# Immunoregulation by TLR-activated TIM-1+ ProB Cells in Transplantation

> **NIH NIH P01** · UNIVERSITY OF PITTSBURGH AT PITTSBURGH · 2021 · $437,500

## Abstract

Abstract (Project 2) 
Rather than use passive transfer of well differentiated adult immunoregulatory cells as a means to create 
transplant tolerance, we have discovered that a population CpG activated pro-B cells are extraordinarily potent, 
far more potent on a per cell basis than Tregs and far easier to prepare. Transfer of only 40,000, but not 
90,000, induces permanent engraftment and probably tolerance in MHC incompatible islet and cardiac allograft 
mouse models. We have discovered that CpG induces expression TIM-1 a potential master switch for 
expression of an immunoregulatory module in mature Bregs (see Project 1/ Kuchroo). We also learned that 
ligation of TIM-1 by anti-TIM-1 mAb induces expression of IL-10, a member of the regulatory model discovered 
by Kuchroo. The stepwise contributions of CpG and activation of the TIM-1 pathway are examined through 
RNAseq and NanoString based analysis. To more precisely characterize the fate and phenotype, proliferative 
and differentiation of CpG activated pro-B cells, lineage-tracking methods are utilized throughout. We know 
that CpG activated pro-B cells proliferate and differentiate after transfer when introduced in optimal, not 
excessive, cell number. Depending upon environmental cues CpG activated pro-B cells can differentiate into a 
variety Bregs but CpG activated pro-B cells from RAG KO mice do not generate Bregs. In this project we will 
analyze the cellular basis of tolerance believing that CpG activated B regs and their progeny create an 
environment conducive to activation, expansion and retention of donor specific Tregs. The specific but 
multifaceted requirements for B cell differentiation, including the nature of the niche impairing expansion and 
differentiation of CpG-ProBs, in the acquisition of tolerance will be analyzed through use of a panel of carefully 
selected gene knockout mice. The role of T and B regs in the induction and maintenance of tolerance will be 
determined through time course protocols that selectively destroy Tregs or the mature progeny of CpG activate 
pro cells. The diversity of CpG-ProB progeny and their expression of a regulatory module that is IL-10 inclusive 
will be examined by single cell RNAseq of CpG-ProBs and their progeny at the transplant.

## Key facts

- **NIH application ID:** 10214480
- **Project number:** 5P01AI129880-04
- **Recipient organization:** UNIVERSITY OF PITTSBURGH AT PITTSBURGH
- **Principal Investigator:** DAVID M ROTHSTEIN
- **Activity code:** P01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $437,500
- **Award type:** 5
- **Project period:** 2018-08-01 → 2023-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10214480

## Citation

> US National Institutes of Health, RePORTER application 10214480, Immunoregulation by TLR-activated TIM-1+ ProB Cells in Transplantation (5P01AI129880-04). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10214480. Licensed CC0.

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