Document Type: Special Articles

Authors

1 Associate Professor, Department of Biology, Faculty of Sciences, Ferdowsi University of Mashhad, Mashhad, Iran.

2 Assistant Professor, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran

3 Professor, Department of Biology, Faculty of Sciences, Ferdowsi University of Mashhad, Mashhad, Iran

4 Department of Biology, Faculty of Sciences, Ferdowsi University of Mashhad, Mashhad, Iran

Abstract

Background:  Astrocytes are cells within the central nervous system which are activated  in a wide spectrum of infections,  and  autoimmune and  neurodegenerative diseases.In pathologic states, they produce  inflammatory cytokines, chemokines,  and  nitric oxide (NO), and sometimes  they induce apoptosis.Their protease-activated receptors (PARs) can be activated by proteases, e.g.thrombin and trypsin,which are important in brain inflammation.The current  study  aimed  to investigate the effects of different concentrations of trypsin (1 to 100U/ml) on cultured astrocytes.Methods: In the present  study, two-day rat infants’ brains were  isolated and homogenized after meninges removal, then  cultivated in DMEM+10% FBS medium. 10 days later, astrocytes were harvested  and recultivated for more purification (up to 95%), using Immunocytochemistry method, in order to be employed   for  tests. They were affected by different concentrations of trypsin (1, 5, 10, 15, 0, 40, 60, 80, and  100 U/ml). To reveal the inflammation progress, NO concentrations (the Griess test) were assessed after 24 and 48 hours.Results:The results showed  that  trypsin concentration up to 20 U/ml caused  a significant increase in NO, in a dose- dependent  manner, on cultured   astrocytes   (P <  0.001). Trypsin  20  U/ml  increased  NO production  fivefold  the control group (P < 0.001). At higher  concentrations than 20 U/ml, NO production diminished  (P < 0.001). At 100 U/ml, NO production was less than the  control  group (P < 0.001).Conclusion: Inflammatory effects of trypsin  5-20 U/ml are probably due to the stimulation of astrocytes’ PAR-2 receptors and the increasing of the activation of NF-κB, PKC, MAPKs. Stimulation  of astrocytes’ PAR-2 receptors causes an increase in iNOS activation which in turn leads to NO production. However, higher  trypsin concentration possibly made astrocyte apoptosis; therefore, NO production diminished. These assumptions need t  be further investigated.