# Silk Fibers-Assisted 3D System for Large-Scale Culture of Human Urine-Derived Stem Cells Suitable for Late Mitotoxicity Testing

> **NIH NIH R03** · WAKE FOREST UNIVERSITY HEALTH SCIENCES · 2021 · $77,500

## Abstract

PROJECT SUMMARY
In vitro 3-D cultures are a promising way to assess delayed mitochondrial toxicity (MtT) caused by antiretroviral
therapy. Standard MtT testing requires large numbers of cells for serial assessments. However, there are no
3D platforms available for mass production of primary human cells for long-term culture. In currently accessible
3D spheroid models, ability to control the geometry of the structures is limited, which does not support reliable
and scaled-up cell production. Furthermore, creating large numbers of primary human cells in 3D spheroid
systems is time-consuming, labor-intensive, and expensive for MtT assessment. Thus, a novel approach to
address each of these issues is urgently demanded. Cells seeding in porous biomaterials used for tissue repair
might provide a solution to these current barriers to progress. Various biomaterials with porous microstructure
have been used in 3D culture for tissue engineering, such as natural materials (spider silk, chitosan,
microspheres made from collagen, gelatin, fibrinogen, hyaluronic acid, alginate) and synthetic materials (PGA,
PLGA, PLLA). However, silk fibroin as a natural biopolymer possesses outstanding characteristics with
biocompatibility, biodegradability, durability, and flexibility for regenerative medicine. Fiber biomatrices with
high porosity have interconnected pore networks, which provide anchoring sites to hold the cells together and
facilitate nutrient and oxygen diffusion and waste removal for efficient cell growth in long-term 3D culture. Our
recent study demonstrated that an in vitro 3D spheroid model of human urine-derived stem cells (USCs) can
be used for nephrotoxicity assays. To extend our ongoing study and bridge the gap between current 3D
spheroid cultures and MtT testing, we will explore a novel technology to provide large numbers of primary
human cells. Thus, the overall goal of this R03 study is to develop a strategy for large-scale production of
human USC in 3D culture suitable for late MtT testing. We hypothesize that an in vitro 3D culture system with
porous silk fiber matrix (SFM) will support adequate USCs (≥ 3 x 106/sample) for long-term growth (≥ 8 weeks),
with stable mitochondrial copy number and function, to eventually be used in late MtT testing. To test this
hypothesis, we propose these aims: Aim 1. Optimize experimental strategies for 3D culture system of silk fiber
network for large-scale production of human USCs; Aim 2. Validate a 3D culture system of USC-SFM and
compare it to 3D spheroid culture. This will be the first study to test a silk fiber network with human USCs for
MtT testing. We will use this newly developed 3D system in our existing study (3D Culture Systems of USCs
for NTRI-Induced Mitotoxicity; R21 AI152832). We expect that 3D culture of USC-SFM as a less labor-
intensive, more efficient, and cost-effective approach will be able to maintain large amounts of human stem
cells within silk fibers with stable mitochondrial quantiti...

## Key facts

- **NIH application ID:** 10326588
- **Project number:** 1R03AI165170-01
- **Recipient organization:** WAKE FOREST UNIVERSITY HEALTH SCIENCES
- **Principal Investigator:** YUANYUAN no ZHANG
- **Activity code:** R03 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $77,500
- **Award type:** 1
- **Project period:** 2021-06-03 → 2023-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10326588

## Citation

> US National Institutes of Health, RePORTER application 10326588, Silk Fibers-Assisted 3D System for Large-Scale Culture of Human Urine-Derived Stem Cells Suitable for Late Mitotoxicity Testing (1R03AI165170-01). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10326588. Licensed CC0.

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