Immunolocalization of serum proteins in living mouse glomeruli under various hemodynamic conditions by “in vivo cryotechnique”

Abstract

Distribution of serum proteins in renal glomeruli is important for histopathology in medical and biological fields, but mechanisms of their passage through glomerular capillary loops (GCL) are still difficult to clarify. We have tried to visualize topographical changes of the serum proteins passing through GCL by "in vivo cryotechnique" in combination with immunohistochemistry. Albumin and immunoglobulin G (IgG), Ig kappa light chain and IgG1 heavy chain were mainly immunolocalized in GCL, but not colocalized with zonula occludens-1 (ZO-1) under normotensive condition. Under heart-arrest condition and in quick-frozen fresh tissues, albumin and kappa light chain were immunolocalized in Bowman's space, indicating their passage caused by the stoppage of blood supply. However, under acute hypertensive condition, they were more clearly immunolocalized along basement membranes and in the Bowman's space, indicating their increased passage through GCL. IgG was also more clearly localized in mesangial areas under acute hypertension, compared with that under the normotensive or heart-arrest condition. This study is the first direct visualization for glomerular passage of serum proteins under abnormal hemodynamic conditions by the "in vivo cryotechnique", and the experimental protocol will be useful for morphofunctional examination of living mouse GCL and immunohistochemical analyses of dynamically changing proteins.