Analysis of polymorphisms in the lactotransferrin gene promoter and dental caries

Abstract

Regarding host aspects, there has been strong evidence for a genetic component in the etiology of caries. The salivary protein lactotransferrin (LTF) exhibits antibacterial activity, but there is no study investigating the association of polymorphisms in the promoter region of LTF gene with caries. The objective of this study was firstly to search the promoter region of the human LTF gene for variations and, if existent, to investigate the association of the identified polymorphisms with dental caries in 12-year-old students. From 687 unrelated, 12-year-old, both sex students, 50 individuals were selected and divided into two groups of extreme phenotypes according to caries experience: 25 students without (DMFT = 0) and 25 with caries experience (DMFT ≥ 4). The selection of individuals with extreme phenotypes augments the chances to find gene variations which could be associated with such phenotypes. LTF gene-putative promoter region (+39 to -1143) of the selected 50 individuals was analyzed by high-resolution melting technique. Fifteen students, 8 without (DMFT = 0) and 7 with caries experience (mean DMFT = 6.28), presented deviations of the pattern curve suggestive of gene variations and were sequenced. However, no polymorphisms were identified in the putative promoter region of the LTF gene.

Figure 1

Comparative sequence analysis of the promoter region amplified by polymerase chain reaction (PCR) whose melting outcome was performed in a single run on a LightCycler 480 instrument (Roche Diagnostics, Mannheim, Germany). Fifteen (15) individuals, being 8 without and 7 with caries experience (mean DMFT = 6.28), were classified as “cases,” being further sequenced.

Figure 2

Comparative sequence analysis of the promoter region amplified by primer pair 5. (a) Consensus sequence of the LTF gene, (b) individual 6 (DMFT = 0), and (c) individual 11 (DMFT = 6). There was not difference between the individual sequences of the promoter region.

Figure 3

Promoter region (+39 to −1143 bp) of the human LTF gene and transcription boxes: (a) M y b ¹, S P1², C/E B P ³, E T s ⁴, and S P1⁵ (region –35 to –85) are involved in LTF expression during myeloid differentiation [82]. (b) E R E ⁶, C O U P ⁷, and G A T A-1⁸ (region –340 to –372), a highly conserved estrogen response element (ERE) overlapping with a chicken ovalbumin upstream promoter (COUP) element [83]. (c) S F R E ⁹  and C O U P ¹⁰ (region – 377 to –394), an extended estrogen response element half site in addition to the ERE, which renders the human LTF gene extremely responsive to estrogen stimulation [83]. (d) T A A C C ¹¹, a highly conserved silencing factor (–774 to –778) that binds the CCAAT displacement protein (CDP/cut) [84].