# Decoding the language of inflammation between central nervous system resident immune cells

> **NIH NIH DP2** · UNIVERSITY OF CALIFORNIA BERKELEY · 2024 · $481,500

## Abstract

SUMMARY
Interactions between central nervous system (CNS)-resident cells are highly heterogeneous; astrocytes and
microglia nourish and protect neurons, while inflammatory subsets drive demyelination and neurodegeneration
in neurologic diseases. However, the molecular mechanisms that control CNS-resident immune cell
interactions remain mostly unknown because methods for defining the specific cell types, pathways and
molecules involved are limited. In this project we apply two novel approaches that we developed to study
astrocyte-microglia interactions during inflammation: 1) an in vivo barcoded rabies tracing strategy that
analyzes cell connections and transcriptomes with single cell resolution, and 2) a droplet-based platform for
genome-wide, unbiased CRISPR/Cas9 screening of interacting cell pairs. These methods provide a unique
opportunity to study pathways used by CNS-resident immune cells to communicate with each other and control
inflammation and neurodegeneration. We propose to:
SPECIFIC AIM 1: Define the transcriptomes of single cells in pro-and anti-inflammatory networks. We
will simultaneously sequence connections and single cell transcriptomes of CNS cells in the experimental
autoimmune encephalomyelitis (EAE) mouse model of multiple sclerosis (MS). These studies will identify pro-
and anti-inflammatory cellular networks and the molecular mechanisms that regulate disease-relevant cell-cell
interactions within these networks.
SPECIFIC AIM 2: Identify novel astrocyte-microglia interactions using a droplet-based platform for
CRISPR/Cas9 forward genetic screens. We will perform unbiased genome-wide screens for genes that
participate in microglia-astrocyte crosstalk. We will co-incubate and microfluidically sort millions of picoliter
water-in-oil droplets containing single microglia harboring a CRISPR/Cas9 library mutation and single
astrocytes carrying a fluorescent reporter. Independent droplets do not mix, providing a powerful platform for
the identification of immune interactions mediated by cell surface and soluble molecules.

## Key facts

- **NIH application ID:** 10640907
- **Project number:** 5DP2AI154435-03
- **Recipient organization:** UNIVERSITY OF CALIFORNIA BERKELEY
- **Principal Investigator:** Iain Clark
- **Activity code:** DP2 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $481,500
- **Award type:** 5
- **Project period:** 2021-06-07 → 2027-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10640907

## Citation

> US National Institutes of Health, RePORTER application 10640907, Decoding the language of inflammation between central nervous system resident immune cells (5DP2AI154435-03). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10640907. Licensed CC0.

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