Plasminogen in proliferative vitreoretinal disorders

Abstract

Objective: Intravitreal fibrin formation is a frequent observation after vitrectomy performed for a variety of vitreoretinal disorders including proliferative vitreoretinopathy (PVR), proliferative diabetic retinopathy (PDR), and endophthalmitis. Plasminogen activators (PA) have been used for the management of this postoperative complication. This approach requires the presence of plasminogen, the substrate for PA mediated fibrinolysis, in the vitreal cavity.

Methods: Quantification of plasminogen in the vitreous of 60 patients with PVR, PDR, and macular pucker was performed by streptokinase mediated activation using a chromogenic substrate. The presence of immunoreactive plasminogen was confirmed by immunoblot analysis of vitreal proteins and immunocytochemistry of surgically removed epiretinal membranes.

Results: Plasminogen levels were dramatically increased in the vitreous of PVR and PDR patients compared with macular pucker patients and normal controls. Staining for plasminogen in epiretinal membranes was confined to the extracellular matrix. Predominant staining of perivascular areas in PDR specimens indicated that breakdown of the blood-retinal barrier is an important source of intravitreal plasminogen in that condition.

Conclusion: Plasminogen may play a role in traction membrane formation in PVR and PDR. Our biochemical analysis of presurgical vitreous indicates that there may be abundant substrate for PA mediated fibrinolysis in the vitreous cavity after vitrectomy.

Figure 1

Total plasminogen levels in the vitreous. Plasminogen levels in the vitreous were measured as described in Methods. Horizontal bars in the boxplots represent the median. The boxplots show upper and lower quartiles. Error bars indicate ranges of values (* = 0.02/** p < 0.01 by Mann-Whitney U test compared with control vitreous).

Figure 2

Detection of vitreal plasminogen by immunoblot analysis. Vitreous and plasma samples were subjected to SDS-PAGE and immunoblot analysis as described in Methods. The predicted band for plasminogen migrates at 85-90 kDa. All vitreous samples were positive, as predicted from the enzymatic assay (Fig 1) (lane 1, molecular weight standards; lanes 2-6, vitreous samples (1:10), tested in duplicates, of human postmortem eyes (2), macular pucker (3), traumatic PVR (4) and PDR (5, 6); 7, pooled plasma from normal patients (1:30).

Figure 3

Detection of immunoreactive plasminogen in epiretinal membranes. Epiretinal membranes from patients with PVR (A, B) or PDR (C, D) were examined fore immunoreactive plasminogen as described in Methods. Positive immunoreactivity is observed throughout the extracellular matrix in a traumatic PVR epiretinal membrane. Cell rich areas do not display strong immunoreactivity (A). No staining is observed with the isotype control antibody (B). A staining pattern similar to (A) is detected in a PDR membrane (C). The tissue adjacent of a blood vessel displays strong immunoreactivity (D).