声明
摘要
Abstract
1 Introduction
1．2．1Effect of MG infection on chicken respiratory tract and inflammatory response
1．2．2Effect of MG infection on chicken like maerophages(HD11 cells)
1．3Effect of MG infection on chicken thymus and bursa of fabricius (BOF)
1．3．1Effect of MG infection on chicken thymus
1．3．2Effect of MG infection on chicken BOF
1．4MG infection，oxidative stress and cell signaling pathways
1．4．1MG infection caused oxidative stress damage in chickens
1．4．2MG infection and TLR2-NF-κB signaling pathway
1．4．3MG infection and NLRP3 inflammasome signaling pathway
1．4．4MG infection and host cell apoptosis
1．4．5MG infection and mitochondrial dynamics proteins
1．4．6MG infection and host cell autophagy
1．4．7MG infection and Nrf2／HO-1 signaling pathway
1．4．8MG infection and energy metabolism
1．5Baicalin and its intervention mechanisms against MG infection
1．5．1Research overview of baicalin
1．5．2Protective effects of baicalin against MG infection in chicken and HD11 cells
1．6Objectives and significance of research
2 Materials and methods
2．1Experimental materials
2．1．1Drugs and chemical reagents
2．1．2 Main equipments
2．1．3Preparation of solutions
2．2Baicalin alleviated MG-induced oxidative stress and inflammation via modulating NLRP3 inflammasome-autophagypathway in HD11 cells
2．2．1Experimental cells
2．2．3Culture of HD11 cells
2．2．6Detection of oxidant status in HD11 cells
2．2．7Detection of reactive oxygen species (ROS) by flow cytometry
2．2．9Transmission electron microscopic examination of HD11 cells
2．2．10Detection of inflammatory markers by ELISA assays
2．2．11Detection of mRNA expression of inflammation-related genes
2．2．12Detection of protein expression of inflammation-related genes
2．2．13Detection of autophagy-related genes in MG-infected HD11 cells
2．3Study on the molecular mechanisms of baicalin on MG-infected chickens
2．3．1Experimental animals
2．3．2Experimental groups and treatments of chickens
2．3．4Detection of oxidant status in tissues samples
2．3．5Detection of inflammatory markers by ELISA assays
2．3．6Detection of ATPase activities
2．3．7Histopathological examination
2．3．8Extraction of total proteins and western blotting
2．3．9Transmission electron microscopic examination of tissue samples
2．3．10Terminal deoxynucleotidyl transferase-mediated dUTP nick endlabeling (TUNEL) assay
2．3．11Immunofluorescence microscopic examination
2．3．13Baicalin inhibited apoptosis in MG-infected chickens
2．3．14Baicalin restored energy metabolism in MG-infected chickens
2．3．15Effects of baicalin and／or MG on mitochondrial dynamics
2．3．16Baicalin activated autophagy in MG-infected chickens
2．3．17Baicalin upregulated Nrf-2／HO-1 signaling pathway in MG-infected chickens
2．4Data analysis
3 Results
3．1．1Effects of MG infection on cells viability
3．1．2Effects of baicalin on cells viability
3．1．4Baicalin alleviated oxidative stress in MG-infected HD11 cells
3．1．5Baicalin alleviated MG-induced ROS in HD11 cells
3．1．6Baicalin alleviated the disruption in mitochondrial membrane potential (ΔΨM)
3．1．7Baicalin alleviated MG-induced ultrastructural changes in HD11 cells
3．1．9Baiealin suppressed TLR2-MYD88-NF-κB pathway at mRNA level
3．1．10Baicalin suppressed TLR2-MYD88-NF-κB pathway at protein level
3．1．11Baicalin suppressed NLRP3 inflammasome pathway at inRNA level
3．1．12Baiealin suppressed NLRP3 inflammasome pathway at protein level
3．1．13Baicalin modulated autophagy in MG-infected HD11 cells at mRNA level
3．1．14Baicalin modulated autophagy in MG-infected HD11 cells atprotein level
3．2Baicalin attenuated MG-induced inflammation and apoptosis by restoring energy metabolism in chicken lungs
3．2．1Baicalin attenuated MG-induced increase in serum inflammatory markers
3．2．2Histopathological assessment of chicken lungs
3．2．3Ultrastruetural examination of chicken lungs
3．2．4 Baicalin attenuated the increased mRNA expressions of NF-κB pathway in lungs tissues
3．2．5Baicalin attenuated the increased protein expressions of NF-κB pathway at protein level
3．2．6Baicalin suppressed MG-induced apoptosis in lung tissues
3．2．7Baicalin attenuated MG-induce decrease in ATPase activities
3．2．8Baiealin restored the mRNA expressions of energy metabolism-related genes
3．2．9Baiealin restored the protein expressions of energy metabolism-related genes
3．3Baicalin mitigated MG-induced oxidative stress and apoptosis in chicken thymus through the Nrf2／HO-1 defence pathway
3．3．1Baicalin ameliorated oxidative stress in chicken thymus tissues
3．3．2Histopathological examination of chicken thymus tissues
3．3．3Ultrastructural examination of chicken thymus tissues
3．3．4Effects of baicalin and／or MG infection on cytokine activities
3．3．5Baicalin inhibited the mRNA expression of apoptosis-related genes
3．3．6Baicalin inhibited the protein expressions of apoptosis-related genes
3．3．7Detection of apoptosis by TUNEL assay
3．3．8Baicalin upregulated the mRNA expression of the Nrf2／HO-1 pathway in chicken thymus
3．3．9Baicalin upregulated the protein expression of the Nrf2／HO-1 pathway in chicken thymus
3．4Baicalin attenuated immune impairment in chicken BOF through modulation of autophagy and inhibited inflammation and apoptosis
3．4．1Histopathological examination of BOF
3．4．2Transmission electron microscopic examination of chicken BOF
3．4．3Baiealin suppressed NF-κB pathway at mRNA level in chicken BOF
3．4．4Baiealin suppressed NF-κB pathway at protein level in chicken BOF
3．4．5Baicalin suppressed proinflammatory cytokines activities in chicken BOF
3．4．6Baicalin attenuated the increased mRNA expressions of apoptosis-related genes
3．4．7Baiealin attenuated the increased protein expressions of apoptosis-related genes
3．4．8Detection of apoptosis by TUNEL assay
3．4．9Baicalin increased the protein expression of CD8+ cells in chicken BOF
3．4．10Effects of baicalin and／or MG infection on mitochondrial dynamics
3．4．11Baicalin modulated autophagy at mRNA level in chicken BOF
3．4．12Baicalin modulated autophagy at protein level in chicken BOF
4 Discussion
4．1．2MG-induced oxidative stress and baicalin interventions
4．2Anti-inflammatory effects of baicalin on MG infection
4．2．2Mechanisms of baiealin intervening MG-induced inflammation
4．3Baicalin alleviated MG-induced apoptosis
4．3．1Mechanisms of apoptosis and diseases
4．3．2Baicalin inhibited MG-induced apoptosis and associated signaling pathways
4．4Baicalin modulated autophagy and mitochondrial dynamics
4．4．1Autophagy machinery and homeostasis
4．4．2Baicalin modulated autophagy during MG infection
4．4．3Baicalin restored the imbalance in mitochondrial dynamics
4．5Baicalin restored energy metabolism in chicken lungs
4．5．1Dysfunction in energy metabolism linked diseases
4．5．2Baicalin attenuated energy metabolism dysfunction
5 Conclusion
Acknowledgement
References
Papers published in the period of Ph．D．education
东北农业大学;
