Contributions
Abstract: EP1599
Type: Poster Sessions
Abstract Category: Therapy - Immunomodulation/Immunosuppression
Background: Targeting of RNA Polymerase-1 (POL1) machinery is a new strategy for suppression of multiple sclerosis (MS) relapse activity. Oral administration of POL1 inhibitor, RAM-589.555 ameliorates PLP-induced experimental autoimmune encephalomyelitis (EAE) by suppression of activated autoreactive lymphocytes. RAM-589.555 can cross the blood brain barrier and access the central nervous system (CNS). However, its immunomodulatory effects on CNS-resident glial cells have not yet been elucidated.
Objectives: To assess whether RAM-589.555 can modulate microglia and astrocytes.
Methods: CD11+ microglia and GLAST+ astrocytes were isolated from mouse pups, cultured according to Miltenyi Biotec protocol and activated with 100 ng/ml lipopolysaccharide (LPS) and tumor necrosis factor a (TNFa), with various concentrations of RAM-589.555 for 48 hrs. Viability and proliferation were examined by XTT and BrdU assays, respectively. Brain derived neurotrophic factor (BDNF) was measured in supernatants by ELISA and the leading marker of POL1 pathway, pre-ribosomal RNA (pre-rRNA) was tested by RT-PCR.
Results: Viability and proliferation of LPS-activated microglia was not affected at 25-200 nM of RAM589.555 while significantly reduced at 400 nM by 70% and 45% (p< 0.05), respectively. Viability and proliferation of TNFa-activated astrocytes were not affected by RAM589.555. Pre-rRNA expression decreased gradually with elevated concentrations of RAM589.555 both in microglia and astrocytes. RAM589.555 did not alter the levels of secreted BDNF by microglia and by astrocytes (6.7±0.8 and 11.8±2.0 pg/ml in un-treated cultures vs. 9±0.7-11±1.9 and 11.6±1.6-14.3±1.8 pg/ml with 25-400 nM RAM589.555 in microglia and astrocytes, respectively; p>0.05).
Conclusions: RAM589.555 is non-detrimental neither for astrocytes nor in 25-200 nM for microglia while highest RAM589.555 dose impairs the microglial viability and proliferation, probably due to higher proliferation rate of microglia. Notably, the neuro-protective potential of microglia and astrocytes is not impaired by RAM589.555.
Disclosure: RZF: nothing to disclose
MG: nothing to disclose
AA:nothing to disclose
Abstract: EP1599
Type: Poster Sessions
Abstract Category: Therapy - Immunomodulation/Immunosuppression
Background: Targeting of RNA Polymerase-1 (POL1) machinery is a new strategy for suppression of multiple sclerosis (MS) relapse activity. Oral administration of POL1 inhibitor, RAM-589.555 ameliorates PLP-induced experimental autoimmune encephalomyelitis (EAE) by suppression of activated autoreactive lymphocytes. RAM-589.555 can cross the blood brain barrier and access the central nervous system (CNS). However, its immunomodulatory effects on CNS-resident glial cells have not yet been elucidated.
Objectives: To assess whether RAM-589.555 can modulate microglia and astrocytes.
Methods: CD11+ microglia and GLAST+ astrocytes were isolated from mouse pups, cultured according to Miltenyi Biotec protocol and activated with 100 ng/ml lipopolysaccharide (LPS) and tumor necrosis factor a (TNFa), with various concentrations of RAM-589.555 for 48 hrs. Viability and proliferation were examined by XTT and BrdU assays, respectively. Brain derived neurotrophic factor (BDNF) was measured in supernatants by ELISA and the leading marker of POL1 pathway, pre-ribosomal RNA (pre-rRNA) was tested by RT-PCR.
Results: Viability and proliferation of LPS-activated microglia was not affected at 25-200 nM of RAM589.555 while significantly reduced at 400 nM by 70% and 45% (p< 0.05), respectively. Viability and proliferation of TNFa-activated astrocytes were not affected by RAM589.555. Pre-rRNA expression decreased gradually with elevated concentrations of RAM589.555 both in microglia and astrocytes. RAM589.555 did not alter the levels of secreted BDNF by microglia and by astrocytes (6.7±0.8 and 11.8±2.0 pg/ml in un-treated cultures vs. 9±0.7-11±1.9 and 11.6±1.6-14.3±1.8 pg/ml with 25-400 nM RAM589.555 in microglia and astrocytes, respectively; p>0.05).
Conclusions: RAM589.555 is non-detrimental neither for astrocytes nor in 25-200 nM for microglia while highest RAM589.555 dose impairs the microglial viability and proliferation, probably due to higher proliferation rate of microglia. Notably, the neuro-protective potential of microglia and astrocytes is not impaired by RAM589.555.
Disclosure: RZF: nothing to disclose
MG: nothing to disclose
AA:nothing to disclose