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Retinal microglial cells increase expression and release of IL-1β when exposed to ATP
Abstract:
Cytokine IL-1β is an early component of inflammatory cascades, with both priming and activation steps required before IL-1β release. Here, the P2X7 receptor (P2X7R) for ATP was shown to both prime and release IL-1β from retinal microglial cells. Isolated retinal microglial cells increased expression of Il1b when stimulated with endogenous receptor agonist extracellular ATP; ATP also rapidly downregulated expression of microglial markers Tmem119 and Cd206. Changes to all three genes were reduced by specific P2X7R antagonist A839977, implicating the P2X7R. Microglial cells expressed the P2X7R on ramifications and responded to receptor agonist BzATP with robust and rapid rises in intracellular Ca 2+ . BzATP increased expression of IL-1β protein colocalizing with CX3CR1-GFP in retinal wholemounts consistent with microglial cells. ATP also triggered release of IL-1β from isolated retinal microglia into the bath; release was inhibited by A839977 and induced by BzATP, supporting a role for the P2X7R in release as well as priming. The IL-1β release triggered by ATP was substantially greater from microglial cells compared to astrocytes from the optic nerve head region. Il1b expression was increased by a transient rise in intraocular pressure and Il1b levels remained elevated 10 days after a single IOP elevation. In summary, this study suggests the P2X7 receptor can both prime IL-1β levels in microglial cells and trigger its release. The P2Y12R was previously identified as a chemoattractant for retinal microglia, suggesting the recruitment of the cells towards the source of released extracellular ATP could position microglia for P2X7R receptor, enabling both priming and release of IL-1β.
Insights
The P2X7 receptor (P2X7R) in retinal microglial cells primes and releases IL-1β, a key inflammatory cytokine. This process is triggered by extracellular ATP and implicated in inflammatory responses within the eye.
Area of Science:
- Ophthalmology
- Neuroscience
- Immunology
Background:
- Interleukin-1 beta (IL-1β) is a critical cytokine in inflammatory processes.
- Its release involves priming and activation steps.
- Microglial cells are key immune cells in the retina.
Purpose of the Study:
- To investigate the role of the P2X7 receptor (P2X7R) in regulating IL-1β release from retinal microglial cells.
- To determine if P2X7R activation by extracellular ATP influences microglial activation and IL-1β expression.
- To explore the involvement of P2X7R in inflammatory responses in the retina.
Main Methods:
- Isolated retinal microglial cells were stimulated with extracellular ATP and its analogs.
- Gene expression of IL-1β, Tmem119, and Cd206 was analyzed.
- P2X7R antagonist A839977 was used to assess receptor specificity.
- Intracellular calcium (Ca 2+) levels were measured in response to P2X7R agonists.
- IL-1β protein release was quantified in isolated microglia and compared to astrocytes.
- Effects of intraocular pressure (IOP) elevation on IL-1β expression were studied.
Main Results:
- Extracellular ATP stimulation increased IL-1β gene expression and decreased microglial markers (Tmem119, Cd206) in retinal microglia.
- These changes were attenuated by the P2X7R antagonist A839977.
- Microglial cells expressed functional P2X7R, responding to BzATP with calcium influx.
- BzATP induced IL-1β protein expression in microglia.
- ATP triggered significant IL-1β release from microglia, an effect inhibited by A839977.
- Microglia released more IL-1β than optic nerve head astrocytes.
- Intraocular pressure elevation increased IL-1β expression, which remained elevated for 10 days.
Conclusions:
- The P2X7 receptor plays a dual role in retinal microglia, both priming IL-1β expression and triggering its release.
- Extracellular ATP acting via P2X7R is a significant mediator of microglial inflammatory responses in the retina.
- These findings suggest a mechanism linking ocular injury (like IOP elevation) to microglial-mediated inflammation via P2X7R signaling.
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