PROTEINASE ACTIVITY IN NORMAL HUMAN TEARS - MALE-FEMALE DIMORPHISM

Recent studies using radial caseinolysis suggested that preoperative p lasmin and plasminogen-activators were predictive indicators of surgic al outcomes. In this study, zymography was used to determine if other caseinolytic proteases were present in normal human tears. Tear sample s were collected and proteins extracted with a buffer of 2% sodium dod ecyl sulfate (SDS), 100 mM Tris/HCl, pH 6.8. Tears were run on casein zymograms (4 mg/ml) which were incubated in 50 mM Tris/HCl, 5 mM CaCl2 , 0.15 M NaCl and NaN3, (pH 7.5) at 37 degrees C for 48 h. After stain ing and destaining, the caseinolytic activity was seen as cleared area s. Plasminogen activator activity was analyzed with zymograms of copol ymerized casein and plasminogen incubated in phosphate buffer saline ( pH 8.0) overnight at room temperature. Results showed that besides a 5 0 kDa plasminogen activator, human tears contained 3 other caseinolyti c activities of molecular weights 90 kDa, 70 kDa and 30 kDa. These wer e serine proteases with optimal activities at pH 7-8 and did not have plasminogen-activator properties. The 70 kDa activity required calcium but the others did not. The 30 kDa caseinolytic activity was gender-r elated as it was readily detected in males and was absent or weakly ex pressed in females.