Effects Of Lactobacillus Plantarum Against The Toxicity Of Typical Perfluoroalkyl Substances And The Involved Protective Mechanisms

As new persistent organic pollutants, perfluorinated and polyfluoroalkyl compounds(PFASs) have been widespread in humans, mammals and the environment with a strong chemical and thermal stability, unbiodegradable, environmental persistence, and highly toxicity, widely distributed in the environment, and bioaccumulation along the food chain enrichment in vivo. It has been attracted widespread concern for environmental pollution. The pollution has been widespread on global ecosystems, and constitutes a serious threat to human and animal health.Nowadays, no clinical drugs for remission PFASs toxic have appeared. Considering the side effects, safety and effectiveness, the only one reported method of bile acid sequestrant is unavailable. Compared to the passive detoxification treatment after poisoning of chelating agent, dietary strategies have a better preventive effect PFASs poisoning. However, reported dietary factor such as antioxidant plant compounds have prevention toxic effects of PFASs, which are required to achieve a greater intake to reach the efficacy. Therefore it will bring unassess risks. Lactic acid bacteria are generally recognized as safe food-grade microorganisms. Previous research showed that the adsorption properties and antioxidant activity indicating that they have the possibility to reduce PFASs toxicity. Based on no related research to make use of lactic acid bacteria to alleviate the toxicity of PFASs, we will build cellular antioxidant model, then screen potential lactic acid bacteria with reduce typical PFASs – PFOA and PFOS and further verify its function and mechanism in animal models. The main results are as follows:In this study, a cellular antioxidant assay(CAA) was used to determine the cellular anti-oxidative properties of different concentrations(107, 108, and 109 CFU/mL). The results showed that a concentration of 109 CFU/mL of the Lactobacillus strains was more effective than 108 CFU/mL or 107 CFU/mL. Then the performance of the CAA assay was compared with that of four chemical antioxidant activity assays, namely, DPPH radical scavenging, hydroxyl radical scavenging, reducing power, and inhibition of linoleic acid peroxidation. Results of the CAA assay were consisted with the final results of four chemical assays. Considering that CAA assay quantifies antioxidant activity in cell cultures, will be a more biologically representative method than the popular chemistry antioxidant capacity measures. Finally RAW264.7, Caco-2, and EA.hy926 cells were used to evaluate the antioxidant activity of Lactobacillus strains, results showed that the three cell lines were related.25 Lactobacillus strains were screened by their PFOA and PFOS tolerate ability, and they were further investigated for their binding ability, acid and bile salt tolerance, CAA and adhesion to Caco-2 cells. Finally, the strain with the highest PFOA binding capability was Lactobacillus plantarum CCFM738, which bound to 49.40%±1.5% of available PFOA in 6h with high CAA value(11.89±1.1?M Quercetin). The strain with the highest PFOS binding capability was Lactobacillus plantarum CCFM737, which bound to 60.00%±1.5% of available PFOA in 9h with high CAA value(9.87±0.98?M Quercetin). Finally, the pH and time effects of L. plantarum CCFM738 bindign to PFOA, L. plantarum CCFM737 binding to PFOS were researched.This study characterized the acute and subchronic toxicities of PFOA and PFOS in male C57BL/6J mice, receptively. The acute oral LD50 value for PFOA and PFOS in C57BL/6J mice was 0.45 g/kg BW and 0.579 g/kg BW, respectively. In the subchronic toxicity study, the mice underwent gavage with 0.075, 0.15, and 0.3 g PFOA/kg BW for 30 days. Compared with the PFOA-untreated mice, the PFOA-treated mice statistically reduced their BW gain and food consumption, decreased kidney and spleen mass, and increased liver mass. Histopathological observation showed that relatively serious damage occurred in the liver and kidney, whereas the spleen of the PFOA-treated mice had no obvious change. PFOA treatment increased the blood biochemical parameters, the N-acetyl-?-D-glucosaminidase of urine, and the production of oxidative stress biomarkers and histopathological changes. Exposure to the subchronic oral toxicity of PFOS at 2.5, 5, and 10 mg PFOS/kg BW/day for 30 days disrupted the homeostasis of antioxidative systems, induced hepatocellular apoptosis, triggered liver injury, and ultimately increased the liver size and relative weight of the mice. PFOS treatment caused liver damage but only slightly affected the kidneys and spleen of the mice.Acute PFOA of C57BL/6J mice model was used to evaluate the effect of L. plantarum CCFM738 on the toxicity of PFOA. Compared to PFOA group, decreased MDA activity, increased antioxidant enzyme activity, decreased serum enzymes, inhibit the occurrence of inflammatory factors, pathological changes weakened, all of these indicated CCFM738 can alleviate liver toxicity caused by acute PFOA damage. Through the establishment of prevention and treatment groups, and found that the prevention group were more effective than treatment group. The reasons are that the treatment group is unabailable in mitigating kidney damage and can not improve the antioxidant ability. Compared with the PFOS group, mice fed L. plantarum CCFM737(1×109 CFU/ day) significantly reduced liver mass, increasing the PFOS content in faeces, reduced liver PFOS content, recovery oxidative stress-related indicators and reduce the pathological damage caused by the acute toxicity of PFOS. However, the urine PFOS contents were not changed in neigher prevention group or treatment groups, which indicated that the targets of CCFM737 for PFOS injury is the liver, not the kidney.The toxicity mechanism of of PFOA was that, on one hand by increasing the activity of MDA induced lipid peroxidation, resulting in a significant increase of ACOX activity and peroxisome increase, On the other hand, through the increase of peroxisome proliferator induced the decline of mouse spleen, immune changes and serum cytokine. And prevention and treatment groups are different. The kidney and spleen mass of prevention groups were hiher than of treatment group. Compared the kindney injury of prevention groups, the treatment groups did not. Compared to the sub-chronic toxicity of PFOA exposure, CCFM738 significantly decreased the MDA activity and ACOX activity, and coupled with adsorption ability, significantly improves the liver and kidney injury, and immune damage caused by atrophy of the spleen.Similar to the PFOA sub-chronic toxicity, PFOS sub-chronic toxicity could increase MDA activity, induce lipid peroxidation, resulting in a further significant increase in ACOX activity, finally induce body injury. The difference between PFOA and PFOS sub-chronic toxicity is that PFOS could stimulate MAPKs signaling pathway, leading to cell apoptosis. However, PFOS can not cause kidney damage. CCFM737 share the common antioxidant activity with CCFM738, and it can stimulate Nrf2 signaling pathway, thereby prompting the body to produce oxidative stress, which is the reason of CCFM737 behave better than quercetin in alleviating oxidative damage caused by PFOS.