# Molecular basis for the loss of differentiation capability in human bone marrow stem cells during expansion

> **NIH NIH R21** · LOUISIANA STATE UNIV A&M COL BATON ROUGE · 2021 · $154,352

## Abstract

Project Summary
The ability of self-renewal and the capability to differentiate into mature cell types are two critical properties of
stem cells. Tissue-derived stem cells (TDSCs) possess multipotent differentiation capability, which is the most
crucial property to make them a valuable cell source for treatments of diseases or injuries. Usually, only limited
quantities of primary stem cells can be directly isolated from tissues; therefore, in vitro expansion of primary stem
cells is needed to obtain large quantities of cells for therapeutic applications. However, in vitro expansion of
TDSCs results in the cells to lose their differentiation capability, which makes a generation of large quantities of
high-potential stem cells difficult. This dramatically hampers the therapeutic applications of the TDSCs. Our long-
term goals are: (a) To elucidate the molecular mechanisms causing the loss of differentiation in TDSCs during
expansion. (b) To develop methods to preserve their differentiation capability, such that large quantities of high-
potency TDSCs can be obtained from in vitro expansion for cell-based therapies. In preliminary studies, we
found that expression of cysteine-rich secretory protein LCCL domain-containing-2 (Crispld2) is dramatically
decreased in major TDSCs when the cells lose their osteogenic capability after in vitro expansion. More
importantly, knockdown of this gene in high potential osteogenic TDSCs causes the cells to lose their osteogenic
differentiation ability. Crispld2 knockout was reported to be embryonic lethal, suggesting its crucial role in
development. Our central hypothesis is that in vitro expansion of TDSCs results in dysregulation of Crispld2,
which in turn leads to loss of their differentiation ability. The hypothesis will be tested in two specific aims with
human bone marrow stem cells (hBMSCs): Aim 1. Identify and characterize transcription factors (TFs) regulating
Crispld2 expression in hBMSCs. Aim 2. Determine if enforced or induced expression of Crispld2 affects
differentiation of hBMSCs. We will combine novel and modern technologies, CRISPR/Cas9, and bioinformatics,
with traditional cell and molecular biology methods, such as cell culture, bioassays, gene expression analysis,
and cell differentiation assays to achieve these Aims. In vitro cell experiments and in vivo animal studies will be
used in our approaches. This proposal is significant because it will lead to overcoming the difficulty in generating
large quantities of TDSCs by understanding the molecular regulation for maintaining the differentiation capability
of the stem cells. We expect to identify TFs (activators and repressors) that regulate Crispld2 expression and
determine whether high-level Crispld2 expression is necessary to preserve the differentiation capability of
TDSCs. Accomplishment of this project will begin to elucidate the regulatory network of Crispld2. Because
Crispld2 is widely expressed in many tissues and has multiple functions,...

## Key facts

- **NIH application ID:** 10240719
- **Project number:** 5R21AR076583-02
- **Recipient organization:** LOUISIANA STATE UNIV A&M COL BATON ROUGE
- **Principal Investigator:** Shaomian Yao
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $154,352
- **Award type:** 5
- **Project period:** 2020-08-17 → 2025-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10240719

## Citation

> US National Institutes of Health, RePORTER application 10240719, Molecular basis for the loss of differentiation capability in human bone marrow stem cells during expansion (5R21AR076583-02). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10240719. Licensed CC0.

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