Fig 1.
Experimental design for gastroprotective evaluation of MEPS.
Fig 2.
Phytochemical profiles of ethanolic and methanolic extracts of P. syriaca (EEPS and MEPS, respectively).
The total phenolic compounds were superior to flavonoid and anthocyanin contents, while methanolic extracts showed higher phytoconstituents than ethanolic partitioning. Values are expressed as mean ± SEM of triplicates. **, P > 0.01.
Fig 3.
Microscopic appearance of dissected organs from rats treated with either normal saline (A), 2 g/kg MEPS (B), or 5 g/kg MEPS (C).
green star, central vein; yellow arrow, sheets of hepatocyte; gray arrow, Kupffer cells; green arrow, sinusoids; yellow star, glomerulus; DCT, distal convoluted tubules; blue star, bowman’s space; PCT, proximal convoluted tubules; black line, glomerulus with bowmen’s capsule (H & E staining, 40x).
Fig 4.
Gastric gross views of rats receiving either physiological saline (A); physiological saline+ absolute ethanol (B); 30 mg/kg lansoprazole+ absolute ethanol (C); 250 mg/kg MEPS + absolute ethanol (D); 500 mg/kg MEPS + absolute ethanol (E).
Ulcer controls exhibited increased redness areas, hemorrhage (bleeding), erosions (shallow breaks on the surface mucosa), and erythema (redness). Such mucosal tissue injury was ameliorated in rats pretreated with lansoprazole or MEPS in a dose-related manner.
Table 1.
Effect of MEPS on some stomach contents.
Fig 5.
Microscopic structure (10x and 40x) of Gastric tissue for rats receiving either physiological saline (A); physiological saline+ absolute ethanol (B); 30 mg/kg lansoprazole+ absolute ethanol (C); 250 mg/kg MEPS + absolute ethanol (D); 500 mg/kg MEPS + absolute ethanol (E).
Ulcer control exhibited epithelial disruption and detachment (yellow arrow), increased vacuoles in the mucosal cell lining (green arrow), and increased inflammatory infiltration (gray arrow). MEPS treatment preserved mucosal integrity and reduced submucosal edema and inflammatory cell infiltration. blue arrow, intact epithelium; Black circle, area of mucosal erosion/ulceration; gray arrow, inflammatory cells; brown arrow, apoptosis injury.
Fig 6.
Microscopic gastric tissue views (10x) showing different levels of PAS in rats physiological saline(A); physiological saline+ absolute ethanol (B); 30 mg/kg lansoprazole+ absolute ethanol (C); 250 mg/kg MEPS + absolute ethanol (D); 500 mg/kg MEPS + absolute ethanol (E).
MEPS-treated rats showed increased expression of PAS stains in gastric tissues, denoting positive modulation of mucopolysaccharides/glycoproteins in gastric tissues that act as a physical barrier against ethanol-mediated gastric mucosal injury.
Fig 7.
Microscopic gastric tissues (10x) presenting different levels of Bax proteins in rats receiving either physiological saline (A); physiological saline+ absolute ethanol (B); 30 mg/kg lansoprazole+ absolute ethanol (C); 250 mg/kg MEPS + absolute ethanol (D); 500 mg/kg MEPS + absolute ethanol (E).
The ulcer control rats exhibited increased Bax protein (brown colored areas) in their stomachs, denoting elevated apoptotic actions that further deteriorate the stomach injuries. While lansoprazole or MEPS (250 and 500 mg/kg) administration reduced Bax protein expression, suppressing ethanol-mediated apoptotic tissue injury occurred. The optical density of Bax-expressing cells was significantly lower in lansoprazole and MEPS-supplemented rats relative to ulcer-inducing rats (F). *, P > 0.05; ***, P > 0.001; ****, P > 0.0001.
Fig 8.
Microscopic gastric tissues (10x) presenting different levels of Bcl-2 proteins in rats receiving either physiological saline (A); physiological saline+ absolute ethanol (B); 30 mg/kg lansoprazole+ absolute ethanol (C); 250 mg/kg MEPS + absolute ethanol (D); 500 mg/kg MEPS + absolute ethanol (E).
The ulcer control rats exhibited reduced Bcl-2 proteins (brown colored areas) in their stomachs, which enhanced apoptotic actions that further deteriorated the stomach injuries. While lansoprazole or MEPS (250 and 500 mg/kg) administration up-regulated Bcl-2 protein expression, suppressing ethanol-mediated apoptotic tissue injury and thus less gastric tissue damage. The optical density of Bcl-2-expressing cells was significantly higher in lansoprazole and supplemented rats relative to ulcer control rats (F). *, P > 0.05; ***, P > 0.001.
Fig 9.
Concentration of MDA and antioxidants in gastric tissues of rats receiving either physiological saline (A); physiological saline+ absolute ethanol (B); 30 mg/kg lansoprazole+ absolute ethanol (C); 250 mg/kg MEPS + absolute ethanol (D); 500 mg/kg MEPS + absolute ethanol (E).
The ulcer control rats exhibited severe ethanol-mediated oxidative tissue injury, denoted by up-regulated MDA and reduced antioxidants. While MEPS supplementation enhanced production of endogenous antioxidants that attenuated oxidative stress in gastric tissues exposed to ethanol-mediated ulceration, as indicated by significantly higher antioxidant enzymes and lower MDA index relative to ulcer controls. *, P > 0.05; **, P > 0.01; ***, P > 0.001; ****, P > 0.0001.
Fig 10.
Concentration of inflammatory cytokines in serum of rats receiving either physiological saline (A); physiological saline+ absolute ethanol (B); 30 mg/kg lansoprazole+ absolute ethanol (C); 250 mg/kg MEPS + absolute ethanol (D); 500 mg/kg MEPS + absolute ethanol (E).
Ethanol oral delivery provoked a significant inflammatory response that delayed ulcer healing. Noticeably, MEPS supplementation attenuated ethanol-mediated inflammation, indicated by significantly pro-inflammatory cytokines and up-regulated IL-10 cytokines, all of which shortened the inflammatory response that accelerated ulcer healing.
Fig 11.
Summary of the possible underlying gastroprotective pathways modulated by MEPS in ethanol-mediated gastropathy in rats.
Ethanol oral delivery reduced the expression of Nrf2, ameliorated the production of SOD, GSH, and HO-1, and up-regulated MDA levels. Conversely, MEPS supplementation down-regulated NF-KB expression and its inflammatory factors (TNF-α and IL-6). Pretreatment with MEPS attenuated ethanol-mediated autophagy by reducing Bax proteins and increasing Bcl-2 proteins. MEPS delivery resisted ethanol-mediated oxidative stress pathways, indicated by activated Nrf2 pathways and their antioxidant mediators SOD, CAT, PGE2, and lowered MDA level.