| Environmental endocrine disruptors(EEDs)are a group of natural chemical compounds which are interfering with the synthesis,secretion,transport,metabolism,binding,action,and elimination of the natural endocrine hormones.Therefore,they can make an active or antagonistic effect in the body.With the development of the world economy,EEDs are in full contact with environmental systems,including animals’ and humans’,through environmental media such as water,soil,food and the atmosphere.Glyphosate(GLP)is a kind of herbicides which have been extensively used in most countries.Its residues in the environment lead to a potential hazard.GLP interfere with endocrine system and show a reproductive toxicity.Bisphenol A(BPA)is a kind of environment estrogenic-compound which widely exists in plastic products and creatures cannot avoid reaching in our lives.It consists in food packages,children’s toys and the articles for daily use.BAP can be directly released into the environment through the production and use process of these products,which cause great harm to animals’ and humans’ health.The changes of intestinal microflora can activate the immune system,which can lead to the expression of pro-inflammation genes and promote liver diseases,including viral hepatitis,alcoholic liver diseases,non-alcoholic fatty liver diseases,cirrhosis and so on.In the present study;we focused on GLP,BPA and PNP-induced liver and small intestinal injuries.We investigated the changes in ion levels and oxidative stress in hepatic and small intestinal tissue.We also analyzed the changes of fecal microbial flora in rats through high-throughput sequencing.The main results are as following:1.Glyphosate affects the expression of oxidative stress,inflammation and lipid peroxidation-related genes in the liver of rats The objective of the study is to investigate the effects of glyphosate on rat liver function and its induction in pathological changes in ion levels and oxidative stress in hepatic tissue.Sprague-Dawley rats were orally treated with 5,50 and 500 mg/kg the GLP,on a daily basis for a period of 35 days.The weight of the body,the related viscera,and small intestine were examined.The histological parameter of the liver tissue was determined by HE staining.The mRNA expressions of oxidative stress,inflammation,lipid peroxidation and apoptosis-regulated genes were measured by real-time qPCR.In addition,we also analyzed the Nrf2 and Cleaved-caspase-3 expression in the liver by immunohistochemical staining and western blotting.The results showed that the body weight,weights of liver,kidney,and spleen decreased(p<0.05).The morphology of liver and kidney changed in the GLP-treated rats.The GLP treatment resulted in increase in the expression of glutamic-oxalacetic transaminase(GOT),glutamic-pyruvic transaminase(GPT)and IL-1βin the serum(p<0.05).Also,the decline of total superoxide dismutase(T-SOD)activity and increased malondialdehyde(MDA)levels in the serum,liver,and kidney indicated the presence of oxidative stress(p<0.05).Moreover,the increase in hydrogen peroxide(H2O2)level and catalase(CAT)activity in the serum and liver,and the decline of glutathione(GSH)and glutathione peroxidase(GSH-Px)activity in the kidney tissue further confirmed the occurrence of oxidative stress(p<0.05).Nrf2 and Cleaved-caspase-3 protein expressed higher in liver tissue in GLP-treated rats(p<0.05).The results of RT-PCR showed that the mRNA expression of IL-1α,IL-1β,IL-6,MAPK3,NF-κB,SIRT1,TNF-α,Keap1,GPX2,Nrf2 and Caspase-3 significantly increased in the GLP-treated group compared with the control group(p<0.05).Furthermore,PPARa,DGAT,SREBP-1c and SCD1 mRNA expression also significantly increased in the GLP-treated group compared with the control group(p<0.05).Conversely,FAS expression displayed no significant differences(p>0.05).These results suggest that glyphosate causes obvious damages in rat liver and causes the imbalances in Al,Fe,Zn and other ion levels in rat liver,kidney,spleen,lung,heart,muscle,brain and fat.Ion imbalance-related oxidative stress might involve in the mechanism of chronic liver injury caused by GLP.2.Effects of glyphosate on small intestinal oxidative damage and fecal microbial composition in rats The aim of this study was to evaluate the effects of glyphosate on small intestinal toxicity and intestinal microbes in rats.Sprague-Dawley rats were treated orally with 5,50 and 500 mg/kg the GLP,on a daily basis for a period of 35 days.After the last 24 hours of intragastric administration,blood samples,urine,feces,stomach,small intestine and colon were collected.HE staining was used to observe the small intestinal histological and the villus height and crypt depth were detected.The contents of various mineral elements in the small intestine were measured.The mRNA expressions of inflammation-regulated genes were measured by RT-PCR.We also used the high throughput sequencing to analyze the gut microbes in feces.The results showed that GLP caused small intestinal mucosal tissue abnormalities,inherent layer disintegration,a large number of neutrophil infiltration of submucosal and villus fall off.The villus height of the small intestine tissue of each GLP group was lower than that of the control group(p<0.05).The crypt significantly increased compared with the control group(p<0.05).Compared with the control group,SOD activity in duodenal tissue decreased significantly in GLP treatment group(p<0.05).CAT activity in jejunum tissue showed a significant increased in GLP treatment group(p<0.05).MDA content showed a significant increased in the ileum tissue in the GLP treatment group(p<0.05).The results of ion concentration test showed that GLP changed the serum concentration of urine,urine,feces and small intestine.The results of RT-PCR showed that the mRNA expression of MAPK3,NF-κB,SIRT1,TNF-α,GPX2 and Caspase-3 significantly increased in the GLP-treated group compared with the control group(p<0.05).The high abundance sequencing results showed that the relative abundance of the tenericutes and spirochaeta bacteria in the GLP-treated group was higher compared with the control group(p<0.05),and GLP significantly decreased the relative abundance offirmicutes(p<0.05).GLP also significantly increased the relative abundance of Prevotella 1,Actinomyces,Treponema 2 and some unclassified flora in the fecal samples of the rats and reduced the relative abundance of Lactobacillus in fecal samples(p<0.05).Our results demonstrate that exposure to GLP induced the small intestinal oxidative stress injury,which may be related to the change of ion concentration and the composition and abundance of rat flora in rats.3.Bisphenol A affects the expression of oxidative stress,inflammation and lipid peroxidation-related genes in the liver of rats The purpose of this study was to investigate BPA-induced hepatic damage of male rats.Twenty Sprague-Dawley male rats were randomly assigned into four experimental groups(0,0.5,5 and 50 mg/kg BPA),for a period of 30 days.24 hours after the final intragastric administration.Rats and organs were weighed,and blood samples were collected.HE and transmission electron microscopy staining were used to observe the hepatic histological.Levels of IL-1β,IL-6 and TNF-a in serum and liver were determined by ELISA.The levels of oxidative stress in serum,liver,and kidney were also measured.The mRNA expressions of oxidative stress,inflammation,lipid peroxidation and apoptosis-regulated genes were measured by RT-PCR.We also analyzed the Nrf2 and Cleaved-caspase-3 expression in the liver by immunohistochemical staining and western blotting.The results showed that the weight of liver,kidney,and right adrenal gland decreased(p<0.05),and the morphology of liver and kidney changed in the BPA-treated rat.The GLP treatment resulted in an increase glutamic-oxalacetic transaminase(GOT)levels in the serum(p<0.05).Also,increased TNF-a level in the liver indicates the presence of oxidative stress(p<0.05).At the same time,catalase(CAT)activity in the serum increased(p<0.05)and glutathione peroxidase(GSH-Px)activity in the liver tissue declined(p<0.05).Nrf2 and Cleaved-caspase-3 protein expression increased in liver tissue in BPA treatment group compared with the control group(p<0.05).The results of RT-PCR showed that the mRNA expression of IL-1α,TNF-α,GPX2,NF-kB,PPARa,SREBP-1c,SCR-1,Nrf2 and Caspase-3 mRNA significantly increased in the BPA-treated group compared with the control group.These results demonstrated that BPA-induced hepatic oxidative stress caused liver damage,while Nrf2 and Cleaved-caspase-3 play an important role in BPA-induced liver damage.4.Effects of bisphenol A on small intestinal oxidative damage and fecal microbial composition in rats To investigate the effect of Bisphenol A(BPA)on rat intestine injury and intestinal microflora.Twenty Sprague-Dawley male rats were randomly assigned into four experimental groups(0,0.5,5 and 50 mg/kg BPA),for a period of 30 days,After the last 24 hours of intragastric administration,blood samples and small intestine tissue were collected.HE and PAS staining were used to observe the small intestinal histology and the villus height,and crypt depth was detected.Transmission electron microscopy(TEM)was used to observe the ultrastructure of the jejunum and detection of oxidative stress parameters in the small intestine.The mRNA expressions of oxidative stress,inflammation,lipid peroxidation and apoptosis-regulated genes were measured by RT-PCR.We also used the high throughput sequencing to analyze the gut microbes in feces.The results of HE staining showed that the mucosal morphology of the small intestine was normal and the villus was complete,BPA caused abnormalities of small intestinal mucosal tissue,villus falls off and necrosis.PAS staining results showed that the number of goblet cells significantly increased compared with the control group(p<0.05).Compared with the control group,the villus height significantly decreased and the crypt depth was significantly increased in the BPA treatment group(p<0.05).In duodenum and jejunum tissue,SOD activity showed a significant decline in GLP treatment group compared with the control group(p<0.05).MDA content significantly increased in the ileum tissue in the GLP treatment group compared with the control group(p<0.05).Compared with the control group,the mRNA expression of IL-1,NF-kB,TNF-α,GPX2,MAPK3,SIRT1,PPAR-a,SREBP-1c,SCD-1,Nrf2 and HO-1 significantly increased in the BPA-treated group(p<0.05),but FAS mRNA expression significantly decreased in the BPA-treated group in the small intestine(p<0.05).The high abundance sequencing results showed that the relative abundance of the firmicutes,tenericutes and proteobacteria bacteria were significantly increased(p<0.05)and the relative abundance of bacteroidetes was significantly decreased in the BPA-treated group compared with the control group(p<0.05).BPA also significantly decreased in the relative abundance of romboutsia,aerococcus,alistipes as well as some unclassified flora in the fecal samples of the rats.BPA increase in the relative abundance of ruminiclostridium 9,escherichia-shigella,collinselaandno-rank Mollicutes in fecal samples can be observed(p<0.05).Our results demonstrate that BPA can induce small intestinal injury and change the intestinal microbes in rats.The damage of small intestine may be related to the composition and abundance of the rat flora.5.Effects of 4-nitrophenol on expression of the ER-a and AhR signaling pathway-related genes in the small intestine of rats The aim of this study was to evaluate the role of the estrogen receptor-a(ER-a)and aryl hydrocarbon receptor(AhR)signaling pathway in regulating the damage response to PNP in the small intestine of rats.Wistar-Imamichi male rats(21 d)were randomly divided into two groups:the control group and PNP group.Each group had three processes that were gavaged with PNP or vehicle daily:single dose(1 d),repeated dose(3 consecutive days)(3 d)and repeated dose with recovery(3 consecutive days and 3 recovery days)(6 d).The weight of the body,the related viscera,and small intestine were examined.Histological parameters of the small intestine and the quantities of mucus proteins secreted by small goblet cells were determined using HE staining and PAS staining.The mRNA expression of AhR,ER-a,CYP1A1 and GST was measured by real-time qPCR.In addition,we also analyzed the AhR,ER-a and CYP1A1 expression in the small intestine by immunohistochemical staining.The small intestines histologically changed in the PNP-treated rats,and the expression of AhR,CYP1A1 and GST was increased.While ER-a was significantly decreased in the small intestine,simultaneously,when rats were exposed to a longer PNP treatment,the damages disappeared.Our results demonstrate that PNP has an effect on the expression of AhR signaling pathway genes,AhR,CYP1A1,GST and ER-a in the rat small intestine. |
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