# The role of TGF beta and SIX2 on SC-beta cell maturation

> **NIH NIH F31** · WASHINGTON UNIVERSITY · 2020 · $43,832

## Abstract

Abstract
The greatest challenge to improving human stem cell derived β (SC-β) cell technologies and in turn impeding
advancements in diabetes cell therapeutics and disease modeling is our lack of knowledge on human β cell
functional maturation. In my recently published first author publication I develop a protocol for the generation of
SC-β cells with insulin secretion dynamics approaching that of primary human islets and reveal that permitting
TGFβ signaling is important for SC-β cell functional maturation. In additional preliminary data, I have identified
that SIX2 is upregulated in TGFβ permitted cells and that SIX2 is necessary for SC-β cell function. In this
project I will investigate TGFβ signaling and SIX2 expression in the context of SC-β differentiation and
functional maturation. I hypothesize that TGFβ signaling via SIX2 transcriptional activity is critical for SC-β
cell development and functional maturation. An understanding of TGFβ signaling in β cell functional maturation
is not clear, with limited literature on the topic and the data being confounding and often contradictory with
some reports claiming TGFβ signaling benefits and other harm β cell function. This proposal will help clarify the
ambiguity associated with TGFβ and functional maturation. SIX2 is a recently identified β cell specific
transcription factor within the islet of Langerhans and is linked to diabetes by GWAS studies. The regulatory
role of SIX2 in pancreatic and β cell development and functional maturation has not been characterized. SIX2
is not expressed in mouse β cells making our SC-β cell differentiation model critical for investigating its role in
pancreatic and β cell differentiation and functional maturation. To investigate the role of TGFβ signaling in SC-
β cell functional maturation I will determine which SMADS and TGFβ ligands drive the functional maturation
observed as well as the downstream mechanism driving the functional maturation using a combination of
knockdown, overexpression, and RNA sequencing techniques. To investigate SIX2 action in SC-β cell
differentiation and functional maturation I will determine SIX2 expressing populations through the differentiation
using immunostaining and a SIX2 knockout human pluripotent stem cell line. I will investigate SIX2 role on SC-
β cell functional maturation using RNA sequencing techniques and robust functional characterization
employing calcium imaging, transmission electron microscopy, and insulin/proinsulin protein content assays.
This work will elucidate the role of TGFβ and SIX2 on SC-β cell functional maturation.
Aim 1. Investigate TGFβ signaling in SC-β cell functional maturation.
Aim 2. Characterize SIX2 in SC-β cell differentiation and functional maturation.

## Key facts

- **NIH application ID:** 10141873
- **Project number:** 1F31DK125068-01A1
- **Recipient organization:** WASHINGTON UNIVERSITY
- **Principal Investigator:** Leonardo Velazco-Cruz
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $43,832
- **Award type:** 1
- **Project period:** 2020-09-01 → 2021-08-06

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10141873

## Citation

> US National Institutes of Health, RePORTER application 10141873, The role of TGF beta and SIX2 on SC-beta cell maturation (1F31DK125068-01A1). Retrieved via AI Analytics 2026-06-03 from https://api.ai-analytics.org/grant/nih/10141873. Licensed CC0.

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