טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentMonga Sheelu
SubjectThe Effect of the Novel TSPO Ligands (MGV-1, 2-Cl-MGV-1,
2,4-Di-Cl-MGV-1, CB86, and CB204) on the
LPS-Induced Inflammatory Pathways
DepartmentDepartment of Medicine
Supervisors ASSOCIATE PROF. Rafael M. Nagler
PROFESSOR EMERITUS Moshe Gavish
Full Thesis textFull thesis text - English Version


Abstract

The 18 kDa translocator protein (TSPO) ligands 2-Cl-MGV-1 and MGV-1 can attenuate cell death of astrocyte-like cells (U118MG) and induce differentiation of neuronal progenitor cells (PC-12). Lipopolysaccharide (LPS) is a bacterial membrane endotoxin that activates cellular inflammatory pathways by releasing pro-inflammatory molecules, including cytokines and chemokines. The aim of the present study was to assess the immuno-modulatory effect of TSPO ligands in activated microglial cells. We demonstrated that the TSPO ligands 2-Cl-MGV-1, MGV-1; 2,4-Di-Cl-MGV-1, CB86 and CB204 can inhibit LPS-induced activation of microglia (BV-2 cell line). Co-treatment of LPS (100 ng/mL) with 2-Cl-MGV-1 and MGV-1 (final concentration- 25 µM) reduces significantly the LPS-induced release of interleukin-6 (IL-6) from 16.9-fold to 2.5-fold, IL-β from 8.3-fold to 1.6-fold, interferon-γ from 16.0-fold to 2.2-fold, and tumor necrosis factor-α from 16.4-fold to 1.8-fold. Exposure to LPS along with the TSPO ligand 2,4-Di-Cl-MGV-1 (25 µM) reduced significantly the release of NO by 24-, IL-6 by 14-, IL-1β by 14-, IFN- γ by 6-, and TNF-α by 29-folds, respectively. In contrast to the anti-neuroinflammatory effect of the TSPO ligands, the effect of diclofenac sodium (DS; 25 µM) did not reach statistical significance. This anti-inflammatory activity seems to be achieved by inhibition of NF-κB p65 activation. Assessment of initiation of ROS generation and cell metabolism shows significant protective effects of these two novel TSPO ligands. The IL-10 and IL-13 levels were not affected by any of the TSPO ligands.

The impact of ligands of the 18 kDa translocator protein (TSPO) on the release of chemokines was not vastly investigated. In the present study, we assessed the effect of our novel TSPO ligands 2-Cl-MGV-1 and 2,4-Di-Cl-MGV-1 compared to the classical TSPO ligand PK11195 on chemokine release in LPS-stimulated BV-2 microglial cells. As per the effect of 2-Cl-MGV-1, CCL2, CCL3, and CCL5 were inhibited by 90%, CCL8 by 97%, and IL-2 by 77% (p<0.05 for all). 2,4-Di-Cl-MGV-1 inhibited CCL2 release by 92%, CCL3 by 91%, CCL5 by 90%, CCL8 by 89% and IL-2 by 80% (p<0.05 for all). PK11195 exhibited weaker inhibitory effects: CCL2 by 22%, CCL3 by 83%, CCL5 by 34%, CCL8 by 41%, and the cytokine IL-2 by 14% (p<0.05 for all). Thus, it appears that the novel TSPO ligands are potent suppressors of LPS-stimulated BV-2 microglial cells, and their inhibitory effect is larger than that PK11195. Thus, it appears that the ligands suppress the LPS-induced activation of some inflammatory responses of microglia. Such immunomodulatory effects may be relevant to the pharmacotherapy of neuro-inflammatory diseases.