Protective effects of oral administration of yeast thioredoxin against gastric mucosal injury

Figures & data

Fig. 1. The digestibility of yeast TRX in the stomach of the mouse.

Notes: Recombinant yeast TRX was administered orally to mice at a dosage of 100 mg/kg body weight in a 0.85% NaCl solution. At 1, 2, 4, 6, and 24 h after administration, the stomachs were excised from the mice. Yeast TRX in the stomach contents were detected by the western blot method (A). Yeast TRX in the gastric walls was detected by immunoprecipitation and the western blot method (B). Ten nanograms of recombinant yeast TRX was loaded as a control for detection.

Fig. 2. The effects of recombinant yeast TRX on ethanol-induced cytotoxicity in RGM-1 cells.

Notes: (A) Time course of the LDH release. RGM-1 cells were incubated with 10% ethanol and 10⁻⁷ m recombinant yeast TRX for the time period required. (B) Relative cytotoxicity. The cells were incubated with 10% ethanol and 10⁻⁸ or 10⁻⁷ m recombinant yeast TRX, or 10⁻⁷ m thermal denatured recombinant yeast TRX for 35 min. Panel (A) shows the result of a representative experiment. In panel (B), the relative cytotoxicity (%) is shown relative to the cytotoxicity of the control cells (set as 100%) as an average value from 3 independent experiments. The values are given as mean ± SD. The data were analyzed using two-sided one-sample Student’s t tests. The * symbol indicates values significantly different from those for cells incubated without yeast TRX at p < 0.05. (C) Time course of the remaining reducing activity. Yeast TRX was incubated with 10% ethanol for the time period required. (D) Remaining reducing activity of native and thermally denatured TRX. Yeast TRX was denatured at 100 °C for 20 min. The remaining activity (%) was measured by an assay of the TRX reducing activity.

Fig. 3. The effects of yeast TRX on WRS-induced gastric ulcers.

Notes: Lesion indexes (A) and the number of gastric ulcer lesions (B) in rats pre-treatment or post-treatment with the administration of recombinant yeast TRX in a 0.85% NaCl solution, OVA in a 0.85% NaCl solution, or a 0.85% NaCl solution alone. The animals employed as a negative control group were administered with a 0.85% NaCl solution alone and not exposed to WRS. The values are given as mean ± SD (n = 5). The ^*symbols indicate values significantly different at p < 0.05. The ^**symbol indicates values significantly different at p < 0.01. ND: not detected. (C) Pictures of excised stomachs from rats in the post-treatment groups.

Fig. 4. The effects of yeast TRX on HCl/ethanol-induced gastric ulcers.

Notes: (A) Lesion indexes of gastric ulcer (upper figure) and pictures of excised stomachs (lower figure) in recombinant yeast TRX in a 0.85% NaCl solution or a 0.85% NaCl solution alone administered rats (n = 10 in each group). (B) Lesion indexes of gastric ulcer (upper figure) and pictures of excised stomachs (lower figure) in recombinant yeast TRX in a 0.85% NaCl solution (n = 6) or OVA in a 0.85% NaCl solution (n = 5) administered rats. The values are given as mean ± SD. The ^*symbol indicates values significantly different at p < 0.05.

Fig. 5. DNA microarray analysis of HCl/ethanol-induced gastric ulcers in rats.

Notes: (A) The method employed for the DNA microarray experiment. Two groups of rats were administered with recombinant yeast TRX in a 0.85% NaCl solution or a 0.85% NaCl solution alone and gastric mucosal injuries were induced by HCl/ethanol. Stomachs were excised from the rats, and separated into ulcerated and non-ulcerated regions. RNA extraction and DNA Microarray procedures were performed for each separate sample. (B) Scatter plot of gene expression values (log² ratios of ulcerated vs. non-ulcerated regions) in two groups. The x-axis: rats that ingested a 0.85% NaCl solution alone, the y-axis: rats that ingested yeast TRX. The red and green spots respectively depict significantly higher and lower expressed genes in rats that ingested yeast TRX, compared to rats that ingested a 0.85% NaCl solution alone.

Table 1. Gene-annotation enrichment analysis based on the GO databases using differentially expressed genes between rats that ingested yeast TRX and rats that ingested a 0.85% NaCl solution alone.

GO No	GO annotation	Fold enrichment	p values
TRX>NaCl (385 genes)
	Localization
GO:0006811	Ion transport	2.16	2.77.E-03
GO:0006816	Calcium ion transport	3.87	1.92E-02
	Developmental process
GO:0048732	Gland development	3.78	1.29E-03
GO:0030879	Mammary gland development	5.66	1.43E-03
GO:0045445	Myoblast differentiation	15.21	1.59E-02
GO:0048625	Myoblast cell fate commitment	50.70	3.87E-02
GO:0001944	Vasculature development	2.57	3.60E-02
GO:0001568	Blood vessel development	2.65	3.14E-02
	Biological regulation
GO:0050878	Regulation of body fluid levels	4.52	1.04.E-02
GO:0046716	Muscle maintenance	16.30	1.39.E-02
GO:0051241	Negative regulation of multicellular organismal process	3.37	1.75.E-02
GO:0030193	Regulation of blood coagulation	8.45	4.81.E-02
GO:0007243	Protein kinase cascade	2.85	8.41.E-03
GO:0000165	MAPKKK cascade	3.72	1.12.E-02
	Metabolic process
GO:0008202	Steroid metabolic process	3.17	2.29.E-02
GO:0006575	Cellular amino acid derivative metabolic process	3.17	2.29.E-02
	Cellular process
GO:0006350	Transcription	1.78	3.40.E-02
	Response to stimulus
GO:0009719	Response to endogenous stimulus	1.86	3.73.E-02
GO:0009725	Response to hormone stimulus	2.09	1.62.E-02
TRX<NaCl (65 genes)
	Biological regulation
GO:0046578	Regulation of Ras protein signal transduction	12.32	3.59.E-03
GO:0046579	Positive regulation of Ras protein signal transduction	54.71	3.46.E-02
GO:0050801	Ion homeostasis	4.63	4.88.E-02
GO:0055074	Calcium ion homeostasis	11.07	4.84.E-03
GO:0010647	Positive regulation of cell communication	5.81	2.75.E-02
GO:0032104	Regulation of response to extracellular stimulus	62.01	3.06.E-02
GO:0000165	MAPKKK cascade	9.76	3.49.E-02
GO:0051336	Regulation of hydrolase activity	7.07	1.64.E-02
GO:0051345	Positive regulation of hydrolase activity	10.11	3.27.E-02
GO:0042127	Regulation of cell proliferation	3.50	4.57.E-02
	Response to stimulus
GO:0045471	Response to ethanol	15.50	1.47.E-02
GO:0009991	Response to extracellular stimulus	6.16	2.36.E-02
GO:0009719	Response to endogenous stimulus	4.06	2.86.E-02