Published online Jan 14, 2022. doi: 10.3748/wjg.v28.i2.242
Peer-review started: April 29, 2021
First decision: June 17, 2021
Revised: July 13, 2021
Accepted: December 31, 2021
Article in press: December 31, 2021
Published online: January 14, 2022
Helicobacter pylori (H. pylori) infection represents a major public health challenge in Sudan. However, functional polymorphisms within the tumor necrosis factor-alpha (TNF-A) promoter are associated with the incidence and progression of H. pylori infection by increasing TNF-α production.
TNF lies within the major histocompatibility complex region of chromosome 6, which is a highly polymorphic region. Therefore, many polymorphisms have been detected in the TNF-A promoter and studying which variants could affect TNF-A gene expression is relevant. However, the influence of these single nucleotide polymorphisms (SNPs) on TNF-α production is not fully known and still a contradictory topic of debate due to the ethnic differences between populations. Furthermore, to our knowledge, there are no previous studies in Sudan that have addressed the association between TNF-A and H. pylori infection or H. pylori-associated diseases.
To functionally characterize the genetic variations in the TNF-A 5’-region (-584 to +107) of Sudanese patients infected with H. pylori and predict if these SNPs could alter the regulatory motifs using bioinformatics analyses. Also, to investigate the mammalian conservation of these SNPs using comparative profiling analysis in 11 species.
An observational study was conducted in the major hospitals in Khartoum state. Genomic DNA was extracted from 122 gastric biopsies of patients who had been referred for endoscopy. Genotyping of the TNF-A-1030 polymorphism was performed using PCR with confronting two-pair primer to investigate its association with H. pylori infection in the Sudanese population. Sanger sequencing was applied to detect SNPs in the 5’-region (-584 to +107) of TNF-A in H. pylori-infected patients; in silico tools were used to predict whether these mutations would alter transcription factor binding sites or composite regulatory elements in this region. In addition, the ECR browser and multiple-sequence local alignment and visualization search engine were used to study the conservation of the detected SNPs among 11 mammalian species.
A total of seven SNPs were observed in the TNF-A 5’-region of Sudanese patients infected with H. pylori. Among them, the SNP (T>A, -76) was located at the in silico-predicted promoter region (-146 to +10), and it was predicted to alter transcription factor binding sites and composite regulatory elements, while the novel mutation (A>T, +27) was detected in the 5’ untranslated region. It could affect the post-transcriptional regulatory pathways. Mammalian conservation was detected for the (-146 to +10) region in chimpanzee (99.4%), rhesus monkey (95.6%), cow (91.8%), domesticated dog (89.3%), mouse (84.3%), rat (82.4%) and opossum (78.0%). Furthermore, genotyping of TNF-A-1030 revealed a lack of significant association between -1030T and susceptibility to H. pylori and gastric cancer in the studied population (P = 0.1756 and P = 0.8116, respectively).
Despite the high level of genetic variation in the TNF-A 5’-region (-584 to +107) of the Sudanese patients, the sequences involved in enhanceosome formation and gene regulation are highly conserved. Remarkably, only a single SNP (-76) was detected in this region. In addition, computational analysis was a valuable method for studying gene expression patterns and insights for further in vitro and in vivo experimental proofs.
Further large cohort studies are needed to assess the association between (T>A, -76) mutation and H. pylori infection (susceptibility and progression). Also, further studies are encouraged to investigate the novel mutation (A>T, +27) in terms of the frequency of the minor allele (T) in the Sudanese population and its functional significance using computational and experimental approaches. Identifying which of these detected variants are functional is of great relevance for discovering new preventive, diagnostic and therapeutic strategies against the incidence and/or progression of multifactorial diseases such as H. pylori infection.