title: In Vivo Imaging of Inflamed Glomeruli Reveals Dynamics of Neutrophil Extracellular Trap Formation in Glomerular Capillaries.
abstract: Neutrophil extracellular traps (NETs) have been documented in glomeruli of patients with glomerulonephritis. However, the dynamics of NET formation in the glomerulus and their functional contribution to acute glomerular injury are poorly understood. Herein, we used in vivo multiphoton microscopy to investigate NET formation in the acutely inflamed glomerulus. Glomerular inflammation was induced using an antibody against the glomerular basement membrane. After induction of inflammation, multiphoton microscopy revealed that approximately 20% of glomeruli contained structures composed of extracellular DNA within the capillaries. These structures were not seen in mice depleted of neutrophils, consistent with them being NETs. Most contained myeloperoxidase, as seen in NETs in other tissues, whereas intraglomerular NETs did not contain significant levels of the histone H2Ax or neutrophil elastase. In vivo imaging revealed that intraglomerular NETs were present only transiently, suggesting that NETs were susceptible to disruption under the high shear conditions in glomerular capillaries. Investigation of NETs under flow conditions in vitro supported this concept. Dissolution of NETs via DNase I did not alter anti-glomerular basement membrane antibody-induced glomerular injury, as assessed via albuminuria, although the degree of microscopic hematuria was reduced by this intervention. These data indicate that in this model of acute, neutrophil-dependent glomerulonephritis, NETs are generated in the glomerular capillaries, where they are short lived and make a modest contribution to glomerular injury.
authors.affiliation: Centre for Inflammatory Diseases, Monash University Department of Medicine, Clayton, Victoria, Australia.
authors.forename: Clare L V
authors.initials: CL
authors.lastname: Westhorpe
authors.affiliation: Centre for Inflammatory Diseases, Monash University Department of Medicine, Clayton, Victoria, Australia.
authors.forename: James E
authors.initials: JE
authors.lastname: Bayard
authors.affiliation: Centre for Inflammatory Diseases, Monash University Department of Medicine, Clayton, Victoria, Australia.
authors.forename: Kim M
authors.initials: KM
authors.lastname: O'Sullivan
authors.affiliation: Centre for Inflammatory Diseases, Monash University Department of Medicine, Clayton, Victoria, Australia.
authors.forename: Pam
authors.initials: P
authors.lastname: Hall
authors.affiliation: Centre for Inflammatory Diseases, Monash University Department of Medicine, Clayton, Victoria, Australia.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Cheng
authors.affiliation: Centre for Inflammatory Diseases, Monash University Department of Medicine, Clayton, Victoria, Australia; Department of Nephrology, Monash Medical Centre, Clayton, Victoria, Australia; Department of Pediatric Nephrology, Monash Medical Centre, Clayton, Victoria, Australia.
authors.forename: A Richard
authors.initials: AR
authors.lastname: Kitching
authors.affiliation: Centre for Inflammatory Diseases, Monash University Department of Medicine, Clayton, Victoria, Australia. Electronic address: michael.hickey@monash.edu.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Hickey
mesh_terms: D000818:Animals; D002196:Capillaries / Q000473:pathology*; D004195:Disease Models, Animal; D065206:Extracellular Traps / Q000378:metabolism*; D005921:Glomerulonephritis / Q000473:pathology*; D007150:Immunohistochemistry; D007678:Kidney Glomerulus / Q000098:blood supply / Q000473:pathology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D036641:Microscopy, Fluorescence, Multiphoton