Dental caries prevalence and severity positively associate with AMY1 gene copy number.

OBJECTIVE: To establish the possible relation between total caries (TC) and caries severity (CS) with the AMY1 gene copy number (AMY1GCN). MATERIALS AND METHODS: This was an observational, cross-sectional, population-based, and association study with 303 participants. Each participant underwent a complete anamnesis and stomatological check-up, and peripheral blood was obtained to extract gDNA. TC and CS were determined as the number of caries at the dental exploration and the number of dental surfaces affected by caries, respectively, and AMY1GCN was determined by qPCR. RESULTS: We found an elevated caries prevalence (92.7%); TC and CS were 8 ± 10 and 10 ± 13 (median ± IR). There were higher TC and CS in those participants with AMY1GCN above the mean value (0.02 and 0.01 p values, respectively). A positive correlation between TC and CS with AMY1GCN (0.11 and 0.125 r values, 0.03 and 0.01 p values, respectively) was found, in addition to an association between TC and CS with AMY1GCN (1.5 and 1.6 OR values, 0.48 and 0.26 p values, respectively). CONCLUSION: TC and CS were positively related to the AMY1GCN. CLINICAL RELEVANCE: Dental caries has a high prevalence and a multifactorial etiology and has been related to a genetic component. Indeed, the salivary enzyme alpha-amylase could play a significant role in caries susceptibility, considering that its codifying gene (AMY1) can show variation in its gene copy number. This can be considered an important factor for the development of caries at a genetic level.

Associations of the activity and concentration of carbonic anhydrase VI with susceptibility to dental caries: A systematic review and meta-analysis.

OBJECTIVES: A number of studies have claimed that carbonic anhydrase VI (CA VI) is associated with dental caries. The aim of this systematic review and meta-analysis was to systematically review and analyze the literature on the association of CA VI (in terms of concentration and activity) with dental caries. MATERIALS AND METHODS: A systematic review was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Relevant search terms were employed to search the following databases: PubMed, Web of Science, Scopus, China National Knowledge Infrastructure (CNKI), and Cochrane Library databases. Eligible publications from inception to August 2022 were included. The relevant records were assessed independently by two reviewers, and a meta-analysis was performed using RevMan 5.3. RESULTS: Out of 237 relevant records from the initial search, 9 met the criteria for this review. The 9 papers, including 477 participants, were qualitatively analyzed. Seven studies with 411 participants (203 caries-free) were included in the meta-analysis on CA VI activity, and 2 studies with 141 participants (71 caries-free) were included in the meta-analysis on CA VI concentration. The results showed that CA VI activity was significantly higher among participants with caries than their caries-free counterparts (standardized mean difference (SMD) = 0.894, 95% confidence interval (CI95% ): 0.386 and 1.392; p < 0.001), whereas the CA VI concentration was significantly lower among participants with caries than their caries-free counterparts (SMD = -0.672, CI95% : -1.011 and -0.332; p < 0.001). CONCLUSIONS: This meta-analysis of a relatively small number of studies suggests that the CA VI concentration is lower and CA VI activity is higher in patients with dental caries than in caries-free individuals; however, further studies are needed to determine the exact role of CA VI in dental caries.

Polymorphism in the CAVI gene, salivary properties and dental caries.

Objectives: Carbonic anhydrase (CA) VI is supposed to take part in pH or buffering capacity regulation, which can influence the caries risk of an individual. Its expression in the saliva can be modified by single nucleotide polymorphism (SNP). The aim was to investigate SNP in the CA VI gene in relation to active dental caries and physiochemical properties of saliva.Materials and methods: One hundred and thirty participants aged 11-16 years were involved. Clinical examinations were carried out using standardized WHO criteria, DMFT/DMFS and white spot lesions score was evaluated. Saliva samples were examined for salivary properties and CA VI concentration. DNA evaluated in the investigation was extracted from the buccal smear. Three SNP within CAVI gene (rs2274327; rs2274328; rs2274333) were selected and genotyping was performed.Results: In the active caries group, the mean CAVI concentration was significantly lower than in caries free group (p = .014). No association between increased or decreased risk of caries and analysed SNPs was found. There were some significant relations concerning SNPs and salivary buffer capacity and flow rate in rs2274327 and rs2274328.Conclusions: Polymorphism in the CAVI gene can affect salivary properties but there is no direct connection with dental caries.

Activity of Carbonic Anhydrase VI is Higher in Dental Biofilm of Children with Caries.

This study investigated pH, activity and concentration of carbonic anhydrase VI (CA VI) in dental biofilm of caries and caries-free children of 7-9 years old. Seventy-four children were selected and divided into two groups. The caries diagnosis was performed according to the WHO criteria, including the early caries lesion. After biofilm collection and pH determination, CA VI concentration and activity were determined by ELISA and Zimography respectively. The data were submitted to a Mann-Whitney test and to Pearson and Spearman correlation analyses. Means and standard deviations of dental caries for the caries group were of 3.162 ± 1.385. The biofilm pH was significantly higher in the caries-free group. The CA VI activity was significantly higher in biofilm of children with caries. The CA VI concentration was significantly higher in biofilm of caries-free children. In caries-free children, there was a moderate negative correlation between CA VI activity and concentration in dental biofilm as well as between pH and CA VI activity. A negative correlation between biofilm pH and CA VI concentration was found in the caries group. In conclusion, CA VI was shown to be more active in the biofilm of school children with caries in order to contribute to neutralization of biofilm acid.

Activity and distribution pattern of enzymes in the in-situ pellicle of children.

OBJECTIVE: This study investigated, for the first time, pellicle enzymes with respect to their activity, distribution and fluorescence pattern in children with different caries experience. DESIGN: In-situ pellicles were collected from 41 children (aged 4-6 years) with different caries status; 17 of them were caries-free (dmf = 0), 12 had dental restorations but no current caries (dmf ≥ 2) and 12 had at least two carious lesions (dmf ≥ 2). Bovine enamel samples were fixed on individual upper jaw braces for pellicle formation. After 30 min of intraoral exposure, the pellicle and saliva samples were analysed for the activities of amylase, lysozyme, peroxidase and glucosyltransferase (GTF). The distribution of these enzymes, including GTF-isoforms B, C and D, and the pellicle ultrastructure were examined by gold-immunolabelling and transmission electron microscopy (TEM). Furthermore, interactions between pellicle enzymes and adherent bacteria were visualised using combined fluorescence and immunofluorescence labelling. RESULTS: There were no significant differences in the pellicle enzyme activities between the study groups. TEM analysis revealed the absence of GTF C and D in the pellicle of caries-active children. Amylase, peroxidase and GTF-isoforms showed a random distribution within the pellicle layer; lysozyme was found in the form of clusters. A similar ultrastructural pattern was observed for all subjects. Fluorescence labelling technique enabled visualisation of all enzymes, except for GTF B. CONCLUSION: Pellicle enzyme activities and ultrastructure are not associated with children's caries status. Further investigation is needed to assess the influence of individual GTF-isoforms on caries susceptibility in children.

[A preliminary study of saliva matrix metalloproteinases (MMP-2 and MMP-9) in children with caries].

OBJECTIVE: To investigate the correlation between matrix metalloproteinase (MMP)-2/MMP-9 levels and childhood caries, and the saliva levels of MMP-2/MMP-9 among healthy children and those with different degrees of dental caries, both before and after treatment. METHODS: In the study, 368 children aged 3 to 5 years were separated into three groups: severe caries group (112 children), mild caries group (98 children) and caries free group (158 children). The children with severe caries were included in treatment group (83 children) after accepting a comprehensive treatment of caries. MMP-2 and MMP-9 levels were determined by enzyme-linked immunosorbent assay (ELISA) and the data were analyzed by the Statistics Package for Social Science (SPSS 13.0). The differences among severe caries group, mild caries group and caries free group were analyzed by SNK-q (Student Newman Keuls). The severe caries group and treatment group were compared by paired t test. The differences between each group were statistically analyzed. RESULTS: There was no significant difference of the age and gender composition among severe caries group, mild caries group, caries free group and treatment group. The MMP-2 level of severe caries group [(141.3±32.5) µg/L] was higher than those of mild caries group [(107.5±21.3) µg/L] and caries free group [(102.8±18.5) µg/L] (P<0.05). There was no significant difference between mild caries and caries free group (P>0.05). After analysis of 83 children in the treatment group, the level of MMP-2 [(120.1±24.8) µg/L] was lower than before [(144.6±30.3) µg/L] (P<0.05), but was higher than that of caries free group (P<0.05). The MMP-9 levels of severe caries group [(445.8±68.1) µg/L] and mild caries group [(428.6±59.2) µg/L] were higher than that of caries free group [(385.4±60.6) µg/L] (P<0.05), but the difference between severe caries group and mild caries group was not significant (P>0.05). After analysis of 83 children in the treatment group, the alteration of MMP-9 [(432.2±64.7) µg/L] was not significant either (P>0.05). CONCLUSION: The saliva levels of MMP-2 and MMP-9 in children with severe caries were higher than those in caries free children, even if the treatment was implemented, which suggests that the MMP-2 and MMP-9 in saliva might be related to the caries in children.

Esterase from a cariogenic bacterium hydrolyzes dental resins.

OBJECTIVES: To identify and characterize specific esterases from S. mutans with degradative activity toward methacrylate-based resin monomers. METHODS: Out of several putative esterases, an esterase encoded in an Open Reading Frame as SMU_118c (The National Center for Biotechnology Information, NCBI), was found to have true hydrolase activities. SMU_118c was cloned, expressed, purified and further characterized for its respective hydrolytic activity towards ester-containing nitrophenyl substrates and the universal resin monomers bis-phenyl-glycidyl-dimethacrylate (bisGMA) and triethyleneglycol dimethacrylate (TEGDMA) at neutral (7.0) or cariogenic (5.5) pH. Mass spectrometry (MS) was used to verify the expression of SMU_118c protein in S. mutans UA159. RESULTS: Similar to the whole cell activity of S. mutans, SMU_118c showed the highest affinity toward para-nitrophenyl acetate (pNPA) and para-nitrophenyl butyrate (pNPB) vs. ortho-nitrophenyl butyrate (oNPB) and butyrylthiocholine iodide (BTC) (p < 0.05). The esterase retained 60% of its activity after 21 days and hydrolyzed bisGMA at a higher rate than TEGDMA at both neutral and cariogenic pH (p < 0.001), similarly to the predominant human salivary esterase degradative activity. MS confirmed that SMU_118c is an intracellular protein in S. mutans UA159 and expressed under pathogenic (pH 5.5) growth conditions. SIGNIFICANCE: The similarity in the activity profile to the whole S. mutans bacterial cell, the stability over time at cariogenic pH, the preference to hydrolyze bisGMA and confirmed expression profile suggest that SMU_118c could be a significant contributor to the whole bacterial degradative activity of S. mutans toward the degradation of resin composites, adhesives and the restoration-tooth interface, potentially accelerating restoration's failure. STATEMENT OF SIGNIFICANCE: The current study builds upon our highly-cited previous study by Bourbia et al., (JDR, 2013) that reported on that the cariogenic bacterium, S. mutans has esterase-like activities that enable the bacterium to degrade dental composites and adhesives. The current submission is the first to report on the isolation and characterization of the specific esterase activity (SMU_118c) from S. mutans that is a significant contributor to the whole bacterial degradative activity toward the hydrolysis of dental resins. This activity compromises the restoration-tooth interface, increases interfacial bacterial microleakage (Kermanshahi et al., JDR 2010), potentially contributing to the pathogenesis of recurrent caries around resin composite restorations. This represent a significant contribution to the field of biomaterials and their clinical performance.

The Effect of Chemically Modified Tetracycline-3 on the Progression of Dental Caries in Rats.

OBJECTIVE: Matrix metalloproteinases (MMPs) exist in human saliva and dentin and play an important role in the degradation of organic matrix in teeth. Chemically modified tetracycline-3 (CMT-3) is an inhibitor of MMPs. CMT-3 has been used experimentally to treat caries since 1999, but no distinction between dental caries prevalence and dentin caries prevalence has been described. METHODS: A total of 65 Sprague-Dawley rats were randomly divided into three groups. The positive control group (25 rats) was inoculated with Streptococcus mutans (ATCC700610) and fed the cariogenic feed of improved Keyes Diet 2000. The CMT-3 group (25 rats) was also inoculated with S. mutans and fed the cariogenic feed of improved Keyes Diet 2000; the surfaces of rats' molars were daily treated with 0.02% CMT-3. The negative control group (15 rats) was only fed the standard rodent chow. At the end of the 10th week, the dental caries prevalence and dentin caries prevalence of each group were calculated, and the regions of caries were assessed. RESULTS: No caries was found in the negative control group. The dental caries prevalence of the CMT-3 and the positive control group was 75.0 and 83.3%, respectively (p > 0.05, Table 2). The dentin caries prevalence of the CMT-3 and the positive control group was 33.3 and 70.8%, respectively (p < 0.05, Table 2). The Keyes scoring of dentin caries in the CMT-3 group was significantly lower than that in the positive control group (p < 0.05, Table 3). CONCLUSIONS: CMT-3 had no effect on the prevalence of dental caries, but could lower the prevalence and slow down the progression of dentin caries.

aMMP-8 in correlation to caries and periodontal condition in adolescents-results of the epidemiologic LIFE child study.

OBJECTIVES: The suitability of a chairside aMMP-8 test in determination of periodontal inflammation and caries in adolescents was assessed. Secondly, the influence of orthodontic treatment on aMMP-8 test result was analyzed. MATERIALS AND METHODS: Within the LIFE Child study, 434 adolescents (10 to 18 years) were included. Clinical dental examinations comprised caries experience (DMF/T-Index), signs of periodontal inflammation (probing pocket depth, PPD; community periodontal index of treatment needs; CPITN) at six index teeth and oral hygiene (OH). Information about orthodontic appliances (OA) and socioeconomic status (SES) were obtained by validated questionnaires. Test's sensitivity and specificity to detect periodontal inflammation and carious lesions were evaluated. The influence of OA on the test result was analyzed (multivariate model). RESULTS: No associations between age, gender, SES or OH, and test outcome were found (p > 0.05). Positive test results correlated to periodontal findings (CPITN, mean PPD; p < 0.001). However, for the detection of ≥ 1 site(s) with PPD ≥ 4 mm, the test's sensitivity and specificity were found to be 61 and 69%, respectively. Multivariate analysis revealed a higher probability for a positive test result in cases of fixed OA (odds ratio 5.02, 95% confidence interval 1.90-13.19). The test had no diagnostic value considering carious lesions. CONCLUSIONS: The chairside aMMP-8 test does not reliably identify adolescents with periodontal inflammation. Positive test results were more frequent in case of OA. CLINICAL RELEVANCE: The chairside aMMP-8 test is no appropriate tool to screen children and adolescents neither for periodontal inflammation nor for carious lesions.

A preliminary study of saliva matrix metalloproteinases (MMP-2 and MMP-9) in children with caries / 北京大学学报(医学版)

OBJECTIVE@#To investigate the correlation between matrix metalloproteinase (MMP)-2/MMP-9 levels and childhood caries, and the saliva levels of MMP-2/MMP-9 among healthy children and those with different degrees of dental caries, both before and after treatment.@*METHODS@#In the study, 368 children aged 3 to 5 years were separated into three groups: severe caries group (112 children), mild caries group (98 children) and caries free group (158 children). The children with severe caries were included in treatment group (83 children) after accepting a comprehensive treatment of caries. MMP-2 and MMP-9 levels were determined by enzyme-linked immunosorbent assay (ELISA) and the data were analyzed by the Statistics Package for Social Science (SPSS 13.0). The differences among severe caries group, mild caries group and caries free group were analyzed by SNK-q (Student Newman Keuls). The severe caries group and treatment group were compared by paired t test. The differences between each group were statistically analyzed.@*RESULTS@#There was no significant difference of the age and gender composition among severe caries group, mild caries group, caries free group and treatment group. The MMP-2 level of severe caries group [(141.3±32.5) μg/L] was higher than those of mild caries group [(107.5±21.3) μg/L] and caries free group [(102.8±18.5) μg/L] (P<0.05). There was no significant difference between mild caries and caries free group (P>0.05). After analysis of 83 children in the treatment group, the level of MMP-2 [(120.1±24.8) μg/L] was lower than before [(144.6±30.3) μg/L] (P<0.05), but was higher than that of caries free group (P<0.05). The MMP-9 levels of severe caries group [(445.8±68.1) μg/L] and mild caries group [(428.6±59.2) μg/L] were higher than that of caries free group [(385.4±60.6) μg/L] (P<0.05), but the difference between severe caries group and mild caries group was not significant (P>0.05). After analysis of 83 children in the treatment group, the alteration of MMP-9 [(432.2±64.7) μg/L] was not significant either (P>0.05).@*CONCLUSION@#The saliva levels of MMP-2 and MMP-9 in children with severe caries were higher than those in caries free children, even if the treatment was implemented, which suggests that the MMP-2 and MMP-9 in saliva might be related to the caries in children.

Postradiation Matrix Metalloproteinase-20 Expression and Its Impact on Dental Micromorphology and Radiation-Related Caries.

Recent evidence suggests that head-and-neck radiotherapy (HNRT) increases active forms of matrix metalloproteinase-20 (MMP-20) in human tooth crowns, degrading the dentin-enamel junction (DEJ) and leading to enamel delamination, which is a pivotal step in the formation of radiation-related caries (RRC). Additional participation of enzymatic degradation of organic matrix components in caries progression was attributed to MMP-20 in dentin. Therefore, the current study tested the hypothesis that MMP-20 is overexpressed in the DEJ, dentin-pulp complex components, and carious dentin of post-HNRT patients, leading to detectable micromorphological changes to the enamel and dentin. Thirty-six teeth were studied, including 19 post-HNRT specimens and 17 nonirradiated controls. Optical light microscopy was used to investigate the micromorphological components of the DEJ, dentin-pulp complex components, and carious dentin. The samples were divided into 2 subgroups: nondemineralized ground sections (n = 20) and demineralized histological sections (n = 16). In addition, immunohistochemical analysis using the immunoperoxidase technique was conducted to semiquantitatively assess MMP-20 expression in the DEJ, dentin-pulp complex components, and carious dentin. No apparent damage to the DEJ microstructure or other dentin-pulp complex components was observed and no statistically significant differences were detected in MMP-20 expression (p > 0.05) between the irradiated and control groups. This study rejected the hypothesis that MMP-20 is overexpressed in the DEJ, dentin-pulp complex components, and carious dentin of post-HNRT patients, leading to detectable micromorphological changes. Hence, direct effects of radiation may not be regarded as an independent factor to explain aggressive clinical patterns of RRC.

Enzymology and Ultrastructure of the in situ Pellicle in Caries-Active and Caries-Inactive Patients.

AIM: The present study aimed to evaluate the impact of caries activity on the key enzymes and the ultrastructure of the in situ pellicle. METHODS: Pellicle formation was performed on bovine enamel slabs. Intraoral exposure (3, 30, and 120 min) was accomplished by 14 caries-active (DMFS: 22.7 ± 12.1) and 13 caries-inactive (DMFS: 1.5 ± 1.8) individuals. The enzyme activities (lysozyme, peroxidase, α-amylase, glycosyltransferase [GTF]) in the in situ pellicle and resting saliva of all participants were analyzed directly after oral exposure. In addition, a simultaneous visualization of these enzymes, extracellular glucans, and adherent bacteria was carried out. Fluorescent patterns were analyzed with fluorescence labeling and 4',6-diamidino-2-phenylindole/concanavalin A staining. In addition, the distribution of GTF B, C, and D and the ultrastructure of the pellicle were examined by gold immunolabeling and transmission electron microscopy with selected samples. RESULTS: Enzyme activities of amylase, peroxidase, lysozyme, and GTF were detected on all enamel slabs in an active conformation. Neither exposure time nor caries activity had an impact on the enzyme activities. Gold immunolabeling indicated that the pellicle of caries-active subjects tends to more GTF D molecules. The pellicles of caries-inactive and -active individuals revealed a similar ultrastructural pattern. CONCLUSION: The enzyme activities as well as the pellicle's ultrastructure are of high similarity in caries-active and -inactive subjects. Thereby, oral exposure time has no significant influence. This reflects a high uniformity during the initial phase of bioadhesion (3-120 min) concerning enzymatic functions. However, there is a tendency towards more GTF D in caries-active individuals.

Relationship among α amylase and carbonic anhydrase VI in saliva, visible biofilm, and early childhood caries: a longitudinal study.

AIM: This longitudinal study investigated the relationship among early childhood caries (ECC), α amylase, carbonic anhydrase VI (CA VI), and the presence of visible biofilm, besides detecting if these variables could predict risk for ECC. DESIGN: One hundred children were divided into two groups: caries group (n = 45) and caries-free group (n = 55). Visible biofilm on maxillary incisors was recorded, followed by caries diagnosis in preschoolers at baseline and at follow-up. Saliva samples were collected, and activities of CA VI and α amylase were determined. Data normality was assessed by Shapiro-Wilk test and then Mann-Whitney, Spearman correlation, and chi-square tests followed by multiple logistic regression analysis (α = 0.05, 95% confidence interval). RESULTS: CA VI activity was significantly higher in saliva of children with caries (P ≤ 0.05), and α amylase activity was significantly higher in saliva of caries-free children (P < 0.0001). Children with α amylase activity in saliva lower than 122.8 U/mL (OR = 3.33 P = 0.042) and visible biofilm on maxillary incisors (OR = 3.6 P = 0.009) were more likely to develop ECC than caries-free children. A negative correlation between caries and α amylase activity was found (P = 0.0008). CONCLUSIONS: The presence of visible biofilm and low salivary activity of α amylase may be considered risk predictors for ECC.

Dentin caries progression and the role of metalloproteinases: an update.

AIM: This review aims to summarise our understanding of the destructive role of acid environment and metalloproteinases in dentin caries progression using a review process. METHOD: The acids resulting from consumption of sugars by acidogenic and aciduric bacteria can cause demineralisation of the tooth surface, but are not able to cause caries-like lesions. The appearance of such lesions requires the activation of enzymatic proteolysis in an acidic environment for degradation of the dentin organic matrix, leading to cavity formation. Bacterial collagenases have long been considered responsible for organic matrix destruction; host cell-derived matrix metalloproteinases (MMPs) have recently been considered to be involved in the dentinal matrix destruction of carious lesions. DISCUSSION AND CONCLUSION: MMPs are initially synthesised as inactive zymogens to be activated in acid environment of dentinal fluid during the carious process, resulting in destruction of the collagenous matrix. The role of acid environment on enamel and dentin demineralisation and the role of salivary and dentinal MMPs in dentin progression of caries has encouraged general dentists to include the monitoring of oral environment not only by control of bacterial oral flora in caries treatment protocol, but mainly by inhibition of dentinal and salivary MMPs through the use of toothpaste and/or mouthwash containing specific active agents.

The Significance of Matrix Metalloproteinases in Oral Diseases.

Matrix metalloproteinases (MMPs) belong to a family of structurally related zinc-dependent proteolytic enzymes that are known to play a key role in the catabolic turnover of extracellular matrix (ECM) components. Research studies to date have indicated that MMPs regulate the activity of several non-ECM bioactive substrates, including growth factors, cytokines, chemokines and cell receptors, which determine the tissue microenvironment. Disruption of the balance between the concentration of active matalloproteinases and their inhibitors (TIMPs) may lead to pathological changes associated with uncontrolled ECM turnover, tissue remodeling, inflammatory response, cell growth and migration. This brief review presents some information on MMPs' role in inflammatory, metabolic and cancer abnormalities related to the salivary glands, as well as MMP-related aspects that lead to the formation of human dentinal caries lesions. In oral diseases, the most relevant biological fluid commonly used for diagnosing periodontal diseases is saliva. In diseased patients with significantly higher levels of MMPs in their saliva than healthy people, most extracellular matrix components undergo digestion to lower molecular weight forms. Conventional treatment successfully reduces the levels of MMPs inhibits the progressive breakdown of gingival and periodontal ligament collagens. Beside inflammatory abnormalities like Sjögren's syndrome (SS), a large group of disorders is comprised of cancers, most of them involving the parotid gland.

Increased activity of the antioxidants systems modulate the oxidative stress in saliva of toddlers with early childhood caries.

OBJECTIVE: This study aimed to evaluate the oxidative stress levels and the enzymatic and non-enzymatic antioxidant systems in saliva of toddlers with severe early childhood caries (S-ECC). DESIGN: Unstimulated saliva samples were collected at the morning from 0 to 3 year-old S-ECC (n=30) or caries-free (CF) children (n=30/group) for evaluation of oxidative stress (OS) and total antioxidant capacity (TAC), which were measured by the ferric reducing antioxidant power (FRAP) assay, as well as to assess the activity of enzymatic (superoxide dismutase, SOD) and non-enzymatic (uric acid, UA) antioxidant systems, respectively. Data were analyzed by Student's t-test (p<0.05). RESULTS: Significantly higher protein levels were observed in saliva of S-ECC children (0.083mg/mL) than in the CF group (0.070mg/mL). Oxidative damage was significantly lower in saliva of S-ECC children (0.0019µmol/L/mg protein) than in CF children (0.0039µmol/L/mg protein), while salivary TAC (61.5µmol/L), SOD activity (36.6 UE/mL) and uric acid (7.05mg/mL) were significantly higher in saliva of S-ECC when compared to the CF group (49.1µmol/L, 26.8 UE/mL and 5.02mg/mL, respectively for TAC, SOD and UA). CONCLUSION: Oxidative stress levels were significantly lower in saliva of S-ECC children, what might be associated with the increased activity of salivary enzymatic (SOD) and non-enzymatic (uric acid) antioxidant systems.

Association study for the role of Matrix metalloproteinases 2 and 3 gene polymorphisms in dental caries susceptibility.

OBJECTIVE: Various exogenous and endogenous risk factors have been described as contributing to dental caries susceptibility. In the last decade it has been established that both pro and active forms of host derived Matrix metalloproteinases (MMPs) are present in the oral cavity. MMPs role in caries development has been hypothesized. The aim of this study was to analyse MMP2 (rs2287074) and MMP3 (rs679620) single nucleotide polymorphisms (SNPs) and their role in caries susceptibility. DESIGN: The two SNPs were analysed by PCR- restriction fragment length polymorphism (RFLP) in a sample of 102 ethnic Bulgarian volunteers (42 males and 60 females), all students in Sofia Medical University. RESULTS: Statistical analysis of the MMP2 SNP showed significant differences for the genotype frequencies between the caries free (CF, DMFT=0) and low caries experience (LCE, DMFT≤5) groups. Analysis for the non-synonymous MMP3 SNP found significant differences between both CF vs caries experience groups (LCE+ high caries experience (HCE, DMFT≥5)) and LCE vs HCE groups. The presence of allele G decreased the risk of HCE about 4 times. CONCLUSIONS: MMP2 and MMP3 genes are likely to be involved in caries susceptibility in our population. However, as dental caries is a multifactorial disorder and several genes are likely to have influence on it, it is reasonable to expect that SNPs, even those proven to be functional like rs679620, potentially play a significant, but not major role in the disease outcome.

Collagen degradation and preservation of MMP-8 activity in human dentine matrix after demineralization.

OBJECTIVE: Dental caries is a process driven by acids produced by oral microorganisms followed by degradation of the dentine collagen matrix by proteolytic enzymes. Matrix metalloproteinases (MMPs) have been suggested to contribute to caries by degrading collagen. The aim of this study was to develop a method for generating demineralized dentine matrix substrate (DDM) maintaining MMP-8 bioactivity and no interference with later assays. Such a substrate would allow study of the effects of various treatments on MMP-8 activity and collagen degradation in demineralized dentine. DESIGN: Human dentine was powderized in a tissue grinder and frozen (-80°C). The powder was demineralized in dialysis tubes, using EDTA or acetic acid. The demineralized dentine matrix (DDM) was harvested and analyzed for collagen content using SDS-PAGE. The DDM was subsequently suspended in PBS or TESCA buffer. Protein, MMP-8 (ELISA) and collagen (HYP) was analyzed directly or after 1 wk. RESULTS: EDTA or acid demineralization of dentine using dialysis yielded a substrate rich in collagen coupled with preserved MMP-8 activity. Collagen degraded in room temperature, assessed by higher HYP amounts in the soluble fraction of DDM after one wk, indicating that the methods used preserved active DDM-components after the demineralization process. CONCLUSIONS: The presented demineralization methods both provided insoluble DDM substrates suitable for further intervention studies. However, it was found that the substrates differed depending on the demineralization method and buffers used. This needs further study to find an optimal technique for generating DDM with retained proteins as well as enzymatic bioactivity.

Analysis of the association between polymorphisms in MMP2, MMP3, MMP9, MMP20, TIMP1, and TIMP2 genes with white spot lesions and early childhood caries.

BACKGROUND: Matrix metalloproteinases and their inhibitors might be involved in enamel formation. AIM: This study aimed to evaluate the association between polymorphisms in MMP2, MMP3, MMP9, MMP20, TIMP1, and TIMP2 with white spot lesions (WSL) and early childhood caries (ECC). DESIGN: A cross-sectional study was performed on 786 children aged from 2 to 6 years in Brazil. After clinical evaluation, they were classified into groups with disease (the presence of WSL and/or ECC) and without disease (the absence of WSL or ECC). Genotyping of the selected polymorphisms was carried out with TaqMan real-time PCR, using genomic DNA extracted from buccal cells. Allele and genotype frequencies were compared between groups. Chi-square test, odds ratio (OR), and logistic regression were used (P ≤ 0.05). RESULTS: The dmft score was 1.3 (SD: 2.4), and 41.34% of the children have at least one caries lesion. In MMP9, the GG genotype was more frequent in the group without disease (P = 0.006). In a recessive model, WSL was associated with the marker rs1711437 in MMP20 (P = 0.019; OR = 1.20, 95% CI 1.02-1.42). The marker rs1784418 in MMP20 showed an association between the allele G distribution for the WSL group (P = 0.020; OR = 0.73, 95% CI 0.55-0.96). CONCLUSION: MMP9 and MMP20 are involved in WSL and ECC development.

Uso de azúcares y edulcorantes en la alimentación del niño. Recomendaciones del Comité de Nutrición de la Asociación Española de Pediatría / Use of sugars and sweeteners in children's diets. Recommendations of the Nutrition Committee of the Spanish Paediatric Association

El término edulcorante hace referencia a aquel aditivo alimentario que confiere un sabor dulce y que, habitualmente, no aporta o proporciona muy poca energía. Se utiliza para endulzar alimentos, medicamentos y complementos alimenticios cuando se persiguen fines no nutritivos. Desde hace años, se han empleado edulcorantes acalóricos como sustitutos de todo o parte del contenido en azúcares en comidas y bebidas. En las últimas décadas, se ha incrementado su consumo para prevenir la caries y para el correcto cumplimiento de la dieta en casos de control del peso corporal, obesidad y diabetes y, en general, como coadyuvantes para conseguir un balance energético adecuado. No obstante, el consumo de alimentos y de bebidas azucaradas y/o edulcoradas es elevado, reflejando o un aporte calórico importante, o un patrón de hábitos alimentarios inadecuados en los niños. Por otro lado, sigue habiendo dudas entre los consumidores sobre los riesgos para la salud asociados al uso de edulcorantes, ya sean artificiales o naturales. El principal interés en investigación sobre seguridad y los posibles usos terapéuticos se centra en los «edulcorantes artificiales». El objetivo de este documento es proporcionar información a los pediatras sobre las características de los distintos edulcorantes para aconsejar en la elección de un determinado edulcorante sobre la base de sus propiedades (AU)

The term «sweetener» refers to a food additive that imparts a sweet flavour and usually provides no or very low energy. It is used to sweeten foods, medicines and food supplements with no nutritional purposes. For years, no-calorie sweeteners have been used as substitutes for all or part of the sugar content in foods and beverages. In recent decades its consumption has risen to prevent tooth decay, or as an aid in weight control, obesity and diabetes and, in general, to achieve an optimal energy balance. However, consumption of sugary or sweetened food and soft drinks is high, making this situation of special interest in calorie intake and in the poor behavioural pattern of eating habits in children. In addition, questions remain among consumers about the risks to health associated with their use, whether they are artificial or natural. The «artificial sweeteners» are the group of greatest interest in research in order to demonstrate their safety and to provide firm data on their possible therapeutic effects. The aim of the present document is to increase information for paediatricians on the characteristics of different sweeteners, and to advise on the choice of sweeteners, based on their properties (AU)