Mulberry and dandelion water extracts prevent alcohol-induced steatosis with alleviating gut microbiome dysbiosis

Abstract

Chronic alcohol intake causes hepatic steatosis and changes the body composition and glucose metabolism. We examined whether water extracts of mulberry (WMB) and white flower dandelion ( Taraxacum coreanum Nakai, WTC) can prevent and/or delay the symptoms of chronic ethanol-induced hepatic steatosis in male Sprague Dawley rats, and explored the mechanisms. Ethanol degradation was examined by orally administering 3 g ethanol/kg bw after giving them 0.3 g/kg bw WMB or WTC. All rats were continuously provided about 7 g ethanol/kg bw/day for four weeks and were given either of 0.1% dextrin (control), WMB, WTC, or water extracts of Hovenia dulcis Thunb fruit (positive-control) in high-fat diets. Area under the curve of serum ethanol levels was lowered in descending order of control, WTC and positive-control, and WMB in acute ethanol challenge. WMB and WTC prevented alcohol intake-related decrease in bone mineral density and lean body mass compared to the control. After glucose challenge, serum glucose levels increased more in the control group than other groups in the first part and the rate of decrease after 40 min was similar among all groups. These changes were associated with decreasing serum insulin levels. WMB had the greatest efficacy for decreasing triglyceride and increasing glycogen deposits. WMB and WTC prevented the disruption of the hepatic cells and nuclei while reducing malondialdehyde contents in rats fed alcohol, but the prevention was not as much as the normal-control. The ratio of Firmicutes to Bacteroidetes in the gut was much higher in the control than the normal-control, but WTC and WMB decreased the ratio compared to the control. WMB and WTC separated the gut microbiota community from the control. In conclusion, WMB and WTC protected against alcoholic liver steatosis by accelerating ethanol degradation and also improved body composition and glucose metabolism while alleviating the dysbiosis of gut microbiome by chronic alcohol intake. Impact statement Excessive alcohol consumption is associated with serious pathologies and is common in much of the world. Pathologies include liver damage, glucose intolerance, and loss of lean body mass and bone mass. These pathologies are mediated by changes in metabolism as well as toxic metabolic byproducts, and possibly by gut dysbiosis. In this study, we demonstrate that aqueous extracts of mulberry and dandelion protected rats against ethanol-induced losses in lean body and bone masses, improved glucose tolerance and partially normalized gut bacterial populations, with mulberry extract being generally more effective. This research suggests that mulberry and dandelion extracts may have the potential to improve some of the pathologies associated with excess alcohol consumption, and that further clinical research is warranted.

Keywords: Alcohol; dandelion; glucose metabolism; gut microbiome; mulberry.

Figure 1.

Serum ethanol concentrations after ethanol administration with water extracts of mulberry and dandelion. Male rats had oral intakes of 3 g ethanol/kg bw by oral gavage at 30 min after the assigned extracts (0.3 g/kg bw) were orally provided. Blood was collected at 30, 60, 190, and 300 min after ethanol administration without providing additional water and food and serum ethanol levels were measured by colorimetry method. Changes in serum ethanol concentrations (a) and area under the curve of serum ethanol concentrations (b) were provided.

Figure 2.

Bone mineral density (BMD) and lean body mass (LBM). Male rats were provided 6% vol/vol ethanol instead of water and 0.1% dextrin (control), water extract of mulberry (WMB), dandelion (WTC), or Hovenia dulcis Thunb fruits (positive-control) in high-fat diet for four weeks. Normal-control had no ethanol with a high-fat diet. At the end of the experimental period, BMD in the lumbar spine and femur (a), LBM in the hip and leg regions (b) and fat mass in the abdomen and leg (c) were measured by DEXA. Each bar and error bar represents means ± SD (n=12). The different letters on the bars represent significant differences among the groups by Tukey’s test at P<0.05.

Figure 3.

Serum glucose levels and areas under the curve of glucose and insulin during the oral glucose tolerance test (OGTT). At the fourth week of administration of 6% vol/vol ethanol instead of water and 0.1% dextrin (control), water extracts of mulberry (WMB), dandelion (WTC) or Hovenia dulcis Thunb fruits (positive-control) in high-fat diet, 2 g of glucose/kg body weight were orally administered at the overnight fasting state. Normal-control had no ethanol with a high-fat diet. The changes in the serum glucose levels (a) and serum insulin levels (c) were measured during the OGTT. The averages of the areas under the curve (AUC) of glucose (b) and insulin (d) were calculated for the first part (0–40 min) and second part (40–120 min) of the OGTT. Each dot and bar represents the mean ± SD (n=12). *Significantly different among all groups in one-way ANOVA at P<0.05. The different letters on the bars represent significant differences among the groups by Tukey’s test at P<0.05.

Figure 4.

Changes in the serum glucose levels during an intraperitoneal insulin tolerance test (IPITT).At the fourth week of administration of 6% vol/vol ethanol instead of water and 0.1% dextrin (control), water extracts of mulberry (WMB), dandelion (WTC) or Hovenia dulcis Thunb fruits (positive-control) in high-fat diet, an IPITT was conducted by intraperitoneally injecting insulin (1 U/kg body weight) after a 6 h fast. Normal-control had no ethanol with a high-fat diet. The serum glucose levels (a) and area under the curve of serum glucose levels (b) were measured. Each dot and bar represents the mean ± SD (n=12). *Significantly different among all groups in one-way ANOVA at P<0.05. The different letters on the bars represent significant differences among the groups by Tukey’s test at P<0.05.

Figure 5.

Hematoxylin-eosin (H-E) and periodic acid–Schiff (PAS) staining in the liver tissue. After collecting the liver, it was paraffin-embedded and the liver sections were stained with H-E and PAS staining. The evaluation scores for H-E were calculated by summing of each item such as the nucleus size and shape, cell size and arrangement and the number of macrophages and those for PAS staining were determined by the glycogen storage (red staining) (a). The images of H-E staining (b) and PAS (c) are provided.

Figure 6.

The profiles of gut microbiomes. At the fourth week of administration of 6% vol/vol ethanol instead of water and 0.1% dextrin (control), water extracts of mulberry (WMB), dandelion (WTC) or Hovenia dulcis Thunb fruits (positive-control), feces were collected and the bacterial DNA was analyzed. Proportion of taxonomic assignments [order] for gut microbiomes (a) was analyzed. Normal-control had no ethanol with a high-fat diet. The fecal bacterial community was shown in principal coordinate analysis (PCoA) (b).