* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Download S. aureus infection
Survey
Document related concepts
Inflammation wikipedia , lookup
Molecular mimicry wikipedia , lookup
Adaptive immune system wikipedia , lookup
Immune system wikipedia , lookup
Adoptive cell transfer wikipedia , lookup
Polyclonal B cell response wikipedia , lookup
Hygiene hypothesis wikipedia , lookup
DNA vaccination wikipedia , lookup
Cancer immunotherapy wikipedia , lookup
Innate immune system wikipedia , lookup
Transcript
Alteration in cytokine and chemokine expression during Staphylococcus aureus wound infections By: Kayla Bounds Definition of a wound • A wound is the disruption of normal anatomic structure and function Lazarus, G.S., et al., 1994. 130(4): p. 489-93 Stages of wound healing Time line for wound healing on a log scale 7-300 days 1-30 days (peak 7 days) 0.15-10 day (peak 1 day) 0-0.1 day Cytokines • What is a cytokine? • Proteins that are important for cell- to- cell signaling and the initiation of the immune response • Cytokines and wound healing • In normal wound healing, cytokines will recruit keratinocytes, fibroblasts, endothelial cells, macrophages, and platelets • In chronic wounds, pro-inflammatory cytokines recruit neutrophils • This results in a state of constant inflammation, preventing proper wound healing Acute wounds • Proceed normally through an orderly and timely regenerative process that results in restoration of anatomic function and integrity http://emedicine.medscape.com/artic le/1277941-overview#aw2aab6b5 Lazarus, G.S., et al., Definitions and guidelines for assessment of wounds and evaluation of healing. Arch Dermatol, 1994. 130(4): p. 489-93 Chronic wounds • Chronic wounds fail to proceed through an orderly and timely process to produce anatomic and functional integrity • Usually remain in the inflammation phase of the healing process Diabetic foot ulcer Biofilmbook.hypertextbookshop.com Venous leg ulcer Biofilmbook.hypertextbookshop.com Pressure ulcer Biofilmbook.hypertextbookshop.com Wound colonization 1° colonizers: Gram positive cocci S. aureus, S. epidermidis, Streptococcus spp., Enterococcus spp. 2 ° colonizers: Gram negative rods P. aeruginosa, E. coli, K. pneumoniae, A. baumannii Wound bed 3 ° colonizers: Anaerobic species Prevotella spp., Porphorymonas spp. Biofilm within wounds Dirckx, P. Center for BiofilmEngineering. Montana State University Hypothesis • Bacterial infection of wounds significantly alter the expression of cytokines and chemokines Objectives 1. To generate a wound using the murine model of wound infection 2. Infect the wound with S. aureus 3. Examine the expression of cytokines 1 day, 3 day, and 7 day post infection Staphylococcus aureus • Catalase-positive, gram- positive • Cocci arranged in clusters • Facultative anaerobe • Capsule and slime layer • Commonly isolated in the nasal passage Berger, J. 2011. Science Photo Lib. Murray, P. et al. 2005. Murine Model of Wound Infection Remove hair from mice Euthanize mice and collect tissue 1.5 x 1.5 cm full-thickness surgical wound covered with OPSITE dressing Inject of S. aureus under dressing Experimental design 1 Uninjured Injured uninfected Collect the tissues 1 day post injury/infected Injured infected Experimental design 2 5 mm-tissue punch from site adjacent to wound RNA later Homogenize tissue in RNA Bee Extract RNA Reverse transcribe to cDNA Use data for analysis Perform real-time PCR using RT² Profiler™ PCR Array Mouse Cytokines and Chemokines kit (QIAGEN) Perform initial realtime PCR using primers for specific cytokines to check RNA quality Cytokines/chemokines and the functional groups Response to injury Immune Responses Hematopoiesis and Production of immune cells Recruitment, activation of immune cells Response to injury Injured/uninjured fold change 1.58 Protein Function Adipoq Metabolism (fat cells) Bmp2 Bone mineralization 1.50 Bmp4 Bone formation 2.00 Bmp6 Response to injury Osteoclast Function, wound healing Central Nervous system Wound repair, matrix maintenance, fibrosis Wound healing 1.59 Spp1 Cntf Tgfb Ccl20 31.96 3.14 1.00 63.70 Response to injury Injured infected/Injured uninfected fold change -3.23 Protein Function Adipoq Metabolism (fat cells) Bmp2 Bone mineralizaiton -4.00 Bmp4 Bone formation -3.23 Bmp6 Response to injury -4.00 Spp1 Osteoclast function, wound healing -4.00 Cntf Central nervous system Wound repair, matrix maintenance, fibrosis Wound healing -4.00 Tgfb Ccl-20 -4.00 -1.27 Recruitment and activation of immune cells Injured/Uninjured fold change 18.45 Protein Function Csf3 G-Csf: activation of granulocytes Cxcl1 PMN chemotaxis 43.91 Cxcl3 PMN chemotaxis 193.55 Cxcl5 PMN chemotaxis 2198.70 Ppbp CXCL-7: PMN chemotaxis 20.43 Ccl2 MCP-1: Monocyte chemotaxis, differentiation 12.60 Ccl3 MIP-1a:Monocyte chemotaxis, macrophage diff. 133.56 Ccl4 MIP-1b: Monocyte chemotaxis, macrophage diff. 100.17 Ccl7 MCP-3: Monocyte chemotaxis, macrophage diff. 6.32 Ltb TNFb :Lymphocyte development in periphery -2.00 Cxcl10 Th1, CD8, NK cell movement -1.28 Ccl17 Th2, Treg trafficking 1.57 Cxcl13 B cell movement 10.02 Recruitment and activation of immune cells Protein Function Csf3 Cxcl1 Cxcl3 Cxcl5 Ppbp G-Csf: activation of granulocytes PMN chemotaxis PMN chemotaxis PMN chemotaxis CXCL-7: PMN chemotaxis Ccl2 MCP-1: Monocyte chemotaxis, differentiation Ccl3 Ccl4 Ccl7 MIP-1a: Monocyte chemotaxis, macrophage diff. MIP-1b: Monocyte chemotaxis, macrophage diff. MCP-3: Monocyte chemotaxis, macrophage diff. Injured infected/injured uninfected fold change 1.25 -1.59 2.50 1.58 -3.23 1.26 1.59 1.26 1.25 Ltb TNFb: Lymphocyte development in periphery 1.99 Cxcl10 Ccl17 Cxcl13 Th1, CD8, NK cell movement Th2, Treg trafficking B cell movement 6.35 5.05 -2.56 Role of cytokines in wound healing in uninfected WOUND Epidermis Blood clot Dermis Capillaries Damaged keratinocytes – release IL-1; and compared with UNINJURED tissue, this Stimulates intact keratinocytes to express: Mcp1 (↑12.6), Cxcl1 (↑43.9), Cxcl12 (↑2.5), Cxcl20 (↑63.7) Recruits neutrophils, which express: Il1 (↑2.5), Il6 (↑33.7), Tnfa (↑2.5) Recruits macrophages, which express: Spp1 (↑32), Il1, Il6, Tnfa Regenerating keratinocytes express: Bmp2 (↑1.5), Bmp4 (↑2), Bmp6 (↑1.6), Bmp7 (↑2.5) Effect of S. aureus infection on expression of cytokines in injured tissue Epidermis Blood clot S. aureus Dermis Capillaries Damaged keratinocytes – release IL-1, BUT, compared to uninfected wound expression of all cytokines expressed in uninfected is depressed or not enhanced Keratinocyte expression of: Mcp1 (1.3), Cxcl1 (−1.6), Cxcl12 (−2), Cxcl20 (−1) Neutrophil expression of: Il1 (−1.6), Il6 (-8.3), Tnfa (-2.6) Macrophage expression of: Spp1 (-4.0), Il1, Il6, Tnfa Keratinocyte regeneration delayed (?): Bmp2 (−1.3), Bmp4 (1.6), Bmp6 (−1), Bmp7 (1.6) Acknowledgements • Committee members: • Dr. Abdul Hamood Lab • • • • Dr. Jeter Dr. Colmer-Hamood Dr. Hamood Dr. Zak • Next Science • Dr. Matt Myantti • Dr. Cassandra Kruczek • • • • Nyarie Dzvova Nithya Mudaliar Adeet Amin Sabrina Siddiqui • Kyle Miller M.S. • Dr. Kendra Rumbaugh Lab