanisms remain undefined. A label-free proteomics approach was employed to identify and evaluate changes in bone protein expression in two mouse strains having different susceptibilities to develop dental fluorosis and to alter bone quality. In vivo bone formation and histomorphometry after F intake were also evaluated and related to the proteome. Resistant 129P3/J and susceptible A/J mice were assigned to three groups given low-F food and water containing 0, 10 or 50 ppmF for 8 weeks. Plasma was evaluated for alkaline phosphatase activity. Femurs, tibiae and lumbar vertebrae were evaluated using micro-CT analysis and mineral apposition rate (MAR) was measured in cortical bone. For quantitative proteomic analysis, bone proteins were extracted and analyzed using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS), followed by label-free semi-quantitative differential expression analysis. Alterations in several bone proteins were found among the F treatment groups within each mouse strain and between the strains for each F treatment group (ratio ≥1.5 or ≤0.5; p<0.05). Although F treatment had no significant effects on BMD or bone histomorphometry in either strain, MAR was higher in the 50 ppmF 129P3/J mice than in the 50 ppmF A/J mice treated with 50 ppmF showing that F increased bone formation in a strain-specific manner. Also, F exposure was associated with dose-specific and strain-specific alterations in expression of proteins involved in osteogenesis and osteoclastogenesis. In conclusion, our findings confirm a genetic influence in bone response to F exposure and point to several proteins that may act as targets for the differential F responses in this tissue.
PLoS ONE 12/2014; 9(12):e114343. DOI:10.1371/journal.pone.0114343 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Objective: To compare the protective effect of Sn- and/or F-containing solutions on enamel and dentin erosion prevention and to analyze the protein profile of the Sn- and F-treated acquired pellicles (AP).
Method: Phase I tested four solutions: SnCl2/NaF, NaF, SnCl2 and deionized water (DIW) (as negative control). Forty bovine enamel and dentin specimens (4´4´2mm3) were prepared and randomly allocated to four groups (n=10). Specimens were incubated in clarified human saliva (CHS) for 24h for pellicle formation and were then subjected to a cycling procedure that included 5-min erosive challenges (0.3% citric acid, pH 2.6); 2-min treatments with the test solution (between 1st, 3rd and 6th cycles); 2-h immersions in CHS, and overnight immersions in CHS. Cycles were repeated 6x/day for 5 days. Surface loss (SL) was determined profilometrically. Phase II: Thirty-two (32) bovine enamel specimens (8´8´2 mm3) (n=8) were similarly prepared and incubated in saliva for 24h and then treated with the same solutions used in phase I for 2min followed by CHS immersions for 2h. This cycle was repeated 3x for 1d. The AP formed on the specimens were collected, digested, and analyzed for specific protein content using liquid chromatography electrospray ionization tandem mass spectrometry (LCESI-MS/MS).
Result: Phase I: for enamel, SnCl2/NaF, SnCl2, NaF solutions provided 89%, 67%, and 42% SL reduction respectively compared with the control, while in dentin they provided 60%, 23%, and 36%, respectively (all p < 0.05). Phase II: Seventy-two (72) common proteins were identified in AP of all groups, 30 exclusive to DIW, 20 to SnCl2/NaF, 19 to NaF, and 13 to SnCl2. SnCl2/NaF increased the abundance of pellicle proteins more than each one alone.
Conclusion: Combination of F and Sn ions provided the best anti-erosive effect. Composition of AP changes with different treatment solutions.
[Show abstract][Hide abstract] ABSTRACT: Our group demonstrated that in vivo acquired enamel pellicle (AEP) is formed by the adsorption of successive layers of protein to hydroxiapatite, which consists of initial binding to enamel and subsequent protein-protein interactions. However, information regarding the type of saliva that contributed for the AEP formation is quite limited. We hypothesized that according to type saliva stimulus, a different AEP proteome will be originated. Objective: AEP formed under different types of salivary stimuli will be compared by using relative proteome quantification based on mass spectrometry. Method: AEP was collected from nine healthy subjects after 2 hours pellicle formation under different type of salivary stimulation: resting condition, mechanical and gustatory stimulation. Pooled samples from each group were subjected to nano-liquidchromatography electro-spray-ionization tandem mass-spectrometry. Results: A total of 122 proteins was identified and 51 proteins were common to all three testedconditions. From those proteins, relative proteome quantification was carried out. Critical AEP proteins such as carbonic anhydrase VI, mucins and cystatins showed significant protein abundance difference among the three conditions. Conclusion: This study is the first to use relative quantitative proteomics based on mass spectrometry to explore different types of saliva stimulus on the AEP formation. Our data suggest that obtained AEP protein profile after each salivary stimulus is similar. In addition, well-recognized enamel demineralization functional AEP proteins such as histatins, statherin and acidic PRP1 were detected in all conditions, which highlighted the mainly protective characteristic of this tooth integument to maintain enamel homeostasis. Support: Natural Sciences and Engineering Research Council of Canada (NSERC grant #371813), and the Canadian Institutes of Health Research, (CIHR grant # 106657 and grant # 97577) WLS is recipient of a CIHR New Investigator Award (grant # 113166), FAPESP
[Show abstract][Hide abstract] ABSTRACT: Candida albicans is the most pathogenic fungal species, commonly colonizing human mucosal surfaces. As a polymorphic species, C. albicans is capable of switching between yeast and hyphal forms, causing an array of mucosal and disseminated infections with high mortality. While the yeast form is most commonly associated with systemic disease, the hyphae are more adept at adhering to and penetrating host tissue and are therefore frequently observed in mucosal fungal infections, most commonly oral candidiasis. Objective:To evaluate the potential of Histatin 5 to protect the Human Oral Epithelium against C. albicans adhesion. Methods:Human Oral Epithelial Tissues (HOET) were incubated with PBS containing histatin 5 for 2 h, followed by incubation with C. albicans for 1 h at 37 °C. The tissues were then washed several times in PBS, transferred to fresh RPMI and incubated for 16 h at 37°C at 5 % CO2. HOET were then prepared for histopathological analysis using light microscopy. In addition, the TUNEL assay was employed to evaluate the apoptosis of epithelial cells using fluorescent microscopy. Proteins released from HOET and from C. albicans-incubated HOET were subjected to relative proteome quantification by mass spectrometry. Results:HOET preincubated with histatin 5 showed a lower rate of C. albicans growth and cell apoptosis when compared to the control groups (HOET alone and HOET incubated with C. albicans). The incubation of HOET with histatin 5 resulted in distinct proteome changes in the incubation buffer compared to both control groups. Conclusions:The data suggest that the coating with histatin 5 is able to reduce C. albicans colonization of epithelial cell surfaces and also protect the basal cell layers from undergoing apoptosis. Supported by:NIH/NIDCR/NIAID grants DE05672, DE07652, AI087803 and AI101067, FAPESP Grants#2011/23540-5, 2011/23543-4 and 2013/15412-2, NSERC grant#371813, CIHR grants#106657/#97577 and, CFI-LOF grant#25116. WLS is recipient of a CIHR New Investigator Award grant#113166.
[Show abstract][Hide abstract] ABSTRACT: Secondary caries at the resin-dentin interface is a major reason for resin composites restorations failure that reduces the service life of the restorations with tremendous financial and health burden on the society. Breakdown of the resin-dentin interface by salivary and bacterial esterases generates the biodegradation by-products (BBPs) triethylene-glycol (TEG) and Bis-hydroxy-propoxy-phenyl-propane (Bis-HPPP).
Objectives: To investigate the effects of TEG and/or Bis-HPPP on the expression of S. mutans virulence genes and related proteins that contribute to its cariogenicity.
Methods: Quantitative real-time PCR and label-free quantitative proteomics analysis were employed to measure gene expression and protein abundance of planktonic and/or biofilm cultures of S. mutans-UA159 in the presence of clinically-relevant concentrations of TEG and/or Bis-HPPP (0.001, 0.01, 0.1 and 1mM) at physiological (7.0) and pathogenic (5.5) pHs.
Results: TEG at 0.01 and/or 0.1mM induced up-regulation of Glucosyltransferases-B and-C, Glucan binding protein-B, and comCDE at pH 5.5, more significantly in biofilm vs. planktonic cells (P<0.001). In contrast, at pH 7.0 TEG had no effect on the expression of those genes in biofilms. Bis-HPPP up-regulated all tested genes except comE at almost all concentrations in biofilm cells vs. no Bis-HPPP control at pH 5.5, while some of the genes were mildly down-regulated by Bis-HPPP in planktonic cultures. At pH 7.0, Bis-HPPP had no effect on biofilms, but planktonic cells had some genes up-regulated at 0.01 or 0.1mM. Quantitative proteomics showed that the protein abundance of gtfB, comD, comE and atpH, were significantly higher in the presence of several concentrations of TEG compared to no TEG control (P<0.05).
Conclusion: Composites and adhesive-derived BBPs increase S. mutans pathogenicity by up-regulating important caries-related genes and their related proteins in in vivo-relevant concentrations, especially under cariogenic pH and growth-mode conditions. These observations could potentially explain the higher incidence of recurrent caries and reduced clinical performance of resin composites.
[Show abstract][Hide abstract] ABSTRACT: Moebius syndrome (MS) is a congenital condition mainly characterized by total or partial palsy of facial and abducens nerves. The involvement of these nerves on MS is responsible for unique features of this syndrome, such as, inability to move the eyes laterally and lack of bi- unilateral facial expression (mask-like face). We hypothesize that these deficiencies can affect the salivary gland, consequently modifying the salivary proteome in these subjects. Moreover, due to the single characteristics of this syndrome, we will also evaluate if these individuals could fit as a suitable in vivo study model to the influence of the nerve in salivary composition. Objectives: Explore the salivary proteome in subjects affected by MS. Methods: Bilateral parotid secretion was obtained from MS by using Lashey cup. In addition, each subject was classified according to the most affected facial side. The samples were analyzed by liquid-chromatography-electrospray-ionization-tandem-mass-spectrometry and PAGEs (native cationic and SDS-PAGE). Results: Classical biochemical protein identification methods such as PAGE were not able to provide a significant differentiation between the affected facial sides; on the other hand, innovative and high-sensitive technology such as ESI-MS/MS provides a more detailed protein composition that can facilitate the identification of unique proteome characteristics according to the side. Conclusions: This study is pioneer in exploring saliva composition/secretion side-effect in vivo model. In addition, the outcomes exposed herein open new avenues for the development of an additional exam that can help to determine the degree of involvement of the salivary glands in this syndrome. Support: Natural Sciences and Engineering Research Council of Canada (NSERC grant #371813), and the Canadian Institutes of Health Research, (CIHR grant # 106657 and grant # 97577) WLS is recipient of a CIHR New Investigator Award (grant # 113166).
[Show abstract][Hide abstract] ABSTRACT: The acquired enamel pellicle (AEP) is important for minimizing the abrasion caused by parafunctional conditions as they occur for instance during bruxism. It is a remarkable feature of the AEP that a protein/peptide film can provide enough protection in normofunction to prevent teeth from abrasion and chipping. Despite its obvious critical role in the protection of tooth surfaces, the essential mechanical properties of the AEP with regard to this protective function have been poorly characterized. Objective: To measure the adhesion force between histatin 5, a primary AEP component, and hydroxyapatite (HA) surfaces. Methods: Both streptavidin-biotinylated histatin 5 and streptavidin-biotinylated human serum albumin were allowed to adsorb on a silica microsphere attached to an atomic force microscope (AFM) cantilever. A Multimode AFM was used to measure the adhesion force between the cantilevers containing protein-functionalized microspheres and the HA surface. All measurements were performed within a fluid cell containing water. The imaging was performed in tapping mode using a Si3N4 AFM cantilever while the adhesion forces were measured in AFM contact mode using the functionalized cantilever system. Results: A collection of force-distance curves (~3,000/replicate) was used to generate histograms that illustrate the adhesion force between the HA surface and histatins 5 or albumin. Histatin 5 (>1.830 nN) exhibited stronger adhesion forces to the HA surface than albumin (>0.282 nN). Conclusion: This study represents an objective approach to adhesion force measurements between histatin 5 and HA, and provides the experimental basis for measuring the same parameters for other AEP constituents. Such knowledge will help in the design of synthetic proteins and peptides with clinical benefits for protective functions of the enamel surface.
Support: NSERC grant #371813, CIHR grants #106657/#97577 CFI-LOF grant #25116. WLS is recipient of a CIHR New Investigator Award grant #113166, NIH/NIDCR/NIAID grants DE05672, DE07652, AI087803 and AI101067.
[Show abstract][Hide abstract] ABSTRACT: The acquired enamel pellicle (AEP) is important for minimizing the abrasion caused by parafunctional conditions as they occur, for instance, during bruxism. It is a remarkable feature of the AEP that a protein/peptide film can provide enough protection in normofunction to prevent teeth from abrasion and wear. Despite its obvious critical role in the protection of tooth surfaces, the essential adhesion features of AEP proteins on the enamel surface are poorly characterized. The objective of this study was to measure the adhesion force between histatin 5, a primary AEP component, and hydroxyapatite (HA) surfaces. Both biotinylated histatin 5 and biotinylated human serum albumin were allowed to adsorb to streptavidin-coated silica microspheres attached to atomic force microscope (AFM) cantilevers. A multimode AFM with a Nanoscope IIIa controller was used to measure the adhesion force between protein-functionalized silica microspheres attached to cantilever tips and the HA surface. The imaging was performed in tapping mode with a Si3N4 AFM cantilever, while the adhesion forces were measured in AFM contact mode. A collection of force-distance curves (~3,000/replicate) was obtained to generate histograms from which the adhesion forces between histatin 5 or albumin and the HA surface were measured. We found that histatin 5 exhibited stronger adhesion forces (90% >1.830 nN) to the HA surface than did albumin (90% > 0.282 nN). This study presents an objective approach to adhesion force measurements between histatin 5 and HA, and provides the experimental basis for measuring the same parameters for other AEP constituents. Such knowledge will help in the design of synthetic proteins and peptides with preventive and therapeutic benefits for tooth enamel.
Journal of dental research 03/2014; 93(5). DOI:10.1177/0022034514526599 · 4.14 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Dental erosion is a multifactorial condition that can result in the loss of tooth structure and function, potentially increasing tooth sensitivity. The exposure of enamel to acids from non-bacterial sources is responsible for the progression of erosion. These erosive challenges are counteracted by the anti-erosive properties of the acquired pellicle (AP), an integument formed in vivo as a result of selective adsorption of salivary proteins on the tooth surface, containing also lipids and glycoproteins. This review provides an in-depth discussion regarding how the physical structure of the AP, along with its composition, contributes to AP anti-erosive properties. The physical properties that contribute to AP protective nature include pellicle thickness, maturation time, and site of development. The pellicle contains salivary proteins embedded within its structure that demonstrate anti-erosive properties; however, rather than individual proteins, protein-protein interactions play a fundamental role in the protective nature of the AP. In addition, dietary and synthetic proteins can modify the pellicle, enhancing its protective efficiency against dental erosion. The salivary composition of the AP and its corresponding protein-profile may be employed as a diagnostic tool, since it likely contains salivary biomarkers for oral diseases that initiate at the enamel surface, including dental erosion. Finally, by modifying the composition and structure of the AP, this protein integument has the potential to be used as a target-specific treatment option for oral diseases related to tooth demineralization.
[Show abstract][Hide abstract] ABSTRACT: To quantify the proteome composition of the GCF in periodontal health (HH) and in sites with different clinical conditions in chronic periodontitis (CP) subjects.
5 subjects with HH and 5 with CP were submitted to full-mouth periodontal examination, and GCF sampling. Sites in the CP group were classified and sampled as periodontitis (P, probing depth, PD>4 mm), gingivitis (G, PD≤3mm with bleeding on probing, BOP), and healthy sites (H, PD≤3mm without BOP). GCF proteins were subjected to liquid chromatography electrospray ionization mass spectrometry for identification, characterization and quantification.
230 proteins were identified; 145 proteins were detected in HH, 214 in P, 154 in G, and 133 in H. Four proteins were exclusively detected at HH, 43 proteins at P, 7 proteins at G, and 1 protein at H. Compared to HH group, 35 and 6 proteins were more abundant in P and G (p<0.001), respectively; and 4, 15 and 37 proteins were less abundant in P, G and H (p≤0.01), respectively.
There are marked differences in the GCF proteome according to disease profile. Comprehension of the role of the identified proteins in the etiopathogenesis of periodontal disease may lead to biomarkers definition.
PLoS ONE 10/2013; 8(10):e75898. DOI:10.1371/journal.pone.0075898 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Acquired enamel pellicle (AEP) is a protein film that forms on the enamel surface of teeth by selective adsorption of proteins and peptides present in the mouth. This protein film forms the interface between enamel and the damage oral biofilm, which modulates the attachment of bacteria found in oral biofilm. The overall goal of this study was to gain insight into the biological formation of the human in vivo AEP. This study hypothesized that AEP is created by the formation of successive protein layers, which consist of initial binding to enamel and subsequent protein-protein interactions. This hypothesis was examined by observing quantitative and qualitative changes in pellicle composition during the first two hours of AEP formation in the oral cavity. Quantitative mass spectrometry approaches were used to generate an AEP protein profile for each time-point studied. Relative proteomic quantification was carried out for the 50 proteins observed in all four time-points. Notably, the abundance of important salivary proteins, such as histatin 1, decrease with increasing of the AEP formation, while other essential proteins such as statherin showed constant relative abundance in all time-points. In summary, this is the first study that investigates the dynamic process to the AEP formation by using proteomic approaches. Our data demonstrated that there are significant qualitative and quantitative proteome changes during the AEP formation, which in turn will likely impact the development of oral biofilms.
PLoS ONE 07/2013; 8(7):e67919. DOI:10.1371/journal.pone.0067919 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Objective
Low-amplitude whole-body vibration has been adopted for the treatment of back pain and spinal disorders. However, there is limited knowledge of the impact of vibration on the intervertebral disc (IVD). This study was undertaken to examine the effects of acute vibration on anabolic and catabolic pathways in the IVD and to characterize the dependence of these changes on time and frequency. Methods
Custom-designed platforms were developed to apply acute vibration to ex vivo and in vivo mouse models. Spinal segments (ex vivo) or mice (in vivo) were subjected to vibration (for 30 minutes at 15-90 Hz with peak acceleration at 0.3g), and IVDs were examined at specific time points after vibration. Gene expression was quantified using real-time polymerase chain reaction, and protein levels were examined by quantitative mass spectrometry and immunofluorescence. ResultsIn the ex vivo model, acute vibration at 15 Hz induced expression of anabolic genes (aggrecan, biglycan, decorin, type I collagen, and Sox9) and suppressed expression of Mmp13, with the most pronounced changes detected 6 hours following vibration. These beneficial effects were frequency dependent and were no longer evident between 45 and 90 Hz. In vivo, the effects on anabolic gene expression were even more robust and were accompanied by decreased expression of Adamts4, Adamts5, and Mmp3. Moreover, significant increases in the protein levels of aggrecan, biglycan, decorin, and type I collagen were detected in vivo. Conclusion
These findings demonstrate dramatic anabolic effects of acute vibration on IVD tissue, responses that are dependent on frequency. The similarity of the in vivo and ex vivo responses indicates that at least some effects of vibration are tissue autonomous.
[Show abstract][Hide abstract] ABSTRACT: Objective:
This study evaluated the use of non-stimulated and stimulated human saliva in commercial test for the detection of dengue NS1 protein during the clinical phase of dengue virus (DENV) infection—an immunochromatographic test allowing rapid detection of the NS1 antigen, “Dengue NS1 Ag STRIP (Bio-Rad)”.
Non stimulated saliva (10 patients) and stimulated saliva (10 patients) samples from 20 patients with acute infection of one of the four dengue serotypes, detected by RT-PCR and/or virus isolation and 20 healthy, non-stimulated saliva (10 patients) and stimulated saliva (10 patients) samples, were used. In a single plastic tube was distributed 50 μl of saliva to be tested with Dengue NS1 Ag STRIP (Bio-Rad) for four groups.
The sensitivity of the Dengue NS1 Ag STRIP to patients with acute infection of one of the four dengue serotypes wasn’t detected. There was no difference for samples of non-stimulated and stimulated saliva in both groups of patients with dengue fever. The specificity was 100% to healthy patients for non-stimulated and stimulated saliva samples.
Our findings could not support the use of diagnostic tools based on the NS1 antigen detection for the diagnosis of acute DENV infection using non-stimulated and stimulated human saliva.
IADR/AADR/CADR General Session and Exhibition 2013; 03/2013