Phenotypes,distribution, and morphological features of antigen-presenting cells in the normal murine iris / 中华实验眼科杂志

ABSTRACT

Background The conventional study of antigen-presenting cells(APCs)in eye relies on in vitro histoimmunochemistry,but its outcome is influenced by many factors.The anterior chamber injection of fluoresceinmarked antibody was used as a new approach before,however,it is liable to lead to injury of cornea.The intravitreal injection of fluorescein-labeled antibody may be important for the in vivo study of the phenotype features of APCs in iris,which is significant for evaluating the function of APCs in immune homeostasis.Objective This study was to investigate the phenotype characters,distribution and morphology of different types of APCs in the normal murine iris.Methods Fifty-one SPF female BALB/c mice(from 6-to 8-week old)were randomized into 17 groups according to the injection of different antibodies.Alexa Fluor 594 or Alexa Fluor 488-tagged ovalbumin (OVA),CD11 c,major histocompatibility complex Ⅱ (MHC-Ⅱ),F4/80,B7-1 and B7-2 monoclonal antibodies or mixtures of two antibodies (2.0 μl)were intravitreally injected at 0.5 mm far from corneal limbus with microneedle under the biomicroscope.The iris tissues were isolated 24 hours after injection.The phenotype characters,precise distribution and morphology of different types of APCs were identified by epifluorescence microscope and laser confocal microscope.In vitro staining was also performed to validate the in vivo staining results.Results After in vivo staining via intravitreal injection,the cell positive for OVA as well as MHC-Ⅱ,F4/80,CD11 c,B7-1 and B7-2 were exhibited with the regular networkline appearance throughout the normal murine iris.Positive cells tagged with Alexa Fluor 594 or Alexa Fluor 488 presented the red or green fluorescence.Double-fluorescein staining showed that about 90％ of F4/80+ cells were OVA+,and MHC-Ⅱ was expressed in about 60％ of F4/80+ cells and CD11c+cells,and about 35％ of F4/80+ cells and CD1 1 c+ cells expressed B7-1 and B7-2 simultaneously,and over 70％ of OVA+ cells were positive to MHC-Ⅱ.These labeled cells were identified as two populations based on their shape.One type was dendritiform cell (DC) with a small cell body and many long dendrites,including OVA+,CD1 1 c+,F4/80+ cells and MHC-Ⅱ + cells ; and the other types were polymorphic population being round,pleomorphic or irregular shape with a large cell body and a few short dendrities,including B7-1 + and B7-2+ cells.Conclusions In vivo intravitreal injection of labeled antibodies can be adapted to visualize the labeled cells in the murine iris.APCs with distinct morphologies,phenotypes and distribution may contribute to the immunologically privileged feature and inflammation of the eye.