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Compiled by: Wen Wen, edited by: Xie Yi, Jiang shunyao.
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Perfluorooctane sulfonic acid (PFOS) is a recently discovered persistent organic pollutant, which is enriched in organism through bioaccumulation and food chain. PFOS can induce a variety of diseases including hepatotoxicity, neurotoxicity and metabolic abnormalities. However, the effect of PFOS on intestinal environment is still unknown. Researchers from the school of food science, Jiangnan University and other research institutions found that PFOS can damage all aspects of the intestinal environment (including microbiota, short chain fatty acids and barrier functions), thus increasing the toxicity related to liver, intestinal and metabolic disorders.
Paper ID
Original name: internal environmental disorders associate with the tissue images induced by perfluorooctane successful exposure
Intestinal environmental diseases are related to tissue damage caused by PFOS exposure
Journal: ecological and environmental safety
IF：4.527
Published on: April 2020
Corresponding author: Zhao Jianxin & Wu Yanmin
Corresponding author: School of food science, Jiangnan University & Affiliated Hospital of Jiangnan University
experimental design
The overall objective of the study was to evaluate the effect of PFOS on C57BL / 6J male mice, and to explore the relationship between tissue damage and intestinal environment. The mice were treated with oral PFOS for 16 days (low dose: 0.3 μ g / g; medium dose: 3 μ g / g; high dose: 30 μ g / g). Liver damage was assessed by examining the inflammatory response in the liver and serum liver enzyme concentration. The metabolic function was evaluated by the level of liver cholesterol and the concentration of glucose, HDL cholesterol, total cholesterol and triglyceride in serum. Intestinal environmental diseases were assessed by assessing intestinal flora, short chain fatty acid (SCFAs) production, inflammatory response, and intestinal tight junction protein expression.
experimental result
PFOS induced changes of body weight, intake and organ index in mice
After 16 days of PFOS treatment, a decrease in body weight and food intake was observed (figures 1a and b). Organ index is an important index of toxicity. The increase of organ index indicates hyperemia, edema, hyperplasia or hypertrophy, while the decrease indicates atrophy or growth retardation. Liver index, kidney index and spleen index were analyzed. The higher the dose of PFOS, the higher the liver index and the kidney index. Only the spleen index of the high dose group decreased significantly (Fig. 1c-e). The results showed that PFOS had obvious hepatotoxicity in mice. Slight atrophy of spleen was observed in high dose PFOS, indicating that the chemical has immunosuppressive effect.
Fig. 1 Effect of PFOS on body weight, intake and organ index of adult male C57BL / 6J mice
PFOS induced liver pathology and inflammation
He staining of liver was used for histopathological analysis. The control group showed normal lobular structure and hepatocyte morphology, orderly arrangement of hepatic cord, normal distribution of cell nucleoplasm. After PFOS treatment, the intercellular space in the low dose group became larger and the degree of confusion was higher; the liver in the middle dose group showed inflammatory cell infiltration, the arrangement of liver cells was more disordered and the edema was more obvious; the structure of liver lobule was not clear in the high dose group, the nucleus of liver was obviously contracted, and no nucleus was found in the liver cord of some liver cells (Figure 2a). The results showed that the hepatocytes showed swelling and nuclear migration in different degrees, and the high dose group showed the most obvious changes.
The increase of liver index and tissue damage is accompanied by the change of liver enzyme function. Liver is the main producer of ALP, AST, ALT and γ - GT. When the liver is inflamed or damaged, elevated serum concentrations of these liver enzymes are observed. PFOS treatment can significantly increase serum ALT, ALP and γ - GT levels, but serum ast has no significant change (Figure 2b-e). Next, we analyzed the effect of PFOS on liver inflammation. PFOS can reduce TNF - α concentration in liver and increase IL-1 β and IL-10 concentration (Fig. 2f-h). The results showed that PFOS could cause hepatomegaly, hepatocytopathy and inflammation.
Fig. 2 Effect of PFOS on liver of adult male C57BL / 6J mice
3. PFOS causes different degrees of oxidative damage and dyslipidemia
Next, the levels of cat, GSH, T-AOC and MDA in the liver were measured to determine whether PFOS could cause oxidative stress in the liver. PFOS can increase MDA, cat, GSH and T-AOC levels in the liver (Figure 3a-d). The above results show that there is a self-protection mechanism which can resist the oxidative damage caused by PFOS.
Next, we will study whether PFOS can destroy lipid metabolism. In the high dose PFOS group, the serum Glu concentration decreased, but in the middle and low dose groups, the serum Glu concentration increased (Fig. 3e). PFOS also reduced serum HDL-C, TC, and TG (Fig. 3f-h). TC concentration in liver increased significantly in the high dose group, but decreased in the middle and low dose groups (Fig. 3I), indicating that PFOS can damage the metabolism of liver cholesterol in the high dose group.
Bile acid synthesis is the main pathway of cholesterol degradation. CYP7A1 is a key enzyme that mediates the conversion of cholesterol into bile acid. With the increase of PFOS dose, the content of CYP7A1 in liver decreased significantly (Fig. 3j). It is suggested that high dose PFOS can increase the accumulation of liver cholesterol by affecting the transport and transformation of cholesterol. In other words, PFOS may cause metabolic damage.
Figure 3 PFOS related oxidative damage and lipid metabolism changes in the liver of adult male C57BL / 6J mice
4 PFOS destroys gut barrier and short chain fatty acid production
The expression of proinflammatory and anti-inflammatory cytokines in colon was analyzed to determine whether PFOS caused intestinal inflammation in mice. Similar to the inflammatory response observed in the liver, medium and high dose PFOS treatment reduced TNF - α concentration in colon tissue and increased IL-1 β and IL-10 concentration (Figure 4a-c). It is suggested that PFOS may cause obvious inflammatory reaction in colon.
Furthermore, the expression of mRNA encoding the tight junction protein in colon was analyzed. After PFOS treatment, the expression of ZO-1, occludin and claudin-1 were significantly lower than that of the control group (Figure 4d-f). The decreased expression of the gene encoding the tight junction protein in the colon indicates that PFOS destroys the intestinal barrier.
The production of SCFAs partly reflects intestinal health. The levels of acetic acid, propionic acid, butyric acid, isobutyric acid and valeric acid in feces were evaluated. Among them, acetic acid is the most abundant, and its concentration in feces is significantly reduced after high-dose PFOS treatment (Figure 4G). In addition, no valeric acid was detected in fecal samples from the high and medium dose groups (Fig. 4h). The above results showed that high dose PFOS could significantly affect the level of acetic acid and valeric acid in feces.
Fig. 4 Effect of PFOS on inflammation, tight junction protein and short chain fatty acid production in colon of adult male C57BL / 6J mice
5 prediction of PFOS destroying intestinal flora and picrust pathway
Since PFOS can affect the number of SCFAs in feces, 16S RNA sequencing was used to examine the effect of PFOS on intestinal flora. The results of α diversity analysis showed that after high-dose PFOS treatment, both the Chao1 index and Simpson index decreased significantly, indicating that the community richness and diversity decreased (Fig. 5a). In addition, PFOS significantly influenced the β diversity of intestinal flora in a dose-dependent manner (Figure 5b). The effects of PFOS on intestinal microflora were analyzed. After PFOS treatment, the abundance of Proteus and sclerenchyma decreased, while that of Bacteroides increased. The ratio (F / b) of sclerella and Bacteroides is an important parameter reflecting intestinal flora disorder, and PFOS can reduce f / b (Fig. 5C). Lefse analyzed the abundance of different intestinal microflora in feces (Figure 5d). PFOS can cause many changes of intestinal flora. At the class level, PFOS significantly reduced the abundance of Proteus gamma; at the class level, PFOS significantly reduced the abundance of Clostridium, Enterobacter and Lactobacillus; at the family level, after PFOS treatment, the abundance of erysipelas decreased, the abundance of physical research bacteria and rumen bacteria increased; at the genus level, PFOS significantly reduced the abundance of Brucella (Fig. 5e). The results of picrust pathway analysis of intestinal flora show that PFOS mainly affects metabolic pathways (such as adipocytokine signaling pathway, sphingolipid biosynthesis and lipoic acid metabolism) and some endocrine systems (such as steroid hormone biosynthesis and flavonoid biosynthesis) (Fig. 5F).
Fig. 5 Effect of PFOS on intestinal flora of adult male C57BL / 6J mice
conclusion
Intestinal flora is a key factor in human metabolism. Many animals, including humans, live in environments that contain various pollutants that may affect the intestinal flora. Exposure to these pollutants may damage the intestinal flora and cause downstream effects. Our results show that the hepatotoxicity and colon toxicity of male C57BL / 6J mice treated with PFOS may be related to the significant changes of intestinal environment.
Original website: https://doi.org/10.1016/j.ecoenv.2020.110590
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