In this study,the effects of cold stress on the expression levels of HSP70 in duodenum,jejunum,ileum and cecum were studied.120 1-day-old male chickens were kept under the temperature of (30± 2) ℃ and were given commercial diet and water.When the chickens were 15 days old,they were randomly divided into 12 groups on average and were kept under the temperature of (12±1) ℃ for acute and chronic cold stress.The duration of acute cold stress were 0,1,3,6,12 and 24 h,and the duration of chronic cold stress were 5,10 and 20 d.0 h was the control group for the acute cold stress,and there were three control groups for chronic cold stress.After the cold stress treatment,the tissues were collected for real-time PCR assessment.The results showed that at the beginning of the acute cold stress and chronic cold strss,HSP70 expression levels were increased significantly(P0.01),but at the end of acute cold stress,the expression levels were decreased significantly(P0.01).The study suggested that HSP70 expressioin levels could be influenced by the cold stress and the expression levels be different by different tissues or different durations of the cold exposure.

Different levels of 600mg/kg,900mg/kg and 1800mg/kg MnCl_ 2 were added into the forage to establish the sub-chronic manganism model.On days 30,60 and 90 post-exposure-to-MnCl_2,the livers of chickens were respectively collected to detect the contents of Mn,Fe,Cu,Zn and Ca by the flame atomic absorption spectrometry.Compared with the control group,the contents of Mn and Cu increased,the content of Zn took on a fluctuated change and the contents of Fe and Ca decreased in each treatment group,which had a time-dose relationship.There were significantly positive correlation between Mn and Cu and significantly negative correlation between Mn and Fe.In conclusion,the excess manganese could change the contents of Mn,Fe,Cu,Zn and Ca in the livers and damage the livers of chickens.

The aim of this study was to investigate the effect of manganese on the hepatic cytochrome P450 enzyme system in cocks.400 fifty-day old Hy-line brown cocks were randomly divided into four groups.The cocks in four groups were respectively fed with the basal diet containing 0,600,900 and 1 800 mg·kg-1 MnCl2 to establish the sub-chronic manganism model.After 30,60 and 90-day treatment,the livers in every group were collected to detect the activity of microsomal cytochrome P450 enzyme system and the transcription level of CYP2H1 gene.The contents of cytochrome P450 and b5,the activities of aminopyrin-N-demethylase(AND) and aniline-4-hydroxylase(AH) were gradually declined with the increase of manganese in diet.The high dose group was significantly lower than the control group while there were no difference between the low and middle dose group and the control group.There were no markedly changes in the activities of NADPH-cytochrome C reductase(CR) and erythromycin-N-demethylase(ERND).The activity of CR declined at 30 and 60 d,but increased at 90 d.Besides,the CR activity of the high dose group was significantly higher than that in the control group(P0.01).The activity of ERND increased at 30 d but no markedly tendency appeared at 60 and 90 d.The ERND activity in the high dose group was significantly higher than that in the low dose group(P0.01).The transcription level of CYP2H1 mRNA in the low dose and high dose group at 30 d,the middle dose group at 60 d,and the low dose group at 90 d were higher than that in the control group.The transcription level of CYP2H1 mRNA in other treatment groups were lower than that in the control group.These results indicated that manganism could significantly changes the activities of microsomal-cytochrome P450s and the transcription level of CYP2H1 mRNA.

To determine the effect of MnCl2 exposure on apoptosis of cock sertoli-germ cell and on expression of Bak and Bcl-x mRNAs,cock sertoli-germ cells were cultivated in DMEM with the final concentrations of 0,2,3 and 4mmol/L MnCl2 respectively for 24h.The apoptosis was determined by TUNEL assay,and the expression amounts of Bak and Bcl-x mRNAs were examined by real-time fluorescence quantitative PCR.In result,compared with control group,the apoptosis index(AI) of cock sertoli-germ cell significantly increased(P0.01),the expression of Bak mRNA increased(P0.01) while the expression of Bcl-x mRNA decreased(P0.01) in 2,3 and 4mmol/L MnCl2 treatment groups.The result indicated that MnCl2 was able to induce apoptosis of cock sertoli-germ cell by up-regulating the expression of Bak mRNA and down-regulating the expression of Bcl-x mRNA.

The aim of this study was to investigate the effect of manganese on the hepatic apoptosis in cocks.400 fifty-day old Hy-line brown cocks were randomly divided into four groups.The 0,600,900 and 1 800 mg·kg-1 MnCl2 were respectively added into the basic forage to establish the subchronic manganism model.After 30,60 and 90-day treatment,the livers in every group were collected to observe hepatic morphological change and detect hepatic apoptosis index and the transcription level of Fas mRNA.Results showed that among 90-day treatment groups,the liver form the highest dose group took on the typical morphologic changes of apoptosis compared with the corresponding control group,and the number of the apoptotic cells gradually increased with the increase of the manganese dose.The transcription level of Fas mRNA also took on the dose-effect relationship at different treatment time.

The aim of this study was to discuss the role of mitochondria mediated apoptosis pathway in Sertoli-germ cell apoptosis induced by manganese in cocks.Cock Sertoli-germ cells were cultivated in the DMEM for 24 h added with MnCl2,whose final concentrations were 0,2,3,and 4 mmol·L-1 respectively.The apoptosis was detected by AO/EB double staining.Mitochondrial transmembrane potential was detected by flow cytometer.Expression of Cytochrome c(Cytc)in cytolymph was detected by western blotting.The activities of Caspase-9,3 were examined by spectrophotography.Compared with control group,the apoptosis index(AI)of cock Sertoli-germ cells was significantly increased(P0.01),mitochondrial transmembrane potential was significantly decreased(P0.01),expression of Cytc in cytolymph was increased,and the activity of Caspase-9,3 were increased(P0.01)in 2,3,and 4 mmol·L-1 MnCl2 group.The apoptosis mediated by mitochondria pathway was one of the reproductive toxic mechanisms caused by manganese.

To explore the roles of mitochondria complex and oxidative stress in the apoptosis of chicken embryonic neurons exposed to MnCl2,the content of ROS and GSH,the activity of mitochondria respiratory chain complex and apoptosis index in neurons cultivated in DMEM with the final concentration of MnCl2 (0,1.5,2.0,2.5mmol/L) for 24h were detected by using the kits.In result,with the increase of the concentration of MnCl2,the activities of mitochondria respiratory chain complexes Ⅰ,Ⅱ,Ⅲ and Ⅳ and the content of GSH decreased.But in 1.5mmol/L MnCl2 environment,the activity of mitochondria respiratory chain complexⅤ decreased,the content of ROS and the apoptosis index increased,and morphological changes of apoptosis were observed in the neurons.These results indicated that MnCl2 could induce the apoptosis of the chicken embryonic neurons which was performed through oxidative stress caused by decreasing the activities of mitochondria respiratory chain complexes.

To investigate the relationship between the methylation of p15 gene and the genesis of kidney tumor in the development of Marek's disease(MD),300 one-day-old chicks were randomly divided into control group and MD group.On 35,40 and 45 days-old,kidneys were collected and then pathologic changes were observed in the experimental replication MD cases and the variations of methylation of p15 gene in the genesis and development of kidney tumor of chicks with MD were detected by the methylated specific PCR.The results showed that the clinical manifestations and pathological changes of the kidney in the MD group were in accord with characteristics of MD,indicating that MD model was successfully generated.No methy-lation of p15 gene was generated in the control group,while 61.7% of methylation of p15 gene was found in the MD group,which was significantly higher(P0.01) than that in the control group in time-dependent manner.It was concluded that the methylation of p15 gene was intimately relevant to the genesis and development of MD.

The aim of this study was to investigate the effect of manganese on the liver in cocks. 400 fifty-day-age Hy-Line brown cocks were randomly divided into four groups. The 0,600,900,1 800 mg/kg MnCl2 were respectively added into the basic diet to establish the subchronic manganism model. After 30,60 and 90 days treatment,the livers in every group were collected. The microstructure and ultrastructure of livers were observed. The antioxidase (GSH-Px,SOD) activities and the MDA content in livers were detected. The livers in the 90-day treatment group took on different degrees of pathological changes by the observation of microstructure and ultrastructure. Additionally,at every time point,the GSH-Px and SOD activities in the liver took on a decreased tendency,and the MDA content increased gradually (P 0.01,P 0.05). The subchronic manganism could cause the pathological changes and decrease antioxidation of the liver in cocks.