Effect of free thymol on differential gene expression in gastric mucosa of the young pig

Abstract

Thymol is the most common molecule in thyme and has been proposed as an oral alternative to antibiotics in the feed of pigs and broilers. The knowledge of the in vivo physiological effects of thymol on tissues is limited, particularly its impact on the gastric mucosa, where it is primarily absorbed when it is orally supplied. In this study, thymol (TH, 50 mg/ kg BW) or a placebo (CO) was introduced directly into the stomach of 8 weaned pigs that were slaughtered 12 h later and sampled for gastric oxyntic and pyloric mucosa. The analysis of whole transcript expression was performed using Affymetrix © Porcine Gene 1.1 ST array strips. Affymetrix Transcripts IDs were associated with 13 406 human gene names based on Sus scrofa Ensemble. Gene Set Enrichment Analysis was performed, comparing TH and CO pigs. For each gene set, the normalized enrichment score (NES) was defined as significant when the false discovery rate % was <25 and the P-value of NES was <0.05. In response to TH, 72 and 19 gene sets were significantly enriched in the oxyntic and pyloric mucosa, respectively. Several gene sets involved in mitosis and its regulation ranked near the top, primarily in the oxyntic mucosa; the gene set DIGESTION ranked first and ninth in the pyloric and oxyntic mucosa, respectively. Within this group, somatostatin (SST), SST receptors, peptide transporter 1 (SLC15A1) and calpain 9 (gastrointestinal tract-specific calpain) were the most strongly upregulated genes. Thymol reduced the enrichment of 120 and 59 gene sets in the oxyntic and pyloric mucosa, respectively. Several gene sets related to ion transport and channeling and aqueous pores across membranes, including short transient receptor potential (TRP) channel 4, potassium voltage-gated channel members 1 and 2, and ryanodine receptors 2 and 3, were less enriched. The downregulation of these genes sensitive to thymol in vitro could depend on the thymol dose and contact with the gastric tissues that causes an adaptive response with their reduced activation. Conversely, the activation of the TRPA1 gene (ranked 1072 and 128 among all the genes in the oxyntic and pyloric mucosa, respectively) indicates the involvement of another TRP-regulating cellular calcium storage. In conclusion, the stimulation of gastric proliferative activity and the control of digestive activity by thymol can influence positively gastric maturation and function in the weaned pigs. These properties should be considered in addition to thymol's antimicrobial properties when supplementation of this molecule in feed is evaluated.

Pharmacological Properties and Molecular Mechanisms of Thymol: Prospects for Its Therapeutic Potential and Pharmaceutical Development

“…Thymol (0–2 mM) exerted its relaxant effect on smooth muscle cells by opposing Ca 2+ activation and ATP dependent process by its potent anti-spasmodic effect (Tamura and Iwamoto, 2004). Thymol (50 mg/kg) was shown to influence gastric maturation and function via stimulation of gastric proliferative activity and the control of digestive activity in weaned pigs (Colombo et al, 2014). …”

TRPA1 and issues relating to animal model selection for extrapolating toxicity data to humans

“…cinnamaldehyde (cinnamon), 132,133 allicin and diallyl disulfide (garlic, onion), AITC (horseradish, mustard, wasabi), carvacrol (oregano, marjoram), eugenol (cloves), oleocanthol (olive oil), methyl salicylate (wintergreen), gingerol (ginger), D 9 -tetrahydrocannabinol (marijuana-derived psychoactive compound) and thymol (thyme). 18,[134][135][136] Thiols react with dialkyl disulfide-S-oxides and dialkyl disulfides to give mixed disulfides. 137 As can be seen in Figure 7, it is probable that a cysteine thiol of TRPA1 might react with allicin and diallyl disulfide to yield the same Sallyl product.…”

“…However, failure to explore the potential value of TRPA1 antagonists against agents such as chlorine gas may be a potentially missed opportunity (though it should be noted that the pig is conspicuous by the paucity of papers on its use as a model in TRPA1 studies). Colombo et al 136 have reported that thymol activates the TRPA1 gene in pig pyloric and oxyntic mucosa, whereas it downregulates genes associated with other TRP channels. Boudry and Perrier 244 also reported that ion secretion in the small intestine of the pig is increased by TRPA1 activators such as cinnamon and thyme.…”

Effect of different feed ingredients and additives on IPEC-J2 cells challenged with an enterotoxigenic Escherichia coli strain

“…Finally, thyme is used as a phytogenic feed additive on the basis of the antibacterial activity of its essential oils (mainly thymol). However, it is also considered to stimulate physiological processes in the pig (Colombo et al 2014). As the presumed bioactive ingredient in thyme is an essential oil, watery extracts may contain only very low amounts and therefore low bioactivity was expected.…”

“…A final experiment was conducted to examine the hypothesis that cell cultures may need a preincubation for the feed additives to act. In vivo, feed additives could be present before pathogenic E. coli reach the intestine and thus, the physiology of small intestinal epithelial cells may have been modified as shown by Colombo et al (2014) for thymol. However, a 24 h preincubation with the tested feed additives did not change the outcome.…”