Mechanisms Of Type A Trichothecenes-induced Anorexia In Mice

Type A trichothecenes?TS?is one of important naturally existing mycotoxins,second metabolite of Fusarium.Type A TS mainly contaminate cereal,especially barley,oats,wheat,corn and product of these cereal.These mycotoxins accumulate in animal and animal food through contaminant feedstuff.Type A TS threats great risks to human and animal health.Type A TS elicites a series of adverse effects,including anoreixa,emesis,growth retardation,immune suppression and neuroendocrine disorder in humans and animals.Type A TS-induced anoreixa is serious food-borne intoxication reaction.Although T-2 toxin induced anorectic responses have been proved,the dosage,toxicity and mechanism of the anorexia induced by type A TS within one species remain unclear.Type A TS mainly includes four toxins:?1?T-2 toxin?T-2?,?2?HT-2 toxin?HT-2?,?3?Diacetoxyscirpenol?DAS?and?4?Neosolaniol?NEO?.T-2,HT-2,DAS and NEO were studied in this experiment and mouse acute feed refusal assay model were established with two exposure methods:oral gavage and intraperitoneal?IP?injection.The no observed adverse effect?NOAEL?and lowest observed adverse effect?LOAEL?of T-2-,HT-2-,DAS-and NEO-induced anorexia were obtained based on the dose effect and duration of food refusal.Testing gut satiety hormones cholecystokinin?CCK?,glucagon-like peptide-1?GLP-1?,glucose dependent insulinotropic polypeptide?GIP?and peptide YY3-36?PYY3-36?in plasma with enzyme linked immunosorbent assay?ELISA?method and relating the relationships between the anorexia induced by type A TS and gut satiety hormones;Similarly,plasma neurotransmitters serotonin?5-HT?and substance P?SP?were tested with ELISA method and the relationship between type A TS-induced anorexia and neurotransmitters were related.The hepatic and splenic mRNA levels of proinflammatory cytokines interleukin-1??IL-1??,interleukin-6?IL-6?and tumour necrosis factor-??TNF-??that were related to appetite were detected with real-time polymerase chain reaction?RT-PCR?and the relationship between the anorectic responses induced by type A TS and cytokines were related.1.Comparison of type A TS induced anorexia with mouse bioassayThis experiment was based on mouse acute feed refusal assay model with two methods oral gavage and IP administration.0,0.01,0.1,0.5 and 1 mg·kg-1·bw dose T-2,HT-2,DAS and NEO were exposed to mice with oral gavage or IP method and accumulated food intake were detected at 0.5,1,2,3,6,16,24,48,72 and 96 h.Depending on the dose effect,duration and toleration of food refusal,the capacity of T-2-,HT-2-,DAS-and NEO-induced anorexia were compared.The NOAEL and LOAEL of T-2-,HT-2-,DAS-and NEO-induced food refusals were obtained by statistically analytic method.The results indicated that:Type A TS could induce transient acute food refusal responses and the responses were quickly occurred with dose-dependent manner.Following oral gavage with T-2,HT-2,DAS and NEO,the NOAEL was 0.01,0.01,0.1 and 0.01 mg·kg-1·bw,respectively,and the LOAEL was 0.1,0.1,0.5 and 0.1 mg·kg-1·bw,respectively;After IP administered to T-2,HT-2,DAS and NEO,the NOAEL was 0.01,0.01,<0.01 and 0,01 mg·kg-1·bw,respectively,and the LOAEL was 0.1,0.1,0.01 and 0.1 mg·kg-1·bw,respectively.According to the NOAEL and LOAEL of these four toxins,the approximate toxicity rank order of type A TS was T-2? HT-2?NEO>DAS for oral exposure and DAS>T-2?HT-2?NEO for IP administration.2.Relationships between the anorexia induced by type ATS and gut satiety hormonesBased on the mouse acute feed refusal assay model,mice were exposed to 1 mg·kg-1·bw dosage T-2,HT-2,DAS and NEO and blood were collected at 0,0.5,2,6 and 24 h following toxins administrations.A direct competition ELISA method was used to test the concentrations of satiety hormones CCK,GLP-1,GIP and PYY3-36 in mice plasma,and the changes of satiety hormones following T-2,HT-2,DAS and NEO exposures were analyzed.Oral gavage with T-2,HT-2,DAS and NEO 1)plasma CCK concentration was peaked at 6 h,6 h,2 h and 2 h,respectively,and duration was 24 h,24 h,>6 h and>6 h,respectively;2)compared with CCK,the change of GLP-1 was relatively weak,and the highest concentration was at 2 h,2 h,6 h and 6 h,respectively,and the duration was 6 h,6 h,>6 h,>6 h,respectively.3)the highest concentration of PYY3₃6 in each toxin group was all found at 2 h,and the duration was>6 h,6 h,6 h and 6 h,respectively;4)in four toxins groups,the peak point of plasma GIP concentration appeared at 2 h,and the duration was 24 h,24 h,6 h,and>6 h,respectively.Following IP administered to T-2,HT-2,DAS and NEO 1)plasma CCK concentrations were all peaked at 6 h and durations were all>24 h in four toxins groups;2)plasma GLP-1 concentrations were peaked at 2 h and durations were all 6 h in four toxins groups;3)plasma PYY3-36 concentration was peaked at 6 h,2 h,2 h and 2 h,respectively,and duration was 24 h,>6 h,2 h and 2 h,respectively;4)the peak point of plasma GIP concentration was 2 h,2 h,0.5 h and 0.5 h,respectively,and duration was 24 h 24 h,6 h and 24 h,respectively.To sum up,plasma gut satiety hormones CCK,GLP-1,GIP and PYY3-36 were elevated following exposed to T-2,HT-2,DAS,and NEO and these elevations were positively correlated to T-2-,HT-2-,DAS-and NEO-induced anorexia.These results indicated that gut satiety hormones CCK,GLP-1,GIP and PYY3-36 played roles in T-2-,HT-2-,DAS-and NEO-induced anorexia.3.Relationships between the anorexia induced by type ATS and neurotransmittersThe design of this experiment was consistent with the design of test ?.A direct competition ELISA method was used to test the concentrations of neurotransmitters 5-HT and SP in mice plasma,and changes of neurotransmitters following T-2,HT-2,DAS and NEO exposure were analyzed.Oral gavage with T-2,HT-2,DAS and NEO 1)plasma 5-HT concentration was peaked at 2 h in each toxin group,and duration was 24 h 24 h,6 h and 6 h,respectively;2)similarly,the highest concentrations of SP were all at 2 h and durations were all at 6 h in four toxins group.Following IP administered to T-2,HT-2,DAS and NEO 1)plasma 5-HT concentration was peaked at 6 h,6 h,2 h and 2 h,respectively,and duration was 24 h,24 h,6 h and 6 h,respectively;2)plasma SP concentration was peaked at 2 h,6 h,2 h and 2 h,respectively,and duration was 24 h,24 h,6 h and 6 h,respectively.In conclusion,plasma neurotransmitters 5-HT and SP were elevated following exposed to T-2,HT-2,DAS and NEO and the elevations were positively correlated to T-2-,HT-2-,DAS-and NEO-induced anorexia.These results indicated that plasma neurotransmitters 5-HT and SP played important roles in T-2-,HT-2-,DAS-and NEO-induced anorexia.4.Relationships between the anorexia induced by type ATS and cytokinesThis study was based on the mouse acute feed refusal assay model and mice were exposed to 1 mg·kg-1·bw dosage T-2,HT-2,DAS and NEO.Mice livers and spleens were collected at 0,0.5,2,6 and 24 h following toxins administrations.The mRNA levels of IL-1?,IL-6 and TNF-? in liver and spleen were measured with RT-PCR method.Hepatic and splenic cytokines IL-1?,IL-6 and TNF-? were markedly upregulated following administrated with T-2,HT-2,DAS and NEO.The tendencies of cytokines upregulated were correlated to T-2-,HT-2-,DAS-and NEO-induced anorexia.The results of this study presented herein indicated that hepatic and splenic cytokines IL-1?,IL-6 and TNF-? played roles in T-2-,HT-2-,DAS-and NEO-induced anorexia.This study is the first to compare the acute anorexia toxicity of type A TS T-2,HT-2,DAS and NEO in one species with two methods.Based on the LOAEL and NOAEL induced by T-2,HT-2,DAS and NEO,the approximate toxicity rank order of type A TS were determined.Appetite regulating factors,including gut satiety hormones,neurotransmitters and cytokines,were observed elevated following T-2,HT-2,DAS and NEO exposures and the elevated tendencies of these factors were positively correlated with T-2-,HT-2-,DAS-and NEO-induced anorexia.In this study,we illustrated part mechanism of type A TS induced anorexia and this gave people a further understanding of type A TS'toxicity.The study provided the basis for the formulation of its safety limit,and laid a foundation for the prevention and control of its toxic effects in the future.