WNT10A and RUNX2 mutations associated with non-syndromic tooth agenesis.

The goal of this study was to examine the prevalence of WNT10A and RUNX2 mutations and assess their potential impact on the phenotype of non-syndromic tooth agenesis. The study included 30 participants with non-syndromic tooth agenesis, divided into hypodontia (n = 24) and oligodontia forms (n = 6), and 42 unaffected family members. Genomic DNA from buccal epithelial cells was used for polymerase chain reaction amplification of functionally important exons of the WNT10A and RUNX2 genes. Direct sequencing reactions were performed to confirm the presence of mutations. The trend of increasing prevalence of WNT10A mutations and a slight increase in the prevalence of RUNX2 mutations were revealed in tooth agenesis cases compared to unaffected family members. There was a higher prevalence of hypodontia than oligodontia, increased frequency of females over males with missing teeth, and a wide phenotypic variability was observed in individuals and families analyzed. The common missense mutations (p.Phe228Ile, p.Arg113Cys, p.Asp217Asn, and p.Gly165Arg) and c.114-56T>C in the WNT10A gene and in-frame-deletion/insertions (11A, 24Q, 30Q), synonymous variant c.240G>A, and 424-33dupC in the RUNX2 gene were identified. These findings highlight an important role of WNT10A and RUNX2 mutations in the genetic etiology of non-syndromic tooth agenesis.

Detection rates of periodontal bacteria and herpesviruses in different forms of periodontal disease.

The aim was to investigate the detection rates of periodontal bacteria (Porphyromonas gingivalis, Tannerella forsythia, Prevotella intermedia, and Aggregatibacter actinomycetemcomitans) and herpesviruses (herpes simplex virus-1 [HSV-1], cytomegalovirus [CMV], and Epstein-Barr virus [EBV]) in different forms and severity of periodontal disease, and to compare them with those in periodontally healthy subjects. One hundred and twenty-nine patients participated in the study: 39 diagnosed with periodontal abscess (PA), 33 with necrotizing ulcerative periodontitis (NUP), 27 with chronic periodontitis (CP), and 30 participants with healthy periodontal tissue represented a healthy control group. All patients with periodontal disease (PA, NUP, and CP) were also divided into two groups according to the severity of their disease: moderate and severe periodontitis. The subgingival samples were collected from the periodontitis active sites and the detection of microorganisms was performed by end-point polymerase chain reaction analyses. The results revealed significantly higher detection rates of P. gingivalis, T. forsythia, and P. intermedia in all three groups of patients with periodontitis than in healthy participants. The highest detection rate of A. actinomycetemcomitans was noticed in CP, which was significantly higher than that in PA, NUP, and healthy control. The occurrence of EBV was significantly higher in NUP than in CP and healthy participants. CMV was detected significantly more frequently in PA and NUP than in CP and healthy participants. Comparisons among healthy participants and patients with moderate and severe periodontitis showed significantly higher detection rates of EBV and CMV in patients with severe forms of periodontitis than in healthy participants and those with moderate periodontitis.

The influence of ageing on the extrapineal melatonin synthetic pathway.

Ageing affects various physiological and metabolic processes in a body and a progressive accumulation of oxidative damage stands out as often used explanation. One of the most powerful scavenger of reactive oxygen species (ROS) in all organs is melatonin. A majority of melatonin supplied to the body via blood originates from the pineal gland. However, we have been interested in a locally produced melatonin. We have used 2.5- and 36-months-old Wistar rats. Tissues were collected and gene expression of AA-NAT and ASMT, the two key enzymes in a synthesis of melatonin, was determined in brain, liver, kidney, heart, skin, and intestine. Since melatonin can influence antioxidant enzymes, the activity of superoxide dismutase (SOD) and catalase (CAT), and the level of GSH were measured in liver. In addition, Copper (Cu), Zinc (Zn), and Manganese (Mn) were also determined in liver since these microelements might affect the activity of antioxidant enzymes. The expression of AA-NAT and ASMT was increased in liver and skin of old animals. A positive correlation in AA-NAT and ASMT expression was observed in liver, intestine and kidney. Moreover, the activity of CAT enzyme in liver was increased while SOD activity was decreased. SOD and CAT were probably affected by the observed decreased amount of Cu, Zn, and Mn in liver of old animals. In our model, extrapineal melatonin pathway in ageing consisted of complex interplay of locally produced melatonin, activities of SOD and CAT, and adequate presence of Cu, Zn and Mn microelements in order to defend organs against oxidative damage.

The role of TERT-CLPTM1L SNPs, hTERT expression and telomere length in the pathogenesis of oral squamous cell carcinoma.

The aim of this study was to assess TERT-CLPTM1L single-nucleotide polymorphisms (SNPs) (rs402710 C/T in the CLPTM1L gene; rs2736100 A/C and rs2736098 G/A in the TERT gene) as risk factors for development of oral squamous cell carcinoma (OSCC), and to investigate the relationship between the analyzed polymorphisms, relative telomere length (RTL), telomerase expression and clinicopathologic characteristics of OSCC in a Serbian population. Paraffin-embedded tumor samples and buccal swabs from cancer-free controls were genotyped using PCR-RFLP, while tumor RTL values and telomerase expression were estimated by real-time PCR and immunohistochemistry, respectively. CLPTM1L rs402710 and TERT rs2736100 polymorphisms were associated with a significantly increased risk of OSCC, and TERT rs2736098 with a significantly decreased risk. No significant association was found between TERT-CLPTM1L polymorphisms, tumor RTL values, telomerase expression, and clinicopathologic features, although a trend towards longer telomeres was evident in telomerase-positive samples and less advanced tumors. Kaplan-Meier survival analysis showed that patients with longer telomeres in their tumors had significantly better overall survival than patients with shorter telomeres. Our research seems to provide strong evidence for an association between CLPTM1L rs402710C/T and TERT rs2736100A/C SNPs and the risk of OSSC, and suggests that higher tumor RTL values and positive hTERT expression may be applicable as early prognostic markers.(J Oral Sci 58, 449-458, 2016).

Estimation of total bacteria by real-time PCR in patients with periodontal disease.

INTRODUCTION: Periodontal diseases are associated with the presence of elevated levels of bacteria within the gingival crevice. OBJECTIVE: The aim of this study was to evaluate a total amount of bacteria in subgingival plaque samples in patients with a periodontal disease. METHODS: A quantitative evaluation of total bacteria amount using quantitative real-time polymerase chain reaction (qRT-PCR) was performed on 20 samples of patients with ulceronecrotic periodontitis and on 10 samples of healthy subjects. The estimation of total bacterial amount was based on gene copy number for 16S rRNA that was determined by comparing to Ct values/gene copy number of the standard curve. RESULTS: A statistically significant difference between average gene copy number of total bacteria in periodontal patients (2.55 x 107) and healthy control (2.37 x 106) was found (p = 0.01). Also, a trend of higher numbers of the gene copy in deeper periodontal lesions (> 7 mm) was confirmed by a positive value of coefficient of correlation (r = 0.073). CONCLUSION: The quantitative estimation of total bacteria based on gene copy number could be an important additional tool in diagnosing periodontitis.

Association of fibronectin with hypogalactosylated immunoglobulin G in gingival crevicular fluid in periodontitis.

BACKGROUND: Fibronectin (FN) can bind to immunoglobulins (Ig), influencing both the normal clearance and abnormal deposition of circulating immune complexes. This study focuses on the possible interaction between FN and IgG present in gingival crevicular fluid (GCF) of periodontitis patients and periodontally healthy controls, with the aim to determine whether such interaction may be connected with the glycosylation profile of IgG and, thus, consequentional in accumulation of proinflammatory IgG in periodontal pockets. METHODS: The study included 30 patients with initial or advanced periodontitis, and 13 periodontally healthy subjects. GCF IgG was purified and analyzed for the presence of FN and its fragments and for galactose expression. RESULTS: IgG isolated from GCF contained FN, which was bound to the IgG heavy chains. IgG from GCF of advanced periodontitis patients contained high levels of hypogalactosylated IgG, and bound more FN than IgG from GCF of the other two groups, which contained low levels of this glycoform. FN was in a degraded form in GCF from all studied patients, and a fragment of 48- to 53-kDa molecular mass seemed to be the sole one linked to IgG. CONCLUSIONS: IgG and the FN fragment of 48 to 53 kDa in GCF of periodontitis patients and periodontally healthy subjects are physically connected. This fragment was linked to the heavy chains of IgG and the reaction seemed to be particularly intensive with IgG from advanced periodontitis, which contained significantly less galactose in its sugar chains.

[Identification of periodontopathogen microorganisms by PCR technique].

INTRODUCTION: Periodontitis is an inflammatory disease of the supporting tissues of teeth and is a major cause of tooth loss in adults. The onset and progression of periodontal disease is attributed to the presence of elevated levels of a consortium of pathogenic bacteria. Gram negative bacteria, mainly strict anaerobes, play the major role. OBJECTIVE: The present study aimed to assess the presence of the main types of microorganisms involved in the aetiopathogenesis of periodontal disease: Porphyromonas gingivalis, Aggregatibacter actinomycetemcomitans, Eikenella corrodens, Treponema denticola, Tanerella forsythia and Prevotella intermedia in different samples collected from the oral cavity of 90 patients diagnosed with periodontitis. METHOD: Bacterial DNA detection was performed in diverse biological materials, namely in dental plaque, gingival tissue and saliva, by means of multiplex PCR, a technique that allows simultaneous identification of two different bacterial genomes. RESULTS: In the dental plaque of the periodontitis patients, Treponema denticola dominated. In the gingival tissue, Tannerella forsythia and Treponema denticola were the microbiota most frequently detected, whilst in saliva Treponema denticola and Eikenella corrodens were found with the highest percentage. CONCLUSION: The identification of microorganisms by multiplex PCR is specific and sensitive. Rapid and precise assessment of different types of periodontopathogens is extremely important for early detection of the infection and consequently for the prevention and treatment of periodontal disease. In everyday clinical practice, for routine bacterial evaluation in patients with periodontal disease, the dental plaque is the most suitable biological material, because it is the richest in periodontal bacteria.

Hypogalactosylation of salivary and gingival fluid immunoglobulin G in patients with advanced periodontitis.

BACKGROUND: Altered glycosylation of immunoglobulin G (IgG) has been found to affect certain immunological activities of IgG and to correlate with increased inflammation in various disease states. This work deals with the changes in distribution and galactosylation of IgG subclasses present in saliva and gingival crevicular fluid (GCF) of patients with initial and advanced periodontitis and of normal controls. METHODS: IgG subclasses were quantified by dot-blot assay, and the degrees of expression of galactose in the total IgG and its individual subclasses were estimated by lectin immunoblot assay after sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) separation of IgG and by capture enzyme-linked immunosorbent assay (ELISA), using biotinylated Ricinus communis (RCA-I) and Bandeiraea simplicifolia (BS-II) lectins. RESULTS: The distribution of IgG subclasses in both fluids was found to differ in periodontal patients compared to normal controls. In the periodontitis saliva and GCF, the IgG2 subclass dominated quantitatively, regardless of periodontal status. However, galactose was found to be expressed in IgG heavy chains in normal controls and patients with initial periodontitis but not, or at barely detectable levels, in advanced periodontitis. CONCLUSION: The results suggest that the shift toward hypogalactosylated glycoforms may occur during the process of inflammation of the gingiva.

[Use of saliva as a diagnostic fluid in dentistry].

Saliva is a secretion of the salivary and mucous glands and is of major importance in the maintainance of oral health. Over the last few decades, saliva has been evaluated as a diagnostic fluid in medicine for determining systemic disease markers as well as for monitoring numerous drugs, narcotics, and hormones. The biochemical analysis of saliva is particularly important in dentistry. The estimation of the risk of appearance and diagnosis of disease, monitoring of disease progression, evaluation of therapy efficacy for caries, periodontitis, premalignant and malignant oral lesions, as well as infectious diseases of the oral cavity, can be assessed by analysing different constituents of saliva. Individuals at risk of caries can be identified using tests that determine saliva flow rate, saliva buffer capacity, and colonisation of the oral cavity by cariogenic bacteria. Today, these rapid and simple diagnostic tests are used routinely in caries risk determination. The study and use of saliva-based diagnostics have increased over the last few decades. Clinical testing of saliva shows much promise. However, there is a need for much additional research in this area, before the true clinical value of saliva as a diagnostic fluid in dentistry can be determined.