Phenotypic Assays to Identify Agents That Induce Reactive Gliosis: A Counter-Screen to Prioritize Compounds for Preclinical Animal Studies

ASSAY and Drug Development Technologies, 2015 · DOI: 10.1089/adt.2015.654 · Published: September 1, 2015

Simple Explanation

After traumatic central nervous system (CNS) injury, astrocytes, a type of brain cell, change in a process called reactive gliosis. This involves changes in their structure and the molecules they produce. While the initial response of astrocytes after injury can be helpful, in the long term, they can release substances that hinder the regrowth of damaged nerve fibers. The study developed assays to identify compounds that alter astrocytes in a manner that mimics the glial reaction to injury, focusing on changes in GFAP morphology and CSPG expression.

Study Duration

Not specified

Participants

Primary astrocyte cultures from postnatal day 6 (P6) rat cortices

Evidence Level

Not specified

Key Findings

1
The GFAP morphology assay is suitable for counter-screening with a Z-factor of 0.44 – 0.03.
2
The CSPG assay is reproducible and informative, but does not satisfy common metrics for a screenable assay.
3
One compound was identified that can enhance axon growth without exacerbating the deleterious characteristics of reactive gliosis.

Research Summary

The study developed two HCA assays to investigate reactive-like changes in astrocytes: one measuring the stellation index (GFAP+ process area/cell body area) and another monitoring the expression of glial scar-associated CSPG epitopes. The stellation index assay proved suitable for screening with a Z-factor of 0.44 – 0.03, while the CSPG assay was reproducible and informative but did not meet common metrics for a screenable assay. A small-scale pilot screen identified a compound, ML-7, that enhanced axon growth without inducing reactive-like astrocyte phenotypes, suggesting its potential for CNS injury therapies.

Practical Implications

Drug Discovery

The assays can be used to screen compound libraries and identify potential drug candidates for CNS injuries that promote axon regeneration without exacerbating reactive gliosis.

Counter-Screening Strategy

The counter-screening strategy may accelerate the development of effective drugs for CNS injury by prioritizing the most promising compounds from a primary screen for preclinical regeneration studies.

Understanding Reactive Gliosis

The assays provide a tool to study the molecular mechanisms underlying reactive gliosis and identify potential targets for therapeutic intervention.

Study Limitations

1
Adult astrocytes are more relevant to regeneration failure but are technically challenging for screening.
2
The CSPG assay has negative Z-factors for the parameters measured, limiting its use as a high-throughput screening tool.
3
Many kinase inhibitors do not cross the blood-brain barrier effectively, complicating the extrapolation of biologically relevant dosages from in vitro experiments.

Your Feedback

Was this summary helpful?

Back to Pharmacology