Current trends of genetics in apical periodontitis research

Genetics is an emerging topic in endodontic research focusing on the host response regarding the pathogenesis of apical periodontitis (AP). A number of genetic epidemiological studies carried out by many investigators worldwide have shown evidence of an association between certain candidate genes and AP. Some studies have been conducted on knockout mice with a deficiency in certain proteins, leading to more or less severe AP, and thus suggesting a pivotal role of these genes in AP pathogenesis. Other research has evaluated the association between genetic polymorphisms in humans with different AP aspects; these studies pointed out that genetic polymorphisms in some candidate genes are involved in inter-individual variations in their response to AP. Therefore, the objective of this report was to provide an updated overview of the genes involved in AP pathogenesis, with a focus on the most relevant candidate genes.


Introduction
Apical periodontitis (AP) is generally a sequel of a root canal system infection. 1Although the infection of the root canal system by pathogens is the main factor involved in its pathogenesis, AP is considered a multifactorial condition, in which other aspects also need to be taken into consideration.Host risk factors such as age, gender and systemic conditions have been evaluated by innumerous researchers for many years. 2,3,4,5,6However, the influence of the host's genetic background on AP pathogenesis is an emerging topic in endodontic research.
So far, these studies have suggested the hypothesis that AP pathogenesis is a result of a complex interplay between microbial factors and genes.Therefore, the objective of this report was to provide an updated overview of the genes involved in AP pathogenesis, with a focus on the most relevant candidate genes.

Literature search strategy
This report systematically reviewed the current trends of genetics in AP pathogenesis research.Studies using KO and wild-type mice as the control, with experimentally induced AP, were included to enable identification of the candidate genes based on animal models.In contrast, only studies evaluating the association between genes (genetic polymorphisms) and AP phenotypes were considered in the task of identification the genes and genetic polymorphisms involved in AP pathogenesis in humans.
A systematic screening of the literature up to March 17, 2018, was undertaken using three electronic databases (PubMed, Scopus and Web of Science), including a hand search of corresponding reference lists and citation searching of key papers without language or date restrictions.Grey literature was also consulted through OpenGrey (http:// www.opengrey.eu).Researchers were contacted to identify additional studies.The MeSH (Medical Subject Headings) terms (www.nlm.nih.gov/mesh/meshhome.html),related terms, and free terms used in the search were "Apical Periodontitis," "Periapical Periodontitis," "Periapical Abscess," "Polymorphism," "Genetic," "Polymorphism, Single Nucleotide," and "Knockout."The Boolean research strategy used is reported in Table 1.
Two authors (E.C.K. and L.A.A.) independently assessed selected titles and abstracts to validate the inclusion criteria.Studies appearing in more than one database were considered only once.In these cases, in which the abstract and the title were not clear, the study was read fully to minimize the possibility of overlooking important studies.The full text of all potentially relevant studies was reviewed.Any disagreement was discussed and solved by consensus between the authors.Data from the included studies were extracted and organized into tables.Figure 1 presents a flowchart of the review process.Twelve studies with KO mice and 14 studies conducted on humans were identified.

Candidate genes involved in AP pathogenesis and phenotype, based on knockout animal studies
In endodontics, animal models-mainly of KO mice-have been a valuable tool to understand the role of genes in the progression of AP.A KO mouse is a genetically modified mouse (Mus musculus), in which researchers have inactivated, or "knocked out," an existing gene.KO mice are a valuable tool for geneticists to understand the role of a gene in embryonic development, normal physiological homeostasis and pathological processes.Since humans and mice share about 99% of the same genes, this animal model is a good analogue for many human biological processes, 30 including AP pathogenesis.For geneticists, the targeted deletion of a gene in a mouse provides an important method to determine the biological role of a specific gene, and is useful in studying the in vivo gene function.It is also the best way to delineate the biological role of a protein. 31ur review included 12 articles that used KO mice as a model to study the role of a specific gene in AP formation and progression, as well as pathogenesis.
In these studies, an experimentally AP was induced in molars for different periods, and compared with AP formation and progression in wild-type mice.These studies are presented in Table 2, together with the description of candidate genes.
After AP induction, KO mice for IL6, 7 OPN, 8 iNOS, 9 TLR22, 10 IL17RA; 11 MyD88 12 and MMP9 13 developed more severe AP than wild-type mice; on the other hand, KO mice for IL22 32 and for CB2 33 developed less severe AP.Animals deficient in PHOX, which mediates the generation of reactive oxygen species, did not present alterations in the AP phenotype. 9hese findings suggest that IL6, IL17AR, OPN, iNOS, MMP9, TLR2 and MyD88 are candidate genes, which might increase the risk for more severe AP phenotypes.On the other hand, IL22 and CB2 may be involved as candidate genes in protection from AP.

Studies evaluating genetic polymorphisms
In the past decade, a growing number of studies have demonstrated that genetic polymorphisms play an important role in AP pathogenesis and phenotype. 16,17,18,19,20,21,22,23,24,25,26,27,28,29The genetic polymorphisms studied in endodontics have so far focused on single nucleotide polymorphisms, also known as SNP.These genetic polymorphisms are the most common form of DNA sequence variation, and account for more than 90% of all variations present in the human genome. 34Genetic polymorphisms could serve as useful genetic markers for identifying genes associated with the pathogenesis of complex alterations, such as AP.Genetic association studies have been conducted to evaluate the association between genetic polymorphisms and different AP phenotypes, and thus determine whether genetic polymorphisms are associated with AP.If there is in fact an association, then the specific allele, genotype or haplotype of a polymorphism is found more often than what is expected by chance in the affected group.Different study designs were used, such as a comparison of 1) acute AP and chronic AP; 2) persistent AP and a periapical healed periodontium; and 3) deep caries lesion with AP and deep caries lesion without AP.
Table 3 presents a list of these studies, together with the genes and polymorphisms evaluated, including the respective study design used.In these studies have provided new etiologic perspectives for AP pathogenesis, with a greater focus on host response.

Final considerations and conclusions
The present review clearly highlights that genetic factors may contribute to inter-individual variation in AP pathogenesis and severity of host response (phenotype).It is important to emphasize that candidate genes were identified based on the observations and results obtained from the KO animal studies.If a deficiency of genes was involved in more or less severe AP, these genes became candidates to be studied in humans.However, it is also important to underscore that studies with animal models are not the only way to select candidate genes to be evaluated in human samples.The functions of the gene or the gene family in question are also important clues.Genes are considered candidate genes for AP pathogenesis, if genetic polymorphisms

CD14 (-260 C/T, rs2569190)
Genotype and allele distribution for the genetic polymorphism in IL6 was associated with symptomatic dental abscesses.

FcgRIIA (R131 or H131, rs1801274)
There was a statistical association observed for the genetic polymorphism in the FcγRIIIb gene (p<0.001).
(2011) 18 34 cases with signs/symptoms of AP and 61 controls showing healing periradicular tissues
Siqueira Junior et al.
(2011) 19 26 cases with PAP and 43 cases with root-canal-treated teeth exhibiting healthy/healing periradicular tissues The genetic polymorphism in FcγRIIIa was not associated with AP.
can be identified and associated with the condition.
There may be one or many candidate genes in each gene family.In addition, a genome-wide association study approach, which uses a high density of genetic markers spread over the entire human genome and a large sample size, could identify potentially new genetic polymorphisms.Another important aspect observed from this review was the lack of validation studies.Only few researchers have evaluated whether genetic polymorphisms in genes involved in AP pathogenesis in animal models are also involved in AP phenotypes in humans.Furthermore, this review identified the absence of proper replication of genetic association studies in different populations.Although replications are considered a key part of genetic epidemiological research, only few research groups aimed to replicate previous studies with results compiled in two meta-analyses, 35,36 which report an association between the genetic polymorphism -308 G>A in TNF-α with acute AP and the polymorphism +3954 C>T in IL1B with persistent AP.
In summary, in the present review, we have systematically collected and screened various studies exploring the role of genes and genetic polymorphisms in AP pathogenesis, thus presenting an updated overview of the field and offering useful information for carrying out future research focusing on the role of various genetic factors.Based on the available data, further identification of candidate genes and their role in AP pathogenesis and phenotype was deemed warranted and necessary.
Moreover, patients with AP may have one or more genetic polymorphisms in several different genes, requiring the translation of new genetic findings into meaningful dental practice or public health applications.

Figure 1 .
Figure 1.Flowchart of the review process.

Table 2 .
List of candidate genes, in alphabetic order, based on the animal knockout (KO) model studies.
Note: *http://www.genecards.org/cgi-bin/carddisp.pl.Only the genes resulting in less or more severe AP are presented here.

Table 3 .
List of articles and genetic polymorphisms evaluated by the authors.