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Autocrine alpha-melanocyte-stimulating hormone inhibits NF-kappaB activation in human glioma
T Ichiyama1, I L Campbell, S Furukawa
1Department of Physiology, University of Texas Southwestern Medical Center at Dallas, Dallas, USA.
Abstract:
The neuropeptide alpha-melanocyte-stimulating hormone (alpha-MSH) modulates inflammation by inhibiting production of proinflammatory cytokines. Using a plasmid vector encoding alpha-MSH, we examined whether autocrine alpha-MSH inhibits activation of the nuclear transcription factor NF-kappaB, a factor that is essential to expression of proinflammatory cytokines, in human glioma cells (A-172). Electrophoretic mobility shift assays of nuclear extracts demonstrated that NF-kappaB activation induced by lipopolysaccharide was inhibited in glioma cells transfected with alpha-MSH vector. Western blot analysis revealed that this inhibition was linked to preservation of expression of IkappaBalpha protein. Chloramphenicol acetyltransferase assay indicated that NF-kappaB-dependent reporter gene expression was suppressed in A-172 cells transfected with alpha-MSH vector. Finally, fluorescence staining confirmed that A-172 cells bear alpha-MSH receptors. The findings are consistent with the idea that, in central nervous system (CNS) inflammation, autocrine alpha-MSH exerts anti-inflammatory actions via modulation of NF-kappaB activation by preservation of IkappaBalpha protein. Based on this action of the peptide, it should be possible to treat neurodegenerative disease, stroke, encephalitis, trauma, and other CNS disorders that have an inflammatory component through gene therapy with alpha-MSH vector.
Insights
Autocrine alpha-melanocyte-stimulating hormone (alpha-MSH) gene therapy inhibits NF-kappaB activation in human glioma cells. This finding suggests potential treatments for neuroinflammatory diseases by preserving IkappaBalpha protein.
Area of Science:
- Neuroscience
- Molecular Biology
- Immunology
Background:
- The neuropeptide alpha-melanocyte-stimulating hormone (alpha-MSH) is known to modulate inflammation by inhibiting proinflammatory cytokine production.
- Nuclear transcription factor kappa B (NF-kappaB) is essential for the expression of proinflammatory cytokines and plays a key role in central nervous system (CNS) inflammation.
Purpose of the Study:
- To investigate whether autocrine alpha-MSH can inhibit NF-kappaB activation in human glioma cells (A-172).
- To explore the potential of gene therapy using alpha-MSH for treating CNS disorders with an inflammatory component.
Main Methods:
- Transfection of A-172 human glioma cells with a plasmid vector encoding alpha-MSH.
- Electrophoretic mobility shift assays (EMSA) to assess NF-kappaB activation.
- Western blot analysis to examine IkappaBalpha protein expression.
- Chloramphenicol acetyltransferase (CAT) assay for reporter gene expression.
- Fluorescence staining to detect alpha-MSH receptors.
Main Results:
- NF-kappaB activation induced by lipopolysaccharide (LPS) was significantly inhibited in glioma cells transfected with the alpha-MSH vector.
- The inhibition of NF-kappaB activation correlated with the preservation of IkappaBalpha protein expression.
- NF-kappaB-dependent reporter gene expression was suppressed in cells transfected with the alpha-MSH vector.
- A-172 cells were confirmed to express alpha-MSH receptors.
Conclusions:
- Autocrine alpha-MSH exerts anti-inflammatory effects in the CNS by modulating NF-kappaB activation through the preservation of IkappaBalpha protein.
- Gene therapy with an alpha-MSH vector holds promise for treating neuroinflammatory conditions such as neurodegenerative diseases, stroke, encephalitis, and trauma.