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Updated: Dec 5, 2025
Characterization and Isolation of Mouse Primary Microglia by Density Gradient Centrifugation
Published on: February 16, 2018
Restoration of PP2A levels in inflamed microglial cells: Important for neuroprotective M2 microglial viability
Md Nematullah1, M N Hoda2, Shwetanjali Nimker3
1Department of Biochemistry, School of Chemical and Life Sciences, Jamia Hamdard, New Delhi 110062, India.
Abstract:
PP2A, a trimeric Serine/Threonine Protein Phosphatase 2A highly expressed in brain, is a master regulator of cellular functions. Reduction in PP2A activity has been linked to progression of microglial mediated neuroinflammatory diseases. Inflammatory conditions are characterized by increased population of CD86+ve M1 cells and a therapeutic strategy to polarize microglial cells towards CD206+ve M2 cells is the need of hour. In this paper we analyzed A: whether the level of PP2A is altered in CD86+ve cells, B: whether FTY720, a known modulator of PP2A, is able to restore the level of PP2A in inflamed CD86+ve cells. Results revealed that PP2A activity was significantly diminished in inflamed cells but the surprising observation was the cell viability of only 35.99% upon FTY720 treatment in inflamed cells lacking basal PP2A activity. A sharp increase at mRNA level of CD95 and ASK-1 indicated that apoptosis occurred in these cells through CD95/ASK-1/JNK pathway. Importantly, flow cytometric analysis revealed apoptosis of not only CD86+ve cells but also CD206+ve cells. Previous studies have reported that FTY720 polarizes microglial cells towards M2 states; however apoptosis of M2 cells was not studied. As western blot analysis revealed that FTY720 failed to completely restore PP2A, another PP2A modulator, Memantine, was used for co-treatment. Upon co-treatment, the level of PP2A was completely restored and also viability of microglial cells was significantly improved with a significant reduction in apoptosis of M2 cells. These findings suggest that co-treatment strategy may prove beneficial to balance M1/M2 microglial population, thereby improving neuronal functions.
Insights
Reduced Serine/Threonine Protein Phosphatase 2A (PP2A) activity in inflamed brain cells impairs viability. Co-treatment with Memantine and FTY720 restored PP2A, improved cell survival, and reduced apoptosis in M1 and M2 microglial cells.
Area of Science:
- Neuroscience
- Cell Biology
- Immunology
Background:
- Serine/Threonine Protein Phosphatase 2A (PP2A) is crucial for brain function and its reduced activity is linked to neuroinflammation.
- Neuroinflammation involves an increase in CD86+ M1 microglial cells, necessitating strategies to promote CD206+ M2 cells.
Purpose of the Study:
- To investigate PP2A alterations in CD86+ M1 cells during inflammation.
- To evaluate FTY720's efficacy in restoring PP2A levels and its impact on cell viability and apoptosis in inflamed microglial cells.
- To explore a co-treatment strategy using Memantine to enhance PP2A restoration and improve microglial cell survival.
Main Methods:
- Analysis of PP2A levels in CD86+ cells.
- Treatment with FTY720 and assessment of cell viability, apoptosis (via CD95/ASK-1/JNK pathway markers), and PP2A restoration.
- Flow cytometry to analyze apoptosis in both CD86+ and CD206+ cells.
- Co-treatment with FTY720 and Memantine, followed by western blot analysis for PP2A levels and assessment of cell viability and apoptosis.
Main Results:
- PP2A activity was significantly diminished in inflamed CD86+ cells.
- FTY720 treatment led to significant apoptosis (35.99% viability) in inflamed cells and apoptosis in both M1 and M2 cells, with incomplete PP2A restoration.
- Co-treatment with FTY720 and Memantine fully restored PP2A levels, significantly improved microglial cell viability, and reduced apoptosis, particularly in M2 cells.
Conclusions:
- FTY720 alone induces apoptosis in both M1 and M2 microglial cells during neuroinflammation.
- Combined treatment with Memantine and FTY720 effectively restores PP2A activity, enhances microglial cell survival, and mitigates apoptosis.
- This co-treatment strategy offers a promising approach to rebalance M1/M2 microglial populations and potentially improve neuronal function in neuroinflammatory diseases.
