Understanding the importance of low-molecular weight (ethylene oxide- and propylene oxide-induced) DNA adducts and mutations in risk assessment: Insights from 15 years of research and collaborative discussions

The interpretation and significance of DNA adduct data, their causal relationship to mutations, and their role in risk assessment have been debated for many years. An extended effort to identify key questions and collect relevant data to address them was focused on the ubiquitous low MW N7-alkyl/hydroxyalkylguanine adducts. Several academic, governmental, and industrial laboratories collaborated to gather new data aimed at better understanding the role and potential impact of these adducts in quantifiable genotoxic events (gene mutations/micronucleus). This review summarizes and evaluates the status of dose–response data for DNA adducts and mutations from recent experimental work with standard mutagenic agents and ethylene oxide and propylene oxide, and the importance for risk assessment. This body of evidence demonstrates that small N7-alkyl/hydroxyalkylguanine adducts are not pro-mutagenic and, therefore, adduct formation alone is not adequate evidence to support a mutagenic mode of action. Quantitative methods for dose–response analysis and derivation of thresholds, benchmark dose (BMD), or other points-of-departure (POD) for genotoxic events are now available. Integration of such analyses of genetox data is necessary to properly assess any role for DNA adducts in risk assessment. Regulatory acceptance and application of these insights remain key challenges that only the regulatory community can address by applying the many learnings from recent research. The necessary tools, such as BMDs and PODs, and the example datasets, are now available and sufficiently mature for use by the regulatory community. Environ. Mol. Mutagen. 60: 100–121, 2019. © 2018 The Authors. Environmental and Molecular Mutagenesis published by Wiley Periodicals, Inc. on behalf of Environmental Mutagen Society.     

Paraffin-embedded tissue as a source of RNA for gene expression analysis in oral malignancy

OBJECTIVE: To assess the feasibility of using archival oral mucosal tissue to examine gene expression at the ribonucleic acid (RNA) level.
MATERIALS AND METHODS: We describe the isolation of RNA from 8 nm sections of formalin-fixed paraffin-embedded oral mucosal tissue. RNA was reverse transcribed and three candidate genes amplified by polymerase chain reaction (PCR). The ribosomal protein S14 gene is a housekeeping gene which has been used as an internal standard in several quantitative PCR protocols. The thymidine kinase (TK) gene is expressed at low levels in most tissues and, with a well-documented genomic organisation, is a useful tool for discrimination between genomic DNA and cDNA. The RIa gene is reported to be overexpressed in many cancer cell lines, in malignant tissue and in vitro transformed cellS. RESULTS: The S14 gene, the TK gene and the RIα gene of the cAMP-dependent protein kinase (PKA) were amplified successfully from formalin-fixed paraffin-embedded tissue sections. The TK primer pair is a useful additional tool in the unambiguous identification of RNA-derived species.
CONCLUSION: RNA suitable for reverse transcribed (RT)-PCR was extracted from archival oral mucosal tissue. This should permit rapid sequence analysis of transcribed tumor suppressor genes and oncogenes in this material. Furthermore, the RT-PCR approach described may allow quantification of gene expression in oral mucosal archival material processed in a standard fashion.     

Aetiological factors for oral manifestations of HIV

OBJECTIVES: Describe the oral diseases in HIV-infected individuals in London, UK and identify social and medical factors related to the presence of specific oral diseases.
DESIGN: Cross-sectional analytic study.
SETTING: Dental clinics.
SUBJECTS: Consecutive sample of 456 patients with HIV infection.
METHODS: Social and medical history and clinical examinations. Univariate and logistic regression analysis.
OUTCOMES: Presence of HIV-associated oral disease.
RESULTS: 80% of patients with AIDS and 50% of patients with HIV had a specific oral disease. The most common diseases were hairy leukoplakia (30%), erythematous candidiasis (24%), pseudomembranous candidiasis (14%), angular chielitis (6%), necrotising periodontal disease (8%) and non-recurrent ulceration (6%).
CONCLUSIONS: The presence of erythematous candidiasis was not related to advanced HIV disease. Pseudomembranous candidiasis, hairy leukoplakia and mucosal ulceration were significantly associated with advanced HIV disease. Smoking was also identified as a strong aetiological factor in oral diseases. Longitudinal studies are required to further explore the prognostic significance of oral diseases in HIV infection.     

Clinical course of chronic periodontitis. II. Incidence, characteristics and time of occurrence of the initial periodontal lesion

AIM: The purpose of this study was to assess the initiation and progression of periodontal disease during adult life. MATERIALS AND METHODS: In a 26-year longitudinal investigation of the initiation and progression of chronic periodontitis that started in 1969 and included 565 men of Norwegian middle class, 223 who had participated in some, but not all, intermediate examinations presented at the last survey in 1995. Fifty-four individuals were available for examination in all seven surveys. RESULTS: Covering the age range from 16 to 60 years, the study showed that at 16 years of age, 5% of the participants had initial loss of periodontal attachment (ILA > or = 2 mm) at one or more sites. Both the subject incidence and the site incidence increased with time, and by 32 years of age, all individuals had one or more sites with loss of attachment. As age progressed, new lesions affected sites, so that as these men approached 60 years of age approximately 50% of all available sites had ILA. An assessment of the intraoral distribution of the first periodontal lesion showed that, regardless of age, molars and bicuspids were most often affected. At and before the age of 40 years, the majority of ILA was found in buccal surfaces in the form of gingival recession. By 50 years, however, a greater proportion of sites presented with attachment loss attributed to pocket formation or a combination of pocket formation and gingival recession. As individuals neared 60 years of age, approximately half of the interproximal areas in posterior teeth had these lesions. CONCLUSION: This investigation has shown that, in a well-maintained population who practises oral home care and has regular check-ups, the incidence of incipient periodontal destruction increases with age, the highest rate occurs between 50 and 60 years, and gingival recession is the predominant lesion before 40 years, while periodontal pocketing is the principal mode of destruction between 50 and 60 years of age.     

Clinical course of chronic periodontitis. I. Role of gingivitis

OBJECTIVES: The purpose of this study was to determine the influence of long-standing gingival inflammation on periodontal attachment loss. On the basis of repeated examinations, the present report describes the influence of gingival inflammation on the initiation of periodontitis from 16 to 59 years of age. MATERIAL AND METHODS: The data originated from a 26-year longitudinal study of Norwegian males, who practiced daily oral home care and received state-of-the-art dental care. The initial examination included 565 individuals. Subsequent examinations took place in 1971, 1973, 1975, 1981, 1988 and 1995. Thus, the study covers the age range of 16-59 years. All tooth sites were divided into four categories according to their history of gingival inflammation over the entire observation period: sites always scoring GI = 0, GI = 1 and GI = 2 sites (GI = gingival index). Sites disclosing various GI scores at different observation periods were not considered. RESULTS: The mean cumulative attachment loss for non-inflamed (GI = 0) sites in individuals approaching 60 years of age was 1.94 mm. Sites always scoring GI = 1 yielded 2.42 mm, and sites that always scored GI = 2 exhibited 3.31 mm of periodontal attachment loss. At interproximal sites of all three groups where gingival trauma was assumed to be minimal or non-existent, only very few sites expressed attachment loss due to gingival recession (2-4%). At interproximal sites always scoring GI = 0, 20% loss of attachment was in the form of pocket formation by 59 years of age. The GI = 1 and the GI = 2 cohorts exhibited attachment loss with pocket formation in 28% and 54%, respectively. CONCLUSION: This study has shown that, as men approach 60 years of age, gingival sites that throughout the 26 years of observation bled on probing had approximately 70% more attachment loss than sites that were consistently non-inflamed (GI = 0). Before 40 years of age, there was a slight increase in periodontal attachment loss due to pocket formation, but after this, the frequency increased significantly. Loss of attachment due to gingival recession was very small in all three groups.The fact that sites with non-inflamed gingiva also exhibited some loss of attachment and pocket formation may be explained by fluctuation in the variations of tissue status during long observation intervals combined with the presence of subclinical inflammation.