Abstract: Ulcerative colitis (UC) is an inflammatory condition of the intestine, resulting from an increase in oxidative stress and pro-inflammatory mediators. In this study, the extract of endophytic bacteriumRhizobium aegyptiacum was prepared for the first time using liquid chromatography-mass spectrometry (LC-MS). In addition, also for the first time, the protective potential ofR. aegyptiacum was revealed using an in vivo rat model of UC. The animals were grouped into four categories: normal control (group I),R. aegyptiacum (group II), acetic acid (AA)‍-induced UC (group III), and R. aegyptiacum-treated AA-induced UC (group IV). In group IV,R. aegyptiacum was administered at 0.2 mg/kg daily for one week before and two weeks after the induction of UC. After sacrificing the rats on the last day of the experiment, colon tissues were collected and subjected to histological, immunohistochemical, and biochemical investigations. There was a remarkable improvement in the histological findings of the colon tissues in group IV, as revealed by hematoxylin and eosin (H&E) staining, Masson’s trichrome staining, and periodic acid-Schiff (PAS) staining. Normal mucosal surfaces covered with a straight, intact, and thin brush border were revealed. Goblet cells appeared magenta in color, and there was a significant decrease in the distribution of collagen fibers in the mucosa and submucosal connective tissues. All these findings were comparable to the respective characteristics of the control group. Regarding cyclooxygenase-2 (COX-2) immunostaining, a weak immune reaction was shown in most cells. Moreover, the colon tissues were examined using a scanning electron microscope, which confirmed the results of histological assessment. A regular polygonal unit pattern was seen with crypt orifices of different sizes and numerous goblet cells. Furthermore, the levels of catalase (CAT), myeloperoxidase (MPO), nitric oxide (NO), interleukin-6 (IL-6), and interlukin-1β (IL-1β) were determined in the colonic tissues of the different groups using colorimetric assay and enzyme-linked immunosorbent assay (ELISA). In comparison with group III, group IV exhibited a significant rise (P<0.05) in the CAT level but a substantial decline (P<0.05) in the NO, MPO, and inflammatory cytokine (IL-6 and IL-1β) levels. Based on reverse transcription-quantitative polymerase chain reaction (RT-qPCR), the tumor necrosis factor-‍α (TNF-‍α) gene expression was upregulated in group III, which was significantly downregulated (P<0.05) by treatment withR. aegyptiacum in group IV. On the contrary, the heme oxygenase-1 (HO-1) gene was substantially upregulated in group IV. Our findings imply that the oral consumption ofR. aegyptiacum ameliorates AA-induced UC in rats by restoring and reestablishing the mucosal integrity, in addition to its anti-oxidant and anti-inflammatory effects. Accordingly,R. aegyptiacum is potentially effective and beneficial in human UC therapy, which needs to be further investigated in future work.

Key words: Inflammatory bowel syndrome; Liquid chromatography-tandem mass spectrometry (LC-MS/MS); Scanning electron microscopy (SEM); Histology; Immunohistochemistry; Reverse transcription-quantitative polymerase chain reaction (RT-qPCR)

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